WorldWideScience

Sample records for broccoli coq5 methyltransferase

  1. Aggregates, broccoli and cauliflower

    Science.gov (United States)

    Grey, Francois; Kjems, Jørgen K.

    1989-09-01

    Naturally grown structures with fractal characters like broccoli and cauliflower are discussed and compared with DLA-type aggregates. It is suggested that the branching density can be used to characterize the growth process and an experimental method to determine this parameter is proposed.

  2. Evaluation of genotypic variation of broccoli (Brassica oleracea var. italic) in response to selenium treatment.

    Science.gov (United States)

    Ramos, Silvio J; Yuan, Youxi; Faquin, Valdemar; Guilherme, Luiz Roberto G; Li, Li

    2011-04-27

    Broccoli (Brassica oleracea var. italic) fortified with selenium (Se) has been promoted as a functional food. Here, we evaluated 38 broccoli accessions for their capacity to accumulate Se and for their responses to selenate treatment in terms of nutritional qualities and sulfur gene expresion. We found that the total Se content varied with over 2-fold difference among the leaf tissues of broccoli accessions when the plants were treated with 20 μM Na(2)SeO(4). Approximately half of total Se accumulated in leaves was Se-methylselenocysteine and selenomethionine. Transcriptional regulation of adenosine 5'-phosphosulfate sulfurylase and selenocysteine Se-methyltransferase gene expression might contribute to the different levels of Se accumulation in broccoli. Total glucosinolate contents were not affected by the concentration of selenate application for the majority of broccoli accessions. Essential micronutrients (i.e., Fe, Zn, Cu, and Mn) remained unchanged among half of the germplasm. Moreover, the total antioxidant capacity was greatly stimulated by selenate in over half of the accessions. The diverse genotypic variation in Se, glucosinolate, and antioxidant contents among accessions provides the opportunity to breed broccoli cultivars that simultaneously accumulate Se and other health benefit compounds.

  3. Effects of ca treatments and temperature on broccoli colour development

    OpenAIRE

    Schouten, R.E.; Zhang, X.; Tijskens, L.M.M.; Kooten, van, G.C.

    2010-01-01

    Broccoli combines high contents of vitamins, fibres and glucosinolates with a low calorie count and is sometimes referred to as the ‘crown jewel of nutrition’. Colour is one of the most important quality attributes of broccoli, and yellowing due to senescence of broccoli florets is the main external quality problem in the broccoli supply chain. Controlled Atmosphere (CA) is a very effective method to maintain broccoli quality but the effects of CA on colour retention have not been studied ext...

  4. Energy efficient drying strategies to retain nutritional components in broccoli broccoli (Brassica oleracea var. italica)

    NARCIS (Netherlands)

    Jin, X.; Sman, van der R.G.M.; Straten, van G.; Boom, R.M.; Boxtel, van A.J.B.

    2014-01-01

    This work concerns the combined optimization of the retention of bioactive components and energy efficiency during drying of broccoli. Kinetics for the degradation of glucosinolates, vitamin C and drying of broccoli are used to calculate optimal drying trajectories for the control variables air flow

  5. Nonadverse effects on allergenicity of isopentenyltransferase-transformed broccoli.

    Science.gov (United States)

    Liao, E C; Chen, J T; Chao, M L; Yu, S C; Chang, C Y; Chu, W S; Tsai, J J

    2013-01-01

    Genetically modified organisms (GMOs) provide modern agriculture with improvements in efficiency and the benefits of enhanced food production; however, the potential impact of GMOs on human health has not yet been clarified. To investigate the allergenicity of isopentenyltransferase (ipt)-transformed broccoli compared with non-GM broccoli. Sera from allergic individuals were used to identify the allergenicity of GM and non-GM broccoli. Immunoglobulin (Ig) binding of different lines of GM and non-GM broccoli was identified using immunoblotting, enzyme-linked immunosorbent assay, and the histamin release assay. Positive reactions to broccoli (Brassica Oleracea) were observed in 7.02% of individuals. Specific IgE to broccoli and total IgE fro allergic individuals were well correlated. The different tests performed showed no significant differences in the allergenicity of conventionally raised and GM broccoli, indicating the absence of unexpected effects on allergenicity in ipt-transformed plants. Using Western blot analysis we detected heterogeneous IgE-reactive allergenic components in broccoli-allergic sera, but no significant differences between GM an non-GM broccoli were observed in serum from the same patients. Our study demonstrates that there are no differences between GM (ipt-transformed) broccoli and non-GM broccoli, as determined by specific IgE in sera from broccoli-allergic patients. This indicates that there were no unexpected effects on allergenicity in this GM broccoli.

  6. Maximizing plant density affects broccoli yield and quality

    Science.gov (United States)

    Increased demand for fresh market bunch broccoli (Brassica oleracea L. var. italica) has led to increased production along the United States east coast. Maximizing broccoli yields is a primary concern for quickly expanding southeastern commercial markets. This broccoli plant density study was carr...

  7. Effects of ca treatments and temperature on broccoli colour development

    NARCIS (Netherlands)

    Schouten, R.E.; Zhang, X.; Tijskens, L.M.M.; Kooten, van O.

    2010-01-01

    Broccoli combines high contents of vitamins, fibres and glucosinolates with a low calorie count and is sometimes referred to as the ‘crown jewel of nutrition’. Colour is one of the most important quality attributes of broccoli, and yellowing due to senescence of broccoli florets is the main external

  8. Agronomy of strip intercropping broccoli with alyssum for biological control of aphids

    Science.gov (United States)

    Organic broccoli growers in California typically control aphids by intercropping broccoli with strips of alyssum (Lobularia maritima (L.) Desv.) which attracts hoverflies (Diptera: Syrphidae) that are important predators of aphids. A three year study with transplanted organic broccoli in Salinas, ...

  9. Machine vision for a selective broccoli harvesting robot

    NARCIS (Netherlands)

    Blok, Pieter M.; Barth, Ruud; Berg, Van Den Wim

    2016-01-01

    The selective hand-harvest of fresh market broccoli is labor-intensive and comprises about 35% of the total production costs. This research was conducted to determine whether machine vision can be used to detect broccoli heads, as a first step in the development of a fully autonomous selective

  10. Glucoraphanin and other glucosinolates in heads of broccoli cultivars

    Science.gov (United States)

    Broccoli (Brassica oleracea L. var. italica) emerged as an increasingly popular vegetable of North American consumers during the second half of the 20th Century, with per captita consumption increasing nearly eight fold during this period. Likewise, production and consumption of broccoli has also i...

  11. Influence of thiopurine methyltransferase gene polymorphism on ...

    Indian Academy of Sciences (India)

    Home; Journals; Journal of Genetics; Volume 96; Issue 6. Influence of thiopurine methyltransferase gene polymorphism on Egyptian children with acute lymphoblastic leukaemia. AZZA A. G. ... thiopurine methyltransferase gene polymorphism; acute lymphoblastic leukaemia; Egyptian children; thiopurine methyltransferase.

  12. Genetic regulation of glucoraphanin accumulation in Beneforté broccoli.

    Science.gov (United States)

    Traka, Maria H; Saha, Shikha; Huseby, Stine; Kopriva, Stanislav; Walley, Peter G; Barker, Guy C; Moore, Jonathan; Mero, Gene; van den Bosch, Frans; Constant, Howard; Kelly, Leo; Schepers, Hans; Boddupalli, Sekhar; Mithen, Richard F

    2013-06-01

    · Diets rich in broccoli (Brassica oleracea var italica) have been associated with maintenance of cardiovascular health and reduction in risk of cancer. These health benefits have been attributed to glucoraphanin that specifically accumulates in broccoli. The development of broccoli with enhanced concentrations of glucoraphanin may deliver greater health benefits. · Three high-glucoraphanin F1 broccoli hybrids were developed in independent programmes through genome introgression from the wild species Brassica villosa. Glucoraphanin and other metabolites were quantified in experimental field trials. Global SNP analyses quantified the differential extent of B. villosa introgression · The high-glucoraphanin broccoli hybrids contained 2.5-3 times the glucoraphanin content of standard hybrids due to enhanced sulphate assimilation and modifications in sulphur partitioning between sulphur-containing metabolites. All of the high-glucoraphanin hybrids possessed an introgressed B. villosa segment which contained a B. villosa Myb28 allele. Myb28 expression was increased in all of the high-glucoraphanin hybrids. Two high-glucoraphanin hybrids have been commercialised as Beneforté broccoli. · The study illustrates the translation of research on glucosinolate genetics from Arabidopsis to broccoli, the use of wild Brassica species to develop cultivars with potential consumer benefits, and the development of cultivars with contrasting concentrations of glucoraphanin for use in blinded human intervention studies. © 2013 The Authors. New Phytologist © 2013 New Phytologist Trust.

  13. Antioxidant effects of broccoli powder extract in goat meat nuggets.

    Science.gov (United States)

    Banerjee, Rituparna; Verma, Arun K; Das, Arun K; Rajkumar, V; Shewalkar, A A; Narkhede, H P

    2012-06-01

    The antioxidant potential of broccoli powder extract (BPE) was determined and evaluated in goat meat nuggets at three different levels 1, 1.5 and 2%, compared with control and butylated hydroxyl toluene (100ppm BHT). Total phenolics in 5mg broccoli powder was higher (Ppower of 10mg broccoli powder was comparable to the 100ppm BHT. Incorporation of 1.5 and 2% BPE decreased (Pgoat meat nuggets without affecting product acceptability. Copyright © 2012 Elsevier Ltd. All rights reserved.

  14. Histone methyltransferases in cancer

    DEFF Research Database (Denmark)

    Albert, Mareike; Helin, Kristian

    2009-01-01

    Cancer is perceived as a heterogeneous group of diseases that is characterized by aberrant patterns of gene expression. In the last decade, an increasing amount of data has pointed to a key role for epigenetic alterations in human cancer. In this review, we focus on a subclass of epigenetic...... regulators, namely histone methyltransferases (HMTs). Several HMTs have been linked to different types of cancer; however, in most cases we only have limited knowledge regarding the molecular mechanisms by which the HMTs contribute to disease development. We summarize the current knowledge regarding some...

  15. Antioxidant activity of fermented broccoli and spinach by Kombucha culture

    Science.gov (United States)

    Artanti, Nina; Susilowati, Agustine; Aspiyanto, Lotulung, Puspa Dewi Narrij; Maryati, Yati

    2017-11-01

    Broccoli (Brassica oleracea L.) and spinach (Amaranthus spp.) are vegetables that known to have many benefit for health. Previous studies on the fermentation of those vegetables using kombucha cultured showed increase in bioactive components such as total polyphenol content. The current studies was performed to evaluate the antioxidant activity of fermented spinach and broccoli before (feed) and after treatment with filtration (retentate and permeate). Filtration was conducted using Stirred Ultrafiltration Cell (SUFC) with UF membrane 100,000 MWCO mode at fixed condition (stirred rotation 300 rpm, room temperature, pressure 40 psia). Antioxidant evaluation was conducted using 2,2-diphenyl-1-picril hydrazyl (DPPH) free radical scavenging activity assay. The results showed that all samples from fermented broccoli showed antioxidant activity (feed 15.82% inhibition and retentate 15.29% inhibition), with the best antioxidant activity was obtained from permeate (75.98% inhibition). Whereas from fermented spinach only permeate showed antioxidant activity (21.84% inhibition) and it significantly lower than broccoli permeate. The mass spectrum of LCMS analysis on broccoli samples showed the present of several mass spectrum with (M+H) range from 148.1 to 442.5 in feed, retentate and permeate. In those samples (M+H) 360.4 always has the highest relative intensity. These results suggest that fermented broccoli has potential for development as functional drink for the source of antioxidant and the permeate obtained from filtration treatment significantly increased the antioxidant activity.

  16. In vivo formation and bioavailability of isothiocyanates from glucosinolates in broccoli as affected by processing conditions

    NARCIS (Netherlands)

    Oliviero, T.; Verkerk, R.; Vermeulen, M.; Dekker, M.

    2014-01-01

    Scope: To study the effect of residual myrosinase (MYR) activity in differently processed broccoli on sulforaphane (SR) and iberin (IB) formation, bioavailability, and excretion in human volunteers. Methods and results: Five different broccoli products were obtainedwith similar glucoraphanin (GR)

  17. Feeding tomato and broccoli powders enriched with bioactives improves bioactivity markers in rats.

    Science.gov (United States)

    Liu, Ann G; Volker, Sonja E; Jeffery, Elizabeth H; Erdman, John W

    2009-08-26

    Many studies have evaluated the cancer -preventive potential of individual bioactives from tomatoes and broccoli, but few have examined them within the context of a whole food. Male Copenhagen rats were fed diets containing 10% standard tomato powder, tomato enriched with lycopene or total carotenoids, standard broccoli floret, broccoli sprouts, or broccoli enriched with indole glucosinolates or selenium for 7 days. All broccoli diets increased the activity of colon quinone reductase (NQO1). Indole glucosinolate-enriched broccoli and selenium-enriched broccoli increased hepatic NQO1 and cytochrome P450 1A activity (P tomato diets down-regulated prostatic glutathione S-transferase P1 mRNA expression. Different tomato diets resulted in altered hepatic accumulation of lycopene, phytofluene, and phytoene. These results demonstrate that the bioactive content of vegetables affects both tissue content of bioactives and activity of detoxification enzymes. Enhancing bioactive content of tomatoes and broccoli may enhance efficacy in the prevention of prostate cancer.

  18. Production and postharvest quality maintenance of single unit and bunching broccoli in Virginia

    OpenAIRE

    Jett, Lewis W.

    1990-01-01

    Broccoli (Brassica oleracea L. var. italica) has become an increasingly popular vegetable with American consumers. Much of the attractiveness of fresh broccoli is derived from this vegetable's high nutrition and excellent organoleptic properties. In a consumer response survey, Virginia Master Gardeners indicated a preference for broccoli that has less stalk and more florets by weight. The objectives of this research were to produce single unit broccoli, and to examine vac...

  19. Development of colour of broccoli heads as affected by controlled atmosphere storage and temperature

    NARCIS (Netherlands)

    Schouten, R.E.; Zhang, X.; Verkerk, R.; Verschoor, J.A.; Otma, E.C.; Tijskens, L.M.M.; Kooten, van O.

    2009-01-01

    Colour is one of the most important quality attributes of broccoli. Yellowing due to senescence of broccoli florets is the main external quality problem. Controlled atmosphere (CA) storage is a very effective method to maintain broccoli quality. The aim of this paper is to characterise the colour

  20. Heterosis for horticultural traits in broccoli.

    Science.gov (United States)

    Hale, Anna L; Farnham, Mark W; Nzaramba, M Ndambe; Kimbeng, Collins A

    2007-08-01

    Over the last three decades, broccoli (Brassica oleracea L., Italica Group) hybrids made by crossing two inbred lines replaced open-pollinated populations to become the predominant type of cultivar. The change to hybrids evolved with little or no understanding of heterosis or hybrid vigor in this crop. Therefore, the purpose of the present study was to determine levels of heterosis expressed by a set of hybrids derived by crossing relatively elite, modern inbreds (n = 9). An additional objective was to determine if PCR-based marker derived genetic similarities among the parents can be useful to predict heterosis in this crop. Thirty-six hybrids derived from a diallel mating design involving nine parents were evaluated for five horticultural characters including the head characteristics of head weight, head stem diameter, and maturity (e.g., days from transplant to harvest), and the plant vigor characteristics of plant height, and plant width in four environments. A total of 409 polymorphic markers were generated by 24 AFLP, 23 SRAP and 17 SSR primer combinations. Euclidean distances between parents were determined based on phenotypic traits. About half of the hybrids exhibited highparent heterosis for head weight (1-30 g) and stem diameter (0.2-3.5 cm) when averaged across environments. Almost all hybrids showed highparent heterosis for plant height (1-10 cm) and width (2-13 cm). Unlike other traits, there was negative heterosis for maturity, indicating that heterosis for this character in hybrids is expressed as earliness. Genetic similarity estimates among the nine parental lines ranged from 0.43 to 0.71 and were significantly and negatively correlated with highparent heterosis for all traits except for stem diameter and days from transplant to harvest. Euclidean distances were not correlated with heterosis. With modern broccoli inbreds, less heterosis was observed for head characteristics than for traits that measured plant vigor. In addition, genetic similarity

  1. Elongation factor methyltransferase 3--a novel eukaryotic lysine methyltransferase.

    Science.gov (United States)

    Zhang, Lelin; Hamey, Joshua J; Hart-Smith, Gene; Erce, Melissa A; Wilkins, Marc R

    2014-08-22

    Here we describe the discovery of Saccharomycescerevisiae protein YJR129Cp as a new eukaryotic seven-beta-strand lysine methyltransferase. An immunoblotting screen of 21 putative methyltransferases showed a loss in the methylation of elongation factor 2 (EF2) on knockout of YJR129C. Mass spectrometric analysis of EF2 tryptic peptides localised this loss of methylation to lysine 509, in peptide LVEGLKR. In vitro methylation, using recombinant methyltransferases and purified EF2, validated YJR129Cp as responsible for methylation of lysine 509 and Efm2p as responsible for methylation at lysine 613. Contextualised on previously described protein structures, both sites of methylation were found at the interaction interface between EF2 and the 40S ribosomal subunit. In line with the recently discovered Efm1 and Efm2 we propose that YJR129C be named elongation factor methyltransferase 3 (Efm3). The human homolog of Efm3 is likely to be the putative methyltransferase FAM86A, according to sequence homology and multiple lines of literature evidence. Copyright © 2014 Elsevier Inc. All rights reserved.

  2. Sulforaphane, a Dietary Component of Broccoli/Broccoli Sprouts, Inhibits Breast Cancer Stem Cells

    Science.gov (United States)

    Li, Yanyan; Zhang, Tao; Korkaya, Hasan; Liu, Suling; Lee, Hsiu-Fang; Newman, Bryan; Yu, Yanke; Clouthier, Shawn G.; Schwartz, Steven J.; Wicha, Max S.; Sun, Duxin

    2010-01-01

    Purpose The existence of cancer stem cells (CSCs) in breast cancer has profound implications for cancer prevention. In this study, we evaluated sulforaphane, a natural compound derived from broccoli/broccoli sprouts, for its efficacy to inhibit breast CSCs and its potential mechanism. Experimental Design Aldefluor assay and mammosphere formation assay were used to evaluate the effect of sulforaphane on breast CSCs in vitro. A NOD/SCID xenograft model was employed to determine whether sulforaphane could target breast CSCs in vivo, as assessed by Aldefluor assay and tumor growth upon cell re-implantation in secondary mice. The potential mechanism was investigated utilizing Western blotting analysis and β-catenin reporter assay. Results Sulforaphane (1~5 μM) decreased aldehyde dehydrogenase (ALDH)-positive cell population by 65%~80% in human breast cancer cells (P sulforaphane for two weeks reduced ALDH-positive cells by more than 50% in NOD/SCID xenograft tumors (P = 0.003). Sulforaphane eliminated breast CSCs in vivo, thereby abrogating tumor growth after re-implantation of primary tumor cells into the secondary mice (P sulforaphane down-regulated Wnt/β-catenin self-renewal pathway. Conclusions Sulforaphane inhibits breast CSCs and down-regulates Wnt/β-catenin self-renewal pathway. These findings support the use of sulforaphane for chemoprevention of breast cancer stem cells and warrant further clinical evaluation. PMID:20388854

  3. Dietary Broccoli Alters Rat Cecal Microbiota to Improve Glucoraphanin Hydrolysis to Bioactive Isothiocyanates.

    Science.gov (United States)

    Liu, Xiaoji; Wang, Yanling; Hoeflinger, Jennifer L; Neme, Bárbara P; Jeffery, Elizabeth H; Miller, Michael J

    2017-03-10

    Broccoli consumption brings many health benefits, including reducing the risk of cancer and inflammatory diseases. The objectives of this study were to identify global alterations in the cecal microbiota composition using 16S rRNA sequencing analysis and glucoraphanin (GRP) hydrolysis to isothiocyanates ex vivo by the cecal microbiota, following different broccoli diets. Rats were randomized to consume AIN93G (control) or different broccoli diets; AIN93G plus cooked broccoli, a GRP-rich powder, raw broccoli, or myrosinase-treated cooked broccoli. Feeding raw or cooked broccoli for four days or longer both changed the cecal microbiota composition and caused a greater production of isothiocyanates ex vivo. A more than two-fold increase in NAD(P)H: quinone oxidoreductase 1 activity of the host colon mucosa after feeding cooked broccoli for seven days confirmed the positive health benefits. Further studies revealed that dietary GRP was specifically responsible for the increased microbial GRP hydrolysis ex vivo, whereas changes in the cecal microbial communities were attributed to other broccoli components. Interestingly, a three-day withdrawal from a raw broccoli diet reversed the increased microbial GRP hydrolysis ex vivo. Findings suggest that enhanced conversion of GRP to bioactive isothiocyanates by the cecal microbiota requires four or more days of broccoli consumption and is reversible.

  4. Effect of industrial freezing on the stability of chemopreventive compounds in broccoli.

    Science.gov (United States)

    Alanís-Garza, Pedro A; Becerra-Moreno, Alejandro; Mora-Nieves, José Luis; Mora-Mora, Juan Pablo; Jacobo-Velázquez, Daniel A

    2015-05-01

    Broccoli (Brassica oleracea L. var. Italica) is largely consumed all over the world and has a high economic importance. Likewise, broccoli contains high levels of glucosinolates, carotenoids and total phenols, which are related with the prevention of chronic diseases. The present project's objective was to evaluate the effect of industrial freezing on the stability of bioactive molecules in seven commercial broccoli cultivars (Tlaloc®, Endurance®, Florapack®, Domador®, Steel®, Iron Man® and Avenger®). In general, industrial freezing increased the extractability of total glucosinolates, whereas total phenols remained constant in most broccoli cultivars. Likewise, broccoli subjected to industrial freezing showed higher levels of total carotenoids (∼60-300% higher) as compared with fresh broccoli. Results suggest that bioactive compounds in frozen broccoli would be more bioavailable than in raw.

  5. The Broccoli Syndrome: Higher Education's Pubdown Of The Minority Student

    Science.gov (United States)

    Tate, Penfield; Delworth, Ursula

    1973-01-01

    The assumption underlying the broccoli syndrome is that minority group students are not familiar with a whole range of Anglo goodies''. This article points out how, as the minority student is enlightened time and again to such facts'', he perceives inherent racism in his enlightener.'' (JC)

  6. Molecular and Biochemical Characterization of Postharvest Senescence in Broccoli

    Science.gov (United States)

    Page, Tania; Griffiths, Gareth; Buchanan-Wollaston, Vicky

    2001-01-01

    Postharvest senescence in broccoli (Brassica oleracea L. var Italica) florets results in phenotypic changes similar to those seen in developmental leaf senescence. To compare these two processes in more detail, we investigated molecular and biochemical changes in broccoli florets stored at two different temperatures after harvest. We found that storage at cooler temperatures delayed the symptoms of senescence at both the biochemical and gene expression levels. Changes in key biochemical components (lipids, protein, and chlorophyll) and in gene expression patterns occurred in the harvested tissue well before any visible signs of senescence were detected. Using previously identified senescence-enhanced genes and also newly isolated, differentially expressed genes, we found that the majority of these showed a similar enhancement of expression in postharvest broccoli as in developmental leaf senescence. At the biochemical level, a rapid loss of membrane fatty acids was detected after harvest, when stored at room temperature. However, there was no corresponding increase in levels of lipid peroxidation products. This, together with an increased expression of protective antioxidant genes, indicated that, in the initial stages of postharvest senescence, an orderly dismantling of the cellular constituents occurs, using the available lipid as an energy source. Postharvest changes in broccoli florets, therefore, show many similarities to the processes of developmental leaf senescence. PMID:11161029

  7. A transdisciplinary approach to developing an eastern broccoli industry

    Science.gov (United States)

    Current economic, social and scientific situations have made it attractive to establish a substantial and profitable broccoli industry in the eastern US, where several obstacles have prevented that from happening previously. Because many barriers must be overcome simultaneously, we assembled a tran...

  8. Yield, irrigation production efficiency and economic return of broccoli ...

    African Journals Online (AJOL)

    ... effect of irrigation methods and irrigation schedules on marketable yield, irrigation oroduction efficiency and economic return of Broccoli (Brassica oleracea. Var. italica). lrrigation was applied when the sum of daily pan evaporation data from United States Weather Bureau (USWB) class- A-open ran reach approximately to ...

  9. Plant regeneration of Brassica oleracea subsp. italica (Broccoli) CV ...

    African Journals Online (AJOL)

    STORAGESEVER

    2009-06-03

    Jun 3, 2009 ... Hypocotyl and shoot tip explants, 5-8 mm in size, were excised from 6-day-old broccoli seedlings. The explants were cultured on. MS medium incorporated with different concentrations of plant growth regulators for shoot proliferation and root formation. For shoot induction and multiplication from hypocotyl ...

  10. The Effect of Steaming on the Glucosinolate Content in Broccoli

    NARCIS (Netherlands)

    Verkerk, R.; Knol, J.J.; Dekker, M.

    2010-01-01

    Total and individual glucosinolates were measured after different duration of steaming broccoli (Brassica oleracea L. var. italica). During steaming, the temperature profile, cell lysis and inactivation of myrosinase were assessed as well. Steaming resulted in high retention of total aliphatic and

  11. Optimization of a plant regeneration protocol for broccoli | Huang ...

    African Journals Online (AJOL)

    The factors which influence the regeneration of broccoli (Brassica oleracea var. italica) were studied using an orthogonal design. The results showed that the major factor was the explant type, followed by naphthylacetic acid (NAA), benzylaminopurine (BAP), sucrose and AgNO3 in turn. The maximum regeneration was on ...

  12. No de novo sulforaphane biosynthesis in broccoli seedlings

    NARCIS (Netherlands)

    Gorissen, Antonie; Kraut, Nicolai U.; de Visser, Ries; de Vries, Marcel; Roelofsen, Han; Vonk, Roel J.

    2011-01-01

    The isothiocyanate sulforaphane, present in significant amounts in broccoli (Brassica oleracea L.) seedlings in the form of its precursor glucoraphanin, has been identified as an inducer of quinine reductase, a phase-II detoxification enzyme known for its anticarcinogenic properties. Its

  13. Molecular and Functional Characterization of Broccoli EMBRYONIC FLOWER 2 Genes

    Science.gov (United States)

    Chen, Long-Fang O.; Lin, Chun-Hung; Lai, Ying-Mi; Huang, Jia-Yuan; Sung, Zinmay Renee

    2012-01-01

    Polycomb group (PcG) proteins regulate major developmental processes in Arabidopsis. EMBRYONIC FLOWER 2 (EMF2), the VEFS domain-containing PcG gene, regulates diverse genetic pathways and is required for vegetative development and plant survival. Despite widespread EMF2-like sequences in plants, little is known about their function other than in Arabidopsis and rice. To study the role of EMF2 in broccoli (Brassica oleracea var. italica cv. Elegance) development, we identified two broccoli EMF2 (BoEMF2) genes with sequence homology to and a similar gene expression pattern to that in Arabidopsis (AtEMF2). Reducing their expression in broccoli resulted in aberrant phenotypes and gene expression patterns. BoEMF2 regulates genes involved in diverse developmental and stress programs similar to AtEMF2 in Arabidopsis. However, BoEMF2 differs from AtEMF2 in the regulation of flower organ identity, cell proliferation and elongation, and death-related genes, which may explain the distinct phenotypes. The expression of BoEMF2.1 in the Arabidopsis emf2 mutant (Rescued emf2) partially rescued the mutant phenotype and restored the gene expression pattern to that of the wild type. Many EMF2-mediated molecular and developmental functions are conserved in broccoli and Arabidopsis. Furthermore, the restored gene expression pattern in Rescued emf2 provides insights into the molecular basis of PcG-mediated growth and development. PMID:22537758

  14. Enhancement of broccoli indole glucosinolates by methyl jasmonate treatment and effects on prostate carcinogenesis.

    Science.gov (United States)

    Liu, Ann G; Juvik, John A; Jeffery, Elizabeth H; Berman-Booty, Lisa D; Clinton, Steven K; Erdman, John W

    2014-11-01

    Broccoli is rich in bioactive components, such as sulforaphane and indole-3-carbinol, which may impact cancer risk. The glucosinolate profile of broccoli can be manipulated through treatment with the plant stress hormone methyl jasmonate (MeJA). Our objective was to produce broccoli with enhanced levels of indole glucosinolates and determine its impact on prostate carcinogenesis. Brassica oleracea var. Green Magic was treated with a 250 μM MeJA solution 4 days prior to harvest. MeJA-treated broccoli had significantly increased levels of glucobrassicin, neoglucobrassicin, and gluconasturtiin (P broccoli powder, or 10% MeJA broccoli powder. Diets were fed throughout the study until termination at 20 weeks of age. Hepatic CYP1A was induced with MeJA broccoli powder feeding, indicating biological activity of the indole glucosinolates. Following ∼ 15 weeks on diets, neither of the broccoli treatments significantly altered genitourinary tract weight, pathologic score, or metastasis incidence, indicating that broccoli powder at 10% of the diet was ineffective at reducing prostate carcinogenesis in the TRAMP model. Whereas broccoli powder feeding had no effect in this model of prostate cancer, our work demonstrates the feasibility of employing plant stress hormones exogenously to stimulate changes in phytochemical profiles, an approach that may be useful for optimizing bioactive component patterns in foods for chronic-disease-prevention studies.

  15. Inhibition of Bladder Cancer by Broccoli Isothiocyanates Sulforaphane and Erucin: Characterization, Metabolism and Interconversion

    OpenAIRE

    Abbaoui, Besma; Riedl, Kenneth M.; Ralston, Robin A.; Thomas-Ahner, Jennifer M.; Schwartz, Steven J.; Clinton, Steven K.; Mortazavi, Amir

    2012-01-01

    Epidemiologic evidence suggests diets rich in cruciferous vegetables, particularly broccoli, are associated with lower bladder cancer risk. Our objectives are to investigate these observations and determine the role of isothiocyanates in primary or secondary bladder cancer prevention. We initially investigate the mechanisms whereby broccoli and broccoli sprout extracts and pure isothiocyanates inhibit normal, non-invasive (RT4) and invasive (J82, UMUC3) human urothelial cell viability. Sulfor...

  16. THE INFLUENCE OF NUTRITION ON THE MAIN PRODUCTION OF BROCCOLI

    Directory of Open Access Journals (Sweden)

    Mădălina Doltu

    2013-07-01

    Full Text Available The research was realized in the unheated greenhouse of Horting Institute Bucharest, in 2012 year. It have aimed the influence of one phase of fertilization with ammonium nitrate (NH4NO3 on inflorescences of broccoli, at a hybrid F1 - Ironman. The fertilizer was used before the formation of main inflorescences, in different doses (100 kg/ha, 150 kg/ha, without fertilizer. Harvesting was done in 4 stages. A phase of fertilization with ammonium nitrate has influenced the main production of broccoli: the variant with fertilization 150 kg/ha has obtained the highest percentage of inflorescences formed, the average weight/inflorescence largest and superior production compared with other variants. Were obtained direct linear correlations between the doses of ammonium nitrate (NH4NO3 and the production aspects followed (percentage of main inflorescences harvested, average weight/inflorescence, total production, significances of the correlations very high.

  17. Polymer coating, germination and vigor of broccoli seeds

    OpenAIRE

    Almeida Celina de; Rocha Sandra Cristina dos Santos; Razera Luiz Fernandes

    2005-01-01

    Brassica oleracea var italica occupies a special place in the internal and external market of vegetables seeds. Vegetables producers demand seeds with high degree of purity, germination and vigor, since seeds' quality is the basis for the success of the production. In this work, broccoli seeds were coated in a spouted bed, by an aqueous suspension of hidroxy-ethyl-cellulose. Effects of the operating variables: spouting air temperature, atomizing air pressure and coating suspension flow rate o...

  18. Remote sensing of water and nitrogen stress in broccoli

    Science.gov (United States)

    Elsheikha, Diael-Deen Mohamed

    Remote sensing is being used in agriculture for crop management. Ground based remote sensing data acquisition system was used for collection of high spatial and temporal resolution data for irrigated broccoli crop. The system was composed of a small cart that ran back and forth on a rail system that was mounted on a linear move irrigation system. The cart was equipped with a sensor that had 4 discrete wavelengths; 550 nm, 660 nm, 720 nm, and 810 nm, and an infrared thermometer, all had 10 nm bandwidth. A global positioning system was used to indicate the cart position. The study consisted of two parts; the first was to evaluate remotely sensed reflectance and indices in broccoli during the growing season, and determine whether remotely sensed indices or standard deviation of indices can distinguish between nitrogen and water stress in broccoli, and the second part of the study was to evaluate remotely sensed indices and standard deviation of remotely sensed indices in broccoli during daily changes in solar zenith angle. Results indicated that nitrogen was detected using Ratio Vegetation index, RVI, Normalized Difference Vegetation Index, NDVI, Canopy Chlorophyll Concentration Index, CCCI, and also using the reflectance in the Near-Infrared, NIR, bands. The Red reflectance band capability of showing stress was not as clear as the previous indices and bands reflectance. The Canopy Chlorophyll Concentration Index, CCCI, was the most successful index. The Crop Water Stress Index was able to detect water stress but it was highly affected by the solar zenith angle change along the day.

  19. Isothiocyanate concentrations and interconversion of sulforaphane to erucin in human subjects after consumption of commercial frozen broccoli compared to fresh broccoli.

    Science.gov (United States)

    Saha, Shikha; Hollands, Wendy; Teucher, Birgit; Needs, Paul W; Narbad, Arjan; Ortori, Catharine A; Barrett, David A; Rossiter, John T; Mithen, Richard F; Kroon, Paul A

    2012-12-01

    Sulforaphane (a potent anticarcinogenic isothiocyanate derived from glucoraphanin) is widely considered responsible for the protective effects of broccoli consumption. Broccoli is typically purchased fresh or frozen and cooked before consumption. We compared the bioavailability and metabolism of sulforaphane from portions of lightly cooked fresh or frozen broccoli, and investigated the bioconversion of sulforaphane to erucin. Eighteen healthy volunteers consumed broccoli soups produced from fresh or frozen broccoli florets that had been lightly cooked and sulforaphane thio-conjugates quantified in plasma and urine. Sulforaphane bioavailability was about tenfold higher for the soups made from fresh compared to frozen broccoli, and the reduction was shown to be due to destruction of myrosinase activity by the commercial blanching-freezing process. Sulforaphane appeared in plasma and urine in its free form and as several thio-conjugates forms. Erucin N-acetyl-cysteine conjugate was a significant urinary metabolite, and it was shown that human gut microflora can produce sulforaphane, erucin, and their nitriles from glucoraphanin. The short period of blanching used to produce commercial frozen broccoli destroys myrosinase and substantially reduces sulforaphane bioavailability. Sulforaphane was converted to erucin and excreted in urine, and it was shown that human colonic flora were capable of this conversion. © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  20. Production of phytotherapeutics from broccoli juice by integrated membrane processes.

    Science.gov (United States)

    Yilmaz, Emel; Bagci, Pelin Onsekizoglu

    2018-03-01

    An integrated membrane process for the recovery and concentration of phytotherapeutics in broccoli juice was investigated to develop a natural product that could be of interest for food and/or pharmaceutical industry. Following a pretreatment step to remove suspended solids, the juice was ultrafiltered through a 50kDa MWCO polyethersulfone membrane. The permeate stream was further concentrated by osmotic distillation up to 42.6°Brix. During ultrafiltration, a complete recovery of sulforaphane, malic acid and citric acid was achieved, where total phenolic content and ascorbic acid were recovered at a ratio of 94.5 and 92.4%, respectively. All the bioactive compounds identified in broccoli juice was well preserved during subsequent 6-fold concentration by osmotic distillation. No significant difference was observed in total antioxidant activity of the juice throughout the operation. The proposed integrated membrane process has shown promising potential to produce a natural concentrate enriched in phytotherapeutics of the broccoli juice. Copyright © 2017 Elsevier Ltd. All rights reserved.

  1. Plant characters of broccoli determinants of head production

    Directory of Open Access Journals (Sweden)

    Fernanda Daniela Brandelero

    2016-06-01

    Full Text Available ABSTRACT: The increasing consumption of single-head broccoli is due to several factors, among them there are food production in minimally processing form and the existence of hybrids that adapt to various climates, in addition to the simple harvesting of this typical architecture.This study aimed to identify the most relevant plant characters of broccoli, represented by growth characters, which are determinant in the production and canopy area. The study was conducted in an experimental area in Pato Branco city, PR. The 11 characters were evaluated for 365 plants, spaced with 0.8x0.5m, on a blank experiment. The characters of group 1 (height, number of leaves, stem height, stem diameter were evaluated on the 21 and 58 day after transplanting (DAT, and the leaf area was evaluated on the 17 and 32 DAT. The characters of group 2 were quantity of fresh head mass and canopy area. At the initial stage of cultivation, on the 17 and 21 DAT, variations in the plants characters did not lead toany variation in production. The higher number of leaves and the larger stem diameter on the 58 DAT determined the greater mass of the broccoli heads.

  2. Characteristics Defining Broccoli Cultivars from Different Seed Producers

    Directory of Open Access Journals (Sweden)

    Elena Ordiales

    2017-01-01

    Full Text Available Broccoli (Brassica oleracea L. var. italica is currently considered a main vegetable food in the markets due to its high nutritional value, containing elevated levels of phytochemicals widely described to have beneficial effects against cancer and other illnesses. Broccoli is an interesting crop with a high commercial value because it complements the tomato industry, the main agricultural income in Badajoz, Spain. Nine varieties of broccoli from four nurseries were evaluated by analyzing both agronomic and quality parameters. Total yield and number of harvests were monitored. Parameters defining quality like diameter, weight, and height of the heads were determined. Granulometry, compactness, and the presence of internal leaves in the heads were also analyzed. Diameter and height of sprouts were complementarily estimated. Principal component analysis was further employed to investigate the relationship between the agronomic variables and the cultivars and nurseries. Results revealed that both first and second principal components explained more than 75% of the variance and grouped data according their cultivar and commercial origin. Additionally, correlations between the scores of those components and the values of the phenotypic parameters suggested that head weights are main determinants of the phenotypic differences observed among the cultivars whereas the presence of internal leaves and granulometry and head weight appear to be key traits defining nurseries.

  3. Antioxidant capacity of broccoli sprouts subjected to gastrointestinal digestion.

    Science.gov (United States)

    Rychlik, Joanna; Olejnik, Anna; Olkowicz, Mariola; Kowalska, Katarzyna; Juzwa, Wojciech; Myszka, Kamila; Dembczyński, Radosław; Moyer, Mary Pat; Grajek, Włodzimierz

    2015-07-01

    Broccoli is a common vegetable recognized as a rich source of antioxidants. To date, research on the antioxidant properties of broccoli, predominantly conducted on extracts, has not considered the lesions of composition and this activity after gastrointestinal digestion. Here the stability of antioxidants during gastrointestinal digestion was evaluated in conjunction with the protective effects of broccoli sprouts (BS) against oxidative stress in human colon cells. The obtained data suggest that, among the biocompounds identified in BS, glucosinolates were mainly degraded under gastrointestinal digestion, while phenolics, particularly hydroxycinnamic acid derivatives, were the most resistant constituents. The antioxidant capacity of BS extract subjected to gastrointestinal digestion was similar to or higher than that determined for non-digested BS. Gastrointestinal digested BS extract exhibited reactive oxygen species (ROS)-inhibitory capacity in NCM460 human colon cells, with 1 mg mL(-1) showing an ROS clearance of 76.59%. A 57.33% reduction in oxidative DNA damage in NCM460 cells due to treatment with digested BS extract was observed. The results lend support to the possible application of BS as a rich source of antioxidants to improve the defensive system against oxidative stress in the human colon mucosa. © 2014 Society of Chemical Industry.

  4. Modifying the processing and handling of frozen broccoli for increased sulforaphane formation.

    Science.gov (United States)

    Dosz, Edward B; Jeffery, Elizabeth H

    2013-09-01

    Frozen broccoli can provide a cheaper product, with a longer shelf life and less preparation time than fresh broccoli. We previously showed that several commercially available frozen broccoli products do not retain the ability to generate the cancer-preventative agent sulforaphane. We hypothesized that this was because the necessary hydrolyzing enzyme myrosinase was destroyed during blanching, as part of the processing that frozen broccoli undergoes. This study was carried out to determine a way to overcome loss of hydrolyzing activity. Industrial blanching usually aims to inactivate peroxidase, although lipoxygenase plays a greater role in product degradation during frozen storage of broccoli. Blanching at 86 °C or higher inactivated peroxidase, lipoxygenase, and myrosinase. Blanching at 76 °C inactivated 92% of lipoxygenase activity, whereas there was only an 18% loss in myrosinase-dependent sulforaphane formation. We considered that thawing frozen broccoli might disrupt membrane integrity, allowing myrosinase and glucoraphanin to come into contact. Thawing frozen broccoli for 9 h did not support sulforaphane formation unless an exogenous source of myrosinase was added. Thermal stability studies showed that broccoli root, as a source of myrosinase, was not more heat stable than broccoli floret. Daikon radish root supported some sulforaphane formation even when heated at 125 °C for 10 min, a time and temperature comparable to or greater than microwave cooking. Daikon radish (0.25%) added to frozen broccoli that was then allowed to thaw supported sulforaphane formation without any visual alteration to that of untreated broccoli. © 2013 Institute of Food Technologists®

  5. Quantitative trait loci mapping of heat tolerance in a doubled haploid population of broccoli using genotyping-by-sequencing

    Science.gov (United States)

    Broccoli is a cool weather vegetable crop with a vernalization requirement to initiate and maintain floral development. Breeding for heat tolerance in broccoli has the potential to both expand viable production areas and extend the growing season. A doubled haploid (DH) population of broccoli (Bras...

  6. Roles of DNA methyltransferases in Arabidopsis development

    African Journals Online (AJOL)

    Jane

    2010-12-13

    Dec 13, 2010 ... RNA-dependent tRNA polymerase; ROS1, repressor of silencing 1; SET, histone methyltransferase; SDC, suppressor of drm1/2 cmt3; siRNA, small interfering RNA; SUP, superman;. SUVH4, su(var)3-9 homolog 4; TRDMT1, tRNA aspartic acid methyltransferase 1; TGS, transcriptional gene silencing; UBA,.

  7. Roles of DNA methyltransferases in Arabidopsis development ...

    African Journals Online (AJOL)

    Mutations that cause severe loss of DNA methylation often leads to abnormal development. In the present review, we summarized recent findings of the three major DNA methyltransferases mutants playing vital role in development of Arabidopsis thaliana. Keywords: DNA methylation, epigenetics, methyltransferase, mutant ...

  8. The effect of quickly fermented pig manure on the broccoli yield parameters and selected soil parameters

    Directory of Open Access Journals (Sweden)

    Peter Kováčik

    2008-01-01

    Full Text Available The effect of Difert (a pig manure fermented by caddices of domestic flies produced on sawdust litter on broccoli yield parameters and selected soil parameters were investigated on gleic fluvisol in a small-plot field trial localized at area of Slovak University of Agriculture in Nitra (48°18´ N, 18°05´ E.The results showed that Difert applied in doses of 250 kg N . ha−1 and 350 kg N . ha−1 acted positively on the yields of fresh primary broccoli roses. However, the results are not statistically significant. Also a positive effect on N, P, K, Ca, Mg contents in broccoli roses was recorded. Difert has a moderate alkalizing effect on soil and increases the content of Cox in the soil. Moreover Difert insignificantly decreased the content of vitamin C and significantly increased the content of nitrates in broccoli roses, but the worst qualitative effect on broccoli parameters was detected by applying mineral N fertilizers, which significantly increased the content of nitrates in broccoli roses and insignificantly decreased the vitamin C content. However, it resulted in the highest broccoli yields. The application of mineral N fertilizers had a negative effect on the total content of carbon in the soil as well.The effect of Difert on broccoli yield and soil parameters refers to the feasibility of reducing the maturing period of the manure from 6 months to 1 week, in order to decrease the manure storage capacities.

  9. Ozonated water and chlorine effects on the antioxidant properties of organic and conventional broccoli during postharvest

    Directory of Open Access Journals (Sweden)

    Giuseppina Pace Pereira Lima

    2014-04-01

    Full Text Available There is growing interest in studies on sanitizers other than chlorine that can maintain the quality of organic products without affecting their phytochemical content. The effects of using chlorinated and ozonized water treatments, as sanitizing procedures, on the post-harvest quality of organic and conventional broccoli (Brassica oleracea L. cv. Italica was evaluated. The biochemical parameters (chlorophyll, polyphenols, flavonoids, vitamin C and antioxidant capacity of the broccoli samples were analyzed at day 0 (arrival of the plant from the field, original features, and 1, 4 and 7 days after harvest. The polyamine analysis was performed on arrival of the plant from the field and on the first and seventh days. The cultivation procedure influenced polyphenol, vitamin C and total chlorophyll content, and the highest value was observed in organic broccoli after the fourth day. Flavenoid content was higher in organic broccoli. The use of ozone appears not to have had an influence on the amount of polyphenolic, flavonoids and vitamin C during storage. Total chlorophyll content was less affected by ozonized water than by the chlorine treatment as at the first and fourth days of storage. The highest content of putrescine was found in conventional broccoli, while the highest levels of spermidine and spermine were found in organic broccoli. Antioxidant capacity was highest in organic broccoli after day 4 of storage and was affected by the bioactive compounds analyzed. Methods of cultivation influenced natural antioxidant and chlorophyll contents in broccoli under cold storage.

  10. Controlling rheology and structure of sweet potato starch noodles with high broccoli powder content by hydrocolloids

    NARCIS (Netherlands)

    Silva, E.; Birkenhake, M.; Scholten, E.; Sagis, L.M.C.; Linden, van der E.

    2013-01-01

    Incorporating high volume fractions of broccoli powder in starch noodle dough has a major effect on its shear modulus, as a result of significant swelling of the broccoli particles. Several hydrocolloids with distinct water binding capacity (locust bean gum (LBG), guar gum, konjac glucomannan (KG),

  11. Bioavailability and kinetics of sulforaphane in humans after consumption of cooked versus raw broccoli

    NARCIS (Netherlands)

    Vermeulen, M.; Klöpping-Ketelaars, I.W.A.A.; Berg, R. van den; Vaes, W.H.J.

    2008-01-01

    The aim of this study was to determine the bioavailability and kinetics of the supposed anticarcinogen sulforaphane, the hydrolysis product of glucoraphanin, from raw and cooked broccoli. Eight men consumed 200 g of crushed broccoli, raw or cooked, with a warm meal in a randomized, free-living, open

  12. Increasing plant density in eastern United States broccoli production systems to maximize marketable head yields

    Science.gov (United States)

    Increased demand for fresh market broccoli (Brassica oleracea L. var. italica) has led to increased production along the eastern seaboard of the United States. Maximizing broccoli yields is a primary concern for quickly expanding eastern commercial markets. Thus, a plant density study was carried ...

  13. Effects of Hot Water Immersion on Storage Quality of Fresh Broccoli Heads

    Directory of Open Access Journals (Sweden)

    Huaqiang Dong

    2004-01-01

    Full Text Available Freshly harvested broccoli heads were immersed for 0, 1, 4 or 8 min into hot water at 45 °C, and then were hydrocooled rapidly for 10 min at 10 °C. Following these treatments, the broccoli were air-dried for 30 min, then packed in commercial polymeric film bags, and, finally, stored for 16 days at –1, 1, and 12 °C. The samples treated with hot water maintained high contents of chlorophyll concentrations, their yellowing rate was delayed, and fungal infection and chilling or freezing injury were inhibited markedly. Compared to non-heat-treated broccoli, a lower level of peroxidase activity with a relatively higher chlorophyll concentration was observed when broccoli were treated with hot water. Among these heat treatments, immersion in hot water for 4 min at 45 °C was the most effective for maintaining the quality of harvested broccoli heads.

  14. DNA damage and repair activity after broccoli intake in young healthy smokers

    DEFF Research Database (Denmark)

    Riso, Patrizia; Martini, Daniela; Møller, Peter

    2010-01-01

    compounds, including smokers. The aim of the study was to evaluate the effect of broccoli intake on biomarkers of DNA damage and repair. Twenty-seven young healthy smokers consumed a portion of steamed broccoli (250 g/day) or a control diet for 10 days each within a crossover design with a washout period...... mRNA expression levels of repair and defence enzymes: 8-oxoguanine DNA glycosylase (OGG1), nucleoside diphosphate linked moiety X-type motif 1 (NUDT1) and heme oxygenase 1 (HO-1). After broccoli consumption, the level of oxidised DNA lesions decreased by 41% (95% confidence interval: 10%, 72......%) and the resistance to H(2)O(2)-induced DNA strand breaks increased by 23% (95% CI: 13%, 34%). Following broccoli intake, a higher protection was observed in subjects with glutathione S-transferase (GST) M1-null genotype. The expression level and activity of repair enzymes was unaltered. In conclusion, broccoli...

  15. Diet rich in high glucoraphanin broccoli reduces plasma LDL cholesterol: Evidence from randomised controlled trials.

    Science.gov (United States)

    Armah, Charlotte N; Derdemezis, Christos; Traka, Maria H; Dainty, Jack R; Doleman, Joanne F; Saha, Shikha; Leung, Wing; Potter, John F; Lovegrove, Julie A; Mithen, Richard F

    2015-05-01

    Cruciferous-rich diets have been associated with reduction in plasma LDL-cholesterol (LDL-C), which may be due to the action of isothiocyanates derived from glucosinolates that accumulate in these vegetables. This study tests the hypothesis that a diet rich in high glucoraphanin (HG) broccoli will reduce plasma LDL-C. One hundred and thirty volunteers were recruited to two independent double-blind, randomly allocated parallel dietary intervention studies, and were assigned to consume either 400 g standard broccoli or 400 g HG broccoli per week for 12 weeks. Plasma lipids were quantified before and after the intervention. In study 1 (37 volunteers), the HG broccoli diet reduced plasma LDL-C by 7.1% (95% CI: -1.8%, -12.3%, p = 0.011), whereas standard broccoli reduced LDL-C by 1.8% (95% CI +3.9%, -7.5%, ns). In study 2 (93 volunteers), the HG broccoli diet resulted in a reduction of 5.1% (95% CI: -2.1%, -8.1%, p = 0.001), whereas standard broccoli reduced LDL-C by 2.5% (95% CI: +0.8%, -5.7%, ns). When data from the two studies were combined the reduction in LDL-C by the HG broccoli was significantly greater than standard broccoli (p = 0.031). Evidence from two independent human studies indicates that consumption of high glucoraphanin broccoli significantly reduces plasma LDL-C. © 2015 The Authors. Molecular Nutrition & Food Research published by Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  16. Supplementation of broccoli or Bifidobacterium longum-fermented broccoli suppresses serum lipid peroxidation and osteoclast differentiation on alveolar bone surface in rats fed a high-cholesterol diet.

    Science.gov (United States)

    Tomofuji, Takaaki; Ekuni, Daisuke; Azuma, Tetsuji; Irie, Koichiro; Endo, Yasumasa; Yamamoto, Tatsuo; Ishikado, Atsushi; Sato, Takehiko; Harada, Kayo; Suido, Hirohisa; Morita, Manabu

    2012-04-01

    High-cholesterol diet enhances osteoclastic activity on alveolar bone by increasing serum lipid peroxidation. We hypothesized that supplementation with dietary antioxidants, such as found in broccoli and its fermented products, might suppress increases in serum lipid peroxidation, contributing to the inhibition of osteoclastic activity after high-cholesterol diet intake. The purpose of the present study was to investigate the effects of broccoli and fermented broccoli consumption on serum lipid peroxidation and osteoclast differentiation in alveolar bone of rats fed a high-cholesterol diet. In this 12-week study, rats were divided into 4 groups (n = 6/group): a control group (fed regular diet) and 3 experimental groups (fed a high-cholesterol [1% wt/wt] diet, or a high-cholesterol diet supplemented with either broccoli powder [5% wt/wt] or Bifidobacterium longum-fermented broccoli powder [5% wt/wt]). Serum hexanoyl-lysine (HEL) levels were measured as a parameter of lipid peroxidation. The number of tartrate-resistant acid phosphatase (TRAP)-positive osteoclasts in alveolar bone was enumerated to evaluate osteoclast differentiation. When compared with regular diet, the high-cholesterol diet increased serum HEL levels and resulted in a higher number of TRAP-positive osteoclasts at 12 weeks. The high-cholesterol diet supplemented with broccoli or B. longum-fermented broccoli showed lower levels of serum HEL and fewer TRAP-positive osteoclasts than the high-cholesterol diet at 12 weeks. In conclusion, consumption of broccoli, or its fermented product, inhibited the effects of a high-cholesterol diet on osteoclast differentiation in rat alveolar bone by suppressing serum lipid peroxidation. Copyright © 2012 Elsevier Inc. All rights reserved.

  17. Quality parameters and antioxidant properties in organic and conventionally grown broccoli after pre-storage hot water treatment.

    Science.gov (United States)

    Zapata, Pedro Javier; Tucker, Gregory A; Valero, Daniel; Serrano, María

    2013-03-30

    Demand for broccoli has increased due to its high content of bioactive compounds. However, broccoli is a perishable commodity with a short shelf life mainly due to dehydration, yellowing and losses of bioactive compounds. Thus, efficient treatments to preserve broccoli quality are needed. The effect of heat treatment on senescence and antioxidant compounds evolution during storage at 20 °C was evaluated in organic and conventionally grown broccoli. Senescence evolved quickly as manifested by floral head yellowing, which was higher in conventional than in organic broccolis, but senescence was significantly delayed by heat treatment. All organic acids, including ascorbic acid, were found at higher concentrations in organic than in conventional broccoli at harvest but decreased during storage in all broccolis. Phenolic concentration and antioxidant activity (in both hydrophilic and lipophilic fractions) also decreased during storage, although these decreases were higher in conventional than in organic broccolis, and no differences were found attributable to heat treatment. Heat treatment was effective in delaying broccoli senescence, manifested by chlorophyll retention. In addition, organic broccoli maintained higher concentrations of bioactive compounds (ascorbic acid and phenolics) and antioxidant potential during storage than conventional broccoli, with higher potential health beneficial effects. © 2012 Society of Chemical Industry.

  18. Mugwort-Mustard Allergy Syndrome due to Broccoli Consumption

    Directory of Open Access Journals (Sweden)

    Yuri Sugita

    2016-01-01

    Full Text Available Pollen-food allergy syndrome (PFAS is a relatively rare form of food allergy which develops in individuals who are sensitized to pollen. Tree pollens, especially birch pollen, frequently induce PFAS; however, the incidence of PFAS due to grass or weed pollens such as ragweed or mugwort is relatively rare. Mugwort-mustard allergy syndrome (MMAS is an example of a PFAS in which individuals sensitized to mugwort may develop an allergy to mustard and experience severe reactions. We herein describe a case of MMAS due to broccoli consumption.

  19. Kinetics of Sulforaphane in Mice after Consumption of Sulforaphane-Enriched Broccoli Sprout Preparation

    Science.gov (United States)

    Li, Yanyan; Zhang, Tao; Li, Xiaoqin; Zou, Peng; Schwartz, Steven J.; Sun, Duxin

    2013-01-01

    Scope Sulforaphane is a natural isothiocyanate in broccoli sprouts with cancer chemopreventive activity. This study is aimed to to use different methods to develop broccoli sprout preparations to compare their ability to deliver sulforaphane to the mice and to evaluate the kinetics and biodistribution of sulforaphane. Methods and Results The sulforaphane-enriched sprout preparation generated by two-step procedure (quick-steaming followed by myrosinase treatment) contained the highest level of sulforaphane, which was 11 and 5 times higher than the freeze-dried fresh broccoli sprouts and the quick-steamed, freeze-dried broccoli sprouts, respectively. After oral administration of 2.5 mg/g body weight of the broccoli sprout preparations, sulforaphane was quickly absorbed and distributed throughout the tissues. The sulforaphane-rich preparation resulted in the highest exposure, with peak plasma sulforaphane concentration of 337 ng/ml, which is 6.0 times and 2.6 times higher compared to the other two preparations. A whole body physiologically-based pharmacokinetic model (developed with ADAPT 5 software) suggests that distribution of sulforaphane is perfusion-limited in all organs. Conclusion This study provides a broccoli sprout preparation that can serve as a good source of sulforaphane, and the model can be utilized to guide the dose design for the use of broccoli sprout preparation in chemoprevention. PMID:23929742

  20. Effect of Selenium Enrichment on the Growth, Photosynthesis and Mineral Nutrition of Broccoli

    Directory of Open Access Journals (Sweden)

    Yousef GHASEMI

    2016-06-01

    Full Text Available Broccoli is placed in primary selenium (Se accumulator group plants, which is considered as an important source of Se for providing human daily need. This experiment used an outdoor hydroponic system to evaluate the effects of Se foliar application at the rates of 0, 10, 50, and 100 μg Se/ml concentrations fortnightly. Among yield parameters, the head weight of broccoli was significantly affected. Se treated broccoli plants produced heavier head than the control; however, head weight among three Se concentrations (Se10, Se50, Se100 was not significantly different. Although most of the chlorophyll fluorescence parameters were not significant, Se treated broccolis maximal fluorescence yield (Fm was higher than the control. Significant increase in chlorophyll content (chlorophyll a, chlorophyll b and total chlorophyll was observed as a result of Se treatments. Different Se concentrations did not have positive or negative effects on nitrogen, phosphorus and potassium uptake. Se treatment at 100 μg Se/ml concentration however, contributed to the highest content of sulfur in broccoli head. Se content of broccoli head increased with the increase in sprayed Se concentrations. The highest concentration of Se (1.41 mg Se/kg dry matter in broccoli head was recorded in Se100, which showed significant difference compared with Se0 and Se10.

  1. Kinetics of sulforaphane in mice after consumption of sulforaphane-enriched broccoli sprout preparation.

    Science.gov (United States)

    Li, Yanyan; Zhang, Tao; Li, Xiaoqin; Zou, Peng; Schwartz, Steven J; Sun, Duxin

    2013-12-01

    Sulforaphane (SF) is a natural isothiocyanate in broccoli sprouts with cancer chemopreventive activity. This study is aimed to use different methods to develop broccoli sprout preparations to compare their ability to deliver SF to the mice and to evaluate the kinetics and biodistribution of SF. The SF-enriched sprout preparation generated by two-step procedure (quick-steaming followed by myrosinase treatment) contained the highest level of SF, which was 11 and 5 times higher than the freeze-dried fresh broccoli sprouts and the quick-steamed, freeze-dried broccoli sprouts, respectively. After oral administration of 2.5 mg/g body weight of the broccoli sprout preparations, SF was quickly absorbed and distributed throughout the tissues. The SF-rich preparation resulted in the highest exposure, with peak plasma SF concentration of 337 ng/mL, which is 6.0 times and 2.6 times higher compared to the other two preparations. A whole body physiologically based pharmacokinetic model (developed with ADAPT 5 software) suggests that distribution of SF is perfusion-limited in all organs. This study provides a broccoli sprout preparation that can serve as a good source of SF, and the model can be utilized to guide the dose designed for the use of broccoli sprout preparation in chemoprevention. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  2. Effect of NaCl treatments on glucosinolate metabolism in broccoli sprouts*

    Science.gov (United States)

    Guo, Rong-fang; Yuan, Gao-feng; Wang, Qiao-mei

    2013-01-01

    To understand the regulation mechanism of NaCl on glucosinolate metabolism in broccoli sprouts, the germination rate, fresh weight, contents of glucosinolates and sulforaphane, as well as myrosinase activity of broccoli sprouts germinated under 0, 20, 40, 60, 80, and 100 mmol/L of NaCl were investigated in our experiment. The results showed that glucoerucin, glucobrassicin, and 4-hydroxy glucobrassicin in 7-d-old broccoli sprouts were significantly enhanced and the activity of myrosinase was inhibited by 100 mmol/L of NaCl. However, the total glucosinolate content in 7-d-old broccoli sprouts was markedly decreased although the fresh weight was significantly increased after treatment with NaCl at relatively low concentrations (20, 40, and 60 mmol/L). NaCl treatment at the concentration of 60 mmol/L for 5 d maintained higher biomass and comparatively higher content of glucosinolates in sprouts of broccoli with decreased myrosinase activity. A relatively high level of NaCl treatment (100 mmol/L) significantly increased the content of sulforaphane in 7-d-old broccoli sprouts compared with the control. These results indicate that broccoli sprouts grown under a suitable concentration of NaCl could be desirable for human nutrition. PMID:23365011

  3. Genetic regulation of glucoraphanin accumulation in Beneforté® broccoli

    Science.gov (United States)

    Traka, Maria H; Saha, Shikha; Huseby, Stine; Kopriva, Stanislav; Walley, Peter G; Barker, Guy C; Moore, Jonathan; Mero, Gene; den Bosch, Frans; Constant, Howard; Kelly, Leo; Schepers, Hans; Boddupalli, Sekhar; Mithen, Richard F

    2013-01-01

    Diets rich in broccoli (Brassica oleracea var italica) have been associated with maintenance of cardiovascular health and reduction in risk of cancer. These health benefits have been attributed to glucoraphanin that specifically accumulates in broccoli. The development of broccoli with enhanced concentrations of glucoraphanin may deliver greater health benefits. Three high-glucoraphanin F1 broccoli hybrids were developed in independent programmes through genome introgression from the wild species Brassica villosa. Glucoraphanin and other metabolites were quantified in experimental field trials. Global SNP analyses quantified the differential extent of B. villosa introgression The high-glucoraphanin broccoli hybrids contained 2.5–3 times the glucoraphanin content of standard hybrids due to enhanced sulphate assimilation and modifications in sulphur partitioning between sulphur-containing metabolites. All of the high-glucoraphanin hybrids possessed an introgressed B. villosa segment which contained a B. villosa Myb28 allele. Myb28 expression was increased in all of the high-glucoraphanin hybrids. Two high-glucoraphanin hybrids have been commercialised as Beneforté® broccoli. The study illustrates the translation of research on glucosinolate genetics from Arabidopsis to broccoli, the use of wild Brassica species to develop cultivars with potential consumer benefits, and the development of cultivars with contrasting concentrations of glucoraphanin for use in blinded human intervention studies. PMID:23560984

  4. Genetics Home Reference: guanidinoacetate methyltransferase deficiency

    Science.gov (United States)

    ... literature. Of these, approximately one-third are of Portuguese origin. Related Information What information about a genetic ... require large amounts of energy, especially the brain. Learn more about the gene associated with guanidinoacetate methyltransferase ...

  5. Heat Transfer during Blanching and Hydrocooling of Broccoli Florets.

    Science.gov (United States)

    Iribe-Salazar, Rosalina; Caro-Corrales, José; Hernández-Calderón, Óscar; Zazueta-Niebla, Jorge; Gutiérrez-Dorado, Roberto; Carrazco-Escalante, Marco; Vázquez-López, Yessica

    2015-12-01

    The objective of this work was to simulate heat transfer during blanching (90 °C) and hydrocooling (5 °C) of broccoli florets (Brassica oleracea L. Italica) and to evaluate the impact of these processes on the physicochemical and nutrimental quality properties. Thermophysical properties (thermal conductivity [line heat source], specific heat capacity [differential scanning calorimetry], and bulk density [volume displacement]) of stem and inflorescence were measured as a function of temperature (5, 10, 20, 40, 60, and 80 °C). The activation energy and the frequency factor (Arrhenius model) of these thermophysical properties were calculated. A 3-dimensional finite element model was developed to predict the temperature history at different points inside the product. Comparison of the theoretical and experimental temperature histories was carried out. Quality parameters (firmness, total color difference, and vitamin C content) and peroxidase activity were measured. The satisfactory validation of the finite element model allows the prediction of temperature histories and profiles under different process conditions, which could lead to an eventual optimization aimed to minimize the nutritional and sensorial losses in broccoli florets. © 2015 Institute of Food Technologists®

  6. Broccoli (Brassica oleracea var. italica head initiation under field conditions

    Directory of Open Access Journals (Sweden)

    Alina Kałużewicz

    2012-12-01

    Full Text Available A two–year study on the influence of temperature on broccoli head initiation was carried out at the ''Marcelin'' experimental station of the Poznań University of Life Sciences. In each year of the study, plants were planted in the field at four dates. The evaluation of the developmental phase of the broccoli shoot apex was based on the analysis of microscope slides. The date of head initiation was assumed as the day on which the first of the examined apices were found to be at the early generative phase. The plant characteristics (number of leaves, leaf area and stem diameter on the date of initiation were also determined. Variation in length of the period from planting to head initiation was found both between dates of planting and between experimental years. The shortest period from planting to initiation was when the plants were planted in April and June (17-18 days in the first year and the longest one for planting in April in the first year of the study (29 days. The length of the period from planting to head initiation depended on mean daily air temperature. The higher the temperature was, the shorter was the period.

  7. Broccoli/weed/soil discrimination by optical reflectance using neural networks

    Science.gov (United States)

    Hahn, Federico

    1995-04-01

    Broccoli is grown extensively in Scotland, and has become one of the main vegetables cropped, due to its high yields and profits. Broccoli, weed and soil samples from 6 different farms were collected and their spectra obtained and analyzed using discriminant analysis. High crop/weed/soil discrimination success rates were encountered in each farm, but the selected wavelengths varied in each farm due to differences in broccoli variety, weed species incidence and soil type. In order to use only three wavelengths, neural networks were introduced and high crop/weed/soil discrimination accuracies for each farm were achieved.

  8. Impact of thermal processing on sulforaphane yield from broccoli (Brassica oleracea L. var. italica)

    Science.gov (United States)

    In broccoli, sulforaphane forms when the glucosinolate glucoraphanin is hydrolyzed by the endogenous plant thiohydrolase myrosinase. A myrosinase cofactor directs hydrolysis away from formation of bioactive sulforaphane and toward an inactive product, sulforaphane nitrile. The cofactor is more hea...

  9. SATURDAY: EPA Administrator to Deliver Remarks at Third Annual Broccoli Festival

    Science.gov (United States)

    WASHINGTON - On Saturday, April 25, EPA Administrator Gina McCarthy will give remarks at the Third Annual Broccoli City Festival's Earth Day celebration. Administrator McCarthy will speak about EPA's efforts to implement President Obama's Climate Ac

  10. The effect of the germination temperature on the phytochemical content of broccoli and rocket sprouts.

    Science.gov (United States)

    Ragusa, Lucia; Picchi, Valentina; Tribulato, Alessandro; Cavallaro, Chiara; Lo Scalzo, Roberto; Branca, Ferdinando

    2017-06-01

    This study investigates the effect of different germination temperatures (10, 20 and 30 °C) on the phytochemical content as well as reducing and antioxidant capacity of broccoli and rocket sprouts. In both seeds and sprouts, the total glucosinolates and ascorbic acid contents did not differ between vegetables, while broccoli exhibited exceptionally higher polyphenols and greater reducing and antioxidant capacity compared to rocket. In both species, an increase in germination temperature positively affected the glucosinolate content. Ascorbic acid increased during germination without a difference among the three tested temperatures. The phenol content in broccoli sprouts increased when they were grown at 30 °C, but the amount decreased at the highest temperatures in rocket. The reducing and antioxidant capacities increased with germination, and higher indexes were detected at 10 °C, particularly in rocket. Different germination temperatures differentiate the health-promoting phytochemical content and antioxidant properties in broccoli and rocket sprouts.

  11. Compositional and proteomic analyses of genetically modified broccoli (Brassica oleracea var. italica) harboring an agrobacterial gene

    National Research Council Canada - National Science Library

    Liu, Mao-Sen; Ko, Miau-Hwa; Li, Hui-Chun; Tsai, Shwu-Jene; Lai, Ying-Mi; Chang, You-Ming; Wu, Min-Tze; Chen, Long-Fang O

    2014-01-01

    .... We also preliminarily assessed safety in mice. Most aspects were comparable between ipt-transgenic broccoli and controls, except for a significant increase in carbohydrate level and a decrease in magnesium content in ipt-transgenic lines 101, 102...

  12. Radiographic images and relationship of the internal morphology and physiological potential of broccoli seeds

    OpenAIRE

    Haynna Fernandes Abud; Silvio Moure Cícero; Francisco Guilhien Gomes Junior

    2017-01-01

    Image analysis using X-rays is an efficient technique for assessing the seed quality of several species, presenting itself as a rapid response method that is simple to execute, reproducible and non-destructive. Thus, this research adjusted a methodology that aims to relate the internal morphology of broccoli seeds to their physiological potential through radiographic analysis of seeds and computerized images of seedlings. The broccoli cultivars used were Piracicaba Precoce and Ramoso Santana ...

  13. The effects of priming on vigor and viability of broccoli (Brassica oleracea var. italica Plenck) seeds

    OpenAIRE

    Jett, Lewis W.

    1994-01-01

    Seed priming is a controlled hydration process, followed by dehydration, that allows pregerminative metabolic activity to proceed without germination. The objective of this research was to investigate the effects of priming on intrinsic characteristics of seed germination including temperature, water, and development, in order to understand how priming affects the germination of broccoli (Brassica oleracea var. italica Plenck) seeds. Priming of broccoli seeds consistently impro...

  14. Colorless Chlorophyll Catabolites in Senescent Florets of Broccoli (Brassica oleracea var. italica)

    OpenAIRE

    Roiser, Matthias H.; M?ller, Thomas; Kr?utler, Bernhard

    2015-01-01

    Typical postharvest storage of broccoli (Brassica oleracea var. italica) causes degreening of this common vegetable with visible loss of chlorophyll (Chl). As shown here, colorless Chl-catabolites are generated. In fresh extracts of degreening florets of broccoli, three colorless tetrapyrrolic Chl-catabolites accumulated and were detected by high performance liquid chromatography (HPLC): two ?nonfluorescent? Chl-catabolites (NCCs), provisionally named Bo-NCC-1 and Bo-NCC-2, and a colorless 1,...

  15. Influence of different blanching methods on colour, ascorbic acid and phenolics content of broccoli

    OpenAIRE

    Severini, C; Giuliani, R.; De Filippis, A; Derossi, A.; T. DE PILLI

    2015-01-01

    Lack of nutrients in cooking water, high energetic costs, high water consumption and recycling are some drawbacks of vegetable blanching. Those disadvantages could be bypassed using microwave blanching. Three blanching methods (microwave, boiling water and steaming) were compared in this study in order to determine their effects on some functional properties of broccoli. In addition, the thermal damage on broccoli colour was evaluated. The effectiveness of each blanching process was performed...

  16. Combinations of tomato and broccoli enhance antitumor activity in dunning r3327-h prostate adenocarcinomas.

    Science.gov (United States)

    Canene-Adams, Kirstie; Lindshield, Brian L; Wang, Shihua; Jeffery, Elizabeth H; Clinton, Steven K; Erdman, John W

    2007-01-15

    The consumption of diets containing 5 to 10 servings of fruits and vegetables daily is the foundation of public health recommendations for cancer prevention, yet this concept has not been tested in experimental models of prostate cancer. We evaluated combinations of tomato and broccoli in the Dunning R3327-H prostate adenocarcinoma model. Male Copenhagen rats (n=206) were fed diets containing 10% tomato, 10% broccoli, 5% tomato plus 5% broccoli (5:5 combination), 10% tomato plus 10% broccoli (10:10 combination) powders, or lycopene (23 or 224 nmol/g diet) for approximately 22 weeks starting 1 month prior to receiving s.c. tumor implants. We compared the effects of diet to surgical castration (2 weeks before termination) or finasteride (5 mg/kg body weight orally, 6 d/wk). Castration reduced prostate weights, tumor areas, and tumor weight (62%, Ptomato reduced tumor weight by 34% (Ptomato and broccoli was more effective at slowing tumor growth than either tomato or broccoli alone and supports the public health recommendations to increase the intake of a variety of plant components.

  17. Microwave assisted dehydration of broccoli by-products and simultaneous extraction of bioactive compounds.

    Science.gov (United States)

    Ferreira, Sónia S; Passos, Cláudia P; Cardoso, Susana M; Wessel, Dulcineia F; Coimbra, Manuel A

    2018-04-25

    Broccoli by-products from frozen-food industry account for 45% of the initial broccoli heads. They consist on stalks, inflorescences, and leaves, blanched and non-blanched, sharing the nutritional value and bioactive compounds of commercial broccoli heads. However, their high perishability prevents further valorisation. Therefore, in this study microwave hydrodiffusion and gravity (MHG) technology was used to dehydrate broccoli by-products and simultaneously recover the water-soluble diffused compounds for food ingredients use. The hydrodiffusion allowed to obtain a dried material with 12% moisture in 43 min when 550 g of broccoli by-products were used, preserving polysaccharides and proteins. Diffused water contained up to 317 µg/mL gallic acid equivalents of phenolic compounds, 11 mg/mL free sugars, 9 mg/mL amino acids, and 356 µg/mL glucosinolates, depending on the type of by-product used. These results show the potential of MHG technology for valorisation of broccoli by-products by its simultaneous stabilization by dehydration and extraction of bioactive compounds. Copyright © 2017 Elsevier Ltd. All rights reserved.

  18. Comparison of the effect of raw and blanched-frozen broccoli on DNA damage in colonocytes.

    Science.gov (United States)

    Lynn, Anthony; Fuller, Zoë; Collins, Andrew R; Ratcliffe, Brian

    2015-07-01

    Consumption of cruciferous vegetables may protect against colorectal cancer. Cruciferous vegetables are rich in a number of bioactive constituents including polyphenols, vitamins and glucosinolates. Before consumption, cruciferous vegetables often undergo some form of processing that reduces their content of bioactive constituents and may determine whether they exert protective effects. The aim of this study was to compare the ability of raw and blanched-frozen broccoli to protect colonocytes against DNA damage, improve antioxidant status and induce xenobiotic metabolizing enzymes (XME). Fifteen Landrace × Large White male pigs were divided into five age-matched and weight-matched sets (79 days, SD 3, and 34·7 kg, SD 3·9, respectively). Each set consisted of siblings to minimize genetic variation. Within each set, pigs received a cereal-based diet, unsupplemented (control) or supplemented with 600 g day(-1) of raw or blanched-frozen broccoli for 12 days. The consumption of raw broccoli caused a significant 27% increase in DNA damage in colonocytes (p = 0·03) relative to the control diet, whereas blanched-frozen broccoli had no significant effect. Both broccoli diets had no significant effect on plasma antioxidant status or hepatic and colonic XME. This study is the first to report that the consumption of raw broccoli can damage DNA in porcine colonocytes. Copyright © 2015 John Wiley & Sons, Ltd.

  19. Compositional and proteomic analyses of genetically modified broccoli (Brassica oleracea var. italica) harboring an agrobacterial gene.

    Science.gov (United States)

    Liu, Mao-Sen; Ko, Miau-Hwa; Li, Hui-Chun; Tsai, Shwu-Jene; Lai, Ying-Mi; Chang, You-Ming; Wu, Min-Tze; Chen, Long-Fang O

    2014-08-28

    Previously, we showed improved shelf life for agrobacterial isopentenyltransferase (ipt) transgenic broccoli (Brassica oleracea var. italica), with yield comparable to commercial varieties, because of the protection mechanism offered by molecular chaperones and stress-related proteins. Here, we used proximate analysis to examine macronutrients, chemical and mineral constituents as well as anti-nutrient and protein changes of ipt-transgenic broccoli and corresponding controls. We also preliminarily assessed safety in mice. Most aspects were comparable between ipt-transgenic broccoli and controls, except for a significant increase in carbohydrate level and a decrease in magnesium content in ipt-transgenic lines 101, 102 and 103, as compared with non-transgenic controls. In addition, the anti-nutrient glucosinolate content was increased and crude fat content decreased in inbred control 104 and transgenic lines as compared with the parental control, "Green King". Gel-based proteomics detected more than 50 protein spots specifically found in ipt-transgenic broccoli at harvest and after cooking; one-third of these proteins showed homology to potential allergens that also play an important role in plant defense against stresses and senescence. Mice fed levels of ipt-transgenic broccoli mimicking the 120 g/day of broccoli eaten by a 60-kg human adult showed normal growth and immune function. In conclusion, the compositional and proteomic changes attributed to the transgenic ipt gene did not affect the growth and immune response of mice under the feeding regimes examined.

  20. Effects of Sub-irrigation at Different Growth Stages on Growth of Pot-cultured Summer-sowing for Fall-harvest Broccoli (Brassica oleracea L.)

    National Research Council Canada - National Science Library

    Yuka Nakano; Kunihiko Okada; Hidekazu Sasaki

    2017-01-01

    ... of sub-irrigation at different growth stages of broccoli and levels of precipitation. Summer-sown fall-harvest broccoli was transplanted in a large pot of andosol or gray lowland soil under rain shelter...

  1. Chemical Probes of Histone Lysine Methyltransferases

    Science.gov (United States)

    2015-01-01

    Growing evidence suggests that histone methyltransferases (HMTs, also known as protein methyltransferases (PMTs)) play an important role in diverse biological processes and human diseases by regulating gene expression and the chromatin state. Therefore, HMTs have been increasingly recognized by the biomedical community as a class of potential therapeutic targets. High quality chemical probes of HMTs, as tools for deciphering their physiological functions and roles in human diseases and testing therapeutic hypotheses, are critical for advancing this promising field. In this review, we focus on the discovery, characterization, and biological applications of chemical probes for HMTs. PMID:25423077

  2. Consumer acceptability and sensory profile of cooked broccoli with mustard seeds added to improve chemoprotective properties.

    Science.gov (United States)

    Ghawi, Sameer Khalil; Shen, Yuchi; Niranjan, Keshavan; Methven, Lisa

    2014-09-01

    Broccoli, a rich source of glucosinolates, is a commonly consumed vegetable of the Brassica family. Hydrolysis products of glucosinolates, isothiocyanates, have been associated with health benefits and contribute to the flavor of Brassica. However, boiling broccoli causes the myrosinase enzyme needed for hydrolysis to denature. In order to ensure hydrolysis, broccoli must either be mildly cooked or active sources of myrosinase, such as mustard seed powder, can be added postcooking. In this study, samples of broccoli were prepared in 6 different ways; standard boiling, standard boiling followed by the addition of mustard seeds, sous vide cooking at low temperature (70 °C) and sous vide cooking at higher temperature (100 °C) and sous vide cooking at higher temperature followed by the addition of mustard seeds at 2 different concentrations. The majority of consumers disliked the mildly cooked broccoli samples (70 °C, 12 min, sous vide) which had a hard and stringy texture. The highest mean consumer liking was for standard boiled samples (100 °C, 7 min). Addition of 1% mustard seed powder developed sensory attributes, such as pungency, burning sensation, mustard odor, and flavor. One cluster of consumers (32%) found mustard seeds to be a good complement to cooked broccoli; however, the majority disliked the mustard-derived sensory attributes. Where the mustard seeds were partially processed, doubling the addition to 2% led to only the same level of mustard and pungent flavors as 1% unprocessed seeds, and mean consumer liking remained unaltered. This suggests that optimization of the addition level of partially processed mustard seeds may be a route to enhance bioactivity of cooked broccoli without compromising consumer acceptability. © 2014 Institute of Food Technologists®

  3. Influences of Cry1Ac broccoli on larval survival and oviposition of diamondback moth.

    Science.gov (United States)

    Yi, Dengxia; Cui, Shusong; Yang, Limei; Fang, Zhiyuan; Liu, Yumei; Zhuang, Mu; Zhang, Yangyong

    2015-01-01

    Larval survival and oviposition behavior of three genotypes of diamondback moth, Plutella xylostella L. (Lepidoptera: Plutellidae), (homozygous Cry1Ac-susceptibile, Cry1Ac-resistant, and their F1 hybrids), on transgenic Bacillus thuringiensis (Bt) broccoli expressing different levels of Cry1Ac protein were evaluated in laboratory. These Bt broccoli lines were designated as relative low, medium, and high, respectively, according to the Cry1Ac content. Untransformed brocccoli plants were used as control. Larval survival of diamondback moth on non-Bt leaves was not significantly different among the three genotypes. The Cry1Ac-resistant larvae could survive on the low level of Bt broccoli plants, while Cry1Ac-susceptible and F1 larvae could not survive on them. The three genotypes of P. xylostella larvae could not survive on medium and high levels of Bt broccoli. In oviposition choice tests, there was no significant difference in the number of eggs laid by the three P. xylostella genotypes among different Bt broccoli plants. The development of Cry1Ac-susceptible and Cry1Ac-resistant P. xylostella on intact Bt plants was also tested in greenhouse. All susceptible P. xylostella larvae died on all Bt plants, while resistant larvae could survive on broccoli, which expresses low Cry1Ac protein under greenhouse conditions. The results of the greenhouse trials were similar to that of laboratory tests. This study indicated that high dose of Bt toxins in broccoli cultivars or germplasm lines is required for effective resistance management. © The Author 2015. Published by Oxford University Press on behalf of the Entomological Society of America.

  4. Using genotyping-by-sequencing to develop broccoli markers for construction of a high-density linkage map and to identify quantitative trait loci associated with heat tolerance

    Science.gov (United States)

    Heat stress reduces the yield and quality of broccoli heads imposing seasonal and geographic limits to broccoli production. For the most part, the risk of producing broccoli with head defects (uneven beads, bracts in heads, etc.) induced by high temperatures has restricted commercial production in t...

  5. Influence of thiopurine methyltransferase gene polymorphism on ...

    Indian Academy of Sciences (India)

    Azza A. G. Tantawy

    2017-11-28

    Nov 28, 2017 ... Journal of Genetics, Vol. 96, No. ... aim of this study was to determine the influence of TPMT gene polymorphism in Egyptian children with acute lymphoblastic leukaemia (ALL). ... Keywords. thiopurine methyltransferase gene polymorphism; acute lymphoblastic leukaemia; Egyptian children; thiopurine.

  6. RESEARCH ARTICLE The influence of thiopurine methyltransferase ...

    Indian Academy of Sciences (India)

    Navya

    2017-02-06

    Feb 6, 2017 ... The influence of thiopurine methyltransferase gene polymorphism on Egyptian children with acute lymphoblastic leukaemia ... influence response and toxicity of therapy in children with acute lymphoblastic leukaemia. (ALL). [1]The ..... Egyptian Journal of Medical Human Genetics. 12, 183–186. 22. Hamdy ...

  7. Inhibition of Bladder Cancer by Broccoli Isothiocyanates Sulforaphane and Erucin: Characterization, Metabolism and Interconversion

    Science.gov (United States)

    Abbaoui, Besma; Riedl, Kenneth M; Ralston, Robin A; Thomas-Ahner, Jennifer M; Schwartz, Steven J; Clinton, Steven K; Mortazavi, Amir

    2013-01-01

    Epidemiologic evidence suggests diets rich in cruciferous vegetables, particularly broccoli, are associated with lower bladder cancer risk. Our objectives are to investigate these observations and determine the role of isothiocyanates in primary or secondary bladder cancer prevention. We initially investigate the mechanisms whereby broccoli and broccoli sprout extracts and pure isothiocyanates inhibit normal, non-invasive (RT4) and invasive (J82, UMUC3) human urothelial cell viability. Sulforaphane (IC50= 5.66±1.2μM) and erucin (IC50= 8.79±1.3μM) are found to be the most potent inhibitors and normal cells are least sensitive. This observation is associated with downregulation of survivin, EGFR and HER2/neu, G2/M cell cycle accumulation and apoptosis. In a murine UMUC3 xenograft model, we fed semipurified diets containing 4% broccoli sprouts, or 2% broccoli sprout isothiocyanate extract; or gavaged pure sulforaphane or erucin (each at 295 μmol/kg, similar to dietary exposure); and report tumor weight reduction of 42% (p=0.02), 42% (p=0.04), 33% (p=0.04) and 58% (pSulforaphane and erucin metabolites are present in mouse plasma (micromolar range) and tumor tissue, with N-acetyl cysteine conjugates as the most abundant. Interconversion of sulforaphane and erucin metabolites was observed. This work supports development of fully characterized, novel food products for phase I/II human studies targeting bladder cancer prevention. PMID:23038615

  8. Optimization of methyl jasmonate application to broccoli florets to enhance health-promoting phytochemical content.

    Science.gov (United States)

    Ku, Kang Mo; Jeffery, Elizabeth H; Juvik, John A

    2014-08-01

    Spray treatment of methyl jasmonate (MeJA) has been shown to increase glucosinolate (GS) concentrations and health-promoting activity in Brassica vegetables. Since there is no reported standardized protocol, several MeJA treatment studies have been conducted to maximize human health bioactivity using the F1 broccoli cultivar 'Green Magic'. Foliar MeJA application 4 days prior to harvest of broccoli at commercial maturity resulted in enhanced total GS concentrations. Although a single application of 250 µmol L(-1) MeJA maximized GS concentrations in broccoli florets, two days of consecutive treatments (4 and 3 days prior to harvest) of 250 µmol L(-1) MeJA further enhanced neoglucobrassicin concentrations and floret extract quinone reductase (QR)-inducing activity. With increasing concentrations of MeJA in spray applications to broccoli florets, concentrations of the glucosinolates glucoraphanin, gluconasturtiin and neoglucobrassicin and the isothiocyanate sulforaphane as well as anticancer and anti-inflammatory bioactivities as measured by QR induction and inhibition of nitric oxide (NO) production respectively were significantly increased. Concentrations of these phytochemicals showed strong positive correlations with QR-inducing and NO-inhibitory activities. These application protocols were found to maximize GS and GS hydrolysis product concentrations and putatively enhance the health-promoting properties of broccoli heads for consumers. © 2013 Society of Chemical Industry.

  9. Growth and yield of broccoli fertilized with doses of velvet bean in greenhouse

    Directory of Open Access Journals (Sweden)

    Ellen Rúbia Diniz

    2015-06-01

    Full Text Available The dose effects of green manure in vegetable crops production are still poorly understood. There are few scientific studies indicate that increasing the dose may influence plants characteristics. The objectives were to evaluate the effect of doses of velvet bean green manure on growth and yield, the partitioning of dry matter and nitrogen (N in plants of broccoli, as well as determine the apparent recovery and physiological efficiency of use N. We established four treatments plus a control. The treatments consisted of four levels of green manure: 0, 3, 6, 9 t ha-1 on a dry matter basis, with the addition of 12 t ha-1 of compost. Doses of green manure applied influenced the growth and yield of broccoli. The distribution of dry matter between plant parts of broccoli is not influenced by the doses of green manure. Doses of green manure influenced the amount of N accumulated in the inflorescence and whole plant of broccoli. The apparent recovery of N from green manure and efficiency of N use by plants of broccoli are positively associated with the applied doses.

  10. Glucosinolate biosynthesis in hairy root cultures of broccoli (Brassica oleracea var. italica).

    Science.gov (United States)

    Kim, Sun-Ju; Park, Woo Tae; Uddin, Md Romij; Kim, Yeon Bok; Nam, Sang-Yong; Jho, Kwang Hyun; Park, Sang Un

    2013-02-01

    Here we present previously unreported glucosinolate production by hairy root cultures of broccoli (B. oleracea var. italica). Growth media greatly influenced the growth and glucosinolate content of hairy root cultures of broccoli. Seven glucosinolates, glucoraphanin, gluconapin, glucoerucin, glucobrassicin, 4-methoxyglucobrassicin, gluconasturtiin, and neoglucobrassicin, were identified by analysis of the broccoli hairy root cultures. Both half and full strength B5 and SH media enabled the highest accumulation of glucosinolates. In most cases, the levels of glucosinolates were higher in SH and BS media. Among the 7 glucosinolates, the accumulation of neoglucobrassicin was very high, irrespective of growth medium. The neoglucobrassicin content was 7.4-fold higher in SH medium than 1/2 MS, in which its level was the lowest. The 1/2 B5 medium supported the production of the highest amounts of glucobrassicin and 4-methoxyglucobrassicin, the levels for which were 36.2- and 7.9- fold higher, respectively, than their lowest content in 1/2 MS medium. The 1/2 SH medium enabled the highest accumulation of glucoraphanin and gluconapin in the broccoli hairy root cultures, whose levels were 1.8- and 4.6-fold higher, respectively, than their lowest content in 1/2 MS medium. Our results suggest that hairy root cultures of broccoli could be a valuable alternative approach for the production of glucosinolate compounds.

  11. Potential of different mechanical and thermal treatments to control off-flavour generation in broccoli puree.

    Science.gov (United States)

    Koutidou, Maria; Grauwet, Tara; Van Loey, Ann; Acharya, Parag

    2017-02-15

    The aim of this study was scientifically investigate the impact of the sequence of different thermo-mechanical treatments on the volatile profile of differently processed broccoli puree, and to investigate if any relationship persists between detected off-flavour changes and microstructural changes as a function of selected process conditions. Comparison of the headspace GC-MS fingerprinting of the differently processed broccoli purees revealed that an adequate combination of processing steps allows to reduce the level of off-flavour volatiles. Moreover, applying mechanical processing before or after the thermal processing at 90°C determines the pattern of broccoli tissue disruption, resulting into different microstructures and various enzymatic reactions inducing volatile generation. These results may aid the identification of optimal process conditions generating a reduced level of off-flavour in processed broccoli. In this way, broccoli can be incorporated as a food ingredient into mixed food products with limited implications on sensorial consumer acceptance. Copyright © 2016 Elsevier Ltd. All rights reserved.

  12. Metabolism of phenolic compounds by Lactobacillus spp. during fermentation of cherry juice and broccoli puree.

    Science.gov (United States)

    Filannino, Pasquale; Bai, Yunpeng; Di Cagno, Raffaela; Gobbetti, Marco; Gänzle, Michael G

    2015-04-01

    This study aimed to investigate the metabolism of phenolic acids and flavonoids during lactic acid fermentation of cherry juice and broccoli puree for potential food and pharmaceutical purposes. When fermenting cherry juice and broccoli puree, Lactobacillus spp. exhibited strain-specific metabolism of phenolic acid derivatives. The metabolism of protocatechuic, caffeic and p-coumaric acids through phenolic acid decarboxylases and reductases differed between mMRS and cherry juice and broccoli puree. The synthesis of reduced compounds was the highest during food fermentations and the substrate seemed to modulate the metabolism of phenolic compounds. The reduction of phenolic acids involves a hydrogen donor and the re-oxidation of the reduced co-factor NADH, which may provide a metabolic advantage through NAD(+) regeneration. Quinic acid reduction may replace fructose and pyruvate as hydrogen acceptors, and it may provide an energetic advantage to heterofermentative bacteria when growing in broccoli puree lacking of fructose. This study demonstrated that phenolics metabolism may confer a selective advantage for lactobacilli in vegetable and fruit fermentation, and the metabolic routes are strongly dependent on the intrinsic factors of substrate. Fermented cherry juice and broccoli puree, due to the selected bacterial bioconversion pathways, are enriched in phenolic derivative with high human bioavailability and biological activity. Copyright © 2014 Elsevier Ltd. All rights reserved.

  13. The Propagation of Variation in Glucosinolate Levels as effected by Controlled Atmosphere and Temperature in a Broccoli Batch

    NARCIS (Netherlands)

    Schouten, R.E.; Zhang, X.; Tijskens, L.M.M.; Kooten, van O.

    2008-01-01

    Broccoli combines high levels of vitamins, fibres and glucosinolates (GLS) with a low calorie count. GLS are precursors for the characteristic broccoli flavour and have anti-carcinogenic properties. This study describes the effect of controlled atmo¬sphere (CA) and temperature on GLS concentrations

  14. Effect of matrix and particle type on rheological, textural and structural properties of broccoli pasta and noodles

    NARCIS (Netherlands)

    Silva, E.; Sagis, L.M.C.; Linden, van der E.; Scholten, E.

    2013-01-01

    Durum wheat semolina (DWS) pasta and sweet potato starch (SPS) noodles were incorporated with dif ferent volume fractions and types of broccoli powder (up to 20% v/v). The incorporation of high volume fractions of broccoli powder produced in-house in SPS noodles increases the modulus of the dough

  15. Using weighted trait indices to select the best performing broccoli hybrids in multi-site and multi-year trials

    Science.gov (United States)

    Understanding and implementing evaluation data from vegetable trials conducted across multiple years and environments by multiple raters presents numerous challenges. In order to select new broccoli hybrids suitable for eastern production, the SCRI East Coast Broccoli Project has conducted over 32 p...

  16. Regional hybrid broccoli trials provide a means to further breeding efforts of this increasingly important vegetable crop

    Science.gov (United States)

    A Coordinated Agricultural Project (CAP) entitled “Establishing an Eastern Broccoli Industry” is funded under the Specialty Crop Research Initiative (SCRI), and a primary component of the project is a system of regional hybrid broccoli trials conducted along the eastern seaboard. Hybrids currently ...

  17. Evaluation of different cooking conditions on broccoli (Brassica oleracea var. italica) to improve the nutritional value and consumer acceptance.

    Science.gov (United States)

    Bongoni, Radhika; Verkerk, Ruud; Steenbekkers, Bea; Dekker, Matthijs; Stieger, Markus

    2014-09-01

    The objective of this study was to gain insights into the effect of the cooking method on the liking as well as the retention of glucosinolates in broccoli. With this knowledge it can be concluded whether the health aspects of broccoli be improved by the cooking method without deteriorating sensory perception. For this, broccoli was cooked by methods commonly applied by consumers: boiling with a cold (water) start; boiling with a hot (water) start; and steaming. Firmness, greenness and amount of total glucosinolates in cooked broccoli were instrumentally determined. Sensory evaluation by untrained consumers (n = 99) for liking and sensory attributes intensity rating were performed on broccoli cooked by steaming and boiling-cold start at three time points, which resulted in 'high', 'medium', 'low' firm broccoli samples. At the end of cooking, steaming showed an increase in the amount of total glucosinolates (+17%). Boiling-hot start (-41%) and boiling-cold start (-50%) showed a decrease in amount of total glucosinolates. Sensory evaluation did not show statistically significant differences between steaming and boiling-cold start in liking at 'high' and 'medium' firmness; and in the attribute intensity ratings (except for juiciness at 'medium' firmness, and flavour at 'medium' and 'low' firmness). This study demonstrates that medium firm broccoli showed optimum liking and that steaming compared to boiled-cold start showed higher amount of glucosinolates. It is concluded that the health aspects of broccoli can be improved without reducing the sensory aspects by optimising the cooking method.

  18. How do High Temperatures Limit Broccoli Production: Is It Possible for this Cole Crop to Take the Heat?

    Science.gov (United States)

    It is generally thought that broccoli does not grow well in most environments during summer months in the eastern United States, because high temperatures can damage the developing heads causing the harvested vegetable to be unmarketable. Identification of broccoli varieties that produce a quality ...

  19. Anticancer and Antioxidant Activity of Bread Enriched with Broccoli Sprouts

    Science.gov (United States)

    Gawlik-Dziki, Urszula; Świeca, Michał; Dziki, Dariusz; Sęczyk, Łukasz; Złotek, Urszula; Różyło, Renata; Kaszuba, Kinga; Ryszawy, Damian; Czyż, Jarosław

    2014-01-01

    This study is focused on antioxidant and anticancer capacity of bread enriched with broccoli sprouts (BS) in the light of their potential bioaccessibility and bioavailability. Generally, bread supplementation elevated antioxidant potential of product (both nonenzymatic and enzymatic antioxidant capacities); however, the increase was not correlated with the percent of BS. A replacement up to 2% of BS gives satisfactory overall consumers acceptability and desirable elevation of antioxidant potential. High activity was especially found for extracts obtained after simulated digestion, which allows assuming their protective effect for upper gastrointestinal tract; thus, the anticancer activity against human stomach cancer cells (AGS) was evaluated. A prominent cytostatic response paralleled by the inhibition of AGS motility in the presence of potentially mastication-extractable phytochemicals indicates that phenolic compounds of BS retain their biological activity in bread. Importantly, the efficient phenolics concentration was about 12 μM for buffer extract, 13 μM for extracts after digestion in vitro, and 7 μM for extract after absorption in vitro. Our data confirm chemopreventive potential of bread enriched with BS and indicate that BS comprise valuable food supplement for stomach cancer chemoprevention. PMID:25050366

  20. Anticancer and Antioxidant Activity of Bread Enriched with Broccoli Sprouts

    Directory of Open Access Journals (Sweden)

    Urszula Gawlik-Dziki

    2014-01-01

    Full Text Available This study is focused on antioxidant and anticancer capacity of bread enriched with broccoli sprouts (BS in the light of their potential bioaccessibility and bioavailability. Generally, bread supplementation elevated antioxidant potential of product (both nonenzymatic and enzymatic antioxidant capacities; however, the increase was not correlated with the percent of BS. A replacement up to 2% of BS gives satisfactory overall consumers acceptability and desirable elevation of antioxidant potential. High activity was especially found for extracts obtained after simulated digestion, which allows assuming their protective effect for upper gastrointestinal tract; thus, the anticancer activity against human stomach cancer cells (AGS was evaluated. A prominent cytostatic response paralleled by the inhibition of AGS motility in the presence of potentially mastication-extractable phytochemicals indicates that phenolic compounds of BS retain their biological activity in bread. Importantly, the efficient phenolics concentration was about 12 μM for buffer extract, 13 μM for extracts after digestion in vitro, and 7 μM for extract after absorption in vitro. Our data confirm chemopreventive potential of bread enriched with BS and indicate that BS comprise valuable food supplement for stomach cancer chemoprevention.

  1. Separation and Purification of Sulforaphane from Broccoli by Solid Phase Extraction

    Science.gov (United States)

    Han, Dandan; Row, Kyung Ho

    2011-01-01

    A simple solid-phase extraction (SPE) method for the determination of sulforaphane in broccoli has been developed. The optimal conditions were found to be use of a silica SPE cartridge, and ethyl acetate and dichloromethane as washing and eluting solvents, respectively, which could eliminate interferences originating from the broccoli matrix. The extracts were sufficiently clean to be directly injected into high-performance liquid chromatography (HPLC) for further chromatographic analysis. Good linearity was obtained from 0.05 to 200 μg/mL (r = 0.998) for sulforaphane with the relative standard deviations less than 3.6%. The mean recoveries of sulforaphane from broccoli were more than 90.8% and the detection limit (S/N = 3:1) was 0.02 μg/mL. The SPE method provides a higher yield of sulforaphane from crude extracts compared to conventional liquid-liquid extraction. PMID:21673926

  2. Separation and Purification of Sulforaphane from Broccoli by Solid Phase Extraction

    Directory of Open Access Journals (Sweden)

    Kyung Ho Row

    2011-03-01

    Full Text Available A simple solid-phase extraction (SPE method for the determination of sulforaphane in broccoli has been developed. The optimal conditions were found to be use of a silica SPE cartridge, and ethyl acetate and dichloromethane as washing and eluting solvents, respectively, which could eliminate interferences originating from the broccoli matrix. The extracts were sufficiently clean to be directly injected into high-performance liquid chromatography (HPLC for further chromatographic analysis. Good linearity was obtained from 0.05 to 200 μg/mL (r = 0.998 for sulforaphane with the relative standard deviations less than 3.6%. The mean recoveries of sulforaphane from broccoli were more than 90.8% and the detection limit (S/N = 3:1 was 0.02 μg/mL. The SPE method provides a higher yield of sulforaphane from crude extracts compared to conventional liquid-liquid extraction.

  3. Structural analysis of an innate immunostimulant from broccoli, Brassica oleracea var. italica.

    Science.gov (United States)

    Urai, Makoto; Kataoka, Keiko; Nishida, Satoshi; Sekimizu, Kazuhisa

    2017-11-22

    Vegetables are eaten as part of a healthy diet throughout the world, and some are also applied topically as a traditional medicine. We evaluated the innate immunostimulating activities of hot water extracts of various vegetables using the silkworm muscle contraction assay system, and found that broccoli, Brassica oleracea var. italica, contains a strong innate immunostimulant. We purified the innate immunostimulant from broccoli, and characterized the chemical structure by chemical analyses and NMR spectroscopy. The innate immunostimulant comprised galacturonic acid, galactose, glucose, arabinose, and rhamnose, and had a pectic-like polysaccharide structure. To determine the structural motif involved in the innate immunostimulating activity, we modified the structure by chemical and enzymatic treatment, and found that the activity was attenuated by pectinase digestion. These findings suggest that a pectic-like polysaccharide purified from broccoli has innate immune-stimulating activity, for which the polygalacturonic acid structure is necessary.

  4. Inhibition of bladder cancer by broccoli isothiocyanates sulforaphane and erucin: characterization, metabolism, and interconversion.

    Science.gov (United States)

    Abbaoui, Besma; Riedl, Kenneth M; Ralston, Robin A; Thomas-Ahner, Jennifer M; Schwartz, Steven J; Clinton, Steven K; Mortazavi, Amir

    2012-11-01

    Epidemiologic evidence suggests diets rich in cruciferous vegetables, particularly broccoli, are associated with lower bladder cancer risk. Our objectives are to investigate these observations and determine the role of isothiocyanates in primary or secondary bladder cancer prevention. We initially investigate the mechanisms whereby broccoli and broccoli sprout extracts and pure isothiocyanates inhibit normal, noninvasive (RT4), and invasive (J82, UMUC3) human urothelial cell viability. Sulforaphane (IC(50) = 5.66 ± 1.2 μM) and erucin (IC(50) = 8.79 ± 1.3 μM) are found to be the most potent inhibitors and normal cells are least sensitive. This observation is associated with downregulation of survivin, epidermal growth factor receptor (EGFR) and human epidermal growth factor receptor 2 (HER2/neu), G(2) /M cell cycle accumulation, and apoptosis. In a murine UMUC3 xenograft model, we fed semipurified diets containing 4% broccoli sprouts, or 2% broccoli sprout isothiocyanate extract; or gavaged pure sulforaphane or erucin (each at 295 μmol/kg, similar to dietary exposure); and report tumor weight reduction of 42% (p = 0.02), 42% (p = 0.04), 33% (p = 0.04), and 58% (p < 0.0001), respectively. Sulforaphane and erucin metabolites are present in mouse plasma (micromolar range) and tumor tissue, with N-acetylcysteine conjugates as the most abundant. Interconversion of sulforaphane and erucin metabolites was observed. This work supports development of fully characterized, novel food products containing broccoli components for phase I/II human studies targeting bladder cancer prevention. © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  5. Antioxidant and antiproliferative activities in different maturation stages of broccoli (Brassica oleracea Italica) biofortified with selenium.

    Science.gov (United States)

    Bachiega, Patricia; Salgado, Jocelem Mastrodi; de Carvalho, João Ernesto; Ruiz, Ana Lúcia T G; Schwarz, Kélin; Tezotto, Tiago; Morzelle, Maressa Caldeira

    2016-01-01

    In this work, three different broccoli maturity stages subjected to biofortification with selenium were evaluated for antioxidant and antiproliferative activities. Antioxidant trials have shown that the maturation stages biofortified with selenium had significantly higher amounts of phenolic compounds and antioxidant activity, especially seedlings. Although non-polar extracts of all samples show antiproliferative activity, the extract of broccoli seedlings biofortified with selenium stood out, presenting cytocidal activity for a glioma line (U251, GI50 28.5 mg L(-1)). Copyright © 2015 Elsevier Ltd. All rights reserved.

  6. DNA Methyltransferase Activity Assays: Advances and Challenges

    OpenAIRE

    Poh, Wan Jun; Wee, Cayden Pang Pee; Gao,Zhiqiang

    2016-01-01

    DNA methyltransferases (MTases), a family of enzymes that catalyse the methylation of DNA, have a profound effect on gene regulation. A large body of evidence has indicated that DNA MTase is potentially a predictive biomarker closely associated with genetic disorders and genetic diseases like cancer. Given the attention bestowed onto DNA MTases in molecular biology and medicine, highly sensitive detection of DNA MTase activity is essential in determining gene regulation, epigenetic modificati...

  7. Nicotinamide -Methyltransferase in Health and Cancer

    Directory of Open Access Journals (Sweden)

    David B Ramsden

    2017-06-01

    Full Text Available Over the past decade, the roles of nicotinamide N -methyltransferase and its product 1-methyl nicotinamide have emerged from playing merely minor roles in phase 2 xenobiotic metabolism as actors in some of the most important scenes of human life. In this review, the structures of the gene, messenger RNA, and protein are discussed, together with the role of the enzyme in many of the common cancers that afflict people today.

  8. Pathogen detection, testing, and control in fresh broccoli sprouts

    Directory of Open Access Journals (Sweden)

    Fahey Jed W

    2006-04-01

    Full Text Available Abstract Background The recent increased interest in consuming green vegetable sprouts has been tempered by the fact that fresh sprouts can in some cases be vehicles for food-borne illnesses. They must be grown according to proper conditions of sanitation and handled as a food product rather than as an agricultural commodity. When sprouts are grown in accordance with the criteria proposed from within the sprout industry, developed by regulatory agencies, and adhered to by many sprouters, green sprouts can be produced with very low risk. Contamination may occur when these guidelines are not followed. Methods A one year program of microbial hold-and-release testing, conducted in concert with strict seed and facility cleaning procedures by 13 U.S. broccoli sprout growers was evaluated. Microbial contamination tests were performed on 6839 drums of sprouts, equivalent to about 5 million consumer packages of fresh green sprouts. Results Only 24 (0.75% of the 3191 sprout samples gave an initial positive test for Escherichia coli O157:H7 or Salmonella spp., and when re-tested, 3 drums again tested positive. Composite testing (e.g., pooling up to 7 drums for pathogen testing was equally sensitive to single drum testing. Conclusion By using a "test-and-re-test" protocol, growers were able to minimize crop destruction. By pooling drums for testing, they were also able to reduce testing costs which now represent a substantial portion of the costs associated with sprout growing. The test-and-hold scheme described herein allowed those few batches of contaminated sprouts to be found prior to packaging and shipping. These events were isolated, and only safe sprouts entered the food supply.

  9. Determination of Sodium, Potassium, Magnesium, and Calcium Minerals Level in Fresh and Boiled Broccoli and Cauliflower by Atomic Absorption Spectrometry

    Science.gov (United States)

    Nerdy

    2018-01-01

    Vegetables from the cabbage family vegetables consumed by many people, which is known healthful, by eaten raw, boiled, or cooked (stir fry or soup). Vegetables like broccoli and cauliflower contain vitamins, minerals, and fiber. This study aims to determine the decrease percentage of sodium, potassium, magnesium, and calcium minerals level caused by boiled broccoli and cauliflower by atomic absorption spectrometry. Boiled broccoli and cauliflower prepared by given boiled treatment in boiling water for 3 minutes. Fresh and boiled broccoli and cauliflower carried out dry destruction, followed by quantitative analysis of sodium, potassium, magnesium, and calcium minerals respectively at a wavelength of 589.0 nm; 766.5 nm; 285.2 nm; and 422.7 nm, using atomic absorption spectrometry methods. After the determination of the sodium, potassium, magnesium, and calcium minerals level followed by validation of analytical methods with accuracy, precision, linearity, range, limit of detection (LOD), and limit of quantitation (LOQ) parameters. Research results show a decrease in the sodium, potassium, magnesium, and calcium minerals level in boiled broccoli and cauliflower compared with fresh broccoli and cauliflower. Validation of analytical methods gives results that spectrometry methods used for determining sodium, potassium, magnesium, and calcium minerals level are valid. It concluded that the boiled gives the effect of decreasing the minerals level significantly in broccoli and cauliflower.

  10. The effect of post-harvest and packaging treatments on glucoraphanin concentration in broccoli (Brassica oleracea var. italica).

    Science.gov (United States)

    Rangkadilok, Nuchanart; Tomkins, Bruce; Nicolas, Marc E; Premier, Robert R; Bennett, Richard N; Eagling, David R; Taylor, Paul W J

    2002-12-04

    The effects of post-harvest and packaging treatments on glucoraphanin (4-methylsulfinylbutyl glucosinolate), the glucosinolate precursor of anticancer isothiocyanate sulforaphane [4-methylsulfinylbutyl isothiocyanate], were examined in broccoli (Brassica oleracea var. italica) during storage times. The results showed that at 20 degrees C, 55% loss of glucoraphanin concentration occurred in broccoli stored in open boxes during the first 3 days of the treatment and 56% loss was found in broccoli stored in plastic bags by day 7. Under both air and controlled atmosphere (CA) storage, glucoraphanin concentration appeared to fluctuate slightly during 25 days of storage and the concentrations under CA was significantly higher than those stored under air treatment. In modified atmosphere packaging (MAP) treatments, glucoraphanin concentration in air control packaging decreased significantly whereas there were no significant changes in glucoraphanin concentration in MAP with no holes at 4 degrees C and two microholes at 20 degrees C for up to 10 days. Decreases in glucoraphanin concentration occurred when the broccoli heads deteriorated. In the present study, the best method for preserving glucoraphanin concentration in broccoli heads after harvest was storage of broccoli in MAP and refrigeration at 4 degrees C. This condition maintained the glucoraphanin concentration for at least 10 days and also maintained the visual quality of the broccoli heads.

  11. Storage related changes of cell wall based dietary fiber components of broccoli (Brassica oleracea var. italica) stems.

    Science.gov (United States)

    Schäfer, Judith; Stanojlovic, Luisa; Trierweiler, Bernhard; Bunzel, Mirko

    2017-03-01

    Storage related changes in the cell wall composition potentially affect the texture of plant-based foods and the physiological effects of cell wall based dietary fiber components. Therefore, a detailed characterization of cell wall polysaccharides and lignins from broccoli stems was performed. Freshly harvested broccoli and broccoli stored at 20°C and 1°C for different periods of time were analyzed. Effects on dietary fiber contents, polysaccharide composition, and on lignin contents/composition were much more pronounced during storage at 20°C than at 1°C. During storage, insoluble dietary fiber contents of broccoli stems increased up to 13%. Storage related polysaccharide modifications include an increase of the portions of cellulose, xylans, and homogalacturonans and a decrease of the neutral pectic side-chains arabinans and galactans. Broccoli stem lignins are generally rich in guaiacyl units. Lignins from freshly harvested broccoli stems contain slightly larger amounts of p-hydroxyphenyl units than syringyl units. Syringyl units are predominantly incorporated into the lignin polymers during storage, resulting in increased acetyl bromide soluble lignin contents. NMR-based analysis of the interunit linkage types of broccoli stem lignins revealed comparably large portions of resinol structures for a guaiacyl rich lignin. Incorporation of syringyl units into the polymers over storage predominantly occurs through β-O-4-linkages. Copyright © 2017 Elsevier Ltd. All rights reserved.

  12. The potential to intensify sulforaphane formation in cooked broccoli (Brassica oleracea var. italica) using mustard seeds (Sinapis alba).

    Science.gov (United States)

    Ghawi, Sameer Khalil; Methven, Lisa; Niranjan, Keshavan

    2013-06-01

    Sulforaphane, a naturally occurring cancer chemopreventive, is the hydrolysis product of glucoraphanin, the main glucosinolate in broccoli. The hydrolysis requires myrosinase isoenzyme to be present in sufficient activity; however, processing leads to its denaturation and hence reduced hydrolysis. In this study, the effect of adding mustard seeds, which contain a more resilient isoform of myrosinase, to processed broccoli was investigated with a view to intensify the formation of sulforaphane. Thermal inactivation of myrosinase from both broccoli and mustard seeds was studied. Thermal degradation of broccoli glucoraphanin was investigated in addition to the effects of thermal processing on the formation of sulforaphane and sulforaphane nitrile. Limited thermal degradation of glucoraphanin (less than 12%) was observed when broccoli was placed in vacuum sealed bag (sous vide) and cooked in a water bath at 100°C for 8 and 12 min. Boiling broccoli in water prevented the formation of any significant levels of sulforaphane due to inactivated myrosinase. However, addition of powdered mustard seeds to the heat processed broccoli significantly increased the formation of sulforaphane. Crown Copyright © 2012. Published by Elsevier Ltd. All rights reserved.

  13. Flaviviral methyltransferase/RNA interaction: structural basis for enzyme inhibition.

    Science.gov (United States)

    Milani, Mario; Mastrangelo, Eloise; Bollati, Michela; Selisko, Barbara; Decroly, Etienne; Bouvet, Mickaël; Canard, Bruno; Bolognesi, Martino

    2009-07-01

    Flaviviruses are the causative agents of severe diseases such as Dengue or Yellow fever. The replicative machinery used by the virus is based on few enzymes including a methyltransferase, located in the N-terminal domain of the NS5 protein. Flaviviral methyltransferases are involved in the last two steps of the mRNA capping process, transferring a methyl group from S-adenosyl-L-methionine onto the N7 position of the cap guanine (guanine-N7 methyltransferase) and the ribose 2'O position of the first nucleotide following the cap guanine (nucleoside-2'O methyltransferase). The RNA capping process is crucial for mRNA stability, protein synthesis and virus replication. Such an essential function makes methyltransferases attractive targets for the design of antiviral drugs. In this context, starting from the crystal structure of Wesselsbron flavivirus methyltransferase, we elaborated a mechanistic model describing protein/RNA interaction during N7 methyl transfer. Next we used an in silico docking procedure to identify commercially available compounds that would display high affinity for the methyltransferase active site. The best candidates selected were tested in vitro to assay their effective inhibition on 2'O and N7 methyltransferase activities on Wesselsbron and Dengue virus (Dv) methyltransferases. The results of such combined computational and experimental screening approach led to the identification of a high-potency inhibitor.

  14. The effects of UV radiation during the vegetative period on antioxidant compounds and postharvest quality of broccoli (Brassica oleracea L.).

    Science.gov (United States)

    Topcu, Yasin; Dogan, Adem; Kasimoglu, Zehra; Sahin-Nadeem, Hilal; Polat, Ersin; Erkan, Mustafa

    2015-08-01

    In this study, the effects of supplementary UV radiation during the vegetative period on antioxidant compounds, antioxidant activity and postharvest quality of broccoli heads during long term storage was studied. The broccolis were grown under three different doses of supplementary UV radiation (2.2, 8.8 and 16.4 kJ/m(2)/day) in a soilless system in a glasshouse. Harvested broccoli heads were stored at 0 °C in modified atmosphere packaging for 60 days. The supplementary UV radiation (280-315 nm) during the vegetative period significantly decreased total carotenoid, the chlorophyll a and chlorophyll b content but increased the ascorbic acid, total phenolic and flavonoid contents of broccolis. All supplementary UV treatments slightly reduced the antioxidant activity of the broccolis, however, no remarkable change was observed between 2.2 and 8.8 kJ/m(2) radiation levels. The sinigrin and glucotropaeolin contents of the broccolis were substantially increased by UV treatments. The prolonged storage period resulted in decreased ascorbic acid, total phenolic and flavonoid contents, as well as antioxidant activity. Discoloration of the heads, due to decreased chlorophyll and carotenoid contents, was also observed with prolonged storage duration. Glucosinolates levels showed an increasing tendency till the 45th day of storage, and then their levels started to decline. The weight loss of broccoli heads during storage progressively increased with storage time in all treatments. Total soluble solids, solids content and titratable acidity decreased continuously during storage. Titratable acidity was not affected by UV radiation doses during the storage time whereas soluble solids and solids content (dry matter) were significantly affected by UV doses. Supplementary UV radiation increased the lightness (L*) and chroma (C*) values of the broccoli heads. Pre-harvest UV radiation during vegetative period seems to be a promising tool for increasing the beneficial health components

  15. Protective effect of broccoli and ferulic acid on imidacloprid-induced ...

    African Journals Online (AJOL)

    Imidacloprid may induce oxidative stress leading to generate free radicals and alternate oxygen free radical scavenging enzyme system. This study aims to investigate the hepatoprotective effect of broccoli water extract and ferulic acid on imidacloprid induced oxidative stress and DNA damage in male albino rats. Rats were ...

  16. Evaluation of the free volume theory to predict moisture transport and quality changes during broccoli drying

    NARCIS (Netherlands)

    Jin, X.; Sman, van der R.G.M.; Boxtel, van A.J.B.

    2010-01-01

    Abstract: Moisture diffusion in porous broccoli florets and stalks is modeled by using the free volume and Maxwell-Eucken theories. These theories are based on the mobility of water and show the variation of the effective diffusion coefficient for a wide range of temperatures and moisture content of

  17. Evaluation of the Free Volume Theory to Predict Moisture Transport and Quality Changes During Broccoli Drying

    NARCIS (Netherlands)

    Jin, X.; Sman, van der R.G.M.; Boxtel, van A.J.B.

    2011-01-01

    Moisture diffusion in porous broccoli florets and stalks is modeled using the free volume and Maxwell-Eucken theories. These theories are based on the mobility of water and concern the variation of the effective diffusion coefficient for a wide range of temperature and moisture content during

  18. Modelling the effect of pH on the colour degradation of blanched broccoli

    NARCIS (Netherlands)

    Tijskens, L.M.M.; Barringer, S.A.; Biekman, E.S.A.

    2001-01-01

    In ready-to-eat salads, a low pH is often applied in the salad dressing. The effect of pH on the colour change of blanched-frozen-and-thawed broccoli was studied. Different acids were used to achieve the pH values in buffer solutions. A model on chlorophyll degradation in fruits and vegetables was

  19. Modelling the change in colour in broccoli and green beans during blanching

    NARCIS (Netherlands)

    Tijskens, L.M.M.; Schijvens, E.P.H.M.; Biekman, E.S.A.

    2001-01-01

    The green colour of vegetables changes considerably during heat treatments like blanching. Green beans from two different countries and growing seasons, and the stems and florets of broccoli were heat-treated from 40 up to 96 °C. The colour was monitored with the CIE-Lab system. Expressing the green

  20. Biotechnological advancement in genetic improvement of broccoli (Brassica oleracea L. var. italica), an important vegetable crop.

    Science.gov (United States)

    Kumar, Pankaj; Srivastava, Dinesh Kumar

    2016-07-01

    With the advent of molecular biotechnology, plant genetic engineering techniques have opened an avenue for the genetic improvement of important vegetable crops. Vegetable crop productivity and quality are seriously affected by various biotic and abiotic stresses which destabilize rural economies in many countries. Moreover, absence of proper post-harvest storage and processing facilities leads to qualitative and quantitative losses. In the past four decades, conventional breeding has significantly contributed to the improvement of vegetable yields, quality, post-harvest life, and resistance to biotic and abiotic stresses. However, there are many constraints in conventional breeding, which can only be overcome by advancements made in modern biology. Broccoli (Brassica oleracea L. var. italica) is an important vegetable crop, of the family Brassicaceae; however, various biotic and abiotic stresses cause enormous crop yield losses during the commercial cultivation of broccoli. Thus, genetic engineering can be used as a tool to add specific characteristics to existing cultivars. However, a pre-requisite for transferring genes into plants is the availability of efficient regeneration and transformation techniques. Recent advances in plant genetic engineering provide an opportunity to improve broccoli in many aspects. The goal of this review is to summarize genetic transformation studies on broccoli to draw the attention of researchers and scientists for its further genetic advancement.

  1. Broccoli Cultivar Performance under Organic and Conventional Management Systems and Implications for Crop Improvement

    NARCIS (Netherlands)

    Renaud, E.N.C.; Lammerts Van Bueren, E.; Caldas Paulo, M.J.; Eeuwijk, van F.A.; Juvik, J.A.; Hutton, M.G.; Myers, J.R.

    2014-01-01

    To determine if present commercial broccoli cultivars meet the diverse needs of organic management systems, such as adaptation to low N input, mechanical weed management, and no chemical pesticide use, and to propose the selection environments for crop improvement for organic production, we compared

  2. Moisture Sorption Isotherms of Broccoli Interpreted with the Flory-Huggins Free Volume Theory

    NARCIS (Netherlands)

    Jin, X.; Sman, van der R.G.M.; Maanen, van J.F.C.; Deventer, van H.C.; Straten, van G.; Boom, R.M.; Boxtel, van A.J.B.

    2014-01-01

    In this work, the Flory Huggins Free Volume theory is used to interpret the sorption isotherms of broccoli from its composition and using physical properties of the components. This theory considers the mixing properties of water, biopolymers and solutes and has the potential to describe the

  3. Impact of different drying trajectories on degradation of nutritional compounds in broccoli (Brassica oleracea var. italica)

    NARCIS (Netherlands)

    Jin, X.; Oliviero, T.; Sman, van der R.G.M.; Verkerk, R.; Dekker, M.; Boxtel, van A.J.B.

    2014-01-01

    This work concerns the degradation of the nutritional compounds glucoraphanin (GR) and vitamin C (Vc), and the inactivation of the enzyme myrosinase (MYR) in broccoli (Brassica oleracea var. italica) during drying with air temperatures in the range of 30e60 C. Dynamic optimization is applied to find

  4. Growth and yield responses of broccoli cultivars to different rates of nitrogen at western Chitwan, Nepal

    DEFF Research Database (Denmark)

    Giri, Raj Kumar; Sharma, Moha Datta; Shakya, Santa Man

    2013-01-01

    A field experiment was conducted with the objective to determine the optimum rate of nitrogen (N) fertilizer for effective growth and yield of two varieties of broccoli in southern plain of Nepal. The experiment was laid out with two-factorial completely random block design (RCBD) comprising two...

  5. Changes of glucosinolates in mixed fresh-cut broccoli and cauliflower florets in modified atmosphere packaging.

    Science.gov (United States)

    Schreiner, M; Peters, P; Krumbein, A

    2007-10-01

    Glucosinolates of broccoli and cauliflower florets were assessed to determine the effect of modified-atmosphere packaging on postharvest glucosinolate dynamics in mixed florets of Brassica vegetables. Mixed-packaged broccoli and cauliflower florets stored in food trays sealed with 2 different microperforated biaxial-oriented polypropylene films for up to 7 d at 8 degrees C were analyzed. Both applied modified atmospheres (1% O(2)+ 21% CO(2); 8% O(2)+ 14% CO(2)) maintained aliphatic glucosinolates in cauliflower florets, whereas in broccoli florets, the aliphatic glucosinolate concentration decreased slightly in each modified atmosphere. In addition, total indole glucosinolate concentration for both broccoli and cauliflower florets was maintained, and even increased in cauliflower florets at 1% O(2)+ 21% CO(2) due to rising neoglucobrassicin concentration. Thus, to simultaneously maintain glucosinolates and external appearance as well as to prevent off-odor, a modified atmosphere of 1% O(2)+ 21% CO(2) provides a suitable environment for storage of this Brassica floret medley for up to 7 d at 8 degrees C.

  6. Multi-phase extraction of glycoraphanin from broccoli using aminium ionic liquid-based silica.

    Science.gov (United States)

    Tian, Minglei; Bi, Wentao; Row, Kyung Ho

    2013-01-01

    Glucosinolates, a class of phytochemicals found in broccoli, have attracted recent interest due to the potential health benefits associated with their dietary intake. Glucoraphanin, the most common glucosinolate in broccoli can be converted to a known cancer chemopreventive agent. Multi-phase extraction in solid-phase extraction cartridges was developed to simultaneously extract and separate this compound. Multi-phase extraction with functionalised ionic liquid-based silica as a sorbent was used to simultaneously extract and separate glucoraphanin from broccoli. The sorbent and broccoli sample were packed into a single cartridge, and a fixed volume of water was then used to extract and remove the target compound from the sample to the sorbent over 15 repetitions. The sorbent was then washed with n-hexane to remove any interference and the target compound was eluted with water-1% acetic acid (vol.). Under the optimised condition, 0.038 mg/g of glucoraphanin was obtained by multi-phase extraction with 0.2 g of sorbent. The adsorption isotherm allowed investigation of the interactions between the sorbent and target compound and provided evidence for the accuracy of this method. The low deviation error, small amount of solvents required, highly selective separation and stability of the method justify further research. Copyright © 2012 John Wiley & Sons, Ltd.

  7. Analysis and Antioxidant Activity of Extracts from Broccoli (Brassica oleracea L.) Sprouts.

    Science.gov (United States)

    Jang, Hae Won; Moon, Joon-Kwan; Shibamoto, Takayuki

    2015-02-04

    Samples prepared from fresh broccoli (Brassica oleracea L.) sprouts by water distillation or freeze-drying were examined for antioxidant activity using three assays. All samples exhibited dose-dependent antioxidant activity. The antioxidant activity ranged from 74.48 ± 0.46% (less volatile sample) to 93.2 ± 0.2% (dichloromethane extract sample) at the level of 500 μg/mL. Both dichloromethane extract samples from a water distillate of broccoli sprouts and freeze-dried broccoli sprouts showed potent antioxidant activity, which was comparable to that of BHT. Among the 43 compounds positively identified by gas chromatography-mass spectrometry, 5-methylthiopentylnitrile (31.64 μg/g) was found in the greatest concentration, followed by 4-methylthiobutylisothiocyanate (14.55 μg/g), 4-methylthiobutylnitrile (10.63 μg/g), 3-methylthiopropylisothiocyanate (3.00 μg/g), and 4-methylpentylisothiocyanate (2.48 μg/g). These isothiocyanates are known to possess antioxidant properties. Possible phenolic antioxidants found are 4-(1-methylpropyl)phenol (0.012 μg/g), 4-methylphenol (0.159 μg/g), and 2-methoxy-4-vinylphenol (0.009 μg/g). The present study demonstrates that broccoli sprouts are a good source of natural antioxidants.

  8. Effects of application timing of saline irrigation water on broccoli production and quality

    Science.gov (United States)

    Irrigation with moderately saline water is a necessity in many semi-arid areas of the Mediterranean Basin, and requires adequate irrigation management strategies. Broccoli (Brassica oleracea var. italica), a crop moderately tolerant to salinity stress, was used to evaluate the effects of the applica...

  9. Impact of thermal processing on sulforaphane yield from broccoli ( Brassica oleracea L. ssp. italica).

    Science.gov (United States)

    Wang, Grace C; Farnham, Mark; Jeffery, Elizabeth H

    2012-07-11

    In broccoli, sulforaphane forms when the glucosinolate glucoraphanin is hydrolyzed by the endogenous plant thiohydrolase myrosinase. A myrosinase cofactor directs hydrolysis away from the formation of bioactive sulforaphane and toward an inactive product, sulforaphane nitrile. The cofactor is more heat sensitive than myrosinase, presenting an opportunity to preferentially direct hydrolysis toward sulforaphane formation through regulation of thermal processing. Four broccoli cultivars were microwave heated, boiled, or steamed for various lengths of time. Production of nitrile during hydrolysis of unheated broccoli varied among cultivars from 91 to 52% of hydrolysis products (Pinnacle > Marathon > Patriot > Brigadier). Boiling and microwave heating caused an initial loss of nitrile, with a concomitant increase in sulforaphane, followed by loss of sulforaphane, all within 1 min. In contrast, steaming enhanced sulforaphane yield between 1.0 and 3.0 min in all but Brigadier. These data are proof of concept that steaming for 1.0-3.0 min provides less nitrile and more sulforaphane yield from a broccoli meal.

  10. Broccoli glucosinolate degradation is reduced performing thermal treatment in binary systems with other food ingredients

    NARCIS (Netherlands)

    Giambanelli, E.; Verkerk, R.; Fogliano, V.; Capuano, E.; Antuono, D' L.F.; Oliviero, T.

    2015-01-01

    Glucosinolate (GL) stability has been widely studied in different Brassica species. However, the matrix effect determined by the presence of other ingredients occurred in many broccoli-based traditional recipes may affect GL thermal degradation. In this study, the matrix effect on GL thermal

  11. Doubled haploid inbred lines USVL048 and USVL131 of heading broccoli

    Science.gov (United States)

    Two inbred lines of heading broccoli (Brassica oleracea L. var. italica), designated USVL048 and USVL131, were released by the Agricultural Research Service of the U.S. Department of Agriculture in 2012. Both of the released lines are doubled haploids originally derived from another culture. As do...

  12. Establishing an eastern broccoli industry: where are we and where do we need to go?

    Science.gov (United States)

    A Coordinated Agricultural Project (CAP) entitled “Establishing an Eastern Broccoli Industry” has been underway since the fall of 2010 and funded under the USDA Specialty Crop Research Initiative (SCRI), which was established as part of the 2008 Farm Bill. This project has brought together research...

  13. Metabolomic assessment reveals a stimulatory effect of calcium treatment on glucosinolates contents in broccoli microgreen

    Science.gov (United States)

    Preharvest calcium application has been shown to increase broccoli microgreen yield and extend shelf life. Here we investigated the effect of calcium application on its metabolome using ultra high-performance liquid chromatography (UHPLC) tandem with mass spectrometry (HRMS). The data collected were...

  14. Effects of pre- and postharvest calcium treatments on shelf life and postharvest quality of broccoli microgreens

    Science.gov (United States)

    Microgreens’ extremely short shelf life limits their commercial usage. The objective of this study is to compare the effect of pre- and post- harvest treatments using different forms of calcium on the postharvest quality and shelf-life of broccoli microgreens. Preharvest spray with calcium lactate, ...

  15. Koeling van bloemkool en broccoli op de veiling "De Tuinbouw" te Grootebroek

    NARCIS (Netherlands)

    Damen, P.M.M.; Hendriks, A.G.M.

    1980-01-01

    In een proef op de veiling "De Tuinbouw" in Grootebroek is zowel bloemkool als broccoli opgeslagen gedurende respectievelijk een en drie dagen bij 2 graden C en 95% r.v. Na koeling zijn de kolen geplaatst in een donkere ruimte met een temperatuur van 18 graden C en een r.v. van 90%. Beoordeeld is op

  16. Using regional broccoli trial data to select experimental hybrids for input into advanced yield trials

    Science.gov (United States)

    A large amount of phenotypic trait data are being generated in regional trials that are implemented as part of the Specialty Crop Research Initiative (SCRI) project entitled “Establishing an Eastern Broccoli Industry”. These data are used to identify the best entries in the trials for inclusion in ...

  17. Purification and characterization of broccoli (Brassica oleracea var. italica) myrosinase (β-thioglucosidase glucohydrolase).

    Science.gov (United States)

    Mahn, Andrea; Angulo, Alejandro; Cabañas, Fernanda

    2014-12-03

    Myrosinase (β-thioglucosidase glucohydrolase, EC 3.2.1.147) from broccoli (Brassica oleracea var. italica) was purified by ammonium sulfate precipitation followed by concanavalin A affinity chromatography, with an intermediate dialysis step, resulting in 88% recovery and 1318-fold purification. These are the highest values reported for the purification of any myrosinase. The subunits of broccoli myrosinase have a molecular mass of 50-55 kDa. The native molecular mass of myrosinase was 157 kDa, and accordingly, it is composed of three subunits. The maximum activity was observed at 40 °C and at pH below 5.0. Kinetic assays demonstrated that broccoli myrosinase is subjected to substrate (sinigrin) inhibition. The Michaelis-Menten model, considering substrate inhibition, gave Vmax equal to 0.246 μmol min(-1), Km equal to 0.086 mM, and K(I) equal to 0.368 mM. This is the first study about purification and characterization of broccoli myrosinase.

  18. BROCCOLI: Software for Fast fMRI Analysis on Many-Core CPUs and GPUs

    Directory of Open Access Journals (Sweden)

    Anders eEklund

    2014-03-01

    Full Text Available Analysis of functional magnetic resonance imaging (fMRI data is becoming ever more computationally demanding as temporal and spatial resolutions improve, and large, publicly available data sets proliferate. Moreover, methodological improvements in the neuroimaging pipeline, such as non-linear spatial normalization, non-parametric permutation tests and Bayesian Markov Chain Monte Carlo approaches, can dramatically increase the computational burden. Despite these challenges, there do not yet exist any fMRI software packages which leverage inexpensive and powerful graphics processing units (GPUs to perform these analyses. Here, we therefore present BROCCOLI, a free software package written in OpenCL (Open Computing Language that can be used for parallel analysis of fMRI data on a large variety of hardware configurations. BROCCOLI has, for example, been tested with an Intel CPU, an Nvidia GPU and an AMD GPU. These tests show that parallel processing of fMRI data can lead to significantly faster analysis pipelines. This speedup can be achieved on relatively standard hardware, but further, dramatic speed improvements require only a modest investment in GPU hardware. BROCCOLI (running on a GPU can perform non-linear spatial normalization to a 1 mm3 brain template in 4-6 seconds, and run a second level permutation test with 10,000 permutations in about a minute. These non-parametric tests are generally more robust than their parametric counterparts, and can also enable more sophisticated analyses by estimating complicated null distributions. Additionally, BROCCOLI includes support for Bayesian first-level fMRI analysis using a Gibbs sampler. The new software is freely available under GNU GPL3 and can be downloaded from github (https://github.com/wanderine/BROCCOLI/.

  19. Antioxidative and antitumor properties of in vitro-cultivated broccoli (Brassica oleracea var. italica).

    Science.gov (United States)

    Cakar, Jasmina; Parić, Adisa; Maksimović, Milka; Bajrović, Kasim

    2012-02-01

    Broccoli [Brassica oleracea L. var. italica Plenck. (Brassicaceae)] contains substantial quantities of bioactive compounds, which are good free radical scavengers and thus might have strong antitumor properties. Enhancing production of plant secondary metabolites could be obtained with phytohormones that have significant effects on the metabolism of secondary metabolites. In that manner, in vitro culture presents good model for manipulation with plant tissues in order to affect secondary metabolite production and thus enhance bioactive properties of plants. Estimation of the antioxidative and antitumor properties of broccoli cultivated in different in vitro conditions. In vitro germinated and cultivated broccoli seedlings, as well as spontaneously developed calli, were subjected to Soxhlet extraction. Antioxidative activity of the herbal extracts was determined using 1,1-diphenyl-2-picrylhydrazyl (DPPH(•)) radical method. Antitumor properties of the extracts were determined using crown-gall tumor inhibition (potato disc) assay. Three, 10, 20, and 30 days old broccoli seedlings, cultivated in vitro on three different Murashige-Skoog media, two types of callus, and seedlings from sterile filter paper were used for extraction. In total, 15 aqueous extracts were tested for antioxidative and antitumor potential. Three day-old seedlings showed the highest antioxidative activity. Eleven out of 15 aqueous extracts demonstrated above 50% of crown-gall tumor inhibition in comparison with the control. Tumor inhibition was in association with types and concentrations of phytohormones presented in growing media. It is demonstrated that phytohormones in plant-growing media could affect the bioactive properties of broccoli either through increasing or decreasing their antioxidative and antitumor potential.

  20. Mutations in cauliflower and sprout broccoli grown from seeds flown in space

    Science.gov (United States)

    Wu, Hong; Huang, Congli; Zhang, Keping; Sun, Yeqing

    2010-11-01

    Cauliflower and sprout broccoli are widely planted vegetables particularly in Fujian Province, China. To study the mutation in these two types of vegetables induced from spaceflight, we flew the seeds on the 20th Chinese recoverable satellite which orbited the Earth for 18 days. After returning to the Earth, the cauliflower seeds were planted for two generations and the sprout broccoli seeds for one generation at the Xiamen Agriculture Research Institute. Of the 12 cauliflowers planted for the first generation, two showed significant phenotypical changes in both the size of the plant and the weight of the flower head. In addition, most of the space flown plants were found to be resistant to the black rot attack in the field. Cauliflowers planted for the second generation from the seeds in one of the two plants that displayed phenotypical changes in the first generation showed similar mutations. For the first generation of sprout broccoli, the rate of emergence from the flown seeds was lower than that of the control by 30%. No significant changes in the phenotype between the sprout broccolis planted from the flown seeds and the control were observed except one of the mutated sprout broccolis showed a change in the appearance in the lesser bud of the chief flower head. Results of the study demonstrated that DNA damages in some of the genes may have occurred in the seeds flown in space, and some of the changes in the genes may have inherited from the first to the second generation. The improved resistance to the black rot attack and increased size of the flower head are apparently beneficial.

  1. Distinction between the Cfr Methyltransferase Conferring Antibiotic Resistance and the Housekeeping RlmN Methyltransferase

    DEFF Research Database (Denmark)

    Atkinson, Gemma C; Hansen, Lykke H; Tenson, Tanel

    2013-01-01

    The cfr gene encodes the Cfr methyltransferase that primarily methylates C-8 in A2503 of 23S rRNA in the peptidyl transferase region of bacterial ribosomes. The methylation provides resistance to six classes of antibiotics of clinical and veterinary importance. The rlmN gene encodes the RlmN meth...

  2. Chemopreventive glucosinolate accumulation in various broccoli and collard tissues: Microfluidic-based targeted transcriptomics for by-product valorization.

    Science.gov (United States)

    Lee, Young-Sang; Ku, Kang-Mo; Becker, Talon M; Juvik, John A

    2017-01-01

    Floret, leaf, and root tissues were harvested from broccoli and collard cultivars and extracted to determine their glucosinolate and hydrolysis product profiles using high performance liquid chromatography and gas chromotography. Quinone reductase inducing bioactivity, an estimate of anti-cancer chemopreventive potential, of the extracts was measured using a hepa1c1c7 murine cell line. Extracts from root tissues were significantly different from other tissues and contained high levels of gluconasturtiin and glucoerucin. Targeted gene expression analysis on glucosinolate biosynthesis revealed that broccoli root tissue has elevated gene expression of AOP2 and low expression of FMOGS-OX homologs, essentially the opposite of what was observed in broccoli florets, which accumulated high levels of glucoraphanin. Broccoli floret tissue has significantly higher nitrile formation (%) and epithionitrile specifier protein gene expression than other tissues. This study provides basic information of the glucosinolate metabolome and transcriptome for various tissues of Brassica oleracea that maybe utilized as potential byproducts for the nutraceutical market.

  3. Effect of water content and temperature on inactivation kinetics of myrosinase in broccoli (Brassica oleracea var. italica)

    NARCIS (Netherlands)

    Oliviero, T.; Verkerk, R.; Boekel, van M.A.J.S.; Dekker, M.

    2014-01-01

    Broccoli belongs to the Brassicaceae plant family consisting of widely eaten vegetables containing high concentrations of glucosinolates. Enzymatic hydrolysis of glucosinolates by endogenous myrosinase (MYR) can form isothiocyanates with health-promoting activities. The effect of water content (WC)

  4. Thermal stability of L-ascorbic acid and ascorbic acid oxidase in broccoli (Brassica oleracea var. italica).

    Science.gov (United States)

    Munyaka, Ann Wambui; Makule, Edna Edward; Oey, Indrawati; Van Loey, Ann; Hendrickx, Marc

    2010-05-01

    The thermal stability of vitamin C (including l-ascorbic acid [l-AA] and dehydroascorbic acid [DHAA]) in crushed broccoli was evaluated in the temperature range of 30 to 90 degrees C whereas that of ascorbic acid oxidase (AAO) was evaluated in the temperature range of 20 to 95 degrees C. Thermal treatments (for 15 min) of crushed broccoli at 30 to 60 degrees C resulted in conversion of l-AA to DHAA whereas treatments at 70 to 90 degrees C retained vitamin C as l-AA. These observations indicated that enzymes (for example, AAO) could play a major role in the initial phase (that is, oxidation of l-AA to DHAA) of vitamin C degradation in broccoli. Consequently, a study to evaluate the temperature-time conditions that could result in AAO inactivation in broccoli was carried out. In this study, higher AAO activity was observed in broccoli florets than stalks. During thermal treatments for 10 min, AAO in broccoli florets and stalks was stable until around 50 degrees C. A 10-min thermal treatment at 80 degrees C almost completely inactivated AAO in broccoli. AAO inactivation followed 1st order kinetics in the temperature range of 55 to 65 degrees C. Based on this study, a thermal treatment above 70 degrees C is recommended for crushed vegetable products to prevent oxidation of l-AA to DHAA, the onset of vitamin C degradation. The results reported in this study are applicable for both domestic and industrial processing of vegetables into products such as juices, soups, and purees. In this report, we have demonstrated that processing crushed broccoli in a temperature range of 30 to 60 degrees C could result in the conversion of l-ascorbic acid to dehydroascorbic (DHAA), a very important reaction in regard to vitamin C degradation because DHAA could be easily converted to other compounds that do not have the biological activity of vitamin C.

  5. RNA-seq analysis of transcriptome and glucosinolate metabolism in seeds and sprouts of broccoli (Brassica oleracea var. italic).

    Science.gov (United States)

    Gao, Jinjun; Yu, Xinxin; Ma, Fengming; Li, Jing

    2014-01-01

    Broccoli (Brassica oleracea var. italica), a member of Cruciferae, is an important vegetable containing high concentration of various nutritive and functional molecules especially the anticarcinogenic glucosinolates. The sprouts of broccoli contain 10-100 times higher level of glucoraphanin, the main contributor of the anticarcinogenesis, than the edible florets. Despite the broccoli sprouts' functional importance, currently available genetic and genomic tools for their studies are very limited, which greatly restricts the development of this functionally important vegetable. A total of ∼85 million 251 bp reads were obtained. After de novo assembly and searching the assembled transcripts against the Arabidopsis thaliana and NCBI nr databases, 19,441 top-hit transcripts were clustered as unigenes with an average length of 2,133 bp. These unigenes were classified according to their putative functional categories. Cluster analysis of total unigenes with similar expression patterns and differentially expressed unigenes among different tissues, as well as transcription factor analysis were performed. We identified 25 putative glucosinolate metabolism genes sharing 62.04-89.72% nucleotide sequence identity with the Arabidopsis orthologs. This established a broccoli glucosinolate metabolic pathway with high colinearity to Arabidopsis. Many of the biosynthetic and degradation genes showed higher expression after germination than in seeds; especially the expression of the myrosinase TGG2 was 20-130 times higher. These results along with the previous reports about these genes' studies in Arabidopsis and the glucosinolate concentration in broccoli sprouts indicate the breakdown products of glucosinolates may play important roles in the stage of broccoli seed germination and sprout development. Our study provides the largest genetic resource of broccoli to date. These data will pave the way for further studies and genetic engineering of broccoli sprouts and will also provide

  6. The potential role of nutritional genomics tools in validating high health foods for cancer control: broccoli as example.

    Science.gov (United States)

    Ferguson, Lynnette R; Schlothauer, Ralf C

    2012-01-01

    Nutritional genomics reflects gene/nutrient interactions, utilising high-throughput genomic tools in nutrition research. The field also considers the contribution of individual genotypes to wellness and the risk of chronic disease (nutrigenetics), and how such genetic predisposition may be modified by appropriate diets. For example, high consumption of brassicaceous vegetables, including broccoli, has regularly associated with low cancer risk. Bioactive chemicals in broccoli include glucosinolates, plant pigments including kaempferol, quercetin, lutein and carotenoids, various vitamins, minerals and amino acids. Cancer prevention is hypothesised to act through various mechanisms including modulation of xenobiotic metabolising enzymes, NF-E2 p45-related factor-2 (Nrf2)-mediated stress-response mechanisms, and protection against genomic instability. Broccoli and broccoli extracts also regulate the progression of cancer through anti-inflammatory effects, effects on signal transduction, epigenetic effects and modulation of the colonic microflora. Human intervention studies with broccoli and related foods, using standard biomarker methodologies, reveal part of a complex picture. Nutrigenomic approaches, especially transcriptomics, enable simultaneous study of various signalling pathways and networks. Phenotypic, genetic and/or metabolic stratification may identify individuals most likely to respond positively to foods or diets. Jointly, these technologies can provide proof of human efficacy, and may be essential to ensure effective market transfer and uptake of broccoli and related foods. Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  7. Vitamin C, Phenolic Compounds and Antioxidant Capacity of Broccoli Florets Grown under Different Nitrogen Treatments Combined with Selenium

    Directory of Open Access Journals (Sweden)

    Peñas Elena

    2018-06-01

    Full Text Available Broccoli consumption is rising worldwide and fertilization is a tool to increase its production. However, little is known about the effect of mineral supplementation to the soil on the bioactive compounds. Therefore, the aim of this investigation was to analyze the content of vitamin C, total phenolic compounds and the antioxidant capacity of broccoli florets cultivated under different nitrogen (N conditions in combination with selenium (IV and VI. Greenhouse experiments were conducted in broccoli grown in commercial soil treated with different N sources [(NH42SO4, NaNO3, NH4NO3 or CO(NH22 at 160 kg N/ha]. In addition, selenium (Se salts [Na2SeO3 (Se IV or Na2SeO4 (Se VI at 10 and 20 kg Se/ha] were applied. There were no evidences of the influence of N treatment on vitamin C content whilst Se (IV or VI uptake led to a significant reduction of this vitamin in broccoli florets, irrespective of the N source. In contrast, total phenolics content and antioxidant capacity underwent a significant increment under N application. However, their combination with Se salts modified total phenolic content and antioxidant capacities in broccoli florets depending on N source and Se doses. Among all the experimental trials, application of NH4NO3 combined with 10 g Se (IV/ha was the elective treatment strategy to produce broccoli florets with higher content of phenolic compounds and antioxidant capacity and, therefore, enhanced functionality.

  8. HPLC Separation of Sulforaphane Enantiomers in Broccoli and Its Sprouts by Transformation into Diastereoisomers Using Derivatization with (S)-Leucine.

    Science.gov (United States)

    Okada, Makiko; Yamamoto, Atsushi; Aizawa, Sen-Ichi; Taga, Atsushi; Terashima, Hiroyuki; Kodama, Shuji

    2017-01-11

    Racemic sulforaphane, which was derivatized with (S)-leucine (l-leucine), was resolved by reversed phase HPLC with UV detection. The optimum mobile phase conditions were found to be 10 mM citric acid (pH 2.8) containing 22% methanol at 35 °C using detection at 254 nm. Sulforaphane enantiomers in florets and stems of five brands of broccoli and leaves and stems of three brands of broccoli sprouts were analyzed by the proposed HPLC method. Both sulforaphane enantiomers were detected in all of the samples. The S/R ratios of sulforaphane in broccoli samples were 1.5-2.6/97.4-98.5% for florets and 5.0-12.1/87.9-95.0% for stems. The S/R ratios in broccoli sprout samples were higher than those in broccoli samples and were found to be 8.3-19.7/80.3-91.7% for leaves and 37.0-41.8/58.2-63.0% for stems. (S)-Sulforaphane detected in the broccoli and its sprout samples was positively identified by separately using an HPLC with a chiral column (Chiralpak AD-RH) and mass spectrometry.

  9. Consumption of selenium-enriched broccoli increases cytokine production in human peripheral blood mononuclear cells stimulated ex vivo, a preliminary human intervention study.

    Science.gov (United States)

    Bentley-Hewitt, Kerry L; Chen, Ronan K-Y; Lill, Ross E; Hedderley, Duncan I; Herath, Thanuja D; Matich, Adam J; McKenzie, Marian J

    2014-12-01

    Selenium (Se) is a micronutrient essential for human health, including immune function. Previous research indicates that Se supplementation may cause a shift from T helper (Th)1- to Th2-type immune responses. We aim to test the potential health promoting effects of Se-enriched broccoli. In a human trial, 18 participants consumed control broccoli daily for 3 days. After a 3-day wash-out period, the participants were provided with Se-enriched broccoli containing 200 μg of Se per serving for 3 days. Plasma and peripheral blood mononuclear cell (PBMC) samples were collected at the start and end of each broccoli feeding period for analysis of total Se and measurement of cytokine production from PBMC stimulated with antigens ex vivo. Plasma Se content remained consistent throughout the control broccoli feeding period and the baseline of the Se-enriched broccoli period (1.22 μmol/L) and then significantly increased following 3 days of Se-enriched broccoli feeding. Interleukin (IL-2, IL-4, IL-5, IL-13, and IL-22) production from PBMC significantly increased after 3 days of Se-enriched broccoli feeding compared with baseline. This study indicates that consumption of Se-enriched broccoli may increase immune responses toward a range of immune challenges. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  10. Influence of Interval Between Postharvest Lettuce Residue Management and Subsequent Seeding of Broccoli on Cabbage Maggot (Diptera: Anthomyiidae) Infestation on Broccoli.

    Science.gov (United States)

    Joseph, Shimat V; Godfrey, Larry D; Bettiga, Christopher

    2017-10-01

    Larval stages of cabbage maggot, Delia radicum (L.) (Diptera: Anthomyiidae), attack the roots of Brassica crops and cause severe economic damage. In the Salinas Valley of California, Brassica crops are often planted after successive lettuce (Lactuca sativa L.) crops. The interval between postharvest soil incorporation of lettuce residue and the subsequent Brassica crop can be as short as 7 d, which could influence D. radicum infestation on broccoli (Brassica oleracea var. italica Plenck). In 2014 and 2015, the effect of intervals between crops (IBC) on D. radicum infestation was evaluated. The treatments were 7, 20, 33, and 48 d IBC, and NL (no lettuce), 7, 21, 36, and 49 d IBC in 2014 and 2015, respectively. Insect counts and feeding damage on broccoli was assessed during 3-6 wk after planting. Adult Delia fly captures were significantly greater at 7 d than 36-49 d IBC in both years. In both years, D. radicum eggs collected were significantly greater at 7 d than at 33 d or 36 d IBC plots. Larvae collected were significantly greater at 7 d IBC than all other treatments in 2014, but not in 2015. Similarly, severity of feeding injury was significantly greater in 7 d than 33 d or 48 d IBC in 2014, but not in 2015. In 2015, broccoli with no prior lettuce had significantly lower Delia flies and D. radicum egg densities than 7 d or 21 d IBC. The implication of these results as a cultural control tactic for D. radicum infestation is discussed. © The Author 2017. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  11. Proteomics and transcriptomics of broccoli subjected to exogenously supplied and transgenic senescence-induced cytokinin for amelioration of postharvest yellowing.

    Science.gov (United States)

    Liu, Mao-Sen; Li, Hui-Chun; Lai, Ying-Mi; Lo, Hsiao-Feng; Chen, Long-Fang O

    2013-11-20

    Previously, we investigated transgenic broccoli harboring senescence-associated-gene (SAG) promoter-triggered isopentenyltransferase (ipt), which encodes the key enzyme for cytokinin (CK) synthesis and mimics the action of exogenous supplied CK in delaying postharvest senescence of broccoli. Here, we used proteomics and transcriptomics to compare the mechanisms of ipt-transgenic and N(6)-benzylaminopurine (BA) CK treatment of broccoli during postharvest storage. The 2 treatments conferred common and distinct mechanisms. BA treatment decreased the quantity of proteins involved in energy and carbohydrate metabolism and amino acid metabolism, and ipt-transgenic treatment increased that of stress-related proteins and molecular chaperones and slightly affected levels of carbohydrate metabolism proteins. Both treatments regulated genes involved in CK signaling, sugar transport, energy and carbohydrate metabolism, amino acid metabolism and lipid metabolism, although ipt-transgenic treatment to a lesser extent. BA treatment induced genes encoding molecular chaperones, whereas ipt-transgenic treatment induced stress-related genes for cellular protection during storage. Both BA and ipt-transgenic treatments acted antagonistically on ethylene functions. We propose a long-term acclimation of metabolism and protection systems with ipt-transgenic treatment of broccoli and short-term modulation of metabolism and establishment of a protection system with both BA and ipt-transgenic treatments in delaying senescence of broccoli florets. Transgenic broccoli harboring senescence-associated-gene (SAG) promoter-triggered isopentenyltransferase (ipt), which encodes the key enzyme for cytokinin (CK) synthesis and N(6)-benzylaminopurine (BA) CK treated broccoli both showed retardation of postharvest senescence during storage. The mechanisms underlying the two treatments were compared. The combination of proteomic and transcriptomic evidences revealed that the 2 treatments conferred common

  12. Influence of different blanching methods on colour, ascorbic acid and phenolics content of broccoli.

    Science.gov (United States)

    Severini, C; Giuliani, R; De Filippis, A; Derossi, A; De Pilli, T

    2016-01-01

    Lack of nutrients in cooking water, high energetic costs, high water consumption and recycling are some drawbacks of vegetable blanching. Those disadvantages could be bypassed using microwave blanching. Three blanching methods (microwave, boiling water and steaming) were compared in this study in order to determine their effects on some functional properties of broccoli. In addition, the thermal damage on broccoli colour was evaluated. The effectiveness of each blanching process was performed measuring the lost of peroxidase activity, that results more rapidly in microwaves and steam treatments (50 and 60 s respectively) than in boiling water treatment (120 s). The colour indexes did not allow to discriminate a significant difference among treatments. The increase of treatment time caused a vitamin C decrease in samples blanched by boiling water and steam; this trend was not observed in microwaved samples. The phenols content did not significantly vary depending both on type and on time of treatment.

  13. Extraction, chemical characterization and biological activity determination of broccoli health promoting compounds.

    Science.gov (United States)

    Ares, Ana M; Nozal, María J; Bernal, José

    2013-10-25

    Broccoli (Brassica oleracea L. var. Italica) contains substantial amount of health-promoting compounds such as vitamins, glucosinolates, phenolic compounds, and dietary essential minerals; thus, it benefits health beyond providing just basic nutrition, and consumption of broccoli has been increasing over the years. This review gives an overview on the extraction and separation techniques, as well as the biological activity of some of the above mentioned compounds which have been published in the period January 2008 to January 2013. The work has been distributed according to the different families of health promoting compounds discussing the extraction procedures and the analytical techniques employed for their characterization. Finally, information about the different biological activities of these compounds has been also provided. Copyright © 2013 Elsevier B.V. All rights reserved.

  14. Colorless chlorophyll catabolites in senescent florets of broccoli (Brassica oleracea var. italica).

    Science.gov (United States)

    Roiser, Matthias H; Müller, Thomas; Kräutler, Bernhard

    2015-02-11

    Typical postharvest storage of broccoli (Brassica oleracea var. italica) causes degreening of this common vegetable with visible loss of chlorophyll (Chl). As shown here, colorless Chl-catabolites are generated. In fresh extracts of degreening florets of broccoli, three colorless tetrapyrrolic Chl-catabolites accumulated and were detected by high performance liquid chromatography (HPLC): two "nonfluorescent" Chl-catabolites (NCCs), provisionally named Bo-NCC-1 and Bo-NCC-2, and a colorless 1,19-dioxobilin-type "nonfluorescent" Chl-catabolite (DNCC), named Bo-DNCC. Analysis by nuclear magnetic resonance spectroscopy and mass spectrometry of these three linear tetrapyrroles revealed their structures. In combination with a comparison of their HPL-chromatographic properties, this allowed their identification with three known catabolites from two other brassicacea, namely two NCCs from oil seed rape (Brassica napus) and a DNCC from degreened leaves of Arabidopsis thaliana.

  15. Ozone dose-response relationships for spring oilseed rape and broccoli

    Science.gov (United States)

    De Bock, Maarten; Op de Beeck, Maarten; De Temmerman, Ludwig; Guisez, Yves; Ceulemans, Reinhart; Vandermeiren, Karine

    2011-03-01

    Tropospheric ozone is an important air pollutant with known detrimental effects for several crops. Ozone effects on seed yield, oil percentage, oil yield and 1000 seed weight were examined for spring oilseed rape ( Brassica napus cv. Ability). For broccoli ( Brassica oleracea L. cv. Italica cv. Monaco) the effects on fresh marketable weight and total dry weight were studied. Current ozone levels were compared with an increase of 20 and 40 ppb during 8 h per day, over the entire growing season. Oilseed rape seed yield was negatively correlated with ozone dose indices calculated from emergence until harvest. This resulted in an R2 of 0.24 and 0.26 ( p broccoli the applied ozone doses had no effect on yield.

  16. Broccoli yield in response to top-dressing fertilization with green manure and biofertilizer

    Directory of Open Access Journals (Sweden)

    Gilberto Bernardo de Freitas

    2011-10-01

    Full Text Available The objective of this work was to evaluate the productive performance of broccoli under different top-dressing organic fertilizations. The experiment was conducted under protected cultivation, in a completely randomized design with four replications, with two plants per experimental unit. Broccoli seedlings were produced in a commercial substrate in styrofoam trays. The seedlings were transplanted to plastic pots containing 10.0 L of substrate made up of subsoil and organic compost at the ratio of 3:1 (v/v, respectively, which is equivalent to about 20.0 t ha-1 of organic compost at planting. After seedling establishment, the top-dressing fertilization treatments were applied: gliricidia biomass associated or not with liquid biofertilizer of cattle manure to the soil and bokashi. Two control treatments were established: one with mineral fertilization recommended for the crop and the other without top-dressing fertilization. The broccoli production was evaluated (commercial standard. Plants that received mineral fertilizer were more productive, however, they were not significantly different (p>0.05, by Dunnet test, from the plants fertilized with 2.5 t ha-1 gliricidiabiomass (dry mass associated with liquid biofertilizer (2.0 L m-2 applied to soil. Top-dressing fertilizations with only gliricidia, at 2.5 and 5.0 t ha-1 of biomass (dry mass, resulted in no significant increase in production of broccoli inflorescence. The use of bokashi in addition to gliricidia biomass and liquid biofertilizer reduced the efficiency of the fertilization compared with plants that received only gliricidia and liquid biofertilizer.

  17. Untargeted Metabolomics Reveals Predominant Alterations in Lipid Metabolism Following Light Exposure in Broccoli Sprouts

    Directory of Open Access Journals (Sweden)

    Mariateresa Maldini

    2015-06-01

    Full Text Available The consumption of vegetables belonging to the family Brassicaceae (e.g., broccoli and cauliflower is linked to a reduced incidence of cancer and cardiovascular diseases. The molecular composition of such plants is strongly affected by growing conditions. Here we developed an unbiased metabolomics approach to investigate the effect of light and dark exposure on the metabolome of broccoli sprouts and we applied such an approach to provide a bird’s-eye view of the overall metabolic response after light exposure. Broccoli seeds were germinated and grown hydroponically for five days in total darkness or with a light/dark photoperiod (16 h light/8 h dark cycle. We used an ultra-performance liquid-chromatography system coupled to an ion-mobility, time-of-flight mass spectrometer to profile the large array of metabolites present in the sprouts. Differences at the metabolite level between groups were analyzed using multivariate statistical analyses, including principal component analysis and correlation analysis. Altered metabolites were identified by searching publicly available and in-house databases. Metabolite pathway analyses were used to support the identification of subtle but significant changes among groups of related metabolites that may have gone unnoticed with conventional approaches. Besides the chlorophyll pathway, light exposure activated the biosynthesis and metabolism of sterol lipids, prenol lipids, and polyunsaturated lipids, which are essential for the photosynthetic machinery. Our results also revealed that light exposure increased the levels of polyketides, including flavonoids, and oxylipins, which play essential roles in the plant’s developmental processes and defense mechanism against herbivores. This study highlights the significant contribution of light exposure to the ultimate metabolic phenotype, which might affect the cellular physiology and nutritional value of broccoli sprouts. Furthermore, this study highlights the

  18. Broccoli sprouts: An exceptionally rich source of inducers of enzymes that protect against chemical carcinogens

    Science.gov (United States)

    Fahey, Jed W.; Zhang, Yuesheng; Talalay, Paul

    1997-01-01

    Induction of phase 2 detoxication enzymes [e.g., glutathione transferases, epoxide hydrolase, NAD(P)H: quinone reductase, and glucuronosyltransferases] is a powerful strategy for achieving protection against carcinogenesis, mutagenesis, and other forms of toxicity of electrophiles and reactive forms of oxygen. Since consumption of large quantities of fruit and vegetables is associated with a striking reduction in the risk of developing a variety of malignancies, it is of interest that a number of edible plants contain substantial quantities of compounds that regulate mammalian enzymes of xenobiotic metabolism. Thus, edible plants belonging to the family Cruciferae and genus Brassica (e.g., broccoli and cauliflower) contain substantial quantities of isothiocyanates (mostly in the form of their glucosinolate precursors) some of which (e.g., sulforaphane or 4-methylsulfinylbutyl isothiocyanate) are very potent inducers of phase 2 enzymes. Unexpectedly, 3-day-old sprouts of cultivars of certain crucifers including broccoli and cauliflower contain 10–100 times higher levels of glucoraphanin (the glucosinolate of sulforaphane) than do the corresponding mature plants. Glucosinolates and isothiocyanates can be efficiently extracted from plants, without hydrolysis of glucosinolates by myrosinase, by homogenization in a mixture of equal volumes of dimethyl sulfoxide, dimethylformamide, and acetonitrile at −50°C. Extracts of 3-day-old broccoli sprouts (containing either glucoraphanin or sulforaphane as the principal enzyme inducer) were highly effective in reducing the incidence, multiplicity, and rate of development of mammary tumors in dimethylbenz(a)anthracene-treated rats. Notably, sprouts of many broccoli cultivars contain negligible quantities of indole glucosinolates, which predominate in the mature vegetable and may give rise to degradation products (e.g., indole-3-carbinol) that can enhance tumorigenesis. Hence, small quantities of crucifer sprouts may protect

  19. Genetic analysis of glucosinolate variability in broccoli florets using genome-anchored single nucleotide polymorphisms.

    Science.gov (United States)

    Brown, Allan F; Yousef, Gad G; Reid, Robert W; Chebrolu, Kranthi K; Thomas, Aswathy; Krueger, Christopher; Jeffery, Elizabeth; Jackson, Eric; Juvik, John A

    2015-07-01

    The identification of genetic factors influencing the accumulation of individual glucosinolates in broccoli florets provides novel insight into the regulation of glucosinolate levels in Brassica vegetables and will accelerate the development of vegetables with glucosinolate profiles tailored to promote human health. Quantitative trait loci analysis of glucosinolate (GSL) variability was conducted with a B. oleracea (broccoli) mapping population, saturated with single nucleotide polymorphism markers from a high-density array designed for rapeseed (Brassica napus). In 4 years of analysis, 14 QTLs were associated with the accumulation of aliphatic, indolic, or aromatic GSLs in floret tissue. The accumulation of 3-carbon aliphatic GSLs (2-propenyl and 3-methylsulfinylpropyl) was primarily associated with a single QTL on C05, but common regulation of 4-carbon aliphatic GSLs was not observed. A single locus on C09, associated with up to 40 % of the phenotypic variability of 2-hydroxy-3-butenyl GSL over multiple years, was not associated with the variability of precursor compounds. Similarly, QTLs on C02, C04, and C09 were associated with 4-methylsulfinylbutyl GSL concentration over multiple years but were not significantly associated with downstream compounds. Genome-specific SNP markers were used to identify candidate genes that co-localized to marker intervals and previously sequenced Brassica oleracea BAC clones containing known GSL genes (GSL-ALK, GSL-PRO, and GSL-ELONG) were aligned to the genomic sequence, providing support that at least three of our 14 QTLs likely correspond to previously identified GSL loci. The results demonstrate that previously identified loci do not fully explain GSL variation in broccoli. The identification of additional genetic factors influencing the accumulation of GSL in broccoli florets provides novel insight into the regulation of GSL levels in Brassicaceae and will accelerate development of vegetables with modified or enhanced GSL

  20. Glucosinolates from pak choi and broccoli induce enzymes and inhibit inflammation and colon cancer differently.

    Science.gov (United States)

    Lippmann, Doris; Lehmann, Carsten; Florian, Simone; Barknowitz, Gitte; Haack, Michael; Mewis, Inga; Wiesner, Melanie; Schreiner, Monika; Glatt, Hansruedi; Brigelius-Flohé, Regina; Kipp, Anna P

    2014-06-01

    High consumption of Brassica vegetables is considered to prevent especially colon carcinogenesis. The content and pattern of glucosinolates (GSLs) can highly vary among different Brassica vegetables and may, thus, affect the outcome of Brassica intervention studies. Therefore, we aimed to feed mice with diets containing plant materials of the Brassica vegetables broccoli and pak choi. Further enrichment of the diets by adding GSL extracts allowed us to analyze the impact of different amounts (GSL-poor versus GSL-rich) and different patterns (broccoli versus pak choi) of GSLs on inflammation and tumor development in a model of inflammation-triggered colon carcinogenesis (AOM/DSS model). Serum albumin adducts were analyzed to confirm the up-take and bioactivation of GSLs after feeding the Brassica diets for four weeks. In agreement with their high glucoraphanin content, broccoli diets induced the formation of sulforaphane-lysine adducts. Levels of 1-methoxyindolyl-3-methyl-histidine adducts derived from neoglucobrassicin were the highest in the GSL-rich pak choi group. In the colon, the GSL-rich broccoli and the GSL-rich pak choi diet up-regulated the expression of different sets of typical Nrf2 target genes like Nqo1, Gstm1, Srxn1, and GPx2. GSL-rich pak choi induced the AhR target gene Cyp1a1 but did not affect Ugt1a1 expression. Both colitis and tumor number were drastically reduced after feeding the GSL-rich pak choi diet while the other three diets had no effect. GSLs can act anti-inflammatory and anti-carcinogenic but both effects depend on the specific amount and pattern of GSLs within a vegetable. Thus, a high Brassica consumption cannot be generally considered to be cancer-preventive.

  1. Ozone effects on yield quality of spring oilseed rape and broccoli

    Science.gov (United States)

    Vandermeiren, Karine; De Bock, Maarten; Horemans, Nele; Guisez, Yves; Ceulemans, Reinhart; De Temmerman, Ludwig

    2012-02-01

    The impact of elevated tropospheric ozone (O 3) on the quality of spring oilseed rape ( Brassica napus cv Ability) and broccoli ( Brassica oleracea L. cv Italic cv Monaco) was assessed during a three year Open - Top Chamber (OTC) experiment. Current ambient O 3 levels were compared to an increase of 20 and 40 ppb during 8 h per day over the entire growing season. The qualitative responses were expressed as a function of the accumulated hourly O 3 concentrations over a threshold of 40 ppb (AOT40) and the phytotoxic O 3 dose above a threshold of 6 nmol s -1 m -2 projected leaf area (POD 6). Our results provide clear evidence that O 3 has an influence on the qualitative attributes of the harvested products of these Brassica species. The responses were comparable whether they were expressed as a function of the accumulated O 3 concentrations or of the modelled O 3 uptake. The protein concentration of oilseed rape seeds and broccoli heads was significantly increased in response to O 3. There was also a shift in the fatty acid composition of the vegetable oil derived from seeds of oilseed rape. Oleic acid (18:1) declined significantly ( p broccoli an important shift occurred from indolic to aliphatic GSLs although the total GSL concentration was not changed. The increase in the aliphatic/indolic GSL ratio ( p broccoli were not influenced by O 3; glutathione (GSH) was slightly increased in response to a higher O 3 uptake ( p < 0.05). The consequences of these changes with regard to food and feed quality and human health are discussed.

  2. Sulforaphane Bioavailability from Glucoraphanin-Rich Broccoli: Control by Active Endogenous Myrosinase.

    Directory of Open Access Journals (Sweden)

    Jed W Fahey

    Full Text Available Glucoraphanin from broccoli and its sprouts and seeds is a water soluble and relatively inert precursor of sulforaphane, the reactive isothiocyanate that potently inhibits neoplastic cellular processes and prevents a number of disease states. Sulforaphane is difficult to deliver in an enriched and stable form for purposes of direct human consumption. We have focused upon evaluating the bioavailability of sulforaphane, either by direct administration of glucoraphanin (a glucosinolate, or β-thioglucoside-N-hydroxysulfate, or by co-administering glucoraphanin and the enzyme myrosinase to catalyze its conversion to sulforaphane at economic, reproducible and sustainable yields. We show that following administration of glucoraphanin in a commercially prepared dietary supplement to a small number of human volunteers, the volunteers had equivalent output of sulforaphane metabolites in their urine to that which they produced when given an equimolar dose of glucoraphanin in a simple boiled and lyophilized extract of broccoli sprouts. Furthermore, when either broccoli sprouts or seeds are administered directly to subjects without prior extraction and consequent inactivation of endogenous myrosinase, regardless of the delivery matrix or dose, the sulforaphane in those preparations is 3- to 4-fold more bioavailable than sulforaphane from glucoraphanin delivered without active plant myrosinase. These data expand upon earlier reports of inter- and intra-individual variability, when glucoraphanin was delivered in either teas, juices, or gelatin capsules, and they confirm that a variety of delivery matrices may be equally suitable for glucoraphanin supplementation (e.g. fruit juices, water, or various types of capsules and tablets.

  3. Broccoli-shaped biosensor hierarchy for electrochemical screening of noradrenaline in living cells.

    Science.gov (United States)

    Emran, Mohammed Y; Mekawy, Moataz; Akhtar, Naeem; Shenashen, Mohamed A; El-Sewify, Islam M; Faheem, Ahmed; El-Safty, Sherif A

    2018-02-15

    Monitoring and determination of ultra-trace concentrations of monoamine neurotransmitter such as noradrenaline (NA) in living cells with simple, sensitive and selective assays are significantly interesting. We design NA-electrode sensing system based on C-, N-doped NiO broccoli-like hierarchy (CNNB). The spherical broccoli-head umbrella architectures associated with nano-tubular arrangements enabled to tailor NA biosensor design. The homogenous doping and anisotropic dispersion of CN nanospheres along the entire NB head nanotubes lead to creating of abundant electroactive sites in the interior tubular vessels and outer surfaces for ultrasensitive detection of NA in living cells such as PC12. The CNNB biosensor electrodes showed efficient electrocatalytic activity, enhanced kinetics for electrooxidation of NA, and fast electron-transfer between electrode-electrolyte interface surfaces, enabling synergistic enhancement in sensitivity, and selectivity at a low-detectable concentration of ∼ 6nM and reproducibility of broccoli-shaped NA-electrodes. The integrated CNNB biosensor electrodes showed evidence of monitoring and screening of NA released from PC12 cells under K + ion-extracellular stimulation process. The unique features of CNNB in terms of NA-selectivity among multi-competitive components, long-term stability during the detection of NA may open their practical, in-vitro application for extracellular monoamine neurotransmitters detection in living cells. Copyright © 2017 Elsevier B.V. All rights reserved.

  4. Stability of Bioactive Compounds in Broccoli as Affected by Cutting Styles and Storage Time.

    Science.gov (United States)

    Torres-Contreras, Ana Mariel; Nair, Vimal; Cisneros-Zevallos, Luis; Jacobo-Velázquez, Daniel A

    2017-04-15

    Broccoli contains bioactive molecules and thus its consumption is related with the prevention of chronic and degenerative diseases. The application of wounding stress to horticultural crops is a common practice, since it is the basis for the fresh-cut produce industry. In this study, the effect of four different cutting styles (CSs) (florets (CS1), florets cut into two even pieces (CS2), florets cut into four even pieces (CS3), and florets processed into chops (CS4)) and storage time (0 and 24 h at 20 °C) on the content of bioactive compounds in broccoli was evaluated. Immediately after cutting, 5-O-caffeoylquinic acid and caffeic acid content increased by 122.4% and 41.6% in CS4 and CS2, respectively. Likewise, after storage, 3-O-caffeoylquinic acid and 5-O-caffeoylquinic acid increased by 46.7% and 98.2%, respectively in CS1. Glucoerucin and gluconasturtiin content decreased by 62% and 50%, respectively in CS3; whereas after storage most glucosinolates increased in CS1. Total isothiocyanates, increased by 133% immediately in CS4, and after storage CS1 showed 65% higher levels of sulforaphane. Total ascorbic acid increased 35% after cutting in CS2, and remained stable after storage. Results presented herein would allow broccoli producers to select proper cutting styles that preserve or increase the content of bioactive molecules.

  5. Validation of the Analytical Method for the Determination of Flavonoids in Broccoli

    Directory of Open Access Journals (Sweden)

    Tuszyńska Magdalena

    2014-09-01

    Full Text Available A simple, accurate and selective HPLC method was developed and validated for determination of quercetin and kaempferol, which are the main flavonols in broccoli. The separation was achieved on a reversed-phase C18 column using a mobile phase composed of methanol/water (60/40 and phosphoric acid 0.2% at a flow rate of 1.0 ml min-1. The detection was carried out on a DAD detector at 370 nm. This method was validated according to the requirements for new methods, which include selectivity, linearity, precision, accuracy, limit of detection and limit of quantitation. The current method demonstrates good linearity, with R2 > 0.99. The recovery is within 98.07-102.15% and 97.92-101.83% for quercetin and kaempferol, respectively. The method is selective, in that quercetin and kaempferol are well separated from other compounds of broccoli with good resolution. The low limit of detection and limit of quantitation of quercetin and kaempferol enable the detection and quantitation of these flavonoids in broccoli at low con–centrations.

  6. Radiographic images and relationship of the internal morphology and physiological potential of broccoli seeds

    Directory of Open Access Journals (Sweden)

    Haynna Fernandes Abud

    2017-11-01

    Full Text Available Image analysis using X-rays is an efficient technique for assessing the seed quality of several species, presenting itself as a rapid response method that is simple to execute, reproducible and non-destructive. Thus, this research adjusted a methodology that aims to relate the internal morphology of broccoli seeds to their physiological potential through radiographic analysis of seeds and computerized images of seedlings. The broccoli cultivars used were Piracicaba Precoce and Ramoso Santana represented by ten lots of each one. The study was conducted using X-ray imaging for seed analysis, in which the area, density and circularity measures were obtained as internal morphology characteristics of seeds. After a germination test, the seedling length was obtained 5 days after sowing. Multivariate analysis was carried out using principal components analysis. It was concluded that the X-ray test is efficient for evaluating the internal morphology of broccoli seeds and associating it with seedling length, thereby classifying seed lots at different levels of vigor.

  7. [Effect of abamectin insecticide on the microbial community in broccoli phyllosphere].

    Science.gov (United States)

    Zhang, Bao-Guo; Tang, Ling; Li, Zu-Ming; Wang, Hui-Li; Xu, Wen-Ting; Zhang, Hong-Xun; Zhuang, Guo-Qiang; Bai, Zhi-Hui

    2009-05-15

    The indigenous microbial communities within the plant phyllosphere are highly diverse and include many different species of bacteria, filamentous fungi, yeasts and algae which play important ecological roles. This study was the first attempt to assess the impact of abamectin treatments on microbial communities of broccoli phyllosphere using two culture-independent techniques of phospholipid fatty acid analysis (PLFA) and terminal restriction fragment length polymorphism (T-RFLP). Results showed that low concentration of abamectin treatments did not affect the microbial biomass and microbial community structure of broccoli phyllosphere significantly. However, high concentration of abamectin treatments significantly change the microbial community structure including a decrease of total and bacterial biomass, and a decrease in the ratio of Gram-positive bacteria to Gram-negative bacteria, but did not change the fungal biomass. Moreover, PLFA suggested that the number of unsaturated and cyclopropane phospholipid fatty acids (PLFAs: 16: 1omega9t, 18: 1omega7, cy17:0 and cy19:0) increased with high concentration abamectin treatment, while the saturated PLFAs i15:0, a15:0, i16:0 and a17:0 decreased. The appearance terminal restriction fragments (T-RFs: 58, 96, 236 and 420 bp) indicated that some bacteria might play a significant role in abamectin degradation in broccoli phylosphere, while the disappeared T-RFs (51, 89, 99, 338, 66, 223 and 482 bp) implied some other bacteria might potentially serve as microbial indicator of abamectin exposure.

  8. Utility of the broccoli sign in the distinction of prolapsed uterine tumor from cervical tumor

    Energy Technology Data Exchange (ETDEWEB)

    Jha, Priyanka; Chang, Stephanie T. [Department of Radiology, University of California San Francisco, 505 Parnassus Avenue, San Francisco, CA 94143-0628 (United States); Rabban, Joseph T. [Department of Anatomic Pathology, University of California San Francisco, 505 Parnassus Avenue, San Francisco, CA 94143-0628 (United States); Chen, Lee-may [Department of Obstetrics, Gynecology, and Reproductive Sciences, University of California San Francisco, 505 Parnassus Avenue, San Francisco, CA 94143-0628 (United States); Yeh, Benjamin M. [Department of Radiology, University of California San Francisco, 505 Parnassus Avenue, San Francisco, CA 94143-0628 (United States); Coakley, Fergus V., E-mail: Fergus.Coakley@radiology.ucsf.edu [Department of Radiology, University of California San Francisco, 505 Parnassus Avenue, San Francisco, CA 94143-0628 (United States)

    2012-08-15

    Objective: To describe the utility, histopathological basis, and clinical correlates of the broccoli sign. Methods: The committee on human research approved this HIPAA compliant study and waived written informed consent. Based on the records of the senior author and our multidisciplinary Gynecologic Oncology Tumor Board, we retrospectively identified thirteen women (mean age of 48.8 years; range, 34-74) with a cervical mass seen at MR imaging (n = 13) or CT (n = 5) that demonstrated the previously reported broccoli sign (i.e., a soft tissue stalk connecting the cervical mass to the uterine cavity) on one or other modality. All available clinical, imaging, and histopathological records were reviewed, with particular emphasis on initially suspected diagnosis, final proven diagnosis, and outcome. Results: Cervical cancer was the initial clinically suspected diagnosis in 6 of 13 patients. Surgical resection demonstrated prolapsed uterine tumor in all patients, consisting of endometrioid adenocarcinoma (n = 7), carcinosarcoma (n = 2), adenosarcoma (n = 1), and leiomyoma (n = 3). Excluding the three patients with leiomyomas, currently, 7 patients with malignant tumors are disease free after a mean interval of 15 months (range, 3-45) and 3 patients have been lost to follow-up. Conclusion: A stalk connecting an apparent cervical mass seen at CT or MR imaging to the endometrial cavity ('broccoli sign') favors the diagnosis of a prolapsed uterine tumor; these prolapsed uterine tumors can often be malignant but appear to have a good prognosis.

  9. Proteomic analysis of broccoli sprouts by iTRAQ in response to jasmonic acid.

    Science.gov (United States)

    Guo, Liping; Wang, Pei; Gu, Zhenxin; Jin, Xiaolin; Yang, Runqiang

    2017-11-01

    Jasmonic acid (JA) is well known as a linolenic acid-derived signal molecule related to the plant response to biotic and abiotic stresses. JA can regulate various plant metabolisms, such as glucosinolate metabolism. In this study, the proteome profiles of broccoli sprouts under JA treatment were analyzed using the iTRAQ-based quantitative proteome approach. A total of 122 differentially expressed proteins participating in a wide range of physiological processes were confidently identified in broccoli sprouts treated with JA. Functional classification analysis showed that photosynthesis and protein synthesis were inhibited by JA treatment, thereby inhibiting sprout growth, while proteins related to carbohydrate catabolism and amino acid metabolism showed an increased expression. Additionally, proteins involved in defense and secondary metabolism were also up-regulated. Proteins related to glucosinolate biosynthesis and degradation were mediated by JA, leading to the accumulation of glucosinolates and sulforaphane. These results indicate that JA stimulated a defense response at the proteome level by redirecting metabolism of growth and physiology in broccoli sprouts. Copyright © 2017 Elsevier GmbH. All rights reserved.

  10. Sulforaphane Bioavailability from Glucoraphanin-Rich Broccoli: Control by Active Endogenous Myrosinase.

    Science.gov (United States)

    Fahey, Jed W; Holtzclaw, W David; Wehage, Scott L; Wade, Kristina L; Stephenson, Katherine K; Talalay, Paul

    2015-01-01

    Glucoraphanin from broccoli and its sprouts and seeds is a water soluble and relatively inert precursor of sulforaphane, the reactive isothiocyanate that potently inhibits neoplastic cellular processes and prevents a number of disease states. Sulforaphane is difficult to deliver in an enriched and stable form for purposes of direct human consumption. We have focused upon evaluating the bioavailability of sulforaphane, either by direct administration of glucoraphanin (a glucosinolate, or β-thioglucoside-N-hydroxysulfate), or by co-administering glucoraphanin and the enzyme myrosinase to catalyze its conversion to sulforaphane at economic, reproducible and sustainable yields. We show that following administration of glucoraphanin in a commercially prepared dietary supplement to a small number of human volunteers, the volunteers had equivalent output of sulforaphane metabolites in their urine to that which they produced when given an equimolar dose of glucoraphanin in a simple boiled and lyophilized extract of broccoli sprouts. Furthermore, when either broccoli sprouts or seeds are administered directly to subjects without prior extraction and consequent inactivation of endogenous myrosinase, regardless of the delivery matrix or dose, the sulforaphane in those preparations is 3- to 4-fold more bioavailable than sulforaphane from glucoraphanin delivered without active plant myrosinase. These data expand upon earlier reports of inter- and intra-individual variability, when glucoraphanin was delivered in either teas, juices, or gelatin capsules, and they confirm that a variety of delivery matrices may be equally suitable for glucoraphanin supplementation (e.g. fruit juices, water, or various types of capsules and tablets).

  11. Effect of visible light treatments on postharvest senescence of broccoli (Brassica oleracea L.).

    Science.gov (United States)

    Büchert, Agustin M; Gómez Lobato, Maria E; Villarreal, Natalia M; Civello, Pedro M; Martínez, Gustavo A

    2011-01-30

    Broccoli (Brassica oleracea L.) is a rapidly perishable vegetable crop. Several postharvest treatments have been applied in order to delay de-greening. Since light has been shown to have an effect on pigment accumulation during development and darkness is known to induce senescence, the effect of continuous and periodic exposure to low-intensity white light at 22 °C on postharvest senescence of broccoli heads was assayed. Exposure to a constant dose of 12 micromol m(-2) s(-1) was selected as the most suitable treatment and was employed for subsequent experiments. During the course of the treatments, hue and L* values as well as chlorophyll content and visual observation of florets indicated an evident delay in yellowing in treated samples compared with controls. No statistically significant differences in total protein content were found, but soluble protein content was higher in treated samples. Total and reducing sugar as well as starch levels decreased during postharvest senescence, with lower values in control samples. The results of this study indicate that storage under continuous low-intensity light is an efficient and low-cost treatment that delays postharvest senescence while maintaining the quality of harvested broccoli florets. 2010 Society of Chemical Industry.

  12. Sulforaphane Bioavailability from Glucoraphanin-Rich Broccoli: Control by Active Endogenous Myrosinase

    Science.gov (United States)

    Fahey, Jed W.; Holtzclaw, W. David; Wehage, Scott L.; Wade, Kristina L.; Stephenson, Katherine K.; Talalay, Paul

    2015-01-01

    Glucoraphanin from broccoli and its sprouts and seeds is a water soluble and relatively inert precursor of sulforaphane, the reactive isothiocyanate that potently inhibits neoplastic cellular processes and prevents a number of disease states. Sulforaphane is difficult to deliver in an enriched and stable form for purposes of direct human consumption. We have focused upon evaluating the bioavailability of sulforaphane, either by direct administration of glucoraphanin (a glucosinolate, or β-thioglucoside-N-hydroxysulfate), or by co-administering glucoraphanin and the enzyme myrosinase to catalyze its conversion to sulforaphane at economic, reproducible and sustainable yields. We show that following administration of glucoraphanin in a commercially prepared dietary supplement to a small number of human volunteers, the volunteers had equivalent output of sulforaphane metabolites in their urine to that which they produced when given an equimolar dose of glucoraphanin in a simple boiled and lyophilized extract of broccoli sprouts. Furthermore, when either broccoli sprouts or seeds are administered directly to subjects without prior extraction and consequent inactivation of endogenous myrosinase, regardless of the delivery matrix or dose, the sulforaphane in those preparations is 3- to 4-fold more bioavailable than sulforaphane from glucoraphanin delivered without active plant myrosinase. These data expand upon earlier reports of inter- and intra-individual variability, when glucoraphanin was delivered in either teas, juices, or gelatin capsules, and they confirm that a variety of delivery matrices may be equally suitable for glucoraphanin supplementation (e.g. fruit juices, water, or various types of capsules and tablets). PMID:26524341

  13. Proteomic analysis of broccoli (Brassica oleracea) under high temperature and waterlogging stresses.

    Science.gov (United States)

    Lin, Hsin-Hung; Lin, Kuan-Hung; Chen, Su-Ching; Shen, Yu-Hsing; Lo, Hsiao-Feng

    2015-12-01

    The production of broccoli (Brassica oleracea) is largely reduced by waterlogging and high temperature stresses. Heat-tolerant and heat-susceptible broccoli cultivars TSS-AVRDC-2 and B-75, respectively, were used for physiological and proteomic analyses. The objective of this study was to identify TSS-AVRDC-2 and B-75 proteins differentially regulated at different time periods in response to waterlogging at 40 °C for three days. TSS-AVRDC-2 exhibited significantly higher chlorophyll content, lower stomatal conductance, and better H2O2 scavenging under stress in comparison to B-75. Two-dimensional liquid phase fractionation analyses revealed that Rubisco proteins in both varieties were regulated under stressing treatments, and that TSS-AVRDC-2 had higher levels of both Rubisco large and small subunit transcripts than B-75 when subjected to high temperature and/or waterlogging. This report utilizes physiological and proteomic approaches to discover changes in the protein expression profiles of broccoli in response to heat and waterlogging stresses. Higher levels of Rubisco proteins in TSS-AVRDC-2 could lead to increased carbon fixation efficiency to provide sufficient energy to enable stress tolerance under waterlogging at 40 °C.

  14. Dietary chemoprevention of PhIP induced carcinogenesis in male Fischer 344 rats with tomato and broccoli.

    Directory of Open Access Journals (Sweden)

    Kirstie Canene-Adams

    Full Text Available The heterocyclic amine, 2-amino-1-methyl-6-phenylimidazo[4,5-B]pyridine (PhIP, found in meats cooked at high temperatures, has been implicated in epidemiological and rodent studies for causing breast, prostate, and colorectal cancers. A previous animal study using a xenograft model has shown that whole tomato and broccoli, when eaten in combination, exhibit a marked effect on tumor reduction compared to when eaten alone. Our aim was to determine if PhIP-induced carcinogenesis can be prevented by dietary consumption of whole tomato + broccoli powders. Male Fischer 344 rats (n = 45 were randomized into the following treatment groups: control (AIN93G diet, PhIP (200 ppm in AIN93G diet for the first 20 weeks of the study, or tomato + broccoli + PhIP (mixed in AIN93G diet at 10% each and fed with PhIP for 20 weeks, and then without PhIP for 32 weeks. Study animals were monitored for 52 weeks and were euthanized as necessary based on a set of criteria for health status and tumor burden. Although there appeared to be some hepatic and intestinal toxicity due to the combination of PhIP and tomato + broccoli, these rodents had improved survival and reduced incidence and/or severity of PhIP-induced neoplastic lesions compared to the PhIP-alone treated group. Rats eating tomato + broccoli exhibited a marked decrease in the number and size of cribiform prostatic intraepitheilial neoplasia/carcinoma in situ (cribiform PIN/CIS lesions and in the incidence of invasive intestinal adenocarcinomas and skin carcinomas. Although the apparent toxic effects of combined PhIP and tomato + broccoli need additional study, the results of this study support the hypothesis that a diet rich in tomato and broccoli can reduce or prevent dietary carcinogen-induced cancers.

  15. Dietary chemoprevention of PhIP induced carcinogenesis in male Fischer 344 rats with tomato and broccoli.

    Science.gov (United States)

    Canene-Adams, Kirstie; Sfanos, Karen S; Liang, Chung-Tiang; Yegnasubramanian, Srinivasan; Nelson, William G; Brayton, Cory; De Marzo, Angelo M

    2013-01-01

    The heterocyclic amine, 2-amino-1-methyl-6-phenylimidazo[4,5-B]pyridine (PhIP), found in meats cooked at high temperatures, has been implicated in epidemiological and rodent studies for causing breast, prostate, and colorectal cancers. A previous animal study using a xenograft model has shown that whole tomato and broccoli, when eaten in combination, exhibit a marked effect on tumor reduction compared to when eaten alone. Our aim was to determine if PhIP-induced carcinogenesis can be prevented by dietary consumption of whole tomato + broccoli powders. Male Fischer 344 rats (n = 45) were randomized into the following treatment groups: control (AIN93G diet), PhIP (200 ppm in AIN93G diet for the first 20 weeks of the study), or tomato + broccoli + PhIP (mixed in AIN93G diet at 10% each and fed with PhIP for 20 weeks, and then without PhIP for 32 weeks). Study animals were monitored for 52 weeks and were euthanized as necessary based on a set of criteria for health status and tumor burden. Although there appeared to be some hepatic and intestinal toxicity due to the combination of PhIP and tomato + broccoli, these rodents had improved survival and reduced incidence and/or severity of PhIP-induced neoplastic lesions compared to the PhIP-alone treated group. Rats eating tomato + broccoli exhibited a marked decrease in the number and size of cribiform prostatic intraepitheilial neoplasia/carcinoma in situ (cribiform PIN/CIS) lesions and in the incidence of invasive intestinal adenocarcinomas and skin carcinomas. Although the apparent toxic effects of combined PhIP and tomato + broccoli need additional study, the results of this study support the hypothesis that a diet rich in tomato and broccoli can reduce or prevent dietary carcinogen-induced cancers.

  16. Dietary Chemoprevention of PhIP Induced Carcinogenesis in Male Fischer 344 Rats with Tomato and Broccoli

    Science.gov (United States)

    Canene-Adams, Kirstie; Sfanos, Karen S.; Liang, Chung-Tiang; Yegnasubramanian, Srinivasan; Nelson, William G.; Brayton, Cory; De Marzo, Angelo M.

    2013-01-01

    The heterocyclic amine, 2-amino-1-methyl-6-phenylimidazo[4,5-B]pyridine (PhIP), found in meats cooked at high temperatures, has been implicated in epidemiological and rodent studies for causing breast, prostate, and colorectal cancers. A previous animal study using a xenograft model has shown that whole tomato and broccoli, when eaten in combination, exhibit a marked effect on tumor reduction compared to when eaten alone. Our aim was to determine if PhIP-induced carcinogenesis can be prevented by dietary consumption of whole tomato + broccoli powders. Male Fischer 344 rats (n = 45) were randomized into the following treatment groups: control (AIN93G diet), PhIP (200 ppm in AIN93G diet for the first 20 weeks of the study), or tomato + broccoli + PhIP (mixed in AIN93G diet at 10% each and fed with PhIP for 20 weeks, and then without PhIP for 32 weeks). Study animals were monitored for 52 weeks and were euthanized as necessary based on a set of criteria for health status and tumor burden. Although there appeared to be some hepatic and intestinal toxicity due to the combination of PhIP and tomato + broccoli, these rodents had improved survival and reduced incidence and/or severity of PhIP-induced neoplastic lesions compared to the PhIP-alone treated group. Rats eating tomato + broccoli exhibited a marked decrease in the number and size of cribiform prostatic intraepitheilial neoplasia/carcinoma in situ (cribiform PIN/CIS) lesions and in the incidence of invasive intestinal adenocarcinomas and skin carcinomas. Although the apparent toxic effects of combined PhIP and tomato + broccoli need additional study, the results of this study support the hypothesis that a diet rich in tomato and broccoli can reduce or prevent dietary carcinogen-induced cancers. PMID:24312188

  17. Dietary broccoli mildly improves neuroinflammation in aged mice but does not reduce lipopolysaccharide-induced sickness behavior.

    Science.gov (United States)

    Townsend, Brigitte E; Chen, Yung-Ju; Jeffery, Elizabeth H; Johnson, Rodney W

    2014-11-01

    Aging is associated with oxidative stress and heightened inflammatory response to infection. Dietary interventions to reduce these changes are therefore desirable. Broccoli contains glucoraphanin, which is converted to sulforaphane (SFN) by plant myrosinase during cooking preparation or digestion. Sulforaphane increases antioxidant enzymes including NAD(P)H quinone oxidoreductase and heme oxygenase I and inhibits inflammatory cytokines. We hypothesized that dietary broccoli would support an antioxidant response in brain and periphery of aged mice and inhibit lipopolysaccharide (LPS)-induced inflammation and sickness. Young adult and aged mice were fed control or 10% broccoli diet for 28 days before an intraperitoneal LPS injection. Social interactions were assessed 2, 4, 8, and 24 hours after LPS, and mRNA was quantified in liver and brain at 24 hours. Dietary broccoli did not ameliorate LPS-induced decrease in social interactions in young or aged mice. Interleukin-1β (IL-1β) expression was unaffected by broccoli consumption but was induced by LPS in brain and liver of adult and aged mice. In addition, IL-1β was elevated in brain of aged mice without LPS. Broccoli consumption decreased age-elevated cytochrome b-245 β, an oxidative stress marker, and reduced glial activation markers in aged mice. Collectively, these data suggest that 10% broccoli diet provides a modest reduction in age-related oxidative stress and glial reactivity, but is insufficient to inhibit LPS-induced inflammation. Thus, it is likely that SFN would need to be provided in supplement form to control the inflammatory response to LPS. Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.

  18. Detection of the Diversity of Cytoplasmic Male Sterility Sources in Broccoli (Brassica Oleracea var. Italica) Using Mitochondrial Markers.

    Science.gov (United States)

    Shu, Jinshuai; Liu, Yumei; Li, Zhansheng; Zhang, Lili; Fang, Zhiyuan; Yang, Limei; Zhuang, Mu; Zhang, Yangyong; Lv, Honghao

    2016-01-01

    Broccoli (Brassica oleracea var. italica) is an important commercial vegetable crop. As part of an efficient pollination system, cytoplasmic male sterility (CMS) has been widely used for broccoli hybrid production. Identifying the original sources of CMS in broccoli accessions has become an important part of broccoli breeding. In this study, the diversity of the CMS sources of 39 broccoli accessions, including 19 CMS lines and 20 hybrids, were analyzed using mitochondrial markers. All CMS accessions contained the ogu orf138-related DNA fragment and the key genes of nap CMS, pol CMS, and tour CMS were not detected. The 39 CMS accessions were divided into five groups using six orf138-related and two simple sequence repeat markers. We observed that ogu CMS R3 constituted 79.49% of the CMS sources. CMS6 and CMS26 were differentiated from the other accessions using a specific primer. CMS32 was distinguished from the other accessions based on a 78-nucleotide deletion at the same locus as the orf138-related sequence. When the coefficient was about 0.90, five CMS accessions (13CMS6, 13CMS23, 13CMS24, 13CMS37, and 13CMS39) exhibiting abnormal floral organs with poor seed setting were grouped together. The polymerase chain reaction amplification profiles for these five accessions differed from those of the other accessions. We identified eight useful molecular markers that can be used to detect CMS types during broccoli breeding. Our data also provide important information relevant to future studies on the possible origins and molecular mechanisms of CMS in broccoli.

  19. Monolignol 4-O-methyltransferases and uses thereof

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Chang-Jun; Bhuiya, Mohammad-Wadud; Zhang, Kewei

    2014-11-18

    Modified (iso)eugenol 4-O-methyltransferase enzymes having novel capacity for methylation of monolignols and reduction of lignin polymerization in plant cell wall are disclosed. Sequences encoding the modified enzymes are disclosed.

  20. The Effect of Differentiated Nutrition on the Content of Antioxidants in Broccoli

    Directory of Open Access Journals (Sweden)

    Anton Uher

    2014-01-01

    Full Text Available The aim of this work is to determine the impact of differentiated nutrition, using different rates of nitrogen and sulphur, on the level of antioxidants, particularly vitamin E1 (α-tocopherol, vitamin C and β-carotene in the broccoli rosette. The experimental broccoli variety was Tiburon F1. It is a strong medium-late variety with a vegetation period of 82 days. In the 3-year field trial we observed the effects of different rates of nitrogen and sulphur on the amount of antioxidant compounds in broccoli. The experiment consisted of four fertilisation treatments: 1 unfertilised control, 2 fertilised with 200 kg N.ha−1, 3 also fertilized with 200 kg N.ha−1 and supplemented with 50 kg S.ha−1, 4 200 kg.ha−1 and 60 kg S.ha−1 was applied. To determine the amount of β-carotene and vitamin E1, the slightly modified method of Olives Barb et al. (2006 was used. The vitamin C content was determined by titration. Nitrogen nutrition has a significant impact not only on the amount of harvested broccoli, but also on the content of β-carotene, vitamin C and vitamin E1. The average content of β-carotene for the entire experimental period ranged from 24.84 mg.kg−1 to 30.13 mg.kg−1 of fresh mass. The content of β-carotene in broccoli rosettes increased as per the following order of treatments: control > N:S (200:50 kg.ha−1 > N:S (200:60 kg.ha−1 > N (kg.ha−1. The β-carotene content increased significantly only in treatment 2 (30.13 mg.kg−1 as compared to all the other treatments. The level of vitamin C revealed the significant effect of fertilisation in all the treatments (567.9–614.2 mg.kg−1 in contrast to the control variant (528.4 mg.kg−1. What is more, in treatment 4 the content of vitamin C increased significantly in contrast to fertilised treatments 2 and 3. The average content of vitamin E1 ranged from 4.33 mg.kg−1 to 4.88 mg.kg−1 of fresh mass. There were no significant differences among the untreated control

  1. Characteristic of phenolic compound and antioxidant activity of fermented broccoli (Brassica oleracea L. ssp.) beverage by lactic acid bacteria (LAB)

    Science.gov (United States)

    Maryati, Yati; Susilowati, Agustine; Melanie, Hakiki; Lotulung, Puspa D.

    2017-01-01

    Broccoli (Brassica oleracea L. ssp.) has a relatively high nutrient content, especially as a source of vitamins, minerals and fiber and contain bioactive compounds that act as antioxidants. In order to increase the nutritional value and innovate new products, fermentation process involving rich-antioxidants lactic acid bacteria (LAB) was done. The aim of this study is to determine the content of bioactive components, such as total polyphenols, total acid and antioxidant activity of the mixed culture of LAB (L. bulgaricus, S. thermophulus, L. acidophilus, Bd. bifidum)-fermented broccoli extracts. Ratio of fermented broccoli extract and concentration of starter cultureLAB was varied in the range of 5, 10, 15 and 20% (v/v), and the alterations of characteristics of the fermented broccoli extract, before and after fermentation (0 and 24 hours), were evaluated. The results showed that fermentation functional beverage broccoli with different concentrations of LAB cultures affect the antioxidant activity, total polyphenols, total acid and total cell of LAB generated. The optimum conditions obtained for the highest antioxidant activity of 6.74%, at aculture concentration of 20% during fermentation time of 24 h with a pH value of 4.29, total sugar of 10.89%, total acids of 0.97%, total polyphenols of 0.076%, and total LAB of 13.02 + 0.05 log cfu /ml.

  2. Antimicrobial activity of broccoli (Brassica oleracea var. italica) cultivar Avenger against pathogenic bacteria, phytopathogenic filamentous fungi and yeast.

    Science.gov (United States)

    Pacheco-Cano, R D; Salcedo-Hernández, R; López-Meza, J E; Bideshi, D K; Barboza-Corona, J E

    2018-01-01

    The objective of this study was to show whether the edible part of broccoli has antibacterial and antifungal activity against micro-organism of importance in human health and vegetable spoilage, and to test if this effect was partially due to antimicrobial peptides (AMPs). Crude extracts were obtained from florets and stems of broccoli cultivar Avenger and the inhibitory effect was demonstrated against pathogenic bacteria (Bacillus cereus, Staphylococcus xylosus, Staphylococcus aureus, Shigella flexneri, Shigella sonnei, Proteus vulgaris), phytopathogenic fungi (Colletotrichum gloeosporioides, Asperigillus niger) and yeasts (Candida albicans and Rhodotorula sp.). It was shown that samples treated with proteolytic enzymes had a reduction of approximately 60% in antibacterial activity against Staph. xylosus, suggesting that proteinaceous compounds might play a role in the inhibitory effect. Antimicrobial components in crude extracts were thermoresistant and the highest activity was observed under acidic conditions. It was shown that antifungal activity of broccoli's crude extracts might not be attributed to chitinases. Organic broccoli cultivar Avenger has antimicrobial activity against pathogenic bacteria, yeast and phytophatogenic fungi. Data suggest that this effect is partially due to AMPs. Broccoli's crude extracts have activity not only against pathogenic bacteria but also against phytophatogenic fungi of importance in agriculture. We suggest for first time that the inhibitory effect is probably due to AMPs. © 2017 The Society for Applied Microbiology.

  3. Assessing the anticancer compounds Se-methylselenocysteine and glucosinolates in Se-biofortified broccoli (brassica oleracea L. var. italica) sprouts and florets

    Science.gov (United States)

    Broccoli (Brassica oleracea L. var. italica) is a rich source of chemopreventive compounds. Here, we evaluated and compared the effect of selenium (Se) treatment on the accumulation of anticancer compound Se-methylselenocysteine (SeMSCys) and glucosinolates in broccoli sprouts and florets. Total Se ...

  4. Influence of dietary blueberry and broccoli on cecal microbiota activity and colon morphology in mdr1a(-/-) mice, a model of inflammatory bowel diseases.

    Science.gov (United States)

    Paturi, Gunaranjan; Mandimika, Tafadzwa; Butts, Christine A; Zhu, Shuotun; Roy, Nicole C; McNabb, Warren C; Ansell, Juliet

    2012-03-01

    Enteric microbiota has been shown to be associated with various pathological conditions such as inflammatory bowel disease (IBD). This study aimed to determine the anti-inflammatory colonic effects of blueberries and broccoli in mdr1a(-/-) mice (IBD mouse model) through modification of microbiota composition in the gastrointestinal tract. The mdr1a(-/-) mice were fed either a control diet or the control diet supplemented with either 10% blueberry or broccoli for 21 wk. We investigated the influence of these diets on cecal microbiota and organic acids, colon morphology, and bacterial translocation to mesenteric lymph nodes. In comparison to mice fed the control diet, blueberry and broccoli supplementation altered cecum microbiota similarly with the exception of Faecalibacterium prausnitzii, which was found to be significantly lower in broccoli-fed mice. High concentrations of butyric acid and low concentrations of succinic acid were observed in the cecum of broccoli-fed mice. Blueberry- and broccoli-supplemented diets increased colon crypt size and the number of goblet cells per crypt. Only the broccoli-supplemented diet significantly lowered colonic inflammation compared to mice fed the control diet. Translocation of total microbes to mesenteric lymph nodes was lower in broccoli-fed mice compared to blueberry and control diet groups. Dietary blueberries and/or broccoli altered the composition and metabolism of the cecal microbiota and colon morphology. Overall, these results warrant further investigation through clinical studies to establish whether the consumption of blueberries and/or broccoli is able to alter the composition and metabolism of large intestine microbiota and promote colon health in humans. Copyright © 2012 Elsevier Inc. All rights reserved.

  5. Effects of industrial pre-freezing processing and freezing handling on glucosinolates and antioxidant attributes in broccoli florets.

    Science.gov (United States)

    Cai, Congxi; Miao, Huiying; Qian, Hongmei; Yao, Leishuan; Wang, Bingliang; Wang, Qiaomei

    2016-11-01

    The effects of industrial pre-freezing processing and freezing handling on the contents of glucosinolates and antioxidants (vitamin C, polyphenols, carotenoid and chlorophyll), as well as the antioxidant capacity in broccoli (Brassica oleracea L. var. italica) florets were investigated in the present study. Our results showed that the glucosinolate accumulations were significantly decreased after pre-freezing processing, whereas elevated levels of phenols, carotenoids, chlorophyll, and also antioxidant capacity were observed in frozen broccoli florets. The contents of vitamin C remained constant during above mentioned processing. In conclusion, the current industrial freezing processing method is a good practice for the preservation of main antioxidant nutrients in broccoli florets, although some improvements in pre-freezing processing, such as steam blanching and ice-water cooling, are needed to attenuate the decrease in glucosinolate content. Copyright © 2016 Elsevier Ltd. All rights reserved.

  6. Effect of Se treatment on glucosinolate metabolism and health-promoting compounds in the broccoli sprouts of three cultivars.

    Science.gov (United States)

    Tian, Ming; Xu, Xiaoyun; Liu, Yanlong; Xie, Lin; Pan, Siyi

    2016-01-01

    Broccoli sprouts are natural functional foods for cancer prevention because of their high glucosinolate (GSL) content and high selenium (Se) accumulation capacity. The regulation mechanism of Se on GSL metabolism in broccoli sprouts was explored. In particular, the effects of Se treatment (100 μmol/L selenite and selenate) on the Se, sulfur (S), glucosinolate and sulforaphane contents; myrosinase activity and health-promoting compounds (ascorbic acid, anthocyanin, total phenolics and flavonoids) of three, 5 day old, cultivars were investigated. The treatment did not influence the total GSL and ascorbic acid contents; significantly increased the myrosinase activity and sulforaphane, anthocyanin and flavonoids contents; and decreased the total phenolics content. The increase in sulforaphane during early growth can be primarily attributed to the increased myrosinase activity caused by Se treatment. Broccoli sprouts with suitable selenite and selenate concentrations, in the early growth days, could be desirable for improved human health. Copyright © 2015 Elsevier Ltd. All rights reserved.

  7. Effect of reactive oxygen species on quality maintenance of broccoli florets with electrostatic atomized water particle treatment.

    Science.gov (United States)

    Nomura, Masahiro; Aiamla-Or, Sukanya; Tanaka, Shuhei; Shigyo, Masayoshi; Masuda, Yukihiro; Yamauchi, Naoki

    2017-12-15

    The effect of electrostatic atomized water particle (EAWP) treatment on quality maintenance during postharvest storage was determined to clarify a physiological role of reactive oxygen species in broccoli florets. The EAWP treatment suppressed the floret yellowing of broccoli during storage. The hydrogen peroxide content increased immediately after EAWP treatment but then remained at a low level during storage. Catalase (CAT) activity increased rapidly with EAWP treatment and was higher than that in the control for the first 4days of storage. The BoCAT gene expression level with EAWP treatment also showed a high trend as compared to the control. The gene expressions of BoRboh, BoCu/ZnSOD, and BoAPX showed almost the same tendency with or without EAWP treatment. These findings suggested that hydrogen peroxide formed by EAWP treatment and regulated by CAT could be relevant to maintaining the quality of broccoli. Copyright © 2017 Elsevier Ltd. All rights reserved.

  8. Comparative miRNAs analysis of Two contrasting broccoli inbred lines with divergent head-forming capacity under temperature stress.

    Science.gov (United States)

    Chen, Chi-Chien; Fu, Shih-Feng; Norikazu, Monma; Yang, Yau-Wen; Liu, Yu-Ju; Ikeo, Kazuho; Gojobori, Takashi; Huang, Hao-Jen

    2015-12-01

    MicroRNAs (miRNAs) play a vital role in growth, development, and stress response at the post-transcriptional level. Broccoli (Brassica oleracea L. var italic) is an important vegetable crop, and the yield and quality of broccoli are decreased by heat stress. The broccoli inbred lines that are capable of producing head at high temperature in summer are unique varieties in Taiwan. However, knowledge of miRNAomes during the broccoli head formation under heat stress is limited. In this study, molecular characterization of two nearly isogenic lines with contrasting head-forming capacity was investigated. Head-forming capacity was better for heat-tolerant (HT) than heat-sensitive (HS) broccoli under heat stress. By deep sequencing and computational analysis, 20 known miRNAs showed significant differential expression between HT and HS genotypes. According to the criteria for annotation of new miRNAs, 24 novel miRNA sequences with differential expression between the two genotypes were identified. To gain insight into functional significance, 213 unique potential targets of these 44 differentially expressed miRNAs were predicted. These targets were implicated in shoot apical development, phase change, response to temperature stimulus, hormone and energy metabolism. The head-forming capacity of the unique HT line was related to autonomous regulation of Bo-FT genes and less expression level of heat shock protein genes as compared to HS. For the genotypic comparison, a set of miRNAs and their targets had consistent expression patterns in various HT genotypes. This large-scale characterization of broccoli miRNAs and their potential targets is to unravel the regulatory roles of miRNAs underlying heat-tolerant head-forming capacity.

  9. DNA Methyltransferase Activity Assays: Advances and Challenges.

    Science.gov (United States)

    Poh, Wan Jun; Wee, Cayden Pang Pee; Gao, Zhiqiang

    2016-01-01

    DNA methyltransferases (MTases), a family of enzymes that catalyse the methylation of DNA, have a profound effect on gene regulation. A large body of evidence has indicated that DNA MTase is potentially a predictive biomarker closely associated with genetic disorders and genetic diseases like cancer. Given the attention bestowed onto DNA MTases in molecular biology and medicine, highly sensitive detection of DNA MTase activity is essential in determining gene regulation, epigenetic modification, clinical diagnosis and therapeutics. Conventional techniques such as isotope labelling are effective, but they often require laborious sample preparation, isotope labelling, sophisticated equipment and large amounts of DNA, rendering them unsuitable for uses at point-of-care. Simple, portable, highly sensitive and low-cost assays are urgently needed for DNA MTase activity screening. In most recent technological advances, many alternative DNA MTase activity assays such as fluorescent, electrochemical, colorimetric and chemiluminescent assays have been proposed. In addition, many of them are coupled with nanomaterials and/or enzymes to significantly enhance their sensitivity. Herein we review the progress in the development of DNA MTase activity assays with an emphasis on assay mechanism and performance with some discussion on challenges and perspectives. It is hoped that this article will provide a broad coverage of DNA MTase activity assays and their latest developments and open new perspectives toward the development of DNA MTase activity assays with much improved performance for uses in molecular biology and clinical practice.

  10. Evaluation of Cabbage- and Broccoli-genetic Resources for Resistance to Clubroot and Fusarium Wilt

    Directory of Open Access Journals (Sweden)

    Ji Hyun Lee

    2014-12-01

    Full Text Available Clubroot and Fusarium wilt of cole crops (Brassica oleracea L. are destructive diseases which for many years has brought a decline in quality and large losses in yields all over the world. The breeding of resistant cultivars is an effective approach to reduce the use of chemical fungicides and minimize crop losses. This study was conducted to evaluate the resistance of 60 cabbage (B. oleracea var. capitata and 6 broccoli (B. oleracea var. italica lines provided by The RDA-Genebank Information Center to clubroot and Fusarium wilt. To investigate resistance to clubroot, seedlings of the genetic resources were inoculated with Plasmodiophora brassicae by drenching the roots with a mixed spore suspension (1 : 1 of two isolates. Of the tested genetic resources, four cabbage lines were moderately resistant and ‘K166220’ represented the highest resistance to P. brassicae. The others were susceptible to clubroot. On the other hand, to select resistant plants to Fusarium wilt, the genetic resources were inoculated with Fusarium oxysporum f. sp. conglutinans by dipping the roots in spore suspension of the fungus. Among them, 17 cabbage and 5 broccoli lines were resistant, 16 cabbage lines were moderately resistant, and the others were susceptible to Fusarium wilt. Especially, three cabbage (‘IT227115’, ‘K161791’, ‘K173350’ and two broccoli (‘IT227100’, ‘IT227099’ lines were highly resistant to the fungus. We suggest that the resistant genetic resources can be used as a basic material for resistant B. oleracea breeding system against clubroot and Fusarium wilt.

  11. DEVELOPMENT OF EST-SSR MARKERS TO ASSESS GENETIC DIVERSITY OF BROCCOLI AND ITS RELATED SPECIES

    Directory of Open Access Journals (Sweden)

    Nur Kholilatul Izzah

    2017-01-01

    Full Text Available Development of Expressed Sequence Tag-Simple Sequence Repeat (EST-SSR markers derived from public database is known to be more efficient, faster and low cost. The objective of this study was to generate a new set of EST-SSR markers for broccoli and its related species and their usefulness for assessing their genetic diversity. A total of 202 Brassica oleracea ESTs were retrieved from NCBI and then assembled into 172 unigenes by means of CAP3 program. Identification of SSRs was carried out using web-based tool, RepeatMasker software. Afterwards, EST-SSR markers were developed using Primer3 program. Among the identified SSRs, trinucleotide repeats were the most common repeat types, which accounted for about 50%. A total of eight primer pairs were successfully designed and yielded amplification products. Among them, five markers were polymorphic and displayed a total of 30 alleles with an average number of six alleles per locus. The polymorphic markers were subsequently used for analyzing genetic diversity of 36 B. oleracea cultivars including 22 broccoli, five cauliflower and nine kohlrabi cultivars based on genetic similarity matrix as implemented in NTSYS program. At similarity coefficient of 61%, a UPGMA clustering dendrogram effectively separated 36 genotypes into three main groups, where 30 out of 36 genotypes were clearly discriminated. The result obtained in the present study would help breeders in selecting parental lines for crossing. Moreover, the novel EST-SSR markers developed in the study could be a valuable tool for differentiating cultivars of broccoli and related species.

  12. Increasing Antioxidant Content of Broccoli Sprouts Using Essential Oils During Cold Storage

    Directory of Open Access Journals (Sweden)

    El-Awady Aml A.

    2016-10-01

    Full Text Available Broccoli sprouts are natural functional foods for cancer prevention because of their high content of glucosinolate and antioxidant. Sprouts and mature broccoli are of potential importance in devising chemoprotective strategies in humans. The aim of the investigation was to study the effect of essential oils on broccoli seed germination, increase their antioxidant content and determine the glucosinolate concentration and other phytochemical parameters in 3-day-old sprouts during cold storage at 4°C and 95% RH for 15 days. The results showed that all treatments of essential oils increased germination index, seed germination percentage, seedling length, seedling vigour index, yield and the antioxidant content of broccoli sprout and reduced the microbial load compared to the control. Fortunately, the coliform bacteria was not detected in all treatments. Different essential oils of fennel, caraway, basil, thyme and sage were tested. The thyme oil was the best treatment, which increased the accumulation of the phenolic compounds and glucosinolate compared to the control at different storage periods. In the sprouts treated with thyme oil treatment and the control, at the end of cold storage, 1.98% and 28.06% of total phenolic content, 1.90% and 20.28% of 2,2-diphenyl-1-picrylhydrazyl (DPPH radical scavenging capacity, 1.39% and 58.33% of flavonoids, 1.93% and 36.25% of vitamin C, 2.95% and 22.02% of anthocyanin and 2.18% and 49.12% of glucosinolate were lost, respectively. A slight reduction differences in all detected compound concentrations occurred between the initial content and the end of storage period because of the application of thyme oil compared to the control. Therefore, the total glucosinolate level in the sprout (27.02 μg/g F.W. was higher than that in the florets (7.37 μg/g F.W.. Glucoraphanin was the most abundant aliphatic glucosinolate present in the sprout and reached the highest value (16.24 μg/g F.W. followed by glucoerucin (5

  13. Modulation of plasma antioxidant levels, glutathione S-transferase activity and DNA damage in smokers following a single portion of broccoli: a pilot study.

    Science.gov (United States)

    Riso, Patrizia; Del Bo', Cristian; Vendrame, Stefano; Brusamolino, Antonella; Martini, Daniela; Bonacina, Gaia; Porrini, Marisa

    2014-02-01

    Broccoli is a rich source of bioactive compounds (i.e. glucosinolates, carotenoids, vitamin C and folate) that may exert an antioxidant effect and reduce oxidative damage. The objective of this pilot study was to investigate the effect of broccoli consumption on carotenoids, vitamin C and folate absorption, glutathione S-transferase (GST) activity, and oxidatively induced DNA damage in male smokers. Ten healthy subjects consumed a single portion of steamed broccoli (250 g) with cooked pasta. Blood was drawn at baseline and at 3, 6 and 24 h from consumption. Broccoli significantly (P ≤ 0.01) increased plasma level of vitamin C and folate (+35% and 70%, respectively) at 3 h, and β-carotene (+8%) at 6 h. A modulation of GST activity occurred in plasma 6 h after broccoli consumption. A significant (P ≤ 0.01) reduction of the levels of H₂O₂-induced DNA damage (-18%) was observed in blood mononuclear cells 24 h after broccoli intake in GSTM1 positive, but not in GSTM1 null subjects. One portion of broccoli increased plasma antioxidant levels, modulated plasma GST activity and improved cell resistance against H₂O₂-induced DNA damage in healthy smokers. These results support the importance of consuming fruit and vegetable regularly. © 2013 Society of Chemical Industry.

  14. A diet rich in high-glucoraphanin broccoli interacts with genotype to reduce discordance in plasma metabolite profiles by modulating mitochondrial function123

    Science.gov (United States)

    Armah, Charlotte N; Traka, Maria H; Dainty, Jack R; Defernez, Marianne; Janssens, Astrid; Leung, Wing; Doleman, Joanne F; Potter, John F

    2013-01-01

    Background: Observational and experimental studies suggest that diets rich in cruciferous vegetables and glucosinolates may reduce the risk of cancer and cardiovascular disease (CVD). Objective: We tested the hypothesis that a 12-wk dietary intervention with high-glucoraphanin (HG) broccoli would modify biomarkers of CVD risk and plasma metabolite profiles to a greater extent than interventions with standard broccoli or peas. Design: Subjects were randomly assigned to consume 400 g standard broccoli, 400 g HG broccoli, or 400 g peas each week for 12 wk, with no other dietary restrictions. Biomarkers of CVD risk and 347 plasma metabolites were quantified before and after the intervention. Results: No significant differences in the effects of the diets on biomarkers of CVD risk were found. Multivariate analyses of plasma metabolites identified 2 discrete phenotypic responses to diet in individuals within the HG broccoli arm, differentiated by single nucleotide polymorphisms associated with the PAPOLG gene. Univariate analysis showed effects of sex (P broccoli arm, the consequence of the intervention was to reduce variation in lipid and amino acid metabolites, tricarboxylic acid (TCA) cycle intermediates, and acylcarnitines between the 2 PAPOLG genotypes. Conclusions: The metabolic changes observed with the HG broccoli diet are consistent with a rebalancing of anaplerotic and cataplerotic reactions and enhanced integration of fatty acid β-oxidation with TCA cycle activity. These modifications may contribute to the reduction in cancer risk associated with diets that are rich in cruciferous vegetables. This trial was registered at clinicaltrials.gov as NCT01114399. PMID:23964055

  15. Dietary Broccoli Lessens Development of Fatty Liver and Liver Cancer in Mice Given Diethylnitrosamine and Fed a Western or Control Diet123

    Science.gov (United States)

    Chen, Yung-Ju; Wallig, Matthew A; Jeffery, Elizabeth H

    2016-01-01

    Background: The high-fat and high-sugar Westernized diet that is popular worldwide is associated with increased body fat accumulation, which has been related to the development of nonalcoholic fatty liver disease (NAFLD). Without treatment, NAFLD may progress to hepatocellular carcinoma (HCC), a cancer with a high mortality rate. The consumption of broccoli in the United States has greatly increased in the last 2 decades. Epidemiologic studies show that incorporating brassica vegetables into the daily diet lowers the risk of several cancers, although, to our knowledge, this is the first study to evaluate HCC prevention through dietary broccoli. Objective: We aimed to determine the impact of dietary broccoli on hepatic lipid metabolism and the progression of NAFLD to HCC. Our hypothesis was that broccoli decreases both hepatic lipidosis and the development of HCC in a mouse model of Western diet–enhanced liver cancer. Methods: Adult 5-wk-old male B6C3F1 mice received a control diet (AIN-93M) or a Western diet (high in lard and sucrose, 19% and 31%, wt:wt, respectively), with or without freeze-dried broccoli (10%, wt:wt). Starting the following week, mice were treated once per week with diethylnitrosamine (DEN; 45 mg/kg body weight intraperitoneally at ages 6, 7, 8, 10, 11, and 12 wk). Hepatic gene expression, lipidosis, and tumor outcomes were analyzed 6 mo later, when mice were 9 mo old. Results: Mice receiving broccoli exhibited lower hepatic triglycerides (P broccoli feeding (P = 0.006), whereas microsomal triglyceride transfer protein was upregulated (P = 0.045), supporting the finding that dietary broccoli decreased hepatic triglycerides. Conclusion: Long-term consumption of whole broccoli countered both NAFLD development enhanced by a Western diet and hepatic tumorigenesis induced by DEN in male B6C3F1 mice. PMID:26865652

  16. Dietary Broccoli Lessens Development of Fatty Liver and Liver Cancer in Mice Given Diethylnitrosamine and Fed a Western or Control Diet.

    Science.gov (United States)

    Chen, Yung-Ju; Wallig, Matthew A; Jeffery, Elizabeth H

    2016-03-01

    The high-fat and high-sugar Westernized diet that is popular worldwide is associated with increased body fat accumulation, which has been related to the development of nonalcoholic fatty liver disease (NAFLD). Without treatment, NAFLD may progress to hepatocellular carcinoma (HCC), a cancer with a high mortality rate. The consumption of broccoli in the United States has greatly increased in the last 2 decades. Epidemiologic studies show that incorporating brassica vegetables into the daily diet lowers the risk of several cancers, although, to our knowledge, this is the first study to evaluate HCC prevention through dietary broccoli. We aimed to determine the impact of dietary broccoli on hepatic lipid metabolism and the progression of NAFLD to HCC. Our hypothesis was that broccoli decreases both hepatic lipidosis and the development of HCC in a mouse model of Western diet-enhanced liver cancer. Adult 5-wk-old male B6C3F1 mice received a control diet (AIN-93M) or a Western diet (high in lard and sucrose, 19% and 31%, wt:wt, respectively), with or without freeze-dried broccoli (10%, wt:wt). Starting the following week, mice were treated once per week with diethylnitrosamine (DEN; 45 mg/kg body weight intraperitoneally at ages 6, 7, 8, 10, 11, and 12 wk). Hepatic gene expression, lipidosis, and tumor outcomes were analyzed 6 mo later, when mice were 9 mo old. Mice receiving broccoli exhibited lower hepatic triglycerides (P broccoli feeding (P = 0.006), whereas microsomal triglyceride transfer protein was upregulated (P = 0.045), supporting the finding that dietary broccoli decreased hepatic triglycerides. Long-term consumption of whole broccoli countered both NAFLD development enhanced by a Western diet and hepatic tumorigenesis induced by DEN in male B6C3F1 mice. © 2016 American Society for Nutrition.

  17. High resolution mass spectrometry studies of sulforaphane and indole-3-carbinol in broccoli.

    Science.gov (United States)

    Kokotou, Maroula G; Revelou, Panagiota-Kyriaki; Pappas, Christos; Constantinou-Kokotou, Violetta

    2017-12-15

    Broccoli is a rich source of bioactive compounds. Among them, sulforaphane and indole-3-carbinol have attracted a lot of attention, since their consumption is associated with reduced risk of cancer. In this work, the development of an efficient and direct method for the simultaneous determination of sulforaphane and indole-3-carbinol in broccoli using UPLC-HRMS/MS is described. The correlation coefficient, and limits of detection (LOD) and quantification (LOQ) were 0.993, 0.77mg/L and 2.35mg/L for sulforaphane and 0.997, 0.42mg/L, 1.29mg/L for indole-3-carbinol, respectively. The content of sulforaphane and indole-3-carbinol varied between 72±9-304±2mg and 77±1-117±3mg per 100g of fresh florets, respectively. Taking into consideration the differences in cultivar, geography, season and environmental factors, the results agreed with values published in the literature using other techniques. Copyright © 2017 Elsevier Ltd. All rights reserved.

  18. Ultrasound assisted immersion freezing of broccoli (Brassica oleracea L. var. botrytis L.).

    Science.gov (United States)

    Xin, Ying; Zhang, Min; Adhikari, Benu

    2014-09-01

    The aim of this study was to research the ultrasound-assisted freezing (UAF) of broccoli. CaCl2 solution was used as freezing medium. The comparative advantage of using UAF over normal freezing on the freezing time, cell-wall bound calcium to total calcium ratio, textural properties, color, drip loss and L-ascorbic acid contents was evaluated. The application of UAF at selected acoustic intensity with a range of 0.250-0.412 W/cm(2) decreased the freezing time and the loss of cell-wall bound calcium content. Compared to normal freezing, the values of textural properties, color, L-ascorbic acid content were better preserved and the drip loss was significantly minimized by the application of UAF. However, when outside that range of acoustic intensity, the quality of the ultrasound-assisted frozen broccoli was inferior compared to that of the normally frozen samples. Selected the appropriate acoustic intensity was very important for the application of UAF. Copyright © 2014 Elsevier B.V. All rights reserved.

  19. Stingless bees damage broccoli inflorescences when collecting fibers for nest building

    Directory of Open Access Journals (Sweden)

    Adriano Jorge Nunes dos Santos

    2012-01-01

    Full Text Available The stingless bee Trigona spinipes (Fabricius, 1793 (Hymenoptera: Apidae is an important pollinator for various crops, but constitutes an occasional pest of other plant species since it causes injury to leaves, stems, flowers and fruits while collecting nest materials. The aim of the present study was to determine the damage caused by T. spinipes to a broccoli (Brassica oleracea L. var. italica, Brassicaceae growing on an organic farm. A significant number of plants (72.5 % presented damaged inflorescences, while 39% of all of the inflorescences suffered some degree of injury. The activities of T. spinipes caused scarifications on the stems of the inflorescences, and these typically evolved to epidermal cicatrices up to 10 mm wide. In some cases, the lesions were sufficiently deep to cause partial destruction of the vascular tissues, and this lead to thinner (< 5 mm diameter floral stems that may collapse. To the best of our knowledge, this is the first report concerning the attack of broccoli plants by T. spinipes. The results obtained should serve to highlight the possibility that stingless bees could be responsible for direct and/or indirect damage to vegetable crops, and to stimulate the development of control strategies for these incidental pests.

  20. Multiwalled carbon nanotubes enter broccoli cells enhancing growth and water uptake of plants exposed to salinity.

    Science.gov (United States)

    Martínez-Ballesta, M Carmen; Zapata, Lavinia; Chalbi, Najla; Carvajal, Micaela

    2016-06-08

    Carbon nanotubes have been shown to improve the germination and growth of some plant species, extending the applicability of the emerging nano-biotechnology field to crop science. In this work, exploitation of commercial multiwalled carbon nanotubes (MWCNTs) in control and 100 mM NaCl-treated broccoli was performed. Transmission electron microscopy demonstrated that MWCNTs can enter the cells in adult plants with higher accumulation under salt stress. Positive effect of MWCNTs on growth in NaCl-treated plants was consequence of increased water uptake, promoted by more-favourable energetic forces driving this process, and enhanced net assimilation of CO2. MWCNTs induced changes in the lipid composition, rigidity and permeability of the root plasma membranes relative to salt-stressed plants. Also, enhanced aquaporin transduction occurred, which improved water uptake and transport, alleviating the negative effects of salt stress. Our work provides new evidences about the effect of MWCNTs on plasma membrane properties of the plant cell. The positive response to MWCNTs in broccoli plants opens novel perspectives for their technological uses in new agricultural practices, especially when 1plants are exposed to saline environments.

  1. Optimized extraction, separation and quantification of twelve intact glucosinolates in broccoli leaves.

    Science.gov (United States)

    Ares, Ana M; Nozal, María J; Bernal, José L; Bernal, José

    2014-01-01

    A new method has been developed and validated to determine twelve intact glucosinolates (glucoiberin, GIB; glucoraphanin, GRA; glucoerucin GER; gluconapin, GNA; glucotropaeolin, GTL; glucobrassicin, GBC; gluconasturtiin, GST; glucoalyssin, ALY; 4-hydroxyglucobrassicin, 4-OH; 4-metoxyglucobrassicin, 4ME; neoglucobrassicin, NEO; sinigrin, SIN) in broccoli leaves using liquid chromatography (LC) coupled to diode array (DAD) and electrospray ionization mass spectrometry (ESI-MS) detection. An extraction procedure has also been proposed and optimized by means of statistical analysis (the Box-Behnken design and analysis of variance); this is based on the deactivation of myrosinase using a microwave and heated water. Low limits of detection and quantification were obtained, ranging from 10 to 72 μg/g with DAD and 0.01 to 0.23 μg/g with ESI-MS, and the resulting recovery values ranged from 87% to 106% in all cases. Finally, glucosinolates were analyzed in broccoli leaf samples from six different cultivars (Ramoso calabrese Parthenon, Marathon, Nubia, Naxos and Viola). Copyright © 2013 Elsevier Ltd. All rights reserved.

  2. Collembola Diversity between Chemical Pesticide and Bioinsecticide in Broccoli Farm (Brassica oleraceae var. italica)

    Science.gov (United States)

    Sanjaya, Y.; Suhara

    2017-02-01

    The existance of Collembola diversity was determined by how land system work. Farming systems with excessive pesticide application can reduce number of Collembola. On the other hand nowaday people aware of environment by using bioinsecticide. The Method were comparing two land system which use Chemical pestisice and Bioinsecticide. Procedure were using Trapping wells (PMS) in three plots; T0: control without treatment, T1: Chemical Insecticide, T2 : Bioinsecticide for 24 hours. The factors that measure are abiotic factors by taking 10 grams of soil planting Broccoli (Brassica oleraceae var Italica), after 24 hours of taking separates it with other land animals, then identifying Collembola species were using Microcam based on identification book of Collembola. The result showed that density and Biodiversity of land system bioinsecticide was the highest value and indic. It was found also that in Broccoli farm dicovered 3 Familia and 8 species of Collembola both litter and soil. Species found that Isotomurus sp, Seira sp, Lepidosira sp, Coecobrya sp, Callyntura sp, Homidia sp, Sallina sp and Ascocytrus sp, three Family is derived from Isotomidae, Entomobryidae and Paronellid.

  3. Glucosinolates in broccoli sprouts (Brassica oleracea var. italica) as conditioned by sulphate supply during germination.

    Science.gov (United States)

    Pérez-Balibrea, Santiago; Moreno, Diego A; García-Viguera, Cristina

    2010-10-01

    Sulphur (S) fertilization is essential for primary and secondary metabolism in cruciferous foods. Deficient, suboptimal, or excessive S affects the growth and biosynthesis of secondary metabolites in adult plants. Nevertheless, there is little information regarding the influence of S fertilization on sprouts and seedlings. An experiment was set up to evaluate the effect of S fertilization, supplied as K(2)SO(4) at 0, 15, 30, and 60 mg/L, on the glucosinolate content of broccoli sprouts during the germination course of 3, 6, 9, and 12 d after sowing. Glucosinolate concentration was strongly influenced by germination, causing a rapid increase during the first 3 d after sowing, and decreasing afterwards. The S supply increased aliphatic and total glucosinolate content at the end of the monitored sprouting period. S-treated sprouts, with S(15), S(30), and S(60) at 9 and 12 d after sowing presented enhanced glucosinolate content. Overall, both germination time and S fertilization were key factors in maximizing the bioactive health-promoting phytochemicals of broccoli. Practical Application: Germination with sulphate is a simple and inexpensive way to obtain sprouts that contain much higher levels of glucosinolates (health promoting compounds), than the corresponding florets from the same seeds.

  4. Pectobacterium carotovorum subsp. carotovorum – the causal agent of broccoli soft rot in Serbia

    Directory of Open Access Journals (Sweden)

    Katarina Gašić

    2014-12-01

    Full Text Available Soft rot symptoms were observed on broccoli plants in several commercial fields in the western part of Serbia. Six strains of bacteria were isolated from diseased tissues and identified as Pectobacterium carotovorum subsp. carotovorum using conventional bacteriological and molecular methods. All strains were non-fluorescent, gram-negative, facultative anaerobes, oxidase-negative and catalase-positive, causing soft rot on potato and carrot slices and did not induce hypersensitive reaction on tobacco leaves. They grew in 5% NaCl and at 37°C, did not produce acid from α-methyl glucoside, sorbitol and maltose, nor reducing substances from sucrose, but utilized lactose and trehalose, and did not produce indole or lecithinase. The investigated strains showed characteristic growth on Logan’s medium and did not produce blue pigmented indigoidine on GYCA medium nor “fried egg” colonies on PDA. The identity of strains was confirmed by ITS-PCR and ITS-RFLP analyses and by sequence analysis of the 16S rRNA gene. In a pathogenicity assay, all strains caused tissue discoloration and soft rot development on inoculated broccoli head tissue fragments.

  5. Identification and characterization of microRNAs related to salt stress in broccoli, using high-throughput sequencing and bioinformatics analysis.

    Science.gov (United States)

    Tian, Yunhong; Tian, Yunming; Luo, Xiaojun; Zhou, Tao; Huang, Zuoping; Liu, Ying; Qiu, Yihan; Hou, Bing; Sun, Dan; Deng, Hongyu; Qian, Shen; Yao, Kaitai

    2014-09-03

    MicroRNAs (miRNAs) are a new class of endogenous regulators of a broad range of physiological processes, which act by regulating gene expression post-transcriptionally. The brassica vegetable, broccoli (Brassica oleracea var. italica), is very popular with a wide range of consumers, but environmental stresses such as salinity are a problem worldwide in restricting its growth and yield. Little is known about the role of miRNAs in the response of broccoli to salt stress. In this study, broccoli subjected to salt stress and broccoli grown under control conditions were analyzed by high-throughput sequencing. Differential miRNA expression was confirmed by real-time reverse transcription polymerase chain reaction (RT-PCR). The prediction of miRNA targets was undertaken using the Kyoto Encyclopedia of Genes and Genomes (KEGG) Orthology (KO) database and Gene Ontology (GO)-enrichment analyses. Two libraries of small (or short) RNAs (sRNAs) were constructed and sequenced by high-throughput Solexa sequencing. A total of 24,511,963 and 21,034,728 clean reads, representing 9,861,236 (40.23%) and 8,574,665 (40.76%) unique reads, were obtained for control and salt-stressed broccoli, respectively. Furthermore, 42 putative known and 39 putative candidate miRNAs that were differentially expressed between control and salt-stressed broccoli were revealed by their read counts and confirmed by the use of stem-loop real-time RT-PCR. Amongst these, the putative conserved miRNAs, miR393 and miR855, and two putative candidate miRNAs, miR3 and miR34, were the most strongly down-regulated when broccoli was salt-stressed, whereas the putative conserved miRNA, miR396a, and the putative candidate miRNA, miR37, were the most up-regulated. Finally, analysis of the predicted gene targets of miRNAs using the GO and KO databases indicated that a range of metabolic and other cellular functions known to be associated with salt stress were up-regulated in broccoli treated with salt. A comprehensive

  6. Sulforaphane formation and bioaccessibility are more affected by steaming time than meal composition during in vitro digestion of broccoli

    NARCIS (Netherlands)

    Sarvan-Kruse, Irmela; Kramer, E.; Bouwmeester, Hans; Dekker, Matthijs; Verkerk, R.

    2017-01-01

    Broccoli is a rich source of the glucosinolate glucoraphanin (GR). After hydrolysis of GR by the endogenous enzyme myrosinase, sulforaphane (SF) or sulforaphane nitrile (SFN) are produced, depending on environmental conditions. How the conversion of GR and bioaccessibility of released breakdown

  7. Biotechnological applications in in vitro plant regeneration studies of broccoli (Brassica oleracea L. var. italica), an important vegetable crop.

    Science.gov (United States)

    Kumar, Pankaj; Srivastava, Dinesh Kumar

    2016-04-01

    Biotechnology holds promise for genetic improvement of important vegetable crops. Broccoli (Brassica oleracea L. var. italica) is an important vegetable crop of the family Brassicaceae. However, various biotic and abiotic stresses cause enormous crop yield losses during commercial cultivation of broccoli. Establishment of a reliable, reproducible and efficient in vitro plant regeneration system with cell and tissue culture is a vital prerequisite for biotechnological application of crop improvement programme. An in vitro plant regeneration technique refers to culturing, cell division, cell multiplication, de-differentiation and differentiation of cells, protoplasts, tissues and organs on defined liquid/solid medium under aseptic and controlled environment. Recent progress in the field of plant tissue culture has made this area one of the most dynamic and promising in experimental biology. There are many published reports on in vitro plant regeneration studies in broccoli including direct organogenesis, indirect organogenesis and somatic embryogenesis. This review summarizes those plant regeneration studies in broccoli that could be helpful in drawing the attention of the researchers and scientists to work on it to produce healthy, biotic and abiotic stress resistant plant material and to carry out genetic transformation studies for the production of transgenic plants.

  8. Salinity’s influence on boron toxicity in broccoli: II. Impacts on boron uptake, uptake mechanisms and tissue ion relations.

    Science.gov (United States)

    Limited research has been conducted on the interactive effects of salinity and boron stresses on plants despite their common occurrence in natural systems. The purpose of this research was to determine and quantify the interactive effects of salinity, salt composition and boron on broccoli (Brassica...

  9. Quantitative trait loci mapping of heat tolerance in broccoli (Brassica oleracea var. italica) using genotyping-by-sequencing

    Science.gov (United States)

    Predicted rising global temperatures due to climate change have generated a demand for crops that are resistant to yield and quality losses from heat stress. Broccoli (Brassica oleracea var. italica) is a cool weather crop with high temperatures during production decreasing both head quality and yie...

  10. Effect of water content and temperature on inactivation kinetics of myrosinase in broccoli (Brassica oleracea var. italica).

    Science.gov (United States)

    Oliviero, T; Verkerk, R; Van Boekel, M A J S; Dekker, M

    2014-11-15

    Broccoli belongs to the Brassicaceae plant family consisting of widely eaten vegetables containing high concentrations of glucosinolates. Enzymatic hydrolysis of glucosinolates by endogenous myrosinase (MYR) can form isothiocyanates with health-promoting activities. The effect of water content (WC) and temperature on MYR inactivation in broccoli was investigated. Broccoli was freeze dried obtaining batches with WC between 10% and 90% (aw from 0.10 to 0.96). These samples were incubated for various times at different temperatures (40-70°C) and MYR activity was measured. The initial MYR inactivation rates were estimated by the first-order reaction kinetic model. MYR inactivation rate constants were lower in the driest samples (10% WC) at all studied temperatures. Samples with 67% and 90% WC showed initial inactivation rate constants all in the same order of magnitude. Samples with 31% WC showed intermediate initial inactivation rate constants. These results are useful to optimise the conditions of drying processes to produce dried broccoli with optimal MYR retention for human health. Copyright © 2014 Elsevier Ltd. All rights reserved.

  11. Chemopreventive glucosinolate accumulation in various broccoli and collard tissues: Microfluidic-based targeted transcriptomics for by-product valorization

    Science.gov (United States)

    Becker, Talon M.; Juvik, John A.

    2017-01-01

    Floret, leaf, and root tissues were harvested from broccoli and collard cultivars and extracted to determine their glucosinolate and hydrolysis product profiles using high performance liquid chromatography and gas chromotography. Quinone reductase inducing bioactivity, an estimate of anti-cancer chemopreventive potential, of the extracts was measured using a hepa1c1c7 murine cell line. Extracts from root tissues were significantly different from other tissues and contained high levels of gluconasturtiin and glucoerucin. Targeted gene expression analysis on glucosinolate biosynthesis revealed that broccoli root tissue has elevated gene expression of AOP2 and low expression of FMOGS-OX homologs, essentially the opposite of what was observed in broccoli florets, which accumulated high levels of glucoraphanin. Broccoli floret tissue has significantly higher nitrile formation (%) and epithionitrile specifier protein gene expression than other tissues. This study provides basic information of the glucosinolate metabolome and transcriptome for various tissues of Brassica oleracea that maybe utilized as potential byproducts for the nutraceutical market. PMID:28945821

  12. Quantitative trait loci mapping of heat tolerance in broccoli (Brassica oleracea var. italica) using genotyping-by-sequencing.

    Science.gov (United States)

    Branham, Sandra E; Stansell, Zachary J; Couillard, David M; Farnham, Mark W

    2017-03-01

    Five quantitative trait loci and one epistatic interaction were associated with heat tolerance in a doubled haploid population of broccoli evaluated in three summer field trials. Predicted rising global temperatures due to climate change have generated a demand for crops that are resistant to yield and quality losses from heat stress. Broccoli (Brassica oleracea var. italica) is a cool weather crop with high temperatures during production decreasing both head quality and yield. Breeding for heat tolerance in broccoli has potential to both expand viable production areas and extend the growing season but breeding efficiency is constrained by limited genetic information. A doubled haploid (DH) broccoli population segregating for heat tolerance was evaluated for head quality in three summer fields in Charleston, SC, USA. Multiple quantitative trait loci (QTL) mapping of 1,423 single nucleotide polymorphisms developed through genotyping-by-sequencing identified five QTL and one positive epistatic interaction that explained 62.1% of variation in heat tolerance. The QTL identified here can be used to develop markers for marker-assisted selection and to increase our understanding of the molecular mechanisms underlying plant response to heat stress.

  13. An overview of health-promoting compounds of broccoli (Brassica oleracea var. italica) and the effect of processing.

    Science.gov (United States)

    Mahn, Andrea; Reyes, Alejandro

    2012-12-01

    Broccoli offers many heath-promoting properties owing to its content of antioxidant and anticarcinogenic compounds. The concentration and bioavailability of polyphenols, glucosinolates, sulforaphane and selenium depend on plant biochemistry, cultivation strategy and type of processing. In this article, the main biochemical properties of broccoli are reviewed regarding their health-promoting effects. Additionally, the way these properties are affected by processing is discussed. Steaming and drying result in an apparent increment of sulforaphane content as well as antioxidant activity, most likely due to an increase of the extractability of antioxidants and sulforaphane. Freezing and boiling diminish polyphenols concentration, mainly due to volatilization and leaching into the cooking water. In view of these results, the optimization of broccoli processing in order to maximize the content of bioactive compounds should be possible. The effect of processing on selenium compounds has been poorly studied so far, and therefore this topic should be investigated in the future. Finally, the effect of operating conditions in different drying processes on the content of bioactive compounds in broccoli should be investigated in a greater depth.

  14. Selenium biofortification of broccoli and carrots grown in soil amended with Se-enriched hyperaccumulator Stanleya pinnata

    Science.gov (United States)

    Amending soils with Se-hyperaccumulator plant derived sources of selenium (Se) may be useful for increasing Se content in food crops in Se-deficient regions of the world. In this study, we evaluated total Se and the different chemical species of Se in broccoli and carrots grown in soils amended with...

  15. QTL mapping for quinone reductase activity in broccoli with Hepa1c1c7 cell lines

    Science.gov (United States)

    Floret tissue from 125 F2:3 broccoli families derived from the cross 'VI-158 x Brocolette Neri E. Cespuglio (BNC)' was harvested in 2009. Tissue was freeze-dried and stored in the dark at -80 until use. Distilled water was added to floret tissue (50 mg/mL) and auto-hydrolyzed for 24 hours in room te...

  16. Temperature and light conditions at different latitudes affect sensory quality of broccoli florets (Brassica oleracea L. var. italica).

    Science.gov (United States)

    Johansen, Tor J; Mølmann, Jørgen Ab; Bengtsson, Gunnar B; Schreiner, Monica; Velasco, Pablo; Hykkerud, Anne L; Cartea, Elena; Lea, Per; Skaret, Josefine; Seljåsen, Randi

    2017-08-01

    Broccoli (Brassica oleracea L. var. italica) is a popular vegetable grown at a wide range of latitudes. Plants were grown in 2009-2011 in pots with standardized soil, irrigation and nutrient supply under natural temperature and light conditions at four locations (42-70° N). A descriptive sensory analysis of broccoli florets was performed by a trained panel to examine any differences along the latitudinal gradient for 30 attributes within appearance, odour, taste/flavour and texture. Average results over three summer seasons in Germany, southern Norway and northern Norway showed that the northernmost location with low temperatures and long days had highest scores for bud coarseness and uniform colour, while broccoli from the German location, with high temperatures and shorter days, had highest intensity of colour hue, whiteness, bitter taste, cabbage flavour, stale flavour and watery flavour. Results from two autumn seasons at the fourth location (42° N, Spain), with low temperatures and short days, tended toward results from the two northernmost locations, with an exception for most texture attributes. Results clearly demonstrate that temperature and light conditions related to latitude and season affect the sensory quality of broccoli florets. Results may be used in marketing special quality regional or seasonal products. © 2016 Society of Chemical Industry. © 2016 Society of Chemical Industry.

  17. The response of broccoli (Brassica oleracea convar. italica) varieties on foliar application of selenium: uptake, translocation, and speciation.

    Science.gov (United States)

    Šindelářová, Kristýna; Száková, Jiřina; Tremlová, Jana; Mestek, Oto; Praus, Lukáš; Kaňa, Antonín; Najmanová, Jana; Tlustoš, Pavel

    2015-01-01

    A model small-scale field experiment was set up to investigate selenium (Se) uptake by four different varieties of broccoli plants, as well as the effect of Se foliar application on the uptake of essential elements for plants calcium (Ca), copper (Cu), iron (Fe), potassium (K), magnesium (Mg), manganese (Mn), phosphorus (P), sulfur (S), and zinc (Zn). Foliar application of sodium selenate (Na2SeO4) was carried out at two rates (25 and 50 g Se/ha), and an untreated control variant was included. Analyses of individual parts of broccoli were performed, whereby it was found that Se in the plant accumulates mainly in the flower heads and slightly less in the leaves, stems, and roots, regardless of the Se rate and broccoli variety. In most cases, there was a statistically significant increase of Se content in all parts of the plant, while there was no confirmed systematic influence of the addition of Se on the changing intake of other monitored elements. Selenization of broccoli leads to an effective increase in the Se content at a rate of 25 g/ha, whereas the higher rate did not result in a substantial increase of Se content compared to the lower rate in all varieties. Therefore, the rate of 25 g/ha can be recommended as effective to produce broccoli with an increased Se content suitable for consumption. Moreover, Se application resulted in an adequate increase of the main organic compounds of Se, such as selenocystine (SeCys2), selenomethionine (SeMet), and Se-methylselenocysteine (Se-MeSeCys).

  18. Isoforms of purified methyltransferase from human blood platelets ...

    African Journals Online (AJOL)

    A membrane-bound protein with N-methyltransferase activity, associated with phospholipid metabolism, has been isolated from purified human blood platelet plasma membranes. The activity of this enzyme has been detected in crude platelet preparations. However, the nature and properties of this enzyme and its ...

  19. Cloning and characterization of a γ-tocopherol methyltransferase ...

    African Journals Online (AJOL)

    ... reading frame of 1041 nucleotides encoding a protein of 39 kD polypeptide. A BoTMT whole cell system was developed for the production of α-tocopherol through the expression of BoTMT in Escherichia coli BL21 (DE3) strain. Keywords: γ-Tocopherol methyltransferase, Chinese cabbage, Perilla frustescens, tocopherol, ...

  20. Label-free electrochemical detection of human methyltransferase from tumors.

    Science.gov (United States)

    Furst, Ariel L; Muren, Natalie B; Hill, Michael G; Barton, Jacqueline K

    2014-10-21

    The role of abnormal DNA methyltransferase activity in the development and progression of cancer is an essential and rapidly growing area of research, both for improved diagnosis and treatment. However, current technologies for the assessment of methyltransferase activity, particularly from crude tumor samples, limit this work because they rely on radioactivity or fluorescence and require bulky instrumentation. Here, we report an electrochemical platform that overcomes these limitations for the label-free detection of human DNA(cytosine-5)-methyltransferase1 (DNMT1) methyltransferase activity, enabling measurements from crude cultured colorectal cancer cell lysates (HCT116) and biopsied tumor tissues. Our multiplexed detection system involving patterning and detection from a secondary electrode array combines low-density DNA monolayer patterning and electrocatalytically amplified DNA charge transport chemistry to measure selectively and sensitively DNMT1 activity within these complex and congested cellular samples. Based on differences in DNMT1 activity measured with this assay, we distinguish colorectal tumor tissue from healthy adjacent tissue, illustrating the effectiveness of this two-electrode platform for clinical applications.

  1. High frequency organogenesis in hypocotyl, cotyledon, leaf and petiole explants of broccoli (Brassica oleracea L. var. italica), an important vegetable crop

    National Research Council Canada - National Science Library

    Kumar, Pankaj; Srivastava, D K

    2015-01-01

    ... organogenesis from hypocotyl, cotyledon, leaf and petiole explants of broccoli (Brassica oleracea L. var. italica cv. Solan green head) has been developed. Hypocotyl and cotyledon explants were used from 10 to 12...

  2. High frequency organogenesis in hypocotyl, cotyledon, leaf and petiole explants of broccoli (Brassica oleracea L. var. italica), an important vegetable crop

    National Research Council Canada - National Science Library

    Kumar, Pankaj; Srivastava, D K

    2015-01-01

    Broccoli (Brassica oleracea L. var. italica) is an important, nutritionally rich vegetable crop, but severely affected by environmental stresses, pests and diseases which cause massive yield and quality losses...

  3. LC-MS/MS quantification of sulforaphane and indole-3-carbinol metabolites in human plasma and urine after dietary intake of selenium-fortified broccoli.

    Science.gov (United States)

    Hauder, Johanna; Winkler, Stefanie; Bub, Achim; Rüfer, Corinna E; Pignitter, Marc; Somoza, Veronika

    2011-08-10

    This study aimed at developing a sensitive LC-MS/MS method for the quantification of sulforaphane (SFN) and indole-3-carbinol metabolites in plasma and urine after dietary intake of regular and selenium-fertilized broccoli using stable isotope dilution analysis. In a three-armed, placebo-controlled, randomized human intervention study with 76 healthy volunteers, 200 g of regular (485 μg of total glucosinolates and fertilized broccoli (589 μg of total glucosinolates and 0.25 μg of selenium per gram fresh weight) was administered daily for 4 weeks. Glucoraphanin and glucobrassicin metabolites quantified in plasma and urine were SFN-glutathione, SFN-cysteine, SFN-cysteinylglycine, SFN-acetylcysteine, and indole-3-carboxaldehyde, indole-3-carboxylic acid, and ascorbigen, respectively. Dietary intake of selenium-fertilized broccoli increased serum selenium concentration analyzed by means of atomic absorption spectroscopy by up to 25% (p urine compared to regular broccoli.

  4. Potential of cultivar and crop management to affect phytochemical content in winter-grown sprouting broccoli (Brassica oleracea L. var. italica).

    Science.gov (United States)

    Reilly, Kim; Valverde, Juan; Finn, Leo; Rai, Dilip K; Brunton, Nigel; Sorensen, Jens C; Sorensen, Hilmer; Gaffney, Michael

    2014-01-30

    Variety and crop management strategies affect the content of bioactive compounds (phenolics, flavonoids and glucosinolates) in green broccoli (calabrese) types, which are cultivated during summer and autumn in temperate European climates. Sprouting broccoli types are morphologically distinct and are grown over the winter season and harvested until early spring. Thus they show considerable potential for development as an import substitution crop for growers and consumers during the 'hungry gap' of early spring. The present study investigated the effect of variety and management practices on phytochemical content in a range of sprouting broccoli varieties. Yields were significantly higher in white sprouting broccoli varieties. Levels of phenolics and flavonoids were in the range 81.64-297.65 and 16.95-104.80 mg 100 g⁻¹ fresh weight, respectively, depending on year and cultivar, and were highest in variety 'TZ 5052' in both years. In-row spacing did not affect flavonoid content. Phenolic and flavonoid content generally increased with increasing floret maturity and levels were high in edible portions of the crop. Crop wastes (leaf and flower) contained 145.9-239.3 and 21.5-116.6 mg 100 g⁻¹ fresh weight total phenolics and flavonoids, respectively, depending on cultivar, tissue and year. Climatic factors had a significant effect on phenolic and flavonoid content. Levels of total and some individual glucosinolates were higher in sprouting broccoli than in the green broccoli variety 'Ironman'. Levels of total phenolics, flavonoids and glucosinolates are higher in sprouting than green broccoli types. Sprouting broccoli represents an excellent source of dietary bioactive compounds. © 2013 Society of Chemical Industry.

  5. Changes in SeMSC, glucosinolates and sulforaphane levels, and in proteome profile in broccoli (Brassica oleracea var. Italica) fertilized with sodium selenate.

    Science.gov (United States)

    Sepúlveda, Ignacio; Barrientos, Herna; Mahn, Andrea; Moenne, Alejandra

    2013-05-07

    The aim of this work was to analyze the effect of sodium selenate fortification on the content of selenomethyl selenocysteine (SeMSC), total glucosinolates and sulforaphane, as well as the changes in protein profile of the inflorescences of broccoli (Brassica oleracea var. Italica). Two experimental groups were considered: plants treated with 100 μmol/L sodium selenate (final concentration in the pot) and control plants treated with water. Fortification began 2 weeks after transplantation and was repeated once a week during 10 weeks. Broccoli florets were harvested when they reached appropriate size. SeMSC content in broccoli florets increased significantly with sodium selenate fortification; but total glucosinolates and sulforaphane content as well as myrosinase activity were not affected. The protein profile of broccoli florets changed due to fortification with sodium selenate. Some proteins involved in general stress-responses were up-regulated, whereas down-regulated proteins were identified as proteins involved in protection against pathogens. This is the first attempt to evaluate the physiological effect of fortification with sodium selenate on broccoli at protein level. The results of this work will contribute to better understanding the metabolic processes related with selenium uptake and accumulation in broccoli.

  6. Influence of thermal processing on hydrolysis and stability of folate poly-gamma-glutamates in broccoli (Brassica oleracea var. italica), carrot (Daucus carota) and tomato (Lycopersicon esculentum).

    Science.gov (United States)

    Munyaka, Ann Wambui; Verlinde, Philippe; Mukisa, Ivan Muzira; Oey, Indrawati; Van Loey, Ann; Hendrickx, Marc

    2010-04-14

    The folate poly-gamma-glutamate profile, their concentrations, and hydrolysis by endogenous gamma-glutamyl hydrolase (GGH) were evaluated in broccoli, carrot and tomato. Further studies on the effect of time and temperature on folate poly-gamma-glutamate hydrolysis and stability were carried out in broccoli since this vegetable showed the highest long-chain and total folate poly-gamma-glutamate concentration. The evolution of l-ascorbic acid, total phenols and Trolox equivalent antioxidant capacity (TEAC) values was evaluated in parallel. Upon thermal inactivation of GGH prior to crushing, it was observed that broccoli, carrot and tomato contained poly-gamma-glutamates with one to seven glutamate residues but differed in the predominant poly-gamma-glutamates. Crushing of raw broccoli, carrot and tomato resulted in significant poly-gamma-glutamate profile changes in broccoli and carrot (indicating GGH-catalyzed hydrolysis) but not in tomato. In this study, the actual crushing of raw broccoli matrix had a greater effect on folate poly-gamma-glutamate hydrolysis than incubation conditions (0-30 min at 25-55 degrees C). During treatments at 25-140 degrees C, folate retention was higher at 80 and 100 degrees C than at the other temperatures. A similar trend in thermal stability was observed for folates, vitamin C, total phenols and TEAC value, an indication that conditions that result in endogenous antioxidants degradation might also result in folate degradation.

  7. Changes in SeMSC, Glucosinolates and Sulforaphane Levels, and in Proteome Profile in Broccoli (Brassica oleracea var. Italica Fertilized with Sodium Selenate

    Directory of Open Access Journals (Sweden)

    Alejandra Moenne

    2013-05-01

    Full Text Available The aim of this work was to analyze the effect of sodium selenate fortification on the content of selenomethyl selenocysteine (SeMSC, total glucosinolates and sulforaphane, as well as the changes in protein profile of the inflorescences of broccoli (Brassica oleracea var. Italica. Two experimental groups were considered: plants treated with 100 mmol/L sodium selenate (final concentration in the pot and control plants treated with water. Fortification began 2 weeks after transplantation and was repeated once a week during 10 weeks. Broccoli florets were harvested when they reached appropriate size. SeMSC content in broccoli florets increased significantly with sodium selenate fortification; but total glucosinolates and sulforaphane content as well as myrosinase activity were not affected. The protein profile of broccoli florets changed due to fortification with sodium selenate. Some proteins involved in general stress-responses were up-regulated, whereas down-regulated proteins were identified as proteins involved in protection against pathogens. This is the first attempt to evaluate the physiological effect of fortification with sodium selenate on broccoli at protein level. The results of this work will contribute to better understanding the metabolic processes related with selenium uptake and accumulation in broccoli.

  8. Molecular identification of carnosine N-methyltransferase as chicken histamine N-methyltransferase-like protein (hnmt-like.

    Directory of Open Access Journals (Sweden)

    Jakub Drozak

    Full Text Available Anserine (beta-alanyl-N(Pi-methyl-L-histidine, a naturally occurring derivative of carnosine (beta-alanyl-L-histidine, is an abundant constituent of skeletal muscles and brain of many vertebrates. Although it has long been proposed to serve as a proton buffer, radicals scavenger and transglycating agent, its physiological function remains obscure. The formation of anserine is catalyzed by carnosine N-methyltransferase which exhibits unknown molecular identity. In the present investigation, we have purified carnosine N-methyltransferase from chicken pectoral muscle about 640-fold until three major polypeptides of about 23, 26 and 37 kDa coeluting with the enzyme were identified in the preparation. Mass spectrometry analysis of these polypeptides resulted in an identification of histamine N-methyltransferase-like (HNMT-like protein as the only meaningful candidate. Analysis of GenBank database records indicated that the hnmt-like gene might be a paralogue of histamine N-methyltransferase gene, while comparison of their protein sequences suggested that HNMT-like protein might have acquired a new activity. Chicken HNMT-like protein was expressed in COS-7 cells, purified to homogeneity, and shown to catalyze the formation of anserine as confirmed by both chromatographic and mass spectrometry analysis. Both specificity and kinetic studies carried out on the native and recombinant enzyme were in agreement with published data. Particularly, several compounds structurally related to carnosine, including histamine and L-histidine, were tested as potential substrates for the enzyme, and carnosine was the only methyl group acceptor. The identification of the gene encoding carnosine N-methyltransferase might be beneficial for estimation of the biological functions of anserine.

  9. Plant regeneration from leaf protoplasts of Brassica oleracea var. italica CV Green Comet broccoli.

    Science.gov (United States)

    Robertson, D; Earle, E D

    1986-02-01

    A procedure is described for regeneration of plants from leaf protoplasts of the hybrid broccoli cultivar, Green Comet (Brassica oleracea var italica). The totipotency of protoplasts isolated from plants regenerated from hypocotyl explants (GCR) was greater than that of protoplasts from plants grown directly from seed (GC). Using medium B developed by Pelletier et al (1983), division efficiencies greater than 70% were obtained in leaf protoplasts isolated from GCR. Approximately 1% of these protoplasts formed calli on solidified medium; 77% of the calli regenerated shoots. In contrast, protoplasts from seed-grown material showed a lower division efficiency (15-22%) and fewer protoplast-derived calli produced shoots. Some of the 178 protoplast-derived plants grown to maturity had variant phenotypes.

  10. Proliferation and glucosinolates accumulation of broccoli adventitious roots in liquid medium

    Science.gov (United States)

    Nhut, Nguyen Minh; Tien, Le Thi Thuy

    2017-09-01

    Cotyledons from 7-day-old in vitro broccoli seedling were used as explant source in adventitious root induction on MS medium supplemented with 30 g/l sucrose, 1.6 mg/l IBA and 7 g/l agar. Adventitious roots from cotyledons were transferred to liquid medium containing the same components as rooting medium for two weeks, then subcultured to MS medium with diferent sugar, macrominerals and casein hydrolysate concentrations. The best adventitious root growth was observed in half-strength MS medium supplemented with 40 g/l sucrose, 600 mg/l casein hydrolysate and 1.6 mg/l IBA (growth index of 4.00 in about 14 culture days with inoculum density of 1.0 g fresh weight / 30 ml of culture medium). The culturing process can be stopped on the 28th day for root biomass and on the 35th day for glucosinolates.

  11. Effects of operating factors on osmotic dehydration of broccoli stalk slices

    Directory of Open Access Journals (Sweden)

    Nora Salina Md Salim

    2016-12-01

    Full Text Available In this study, effects of sucrose concentration (40–60% w/v, immersion time (1–5 h, and temperature (25–65°C on water loss (WL and solute gain (SG during osmotic dehydration of broccoli stalk slices were quantitatively investigated using response surface methodology. It was found that concentration of sucrose solution, immersion time, and temperature affected the WL during osmotic dehydration process. Significant factors affecting the SG are the temperature and immersion time. The operating conditions to obtain water removal of 62% with solute intake of 7% were found to be at a sucrose concentration of 56% w/v, temperature of 42°C, and immersion time of 4 h.

  12. Selenium-Induced Toxicity Is Counteracted by Sulfur in Broccoli (Brassica oleracea L. var. italica).

    Science.gov (United States)

    Tian, Ming; Hui, Maixia; Thannhauser, Theodore W; Pan, Siyi; Li, Li

    2017-01-01

    Selenium (Se) is an essential micronutrient for humans. Increasing Se content in food crops offers an effective approach to enhance the consumption of Se in human diets. A thoroughly understanding of the effects of Se on plant growth is important for Se biofortification in food crops. Given that Se is an analog of sulfur (S) and can be toxic to plants, its effect on plant growth is expected to be greatly affected by S nutrition. However, this remains to be further understood. Here, we evaluated the influence of Se treatments on broccoli (Brassica oleracea L. var. italica) growth when S was withheld from the growth nutrient solution. We found that Se was highly toxic to plants when S nutrition was poor. In contrast to Se treatments with adequate S nutrition that slightly reduced broccoli growth, the same concentration of Se treatments without S supplementation dramatically reduced plant sizes. Higher Se toxicity was observed with selenate than selenite under low S nutrition. We examined the bases underlying the toxicity. We discovered that the high Se toxicity in low S nutrition was specifically associated with an increased ratio of Se in proteins verse total Se level, enhanced generation of reactive oxygen species, elevated lipid peroxidation causing increased cell membrane damage, and reduced antioxidant enzyme activities. Se toxicity could be counteracted with increased supplementation of S, which is likely through decreasing non-specific integration of Se into proteins and altering the redox system. The present study provides information for better understanding of Se toxicity and shows that adequate S nutrition is important to prevent Se toxicity during biofortification of crops by Se fertilization.

  13. Consumer behaviour towards vegetables: a study on domestic processing of broccoli and carrots by Dutch households.

    Science.gov (United States)

    Bongoni, R; Verkerk, R; Dekker, M; Steenbekkers, L P A

    2015-06-01

    Preferences for sensory properties (e.g. taste and texture) are assumed to control cooking behaviour with respect to vegetables. Conditions such as the cooking method, amount of water used and the time-temperature profile determine the nutritional quality (e.g. vitamins and phytochemicals) of cooked vegetables. Information on domestic processing and any underlying motives can be used to inform consumers about cooking vegetables that are equally liked and are nutrient-rich. Two online self-reporting questionnaires were used to identify domestic processing conditions of broccoli and carrots by Dutch households. Questions on various aspects of domestic processing and consumer motives were included. Descriptive data analysis and hierarchical cluster analysis were performed for both vegetables, separately, to group consumers with similar motives and behaviour towards vegetables. Approximately 70% of consumers boiled vegetables, 8-9% steamed vegetables, 10-15% stir fried raw vegetables and 8-10% stir fried boiled vegetables. Mainly texture was used as a way to decide the 'doneness' of the vegetables. For both vegetables, three clusters of consumers were identified: texture-orientated, health-orientated, or taste-orientated. The texture-orientated consumers are identified as the most prevalent (56-59%) group in the present study. Statistically significant associations are found between domestic processing conditions and clusters, whereas no such association are found between demographic details and clusters. A wide variation in domestic processing of broccoli and carrots is found in the present study. Mainly sensory properties (i.e. texture and taste) determined the domestic processing conditions. The findings of the present study can be used to optimise cooking to yield vegetables that meet consumer's specific sensory preference and are higher in nutrients, and as well as to communicate with target consumer groups. © 2014 The British Dietetic Association Ltd.

  14. Selenium-Induced Toxicity Is Counteracted by Sulfur in Broccoli (Brassica oleracea L. var. italica

    Directory of Open Access Journals (Sweden)

    Ming Tian

    2017-08-01

    Full Text Available Selenium (Se is an essential micronutrient for humans. Increasing Se content in food crops offers an effective approach to enhance the consumption of Se in human diets. A thoroughly understanding of the effects of Se on plant growth is important for Se biofortification in food crops. Given that Se is an analog of sulfur (S and can be toxic to plants, its effect on plant growth is expected to be greatly affected by S nutrition. However, this remains to be further understood. Here, we evaluated the influence of Se treatments on broccoli (Brassica oleracea L. var. italica growth when S was withheld from the growth nutrient solution. We found that Se was highly toxic to plants when S nutrition was poor. In contrast to Se treatments with adequate S nutrition that slightly reduced broccoli growth, the same concentration of Se treatments without S supplementation dramatically reduced plant sizes. Higher Se toxicity was observed with selenate than selenite under low S nutrition. We examined the bases underlying the toxicity. We discovered that the high Se toxicity in low S nutrition was specifically associated with an increased ratio of Se in proteins verse total Se level, enhanced generation of reactive oxygen species, elevated lipid peroxidation causing increased cell membrane damage, and reduced antioxidant enzyme activities. Se toxicity could be counteracted with increased supplementation of S, which is likely through decreasing non-specific integration of Se into proteins and altering the redox system. The present study provides information for better understanding of Se toxicity and shows that adequate S nutrition is important to prevent Se toxicity during biofortification of crops by Se fertilization.

  15. 1 Protein Methyltransferases: Their Distribution Among the Five Structural Classes of AdoMet-Dependent Methyltransferases.

    Science.gov (United States)

    Schubert, Heidi L; Blumenthal, Robert M; Cheng, Xiaodong

    2006-01-01

    S-adenosyl-l-methionine (AdoMet) dependent methyltransferases (MTases) are involved in biosynthesis, signal transduction, protein repair, chromatin regulation, and gene silencing. Five different structural folds (designated I through V) have been described that bind AdoMet and catalyze methyltransfer to diverse substrates, although the great majority of known MTases have the Class I fold. Even within a particular MTase class the amino-acid sequence similarity can be as low as 10%. Thus, the structural and catalytic requirements for methyltransfer from AdoMet appear to be remarkably flexible. MTases that act on protein substrates have been found to date among three of the five structural classes (I, the classical fold; III, the corrin MTase fold; and V, the SET fold). "There are many paths to the top of the mountain, but the view is always the same."-Chinese proverb The Columbia World of Quotations, New York, Columbia University Press, 1996. Copyright © 2006 Elsevier Inc. All rights reserved.

  16. Zika Virus Methyltransferase: Structure and Functions for Drug Design Perspectives.

    Science.gov (United States)

    Coutard, Bruno; Barral, Karine; Lichière, Julie; Selisko, Barbara; Martin, Baptiste; Aouadi, Wahiba; Lombardia, Miguel Ortiz; Debart, Françoise; Vasseur, Jean-Jacques; Guillemot, Jean Claude; Canard, Bruno; Decroly, Etienne

    2017-03-01

    The Flavivirus Zika virus (ZIKV) is the causal agent of neurological disorders like microcephaly in newborns or Guillain-Barre syndrome. Its NS5 protein embeds a methyltransferase (MTase) domain involved in the formation of the viral mRNA cap. We investigated the structural and functional properties of the ZIKV MTase. We show that the ZIKV MTase can methylate RNA cap structures at the N-7 position of the cap, and at the 2'-O position on the ribose of the first nucleotide, yielding a cap-1 structure. In addition, the ZIKV MTase methylates the ribose 2'-O position of internal adenosines of RNA substrates. The crystal structure of the ZIKV MTase determined at a 2.01-Å resolution reveals a crystallographic homodimer. One chain is bound to the methyl donor ( S -adenosyl-l-methionine [SAM]) and shows a high structural similarity to the dengue virus (DENV) MTase. The second chain lacks SAM and displays conformational changes in the αX α-helix contributing to the SAM and RNA binding. These conformational modifications reveal a possible molecular mechanism of the enzymatic turnover involving a conserved Ser/Arg motif. In the second chain, the SAM binding site accommodates a sulfate close to a glycerol that could serve as a basis for structure-based drug design. In addition, compounds known to inhibit the DENV MTase show similar inhibition potency on the ZIKV MTase. Altogether these results contribute to a better understanding of the ZIKV MTase, a central player in viral replication and host innate immune response, and lay the basis for the development of potential antiviral drugs. IMPORTANCE The Zika virus (ZIKV) is associated with microcephaly in newborns, and other neurological disorders such as Guillain-Barre syndrome. It is urgent to develop antiviral strategies inhibiting the viral replication. The ZIKV NS5 embeds a methyltransferase involved in the viral mRNA capping process, which is essential for viral replication and control of virus detection by innate immune

  17. The Impact of Broccoli II & Tomato II on European patents in conventional breeding, GMO’s and Synthetic Biology:

    DEFF Research Database (Denmark)

    Minssen, Timo; Nordberg, Ana

    2015-01-01

    On 25 March 2015, the Enlarged Board of Appeal of the European Patent Office (EBA) finally delivered its’ much awaited decisions on the consolidated referrals G2/12 (“Tomato II”) and G2/13 (“Broccoli II”). The EBA affirmed that products, namely plants or parts thereof, obtained by essentially...... biological processes are – unlike individual plant varieties – principally patentable under the European Patent Convention (EPC). This decision leaves considerable leeway for patenting novel and inventive plants and products thereof, which have been produced by “conventional” methods including breeding steps....... The EBA has also clarified that this applies irrespective of if such claims are formulated in a product-by-process format or as a per se product . Moreover, the combined effect of Broccoli & Tomato I & II opens new opportunities for patenting GMOs - provided that all other patent criteria are also met...

  18. The isolation and purification of glucoraphanin from broccoli seeds by solid phase extraction and preparative high performance liquid chromatography.

    Science.gov (United States)

    Rochfort, Simone; Caridi, Domenico; Stinton, Melanie; Trenerry, V Craige; Jones, Rod

    2006-07-07

    Plant foods contain not only essential nutrients, e.g. protein, amino acids, vitamins and minerals, but also phytochemicals that have added health benefits. One such class of phytochemicals are the glucosinolates. Glucosinolates, particularly glucoraphanin, are predominant in plants of the Brassica genus, most notably in vegetables such as broccoli. There is a growing interest in the role glucoraphanin plays in chemoprotection and as a result there is a requirement to accurately determine the levels of glucoraphanin in vegetable products. Reverse phase ion pair high performance liquid chromatography (HPLC) is the method of choice; however, this work has been hindered by the lack of available standard reference materials. Broccoli seeds, which are particularly rich in glucoraphanin (20-50 mg/g), have proved to be ideal for the isolation of glucoraphanin on the preparative scale. A novel preparative scale HPLC method with simple compound recovery has been developed to meet the need for a glucoraphanin standard.

  19. Rapid estimation of glucosinolate thermal degradation rate constants in leaves of Chinese kale and broccoli (Brassica oleracea) in two seasons.

    Science.gov (United States)

    Hennig, Kristin; Verkerk, Ruud; Bonnema, Guusje; Dekker, Matthijs

    2012-08-15

    Kinetic modeling was used as a tool to quantitatively estimate glucosinolate thermal degradation rate constants. Literature shows that thermal degradation rates differ in different vegetables. Well-characterized plant material, leaves of broccoli and Chinese kale plants grown in two seasons, was used in the study. It was shown that a first-order reaction is appropriate to model glucosinolate degradation independent from the season. No difference in degradation rate constants of structurally identical glucosinolates was found between broccoli and Chinese kale leaves when grown in the same season. However, glucosinolate degradation rate constants were highly affected by the season (20-80% increase in spring compared to autumn). These results suggest that differences in glucosinolate degradation rate constants can be due to variation in environmental as well as genetic factors. Furthermore, a methodology to estimate rate constants rapidly is provided to enable the analysis of high sample numbers for future studies.

  20. Methyl jasmonate and 1-methylcyclopropene treatment effects on quinone reductase inducing activity and post-harvest quality of broccoli.

    Directory of Open Access Journals (Sweden)

    Kang Mo Ku

    Full Text Available Effect of pre-harvest methyl jasmonate (MeJA and post-harvest 1-methylcyclopropene (1-MCP treatments on broccoli floret glucosinolate (GS concentrations and quinone reductase (QR, an in vitro anti-cancer biomarker inducing activity were evaluated two days prior to harvest, at harvest and at 10, 20, and 30 days of post-harvest storage at 4 °C. MeJA treatments four days prior to harvest of broccoli heads was observed to significantly increase floret ethylene biosynthesis resulting in chlorophyll catabolism during post-harvest storage and reduced product quality. Post-harvest treatment with 1-methylcyclopropene (1-MCP, which competitively binds to protein ethylene receptors, maintained post-harvest floret chlorophyll concentrations and product visual quality in both control and MeJA-treated broccoli. Transcript abundance of BoPPH, a gene which is responsible for the synthesis of pheophytinase, the primary enzyme associated with chlorophyll catabolism in broccoli, was reduced by 1-MCP treatment and showed a significant, negative correlation with floret chlorophyll concentrations. The GS, glucobrassicin, neoglucobrassicin, and gluconasturtiin were significantly increased by MeJA treatments. The products of some of the GS from endogenous myrosinase hydrolysis [sulforaphane (SF, neoascorbigen (NeoASG, N-methoxyindole-3-carbinol (NI3C, and phenethyl isothiocyanate (PEITC] were also quantified and found to be significantly correlated with QR. Sulforaphane, the isothiocyanate hydrolysis product of the GS glucoraphanin, was found to be the most potent QR induction agent. Increased sulforaphane formation from the hydrolysis of glucoraphanin was associated with up-regulated gene expression of myrosinase (BoMyo and the myrosinase enzyme co-factor gene, epithiospecifier modifier1 (BoESM1. This study demonstrates the combined treatment of MeJA and 1-MCP increased QR activity without post-harvest quality loss.

  1. Methyl jasmonate and 1-methylcyclopropene treatment effects on quinone reductase inducing activity and post-harvest quality of broccoli.

    Science.gov (United States)

    Ku, Kang Mo; Choi, Jeong Hee; Kim, Hyoung Seok; Kushad, Mosbah M; Jeffery, Elizabeth H; Juvik, John A

    2013-01-01

    Effect of pre-harvest methyl jasmonate (MeJA) and post-harvest 1-methylcyclopropene (1-MCP) treatments on broccoli floret glucosinolate (GS) concentrations and quinone reductase (QR, an in vitro anti-cancer biomarker) inducing activity were evaluated two days prior to harvest, at harvest and at 10, 20, and 30 days of post-harvest storage at 4 °C. MeJA treatments four days prior to harvest of broccoli heads was observed to significantly increase floret ethylene biosynthesis resulting in chlorophyll catabolism during post-harvest storage and reduced product quality. Post-harvest treatment with 1-methylcyclopropene (1-MCP), which competitively binds to protein ethylene receptors, maintained post-harvest floret chlorophyll concentrations and product visual quality in both control and MeJA-treated broccoli. Transcript abundance of BoPPH, a gene which is responsible for the synthesis of pheophytinase, the primary enzyme associated with chlorophyll catabolism in broccoli, was reduced by 1-MCP treatment and showed a significant, negative correlation with floret chlorophyll concentrations. The GS, glucobrassicin, neoglucobrassicin, and gluconasturtiin were significantly increased by MeJA treatments. The products of some of the GS from endogenous myrosinase hydrolysis [sulforaphane (SF), neoascorbigen (NeoASG), N-methoxyindole-3-carbinol (NI3C), and phenethyl isothiocyanate (PEITC)] were also quantified and found to be significantly correlated with QR. Sulforaphane, the isothiocyanate hydrolysis product of the GS glucoraphanin, was found to be the most potent QR induction agent. Increased sulforaphane formation from the hydrolysis of glucoraphanin was associated with up-regulated gene expression of myrosinase (BoMyo) and the myrosinase enzyme co-factor gene, epithiospecifier modifier1 (BoESM1). This study demonstrates the combined treatment of MeJA and 1-MCP increased QR activity without post-harvest quality loss.

  2. Small RNA Sequencing Reveals Differential miRNA Expression in the Early Development of Broccoli (Brassica oleracea var. italica) Pollen.

    Science.gov (United States)

    Li, Hui; Wang, Yu; Wu, Mei; Li, Lihong; Jin, Chuan; Zhang, Qingli; Chen, Chengbin; Song, Wenqin; Wang, Chunguo

    2017-01-01

    Pollen development is an important and complex biological process in the sexual reproduction of flowering plants. Although the cytological characteristics of pollen development are well defined, the regulation of its early stages remains largely unknown. In the present study, miRNAs were explored in the early development of broccoli (Brassica oleracea var. italica) pollen. A total of 333 known miRNAs that originated from 235 miRNA families were detected. Fifty-five novel miRNA candidates were identified. Sixty of the 333 known miRNAs and 49 of the 55 predicted novel miRNAs exhibited significantly differential expression profiling in the three distinct developmental stages of broccoli pollen. Among these differentially expressed miRNAs, miRNAs that would be involved in the developmental phase transition from uninucleate microspores to binucleate pollen grains or from binucleate to trinucleate pollen grains were identified. miRNAs that showed significantly enriched expression in a specific early stage of broccoli pollen development were also observed. In addition, 552 targets for 127 known miRNAs and 69 targets for 40 predicted novel miRNAs were bioinformatically identified. Functional annotation and GO (Gene Ontology) analysis indicated that the putative miRNA targets showed significant enrichment in GO terms that were related to plant organ formation and morphogenesis. Some of enriched GO terms were detected for the targets directly involved in plant male reproduction development. These findings provided new insights into the functions of miRNA-mediated regulatory networks in broccoli pollen development.

  3. Resistin mediates tomato and broccoli extract effects on glucose homeostasis in high fat diet-induced obesity in rats.

    Science.gov (United States)

    Aborehab, Nora M; El Bishbishy, Mahitab H; Waly, Nermien E

    2016-07-18

    Resistin is an adipocyte hormone that regulates glucose metabolism. Elevated levels of resistin may cause insulin resistance. This may link obesity, and increased fat mass to type II diabetes and insulin resistance. We hypothesized that treatment with tomato and broccoli extracts regulates glucose homeostasis via modulation of resistin levels in high fat diet-induced obesity rats (HFD). Forty-eight male albino rats were divided into 8 groups as follows: control, HFD, stop fat diet (SD), Tomato 200 mg/kg (T200), Tomato 400 mg/kg (T400), Broccoli 200 mg/kg (B200), Broccoli 400 mg/kg (B400), and Chromax (CX). Treatment continued for 1 month. Serum levels of resistin, leptin, adiponectin, glucose and insulin were measured using ELISA and spectrophotometry. Serum levels of resistin were significantly reduced in the T 200, T 400, B 200, B 400 and CX groups to: 4.13 ± 0.22 ng/ml, 1.51 ± 0.04 ng/ml, 4.13 ± 0.22 ng/ml, 2.32 ± 0.15 ng/ml and 1.37 ± 0.03 ng/ml, respectively, compared to HFD group and SD group (P value broccoli extract treatment regulates glucose homeostasis via reduction of serum resistin and may be a useful non-pharmacological therapy for obesity.

  4. Continued selenium biofortification of carrots and broccoli grown in soils once amended with Se-enriched S. pinnata

    Directory of Open Access Journals (Sweden)

    Gary S. Bañuelos

    2016-08-01

    Full Text Available Selenium (Se biofortification has been practiced in Se-deficient regions throughout the world primarily by adding inorganic sources of Se to the soil. Considering the use of adding organic sources of Se could be useful as an alternative Se amendment for the production of Se-biofortified food crops. In this multi-year micro-plot study, we investigate growing carrots and broccoli in soils that had been previously amended with Se-enriched Stanleya pinnata Pursh (Britton three and four years prior to planting one and two, respectively. Results showed that total and extractable Se concentrations in soils (0-30 cm were 1.65 mg kg-1 and 88 µg L-1, and 0.92 mg kg-1 and 48.6 µg L-1 at the beginning of the growing season for planting one and two, respectively. After each respective growing season, total Se concentrations in the broccoli florets and carrots ranged from 6.99 to 7.83 mg kg-1 and 3.15 to 6.25 mg kg-1 in planting one and two, respectively. In broccoli and carrot plant tissues, SeMet (selenomethionine was the predominant selenoamino acid identified in Se aqueous extracts. In postharvest soils from planting one, phospholipid analyses (PLFA showed that amending the soil with S. pinnata exerted no effect on the microbial biomass, AMF (arbuscular mycorrhizal fungi, actinomycetes and Gram-positive and bacterial PLFA at both 0-5 and 0-30 cm, respectively, three years later. Successfully producing Se-enriched broccoli and carrots three and four years later after amending soil with Se-enriched S. pinnata clearly demonstrates its potential source as an organic Se enriched fertilizer for Se-deficient regions.

  5. Effect of nonwoven jute agrotextile mulch on soil health and productivity of broccoli (Brassica oleracea L.) in lateritic soil.

    Science.gov (United States)

    Manna, Koushik; Kundu, Manik Chandra; Saha, Biplab; Ghosh, Goutam Kumar

    2018-01-16

    A field experiment was conducted in winter season of 2015-2016 in the dry lateritic soil of Eastern India to study the effect of different thicknesses of nonwoven jute agrotextile mulches (NJATM) along with other mulches on soil health, growth and productivity of broccoli (Brassica oleracea L.). The experiment was conducted in randomized block design with six treatments viz., T 1 (control, i.e. no mulching), T 2 (300 gsm NJATM), T 3 (350 gsm NJATM), T 4 (400 gsm NJATM), T 5 (rice straw) and T 6 (black polythene mulch), each of which was replicated four times. The highest average curd weight (355.25 g) and yield (8.53 t ha -1 ) of broccoli were recorded in T 3 treatment. The lowest density of broad leaved weed, sedges and grasses were recorded in T 6 treatment which was statistically at par with T 4 . All the treatments composing of NJATM increased the population of all the soil microbes except bacteria in the root rhizosphere of broccoli from their initial population. On average, the highest population of fungi (54.0 × 10 3  cfu per g) and actinomycetes (134.75 × 10 3  cfu per g) was recorded with T 3 and T 4 treatments respectively in the post-harvest soil. The soil moisture was conserved in all treatments compared to control showing highest moisture content in T 4 treatment. Organic carbon and available N, P and K contents of soil were increased in all mulch treated plots compared to control, and their initial value and their highest value were recorded in T 3 . The NJATM of 350 gsm thickness was very effective compared to other mulches in increasing the growth and productivity of broccoli by suppressing weeds, increasing moisture, microbial population and nutrient content of the lateritic soil.

  6. Rapid and sustainable detoxication of airborne pollutants by broccoli sprout beverage: results of a randomized clinical trial in China.

    Science.gov (United States)

    Egner, Patricia A; Chen, Jian-Guo; Zarth, Adam T; Ng, Derek K; Wang, Jin-Bing; Kensler, Kevin H; Jacobson, Lisa P; Muñoz, Alvaro; Johnson, Jamie L; Groopman, John D; Fahey, Jed W; Talalay, Paul; Zhu, Jian; Chen, Tao-Yang; Qian, Geng-Sun; Carmella, Steven G; Hecht, Stephen S; Kensler, Thomas W

    2014-08-01

    Broccoli sprouts are a convenient and rich source of the glucosinolate, glucoraphanin, which can generate the chemopreventive agent, sulforaphane, an inducer of glutathione S-transferases (GST) and other cytoprotective enzymes. A broccoli sprout-derived beverage providing daily doses of 600 μmol glucoraphanin and 40 μmol sulforaphane was evaluated for magnitude and duration of pharmacodynamic action in a 12-week randomized clinical trial. Two hundred and ninety-one study participants were recruited from the rural He-He Township, Qidong, in the Yangtze River delta region of China, an area characterized by exposures to substantial levels of airborne pollutants. Exposure to air pollution has been associated with lung cancer and cardiopulmonary diseases. Urinary excretion of the mercapturic acids of the pollutants, benzene, acrolein, and crotonaldehyde, were measured before and during the intervention using liquid chromatography tandem mass spectrometry. Rapid and sustained, statistically significant (P ≤ 0.01) increases in the levels of excretion of the glutathione-derived conjugates of benzene (61%), acrolein (23%), but not crotonaldehyde, were found in those receiving broccoli sprout beverage compared with placebo. Excretion of the benzene-derived mercapturic acid was higher in participants who were GSTT1-positive than in the null genotype, irrespective of study arm assignment. Measures of sulforaphane metabolites in urine indicated that bioavailability did not decline over the 12-week daily dosing period. Thus, intervention with broccoli sprouts enhances the detoxication of some airborne pollutants and may provide a frugal means to attenuate their associated long-term health risks. ©2014 American Association for Cancer Research.

  7. Methyl Jasmonate and 1-Methylcyclopropene Treatment Effects on Quinone Reductase Inducing Activity and Post-Harvest Quality of Broccoli

    Science.gov (United States)

    Ku, Kang Mo; Choi, Jeong Hee; Kim, Hyoung Seok; Kushad, Mosbah M.; Jeffery, Elizabeth H.; Juvik, John A.

    2013-01-01

    Effect of pre-harvest methyl jasmonate (MeJA) and post-harvest 1-methylcyclopropene (1-MCP) treatments on broccoli floret glucosinolate (GS) concentrations and quinone reductase (QR, an in vitro anti-cancer biomarker) inducing activity were evaluated two days prior to harvest, at harvest and at 10, 20, and 30 days of post-harvest storage at 4 °C. MeJA treatments four days prior to harvest of broccoli heads was observed to significantly increase floret ethylene biosynthesis resulting in chlorophyll catabolism during post-harvest storage and reduced product quality. Post-harvest treatment with 1-methylcyclopropene (1-MCP), which competitively binds to protein ethylene receptors, maintained post-harvest floret chlorophyll concentrations and product visual quality in both control and MeJA-treated broccoli. Transcript abundance of BoPPH, a gene which is responsible for the synthesis of pheophytinase, the primary enzyme associated with chlorophyll catabolism in broccoli, was reduced by 1-MCP treatment and showed a significant, negative correlation with floret chlorophyll concentrations. The GS, glucobrassicin, neoglucobrassicin, and gluconasturtiin were significantly increased by MeJA treatments. The products of some of the GS from endogenous myrosinase hydrolysis [sulforaphane (SF), neoascorbigen (NeoASG), N-methoxyindole-3-carbinol (NI3C), and phenethyl isothiocyanate (PEITC)] were also quantified and found to be significantly correlated with QR. Sulforaphane, the isothiocyanate hydrolysis product of the GS glucoraphanin, was found to be the most potent QR induction agent. Increased sulforaphane formation from the hydrolysis of glucoraphanin was associated with up-regulated gene expression of myrosinase (BoMyo) and the myrosinase enzyme co-factor gene, epithiospecifier modifier1 (BoESM1). This study demonstrates the combined treatment of MeJA and 1-MCP increased QR activity without post-harvest quality loss. PMID:24146962

  8. Plant isoflavone and isoflavanone O-methyltransferase genes

    Science.gov (United States)

    Broeckling, Bettina E.; Liu, Chang-Jun; Dixon, Richard A.

    2014-08-19

    The invention provides enzymes that encode O-methyltransferases (OMTs) from Medicago truncatula that allow modification to plant (iso)flavonoid biosynthetic pathways. In certain aspects of the invention, the genes encoding these enzymes are provided. The invention therefore allows the modification of plants for isoflavonoid content. Transgenic plants comprising such enzymes are also provided, as well as methods for improving disease resistance in plants. Methods for producing food and nutraceuticals, and the resulting compositions, are also provided.

  9. An Arabidopsis thaliana methyltransferase Capable of Methylating Farnesoic Acid

    Energy Technology Data Exchange (ETDEWEB)

    Yang,Y.; Yuan, J.; Ross, J.; Noel, J.; Pichersky, E.

    2006-01-01

    We previously reported the identification of a new family of plant methyltransferases (MTs), named the SABATH family, that use S-adenosyl-l-methionine (SAM) to methylate a carboxyl moiety or a nitrogen-containing functional group on a diverse array of plant compounds. The Arabidopsis genome alone contains 24 distinct SABATH genes. To identify the catalytic specificities of members of this protein family in Arabidopsis, we screened recombinantly expressed and purified enzymes with a large number of potential substrates. Here, we report that the Arabidopsis thaliana gene At3g44860 encodes a protein with high catalytic specificity towards farnesoic acid (FA). Under steady-state conditions, this farnesoic acid carboxyl methyltransferase (FAMT) exhibits K{sub M} values of 41 and 71 {mu}M for FA and SAM, respectively. A three-dimensional model of FAMT constructed based upon similarity to the experimentally determined structure of Clarkia breweri salicylic acid methyltransferase (SAMT) suggests a reasonable model for FA recognition in the FAMT active site. In plants, the mRNA levels of At3g44860 increase in response to the exogenous addition of several compounds previously shown to induce plant defense responses at the transcriptional level. Although methyl farnesoate (MeFA) has not yet been detected in Arabidopsis, the presence of a FA-specific carboxyl methyltransferase in Arabidopsis capable of producing MeFA, an insect juvenile hormone made by some plants as a presumed defense against insect herbivory, suggests that MeFA or chemically similar compounds are likely to serve as new specialized metabolites in Arabidopsis.

  10. Histone H3 lysine 4 methyltransferase KMT2D.

    Science.gov (United States)

    Froimchuk, Eugene; Jang, Younghoon; Ge, Kai

    2017-09-05

    Histone-lysine N-methyltransferase 2D (KMT2D), also known as MLL4 and MLL2 in humans and Mll4 in mice, belongs to a family of mammalian histone H3 lysine 4 (H3K4) methyltransferases. It is a large protein over 5500 amino acids in size and is partially functionally redundant with KMT2C. KMT2D is widely expressed in adult tissues and is essential for early embryonic development. The C-terminal SET domain is responsible for its H3K4 methyltransferase activity and is necessary for maintaining KMT2D protein stability in cells. KMT2D associates with WRAD (WDR5, RbBP5, ASH2L, and DPY30), NCOA6, PTIP, PA1, and H3K27 demethylase UTX in one protein complex. It acts as a scaffold protein within the complex and is responsible for maintaining the stability of UTX. KMT2D is a major mammalian H3K4 mono-methyltransferase and co-localizes with lineage determining transcription factors on transcriptional enhancers. It is required for the binding of histone H3K27 acetyltransferases CBP and p300 on enhancers, enhancer activation and cell-type specific gene expression during differentiation. KMT2D plays critical roles in regulating development, differentiation, metabolism, and tumor suppression. It is frequently mutated in developmental diseases, such as Kabuki syndrome and congenital heart disease, and various forms of cancer. Further understanding of the mechanism through which KMT2D regulates gene expression will reveal why KMT2D mutations are so harmful and may help generate novel therapeutic approaches. Published by Elsevier B.V.

  11. Effects of broccoli extract on biodistribution and labeling blood components with {sup 99m}Tc-GH

    Energy Technology Data Exchange (ETDEWEB)

    Cekic, Betul; Muftuler, Fazilet Zumrut Biber; Kilcar, Ayfer Yurt; Ichedef, Cigdem; Unak, Perihan [Ege University, Izmir (Turkey). Inst. of Nuclear Sciences. Dept. of Nuclear Applications

    2011-09-15

    Purpose: people consume vegetables without the knowledge of the side effects of the biological and chemical contents and interactions between radiopharmaceuticals and herbal extract. To this end, current study is focused on the effects of broccoli extract on biodistribution of radiolabeled glucoheptonate ({sup 99m}Tc-GH) and radiolabeling of blood components. Methods: GH was labeled with {sup 99m}Tc. Quality control studies were done utilizing TLC method. Biodistribution studies were performed on male rats which were treated via gavage with either broccoli extract or SF as control group for 15 days. Blood samples were withdrawn from rats' heart. Radiolabeling of blood constituents performed incubating with GH, SnCl{sub 2} and {sup 99m} Tc. Results: radiochemical yield of {sup 99m}Tc-GH is 98.46{+-}1.48 % (n=8). Biodistribution studies have shown that according to the control, the treated group with broccoli has approximately 10 times less uptake in kidney. The percentage of the radioactivity ratios of the blood components is found to be same in both groups. Conclusions: although there is no considerable effect on the radiolabeling of blood components, there is an outstanding change on the biodistribution studies especially on kidneys. The knowledge of this change on kidney uptake may contribute to reduce the risk of misdiagnosis and/or repetition of the examinations in Nuclear Medicine. (author)

  12. Impact of pre-harvest light spectral properties on health- and sensory-related compounds in broccoli florets.

    Science.gov (United States)

    Steindal, Anne Linn Hykkerud; Johansen, Tor J; Bengtsson, Gunnar B; Hagen, Sidsel F; Mølmann, Jørgen A B

    2016-04-01

    Plants grown at different latitudes experience differences in light spectral composition. Broccoli (Brassica oleracea L. var italica) plants were grown in climate-controlled chambers under supplemental wavelengths (red, far-red, red + far-red or blue) from light-emitting diodes (LEDs). The light treatments were combined with two cold climate temperatures (12 and 15 °C) during broccoli head formation to investigate the effects on morphology and content of health- and sensory-related compounds: glucosinolates, flavonols, ascorbic acid and soluble sugars. Supplemental far-red and red + far-red light led to elongated plants and the lowest total glucosinolate content in broccoli florets. The content of quercetin was highest with supplemental red light. Vitamin C was not significantly affected by the light treatments, but 12 °C gave a higher content than 15 °C. The effects of supplemental red and far-red light suggest an involvement of phytochromes in the regulation of glucosinolates and flavonols. A shift in red:far-red ratio could cause changes in their content besides altering the morphology. The sugar and vitamin C content appears to be unaffected by these light conditions. Supplemental blue light had little effect on plant morphology and content of the health- and sensory related compounds. © 2015 Society of Chemical Industry.

  13. Optimization of pulsed electric field pre-treatments to enhance health-promoting glucosinolates in broccoli flowers and stalk.

    Science.gov (United States)

    Aguiló-Aguayo, Ingrid; Suarez, Manuel; Plaza, Lucia; Hossain, Mohammad B; Brunton, Nigel; Lyng, James G; Rai, Dilip K

    2015-07-01

    The effect of pulsed electric field (PEF) treatment variables (electric field strength and treatment time) on the glucosinolate content of broccoli flowers and stalks was evaluated. Samples were subjected to electric field strengths from 1 to 4 kV cm(-1) and treatment times from 50 to 1000 µs at 5 Hz. Data fitted significantly (P < 0.0014) the proposed second-order response functions. The results showed that PEF combined treatment conditions of 4 kV cm(-1) for 525 and 1000 µs were optimal to maximize glucosinolate levels in broccoli flowers (ranging from 187.1 to 212.5%) and stalks (ranging from 110.6 to 203.0%) respectively. The predicted values from the developed quadratic polynomial equation were in close agreement with the actual experimental values, with low average mean deviations (E%) ranging from 0.59 to 8.80%. The use of PEF processing at moderate conditions could be a suitable method to stimulate production of broccoli with high health-promoting glucosinolate content. © 2014 Society of Chemical Industry.

  14. Cytochrome P450 IA2 activity in man measured by caffeine metabolism: effect of smoking, broccoli and exercise.

    Science.gov (United States)

    Vistisen, K; Loft, S; Poulsen, H E

    1991-01-01

    The activity of 3 enzymes related to the bioactivation of toxic compounds and the development of cancer--cytochrome P450 IA2, N-acetyl transferase (NAT), and xanthine oxidase (XO)--can be measured from the ratios of formed metabolites excreted into urine. In the 3 experiments that comprised this study, subjects received at least 1 cup of coffee 2- 6 hours before spot urine samples were taken. The subjects included 335 healthy male and female volunteers who provided information on tobacco, caffeine, and broccoli intake in the preceding 2 weeks, 23 healthy men who exercised 8 hours/day for 30 days, and 9 subjects whose diet included green beans and broccoli. As expected, the ratio reflecting P450 IA2 activity was 66% and 70% higher, respectively, in men and women who smoked at least 10 cigarettes/day compared to male and female nonsmokers. The XO ratio also was significantly increased in smokers. 30 days of vigorous physical exercise increased the P450 IA2 ratio by 50% and the XO ratio by over 100%. Broccoli induced a 19% increase in P450 IA2 activity, while pregnancy and oral contraceptive use reduced this ratio by 29% and 20%, respectively. Since these ratios appear to yield reliable indicators of enzyme activity, prospective studies of their association with cancer development are recommended.

  15. Comparative study of the cell wall composition of broccoli, carrot, and tomato: structural characterization of the extractable pectins and hemicelluloses.

    Science.gov (United States)

    Houben, Ken; Jolie, Ruben P; Fraeye, Ilse; Van Loey, Ann M; Hendrickx, Marc E

    2011-07-01

    This study delivers a comparison of the pectic and hemicellulosic cell wall polysaccharides between the commonly used vegetables broccoli (stem and florets separately), carrot, and tomato. Alcohol-insoluble residues were prepared from the plant sources and sequentially extracted with water, cyclohexane-trans-1,2-diamine tetra-acetic acid, sodium carbonate, and potassium hydroxide solutions, to obtain individual fractions, each containing polysaccharides bound to the cell wall in a specific manner. Structural characterization of the polysaccharide fractions was conducted using colorimetric and chromatographic approaches. Sugar ratios were defined to ameliorate data interpretation. These ratios allowed gaining information concerning polysaccharide structure from sugar composition data. Structural analysis of broccoli revealed organ-specific characteristics: the pectin degree of methoxylation (DM) of stem and florets differed, the sugar composition data inferred differences in polymeric composition. On the other hand, the molar mass (MM) distribution profiles of the polysaccharide fractions were virtually identical for both organs. Carrot root displayed a different MM distribution for the polysaccharides solubilized by potassium hydroxide compared to broccoli and tomato, possibly due to the high contribution of branched pectins to this otherwise hemicellulose-enriched fraction. Tomato fruit showed the pectins with the broadest range in DM, the highest MM, the greatest overall linearity and the lowest extent of branching of rhamnogalacturonan I, pointing to particularly long, linear pectins in tomato compared with the other vegetable organs studied, suggesting possible implications toward functional behavior. Copyright © 2011 Elsevier Ltd. All rights reserved.

  16. Variation in bioactive content in broccoli (Brassica oleracea var. italica) grown under conventional and organic production systems.

    Science.gov (United States)

    Valverde, Juan; Reilly, Kim; Villacreces, Salvador; Gaffney, Michael; Grant, James; Brunton, Nigel

    2015-04-01

    Broccoli and other cruciferous vegetables contain a number of bioactive compounds, in particular glucosinolates and polyphenols, which are proposed to confer health benefits to the consumer. Demand for organic crops is at least partly based on a perception that organic crops may contain higher levels of bioactive compounds; however, insufficient research has been carried out to either support or refute such claims. In this study we examined the effect of conventional, organic, and mixed cultivation practices on the content of total phenolics, total flavonoids, and total and individual glucosinolates in two varieties of broccoli grown over 2 years in a split-plot factorial systems comparison trial. Levels of total phenolics and total flavonoids showed a significant year-on-year variation but were not significantly different between organic and conventional production systems. In contrast, levels of the indolyl glucosinolates glucobrassicin and neoglucobrassicin were significantly higher (P organic compared to fully conventional management. Organic cultivation practices resulted in significantly higher levels of glucobrassicin and neoglucobrassicin in broccoli florets; however, other investigated compounds were unaffected by production practices. © 2014 Society of Chemical Industry.

  17. Thermally induced degradation of sulfur-containing aliphatic glucosinolates in broccoli sprouts (Brassica oleracea var. italica) and model systems.

    Science.gov (United States)

    Hanschen, Franziska S; Platz, Stefanie; Mewis, Inga; Schreiner, Monika; Rohn, Sascha; Kroh, Lothar W

    2012-03-07

    Processing reduces the glucosinolate (GSL) content of plant food, among other aspects due to thermally induced degradation. Since there is little information about the thermal stability of GSL and formation of corresponding breakdown products, the thermally induced degradation of sulfur-containing aliphatic GSL was studied in broccoli sprouts and with isolated GSL in dry medium at different temperatures as well as in aqueous medium at different pH values. Desulfo-GSL have been analyzed with HPLC-DAD, while breakdown products were estimated using GC-FID. Whereas in the broccoli sprouts structural differences of the GSL with regard to thermal stability exist, the various isolated sulfur-containing aliphatic GSL degraded nearly equally and were in general more stable. In broccoli sprouts, methylsulfanylalkyl GSL were more susceptible to degradation at high temperatures, whereas methylsulfinylalkyl GSL were revealed to be more affected in aqueous medium under alkaline conditions. Besides small amounts of isothiocyanates, the main thermally induced breakdown products of sulfur-containing aliphatic GSL were nitriles. Although they were most rapidly formed at comparatively high temperatures under dry heat conditions, their highest concentrations were found after cooking in acidic medium, conditions being typical for domestic processing.

  18. Structural characterization of the mitomycin 7-O-methyltransferase.

    Science.gov (United States)

    Singh, Shanteri; Chang, Aram; Goff, Randal D; Bingman, Craig A; Grüschow, Sabine; Sherman, David H; Phillips, George N; Thorson, Jon S

    2011-07-01

    Mitomycins are quinone-containing antibiotics, widely used as antitumor drugs in chemotherapy. Mitomycin-7-O-methyltransferase (MmcR), a key tailoring enzyme involved in the biosynthesis of mitomycin in Streptomyces lavendulae, catalyzes the 7-O-methylation of both C9β- and C9α-configured 7-hydroxymitomycins. We have determined the crystal structures of the MmcR-S-adenosylhomocysteine (SAH) binary complex and MmcR-SAH-mitomycin A (MMA) ternary complex at resolutions of 1.9and 2.3 Å, respectively. The study revealed MmcR to adopt a common S-adenosyl-L-methionine-dependent O-methyltransferase fold and the presence of a structurally conserved active site general acid-base pair is consistent with a proton-assisted methyltransfer common to most methyltransferases. Given the importance of C7 alkylation to modulate mitomycin redox potential, this study may also present a template toward the future engineering of catalysts to generate uniquely bioactive mitomycins. Copyright © 2011 Wiley-Liss, Inc.

  19. Structural characterization of the mitomycin 7-O-methyltransferase

    Energy Technology Data Exchange (ETDEWEB)

    Singh, Shanteri; Chang, Aram; Goff, Randal D.; Bingman, Craig A.; Grüschow, Sabine; Sherman, David H.; Phillips, Jr., George N.; Thorson, Jon S. (Michigan); (UW)

    2014-10-02

    Mitomycins are quinone-containing antibiotics, widely used as antitumor drugs in chemotherapy. Mitomycin-7-O-methyltransferase (MmcR), a key tailoring enzyme involved in the biosynthesis of mitomycin in Streptomyces lavendulae, catalyzes the 7-O-methylation of both C9{beta}- and C9{alpha}-configured 7-hydroxymitomycins. We have determined the crystal structures of the MmcR-S-adenosylhomocysteine (SAH) binary complex and MmcR-SAH-mitomycin A (MMA) ternary complex at resolutions of 1.9 and 2.3 {angstrom}, respectively. The study revealed MmcR to adopt a common S-adenosyl-L-methionine-dependent O-methyltransferase fold and the presence of a structurally conserved active site general acid-base pair is consistent with a proton-assisted methyltransfer common to most methyltransferases. Given the importance of C7 alkylation to modulate mitomycin redox potential, this study may also present a template toward the future engineering of catalysts to generate uniquely bioactive mitomycins.

  20. Regulation of Catechol-O-Methyltransferase Expression in Human Myometrial Cells

    Science.gov (United States)

    Wentz, Melissa J.; Jamaluddin, Mohammad; Garfield, Robert E.; Al-Hendy, Ayman

    2014-01-01

    OBJECTIVE The catechol-O-methyltransferase enzyme catalyzes the methylation of the catechol estrogens, 2- or 4-hydroxyestrogen, to 2- or 4-methoxyestrogen. Both the hydroxy estrogens and methoxy estrogens were shown to modulate the effects of estrogen. Because catechol-O-methyltransferase activity controls levels of these metabolites, it may help regulate the cellular estrogenic milieu. In this study, we examined the regulation of catechol-O-methyltransferase expression in human myometrial cells. METHODS Catechol-O-methyltransferase expression was assessed by reverse transcription–polymerase chain reaction, Western blot, and luciferase assays in human myometrial cells after treatment with estrogen or progesterone. Catechol-O-methyltransferase expression was measured in cells after treatment with tumor necrosis factor alpha (TNFα) alone or with lactacystin, a protea-some inhibitor. Luciferase assays were also conducted using human myometrial cells containing an estrogen response element–luciferase reporter gene to measure levels of estrogen-mediated transactivation after treatment with estrogen and increasing concentrations of 2-hydroxestrogen. RESULTS Catechol-O-methyltransferase expression was down-regulated by progesterone or estrogen. Tumor necrosis factor alpha upregulated catechol-O-methyltransferase expression, whereas cotreatment with lactacystin attenuated this response, suggesting that TNFα activated nuclear factor kappa B to induce catechol-O-methyltransferase expression. Increased concentrations of 2-hydroxyestrogen attenuated estrogen-mediated transcription in the myometrial cells. CONCLUSION Catechol-O-methyltransferase expression may be regulated in the myometrium to control the local action of estrogen. Low levels of catechol-O-methyltransferase in the myometrium would result in an accumulation of 2-hydroxyestrogen and may antagonize the local effect of estrogen. High levels of catechol-O-methyltransferase in the myometrium would result in

  1. Small bowel obstruction from potato and broccoli phytobezoar mimicking mesenteric ischemia.

    Science.gov (United States)

    Serrano, Karen D; Tupesis, Janis P

    2013-01-01

    Bezoars are concretions of undigested foreign material that form in the gastrointestinal tract. Rare in humans, they are nonetheless a well-documented cause of intraluminal bowel obstruction. The objectives of this case report include describing an unusual presentation of small bowel obstruction due to phytobezoar, which mimicked mesenteric ischemia, and highlighting the risk factors, presentation, and management of bezoars, in addition to covering historical beliefs regarding bezoars. Here we report a 64-year-old man who presented to the Emergency Department with chest pain, vomiting, and hypotension. Initial work-up was directed at ruling out cardiac causes and aortic catastrophe such as aortic dissection or ruptured abdominal aortic aneurysm. Computed tomography angiography of the chest and abdomen showed findings suggestive of mesenteric ischemia and small bowel obstruction. However, exploratory laparotomy revealed intraluminal small bowel obstruction from a phytobezoar consisting of undigested chunks of potato, brussels sprouts, and broccoli. Although rare in humans, bezoars are a documented cause of small bowel obstruction, and should be considered when intraluminal bowel obstruction occurs. Bezoars causing small bowel obstruction require surgical treatment. Copyright © 2013 Elsevier Inc. All rights reserved.

  2. Slugs control methods (Deroceras sp. Müller in lettuce and broccoli crops

    Directory of Open Access Journals (Sweden)

    Salazar G. Claudia

    2011-08-01

    Full Text Available

    At the Botana experimental farm of Universidad de Narino, the effect of different management practices of slugs was evaluated, using a randomized block design with five treatments (beer traps, milk traps, sisal bags, metaldehyde, and a control and six repetitions. The evaluated variables were: captured individuals, affected plants and damage index that were estimated by analysis of variance and Tukey comparison tests. The results show highly significant differences between treatments in different phenological crops stages, the most effective were the metaldehyde and the beer trap with a caught individual number of 89 for lettuce and 126 for broccoli, being the treatments with lower percentages of affected plants. The damage index was evaluated based on percentages and proposed scales to observed crops damage, considering that the treatments described above stood out with mild damage index. This allows involved the beer treatment in an integrated management program for slugs as an alternative for reducing the pest population.

  3. Effects of Replacement of Concentrate Mixture by Broccoli Byproducts on Lactating Performance in Dairy Cows

    Directory of Open Access Journals (Sweden)

    X. W. Yi

    2015-10-01

    Full Text Available The objective of the present study was to determine the effects of feeding pelletized broccoli byproducts (PBB on milk yield and milk composition in dairy cows. In Trial 1, an in vitro gas test determined the optimal replacement level of PBB in a concentrate mixture in a mixed substrate with Chinese wild ryegrass hay (50:50, w/w at levels of 0, 10%, 20%, 30%, or 40% (dry matter basis. When the concentrate was replaced by PBB at a level of 20%, no adverse effects were found on the gas volume or its rate constant during ruminal fermentation. In trial 2, 24 lactating cows (days in milk = 170.4±35; milk yield = 30±3 kg/d; body weight = 580 ±13 kg were divided into 12 blocks based on day in milk and milk yield and randomly allocated to two dietary treatments: a basic diet with or without PBB replacing 20% of the concentrate mixture. The feeding trial lasted for 56 days; the first week allowed for adaptation to the diet. The milk composition was analyzed once a week. No significant difference in milk yield was observed between the two groups (23.5 vs 24.2 kg. A significant increase was found in milk fat content in the PBB group (p0.05. These results indicated that PBB could be included in dairy cattle diets at a suitable level to replace concentrate mixture without any adverse effects on dairy performance.

  4. Negative in vitro selection identifies the rRNA recognition motif for ErmE methyltransferase

    DEFF Research Database (Denmark)

    Nielsen, A K; Douthwaite, S; Vester, B

    1999-01-01

    Erm methyltransferases modify bacterial 23S ribosomal RNA at adenosine 2058 (A2058, Escherichia coli numbering) conferring resistance to macrolide, lincosamide, and streptogramin B (MLS) antibiotics. The motif that is recognized by Erm methyltransferases is contained within helix 73 of 23S r...

  5. Association of myasthenia gravis with polymorphisms in the gene of histamine N-methyltransferase

    DEFF Research Database (Denmark)

    Kellermayer, Blanka; Polgar, Noemi; Pal, Jozsef

    2013-01-01

    Histamine N-methyltransferase (HNMT) is the main metabolizing enzyme of histamine. Histamine modulates immune responses and plays a role in the pathogenesis of autoimmune disorders.......Histamine N-methyltransferase (HNMT) is the main metabolizing enzyme of histamine. Histamine modulates immune responses and plays a role in the pathogenesis of autoimmune disorders....

  6. Archease from Pyrococcus abyssi improves substrate specificity and solubility of a tRNA m5C methyltransferase

    DEFF Research Database (Denmark)

    Auxilien, Sylvie; El Khadali, Fatima; Rasmussen, Anette

    2007-01-01

    reading frame (PAB1947), which is shown here to encode a tRNA m(5)C methyltransferase. In vitro, the purified recombinant methyltransferase catalyzes m(5)C formation at several cytosines within tRNAs with preference for C49. The specificity of the methyltransferase is increased by the archease...

  7. Coordinate regulation of DNA methyltransferase expression during oogenesis

    Directory of Open Access Journals (Sweden)

    Bestor Timothy H

    2007-04-01

    Full Text Available Abstract Background Normal mammalian development requires the action of DNA methyltransferases (DNMTs for the establishment and maintenance of DNA methylation within repeat elements and imprinted genes. Here we report the expression dynamics of Dnmt3a and Dnmt3b, as well as a regulator of DNA methylation, Dnmt3L, in isolated female germ cells. Results Our results indicate that these enzymes are coordinately regulated and that their expression peaks during the stage of postnatal oocyte development when maternal methylation imprints are established. We find that Dnmt3a, Dnmt3b, Dnmt3L and Dnmt1o transcript accumulation is related to oocyte diameter. Furthermore, DNMT3L deficient 15 dpp oocytes have aberrantly methylated Snrpn, Peg3 and Igf2r DMRs, but normal IAP and LINE-1 methylation levels, thereby highlighting a male germ cell specific role for DNMT3L in the establishment of DNA methylation at repeat elements. Finally, real-time RT-PCR analysis indicates that the depletion of either DNMT3L or DNMT1o in growing oocytes results in the increased expression of the de novo methyltransferase Dnmt3b, suggesting a potential compensation mechanism by this enzyme for the loss of one of the other DNA methyltransferases. Conclusion Together these results provide a better understanding of the developmental regulation of Dnmt3a, Dnmt3b and Dnmt3L at the time of de novo methylation during oogenesis and demonstrate that the involvement of DNMT3L in retrotransposon silencing is restricted to the male germ line. This in turn suggests the existence of other factors in the oocyte that direct DNA methylation to transposons.

  8. A SABATH Methyltransferase from the moss Physcomitrella patens catalyzes

    Energy Technology Data Exchange (ETDEWEB)

    Zhao, Nan [ORNL; Ferrer, Jean-Luc [Universite Joseph Fourier, France; Moon, Hong S [Department of Plant Sciences, University of Tennessee; Kapteyn, Jeremy [Institute of Biological Chemistry, Washington State University; Zhuang, Xiaofeng [Department of Plant Sciences, University of Tennessee; Hasebe, Mitsuyasu [Laboratory of Evolutionary Biology, National Institute for Biology, 38 Nishigounaka; Stewart, Neal C. [Department of Plant Sciences, University of Tennessee; Gang, David R. [Institute of Biological Chemistry, Washington State University; Chen, Feng [University of Tennessee, Knoxville (UTK)

    2012-01-01

    Known SABATH methyltransferases, all of which were identified from seed plants, catalyze methylation of either the carboxyl group of a variety of low molecular weight metabolites or the nitrogen moiety of precursors of caffeine. In this study, the SABATH family from the bryophyte Physcomitrella patens was identified and characterized. Four SABATH-like sequences (PpSABATH1, PpSABATH2, PpSABATH3, and PpSABATH4) were identified from the P. patens genome. Only PpSABATH1 and PpSABATH2 showed expression in the leafy gametophyte of P. patens. Full-length cDNAs of PpSABATH1 and PpSABATH2 were cloned and expressed in soluble form in Escherichia coli. Recombinant PpSABATH1 and PpSABATH2 were tested for methyltransferase activity with a total of 75 compounds. While showing no activity with carboxylic acids or nitrogen-containing compounds, PpSABATH1 displayed methyltransferase activity with a number of thiols. PpSABATH2 did not show activity with any of the compounds tested. Among the thiols analyzed, PpSABATH1 showed the highest level of activity with thiobenzoic acid with an apparent Km value of 95.5 lM, which is comparable to those of known SABATHs. Using thiobenzoic acid as substrate, GC MS analysis indicated that the methylation catalyzed by PpSABATH1 is on the sulfur atom. The mechanism for S-methylation of thiols catalyzed by PpSABATH1 was partially revealed by homology-based structural modeling. The expression of PpSABATH1 was induced by the treatment of thiobenzoic acid. Further transgenic studies showed that tobacco plants overexpressing PpSABATH1 exhibited enhanced tolerance to thiobenzoic acid, suggesting that PpSABATH1 have a role in the detoxification of xenobiotic thiols.

  9. Chromosomal replication incompatibility in Dam methyltransferase deficient Escherichia coli cells

    DEFF Research Database (Denmark)

    Freiesleben, Ulrik Von

    1996-01-01

    Dam methyltransferase deficient Escherichia coli cells containing minichromosomes were constructed. Free plasmid DNA could not be detected in these cells and the minichromosomes were found to be integrated in multiple copies in the origin of replication (oriC) region of the host chromosome...... and functional oriC sequence. The seqA2 mutation was found to overcome the incompatibility phenotype by increasing the cellular oriC copy nnumber 3-fold thereby allowing minichromosomes to coexist with the chromosome. The replication pattern of a wild type strain with multiple integrated minichromosomes...

  10. Concentrating biomass of fermented broccoli (Brassica oleracea) and spinach (Amaranthus sp.) by ultrafiltration for source of organic acids and natural antioxidant

    Science.gov (United States)

    Aspiyanto, Susilowati, Agustine; Lotulung, Puspa D.; Maryati, Yati

    2017-11-01

    Organic acids and polyphenol from fermentation of green vegetables by Kombucha culture are novelty functional food to achieve prebiotic and natural antioxidant. Ultrafiltration (UF) mode was performed to concentrate biomass of fermented broccoli (Brassica oleracea L.) and spinach (Amaranthus spp.) at stirrer rotation speed of 200, 300 and 400 rpm, room temperature and trans membrane pressure 40 psia for 30 minutes. Based on total organic acids, experiment activity showed that the best treatment on biomass of fermented broccoli and spinach were reached at stirrer rotation speed of 400 rpm and 300 rpm, respectively. In this condition, fermented broccoli and spinach concentrates gave total acids 0.83 % and 0.81 %, total polyphenol 0.06 % and 0.11 %, reducing sugar 63.95 mg/mL and 20.54 mg/mL, total sugars 2.43 ug/mL and 2.28 ug/mL, total solids 6.42 % and 7.17 %, respectively. Compared with feed, the optimum condition on fermented spinach and broccoli concentrates increased total acids 13.33 % and 10 %, however decreased total polyphenol 34.1 % and 41 %. Identification on monomer from fermented spinach and broccoli at optimum condition on lactic acid were dominated by monomers with molecular weights (MWs) 252.19 and 252.36 Dalton (Da.), and monomer of polyphenol dominated by monomer with MWs 193.17 and 193.22 Da. and relative intensity 100 %. Fermented broccoli has potency as prebiotic, meanwhile fermented spinach has potency as anti oxidant.

  11. Yield, quality, and concentration of seven heavy metals in cabbage and broccoli grown in sewage sludge and chicken manure amended soil.

    Science.gov (United States)

    Antonious, George F; Kochhar, Tejinder S; Coolong, Timothy

    2012-01-01

    The mobility of heavy metals from soil into the food chain and their subsequent bioaccumulation has increased the attention they receive as major environmental pollutants. The objectives of this investigation were to: i) study the impact of mixing native agricultural soil with municipal sewage sludge (SS) or chicken manure (CM) on yield and quality of cabbage and broccoli, ii) quantify the concentration of seven heavy metals (Cd, Cr, Mo, Cu, Zn, Pb, and Ni) in soil amended with SS or CM, and iii) determine bioavailability of heavy metals to cabbage leaves and broccoli heads at harvest. Analysis of the two soil amendments used in this investigation indicated that Cr, Ni, Cu, Zn, Mo, Cd, Pb, and organic matter content were significantly greater (P grown in soil amended with CM was low compared to plants grown in no-mulch soil. No significant differences were found in Cd and Pb accumulation between cabbage and broccoli. Concentrations of Ni, Cu, Zn, and Mo were greater in broccoli than cabbage. Total metals and plant available metals were also determined in the native and amended soils. Results indicated that the concentration of heavy metals in soils did not necessary reflect metals available to plants. Regardless of soil amendments, the overall bioaccumulation factor (BAF) of seven heavy metals in cabbage leaves and broccoli heads revealed that cabbage and broccoli were poor accumulators of Cr, Ni, Cu, Cd, and Pb (BAF 1 for Zn and Mo. Elevated Ni and Mo bioaccumulation factor (BAF >1) of cabbage grown in chicken manure mixed soil is a characteristic that would be less favorable when cabbage is grown on sites having high concentrations of these two metals.

  12. Effect of cooking on the concentration of bioactive compounds in broccoli (Brassica oleracea var. Avenger) and cauliflower (Brassica oleracea var. Alphina F1) grown in an organic system.

    Science.gov (United States)

    Dos Reis, Luzia Caroline Ramos; de Oliveira, Viviani Ruffo; Hagen, Martine Elisabeth Kienzle; Jablonski, André; Flôres, Simone Hickmann; de Oliveira Rios, Alessandro

    2015-04-01

    Brassica vegetables have been shown to have antioxidant capacities due to the presence of carotenoids, flavonoids and vitamins. This study evaluates the influence of different processing conditions (boiling, steaming, microwaving and sous vide) on the stability of flavonoids, carotenoids and vitamin A in broccoli and cauliflower inflorescences grown in an organic system. Results indicated that sous vide processing resulted in greater antioxidant capacity and that all processes contributed in some way to an increased content of antioxidant compounds in both cauliflower and broccoli. Copyright © 2014 Elsevier Ltd. All rights reserved.

  13. Effects of combined treatments of irradiation and antimicrobial coatings on reduction of food pathogens in broccoli florets

    Energy Technology Data Exchange (ETDEWEB)

    Takala, P.N.; Salmieri, S.; Vu, K.D. [INRS-Institut Armand-Frappier, Research Laboratories in Sciences Applied to Food, Canadian Irradiation Centre, 531, Blvd des Prairies, Laval, QC, H7V 1B7 (Canada); Lacroix, M., E-mail: Monique.Lacroix@iaf.inrs.ca [INRS-Institut Armand-Frappier, Research Laboratories in Sciences Applied to Food, Canadian Irradiation Centre, 531, Blvd des Prairies, Laval, QC, H7V 1B7 (Canada)

    2011-12-15

    The effect of combined treatment of antimicrobial coatings and {gamma}-radiation on reduction of food pathogens such as Listeria monocytogenes, Escherichia coli, and Salmonella Typhimurium was evaluated in broccoli florets. Broccoli florets were inoculated with pathogenic bacteria at 10{sup 6} CFU/g. Inoculated florets were then coated with methylcellulose-based coating containing various mixtures of antimicrobial agents: organic acids (OAs) plus lactic acid bacteria metabolites (LABs), OA plus citrus extract (CE), OA plus CE plus spice mixture (SM), and OA plus rosemary extract (RE). Coated florets were irradiated with various doses (0-3.3 kGy), and microbial analyses were used to calculate the D{sub 10} value and radiosensitive relative. The coating containing OA plus CE was the most effective formulation for increasing the sensitization of Escherichia coli by 2.4 times as compared to the control without the antimicrobial coating. For Salmonella Typhimurium, coating containing OA plus LAB was the most effective formulation, increasing radiosensitivity by 2.4 times as well. All antimicrobial coatings had almost the same effect of increasing the sensitivity of Listeria monocytogenes (from 1.31 to 1.45 times) to {gamma}-irradiation. - Highlights: > Demonstrate scientifically the synergistic effect of the combined treatment of gamma-irradiation and natural antimicrobial coating in reduction of food pathogens in broccoli. > The coating containing organic acids plus citrus extract was the most efficient formulation for increasing the sensitization of E. coli by 2.40 times as compared to the control. > The coating containing organic acids plus lactic acid bacteria metabolites was the most effective formulation causing the sensitization of S. Typhimurium to {gamma}-irradiation by 2.4 times. > Potential in application of developed formulations to protect food products against food pathogens.

  14. Organelle Simple Sequence Repeat Markers Help to Distinguish Carpelloid Stamen and Normal Cytoplasmic Male Sterile Sources in Broccoli

    Science.gov (United States)

    Shu, Jinshuai; Liu, Yumei; Li, Zhansheng; Zhang, Lili; Fang, Zhiyuan; Yang, Limei; Zhuang, Mu; Zhang, Yangyong; Lv, Honghao

    2015-01-01

    We previously discovered carpelloid stamens when breeding cytoplasmic male sterile lines in broccoli (Brassica oleracea var. italica). In this study, hybrids and multiple backcrosses were produced from different cytoplasmic male sterile carpelloid stamen sources and maintainer lines. Carpelloid stamens caused dysplasia of the flower structure and led to hooked or coiled siliques with poor seed setting, which were inherited in a maternal fashion. Using four distinct carpelloid stamens and twelve distinct normal stamens from cytoplasmic male sterile sources and one maintainer, we used 21 mitochondrial simple sequence repeat (mtSSR) primers and 32 chloroplast SSR primers to identify a mitochondrial marker, mtSSR2, that can differentiate between the cytoplasm of carpelloid and normal stamens. Thereafter, mtSSR2 was used to identify another 34 broccoli accessions, with an accuracy rate of 100%. Analysis of the polymorphic sequences revealed that the mtSSR2 open reading frame of carpelloid stamen sterile sources had a deletion of 51 bases (encoding 18 amino acids) compared with normal stamen materials. The open reading frame is located in the coding region of orf125 and orf108 of the mitochondrial genomes in Brassica crops and had the highest similarity with Raphanus sativus and Brassica carinata. The current study has not only identified a useful molecular marker to detect the cytoplasm of carpelloid stamens during broccoli breeding, but it also provides evidence that the mitochondrial genome is maternally inherited and provides a basis for studying the effect of the cytoplasm on flower organ development in plants. PMID:26407159

  15. Potency of microfiltration membrane process in purifying broccoli (Brassica oleracea L.) fermented by lactic acid bacteria (LAB) as functional food

    Science.gov (United States)

    Susilowati, Agustine; Aspiyanto, Maryati, Yati; Melanie, Hakiki; Lotulung, Puspa D.

    2017-01-01

    Purifying broccoli (Brassica oleracea L.) fermented by Lactic Acid Bacteria (LAB) using mixture of L. bulgaricus, S. thermopillus, L. acidophillusand Bifidobacteriumbifidum and fructooligosaccharides (FOS) as carbon source have been performed to recover biomass concentrate for probiotic and antioxidant. Purification of fermented broccoli was conducted through microfiltration (MF) membrane of 0.15 µm at stirrer rotation speed 400 rpm, room temperature and pressure 40 psia for 30 minutes. Fermented broccoli produced via fermentation process with fermentation time 0 (initial) and 48 hours, and LAB concentration 10% and 20% (v/v) represented as biomass of A, B, C and D. The experimental result showed that based on selectivity of total organic acids, separating optimization was achieved at biomass D (fermentation time 48 hours and mixed LAB culture concentration 20%). Concentrate composition produced in this condition were total acids 6.04%, total solids 24.31%, total polyphenol 0.0252%, reducing sugar 68.25 mg/mL, total sugars 30.89 mg/mL, and dissolved protein 28.54 mg/mL with pH 3.94. In this condition, recovery of biomass concentrate of D for total acids 5.64 folds, total solids 1.82 folds, total polyphenol 3.03 folds, reducing sugar 1.16 folds, total sugars 1.19 folds, and dissolved protein 0.67 folds compared with feed (initial process). Identification of monomer of biomass concentrate D as polyphenol derivatives at T2,01 and T3.01 gave monomer with molecular weight (MW) 192.78 Dalton (Da.), and monomer with MW 191.08, 191.49 and 192.07 Da., while lactic acid derivatives showed MW 251.13, 251.6 and 252.14, and monomer with MW 250.63, 252.14 and 254.22 Da.

  16. Dynamics and reactivity in Thermus aquaticus N6-adenine methyltransferase.

    Science.gov (United States)

    Aranda, Juan; Zinovjev, Kirill; Roca, Maite; Tuñón, Iñaki

    2014-11-19

    M.TaqI is a DNA methyltransferase from Thermus aquaticus that catalyzes the transfer of a methyl group from S-adenosyl-L-methionine to the N6 position of an adenine, a process described only in prokaryotes. We have used full atomistic classical molecular dynamics simulations to explore the protein-SAM-DNA ternary complex where the target adenine is flipped out into the active site. Key protein-DNA interactions established by the target adenine in the active site are described in detail. The relaxed structure was used for a combined quantum mechanics/molecular mechanics exploration of the reaction mechanism using the string method. According to our free energy calculations the reaction takes place through a stepwise mechanism where the methyl transfer precedes the abstraction of the proton from the exocyclic amino group. The methyl transfer is the rate-determining step, and the obtained free energy barrier is in good agreement with the value derived from the experimental rate constant. Two possible candidates to extract the leftover proton have been explored: a water molecule found in the active site and Asn105, a residue activated by the hydrogen bonds formed through the amide hydrogens. The barrier for the proton abstraction is smaller when Asn105 acts as a base. The reaction mechanisms can be different in other N6-DNA-methyltransferases, as determined from the exploration of the reaction mechanism in the Asn105Asp M.TaqI mutant.

  17. Structural biology of human H3K9 methyltransferases.

    Directory of Open Access Journals (Sweden)

    Hong Wu

    2010-01-01

    Full Text Available SET domain methyltransferases deposit methyl marks on specific histone tail lysine residues and play a major role in epigenetic regulation of gene transcription. We solved the structures of the catalytic domains of GLP, G9a, Suv39H2 and PRDM2, four of the eight known human H3K9 methyltransferases in their apo conformation or in complex with the methyl donating cofactor, and peptide substrates. We analyzed the structural determinants for methylation state specificity, and designed a G9a mutant able to tri-methylate H3K9. We show that the I-SET domain acts as a rigid docking platform, while induced-fit of the Post-SET domain is necessary to achieve a catalytically competent conformation. We also propose a model where long-range electrostatics bring enzyme and histone substrate together, while the presence of an arginine upstream of the target lysine is critical for binding and specificity.This article can also be viewed as an enhanced version in which the text of the article is integrated with interactive 3D representations and animated transitions. Please note that a web plugin is required to access this enhanced functionality. Instructions for the installation and use of the web plugin are available in Text S1.

  18. Crystal structure of arginine methyltransferase 6 from Trypanosoma brucei.

    Directory of Open Access Journals (Sweden)

    Chongyuan Wang

    Full Text Available Arginine methylation plays vital roles in the cellular functions of the protozoan Trypanosoma brucei. The T. brucei arginine methyltransferase 6 (TbPRMT6 is a type I arginine methyltransferase homologous to human PRMT6. In this study, we report the crystal structures of apo-TbPRMT6 and its complex with the reaction product S-adenosyl-homocysteine (SAH. The structure of apo-TbPRMT6 displays several features that are different from those of type I PRMTs that were structurally characterized previously, including four stretches of insertion, the absence of strand β15, and a distinct dimerization arm. The comparison of the apo-TbPRMT6 and SAH-TbPRMT6 structures revealed the fine rearrangements in the active site upon SAH binding. The isothermal titration calorimetry results demonstrated that SAH binding greatly increases the affinity of TbPRMT6 to a substrate peptide derived from bovine histone H4. The western blotting and mass spectrometry results revealed that TbPRMT6 methylates bovine histone H4 tail at arginine 3 but cannot methylate several T. brucei histone tails. In summary, our results highlight the structural differences between TbPRMT6 and other type I PRMTs and reveal that the active site rearrangement upon SAH binding is important for the substrate binding of TbPRMT6.

  19. Hypnotizability and Catechol-O-Methyltransferase (COMT polymorphysms in Italians

    Directory of Open Access Journals (Sweden)

    Silvano ePresciuttini

    2014-01-01

    Full Text Available Higher brain dopamine content depending on lower activity of Catechol-O-Methyltransferase (COMT in subjects with high hypnotisability scores (highs has been considered responsible for their attentional characteristics. However, the results of the previous genetic studies on association between hypnotisability and the Catechol-O-Methyltransferase (COMT single nucleotide polymorphism (SNP rs4680 (Val158Met were inconsistent. Here, we used a selective genotyping approach to re-evaluate the association between hypnotisability and COMT in the context of a two-SNP haplotype analysis, considering not only the Val158Met polymorphism, but also the closely located rs4818 SNP. An Italian sample of 53 highs, 49 low hypnotizable subjects (lows and 57 controls, were genotyped for a segment of 805 bp of the COMT gene, including Val158Met and the closely located rs4818 SNP. Our selective genotyping approach had 97.1% power to detect the previously reported strongest association at the significance level of 5%. We found no evidence of association at the SNP, haplotype and diplotype levels. Thus, our results challenge the dopamine-based theory of hypnosis and indirectly support recent neuropsychological and neurophysiological findings reporting the lack of any association between hypnotisability and focused attention abilities.

  20. Roles of the EZH2 histone methyltransferase in cancer epigenetics.

    Science.gov (United States)

    Simon, Jeffrey A; Lange, Carol A

    2008-12-01

    EZH2 is the catalytic subunit of Polycomb repressive complex 2 (PRC2), which is a highly conserved histone methyltransferase that targets lysine-27 of histone H3. This methylated H3-K27 chromatin mark is commonly associated with silencing of differentiation genes in organisms ranging from plants to flies to humans. Studies on human tumors show that EZH2 is frequently over-expressed in a wide variety of cancerous tissue types, including prostate and breast. Although the mechanistic contributions of EZH2 to cancer progression are not yet determined, functional links between EZH2-mediated histone methylation and DNA methylation suggest partnership with the gene silencing machinery implicated in tumor suppressor loss. Here we review the basic molecular biology of EZH2 and the findings that implicate EZH2 in different cancers. We also discuss EZH2 connections to other silencing enzymes, such as DNA methyltransferases and histone deacetylases, and we consider progress on deciphering mechanistic consequences of EZH2 overabundance and its potential roles in tumorigenesis. Finally, we review recent findings that link EZH2 roles in stem cells and cancer, and we consider prospects for integrating EZH2 blockade into strategies for developing epigenetic therapies.

  1. Dietary Chemoprevention of PhIP Induced Carcinogenesis in Male Fischer 344 Rats with Tomato and Broccoli

    OpenAIRE

    Canene-Adams, Kirstie; Sfanos, Karen S.; Liang, Chung-Tiang; Yegnasubramanian, Srinivasan; Nelson, William G.; Brayton, Cory; De Marzo, Angelo M.

    2013-01-01

    The heterocyclic amine, 2-amino-1-methyl-6-phenylimidazo[4,5-B]pyridine (PhIP), found in meats cooked at high temperatures, has been implicated in epidemiological and rodent studies for causing breast, prostate, and colorectal cancers. A previous animal study using a xenograft model has shown that whole tomato and broccoli, when eaten in combination, exhibit a marked effect on tumor reduction compared to when eaten alone. Our aim was to determine if PhIP-induced carcinogenesis can be prevente...

  2. Phase 1 Study of a Sulforaphane-Containing Broccoli Sprout Homogenate for Sickle Cell Disease.

    Directory of Open Access Journals (Sweden)

    Jennifer F Doss

    Full Text Available Sickle cell disease (SCD is the most common inherited hemoglobinopathy worldwide. Our previous results indicate that the reduced oxidative stress capacity of sickle erythrocytes may be caused by decreased expression of NRF2 (Nuclear factor (erythroid-derived 2-like 2, an oxidative stress regulator. We found that activation of NRF2 with sulforaphane (SFN in erythroid progenitors significantly increased the expression of NRF2 targets HMOX1, NQO1, and HBG1 (subunit of fetal hemoglobin in a dose-dependent manner. Therefore, we hypothesized that NRF2 activation with SFN may offer therapeutic benefits for SCD patients by restoring oxidative capacity and increasing fetal hemoglobin concentration. To test this hypothesis, we performed a Phase 1, open-label, dose-escalation study of SFN, contained in a broccoli sprout homogenate (BSH that naturally contains SFN, in adults with SCD. The primary and secondary study endpoints were safety and physiological response to NRF2 activation, respectively. We found that BSH was well tolerated, and the few adverse events that occurred during the trial were not likely related to BSH consumption. We observed an increase in the mean relative whole blood mRNA levels for the NRF2 target HMOX1 (p = 0.02 on the last day of BSH treatment, compared to pre-treatment. We also observed a trend toward increased mean relative mRNA levels of the NRF2 target HBG1 (p = 0.10 from baseline to end of treatment, but without significant changes in HbF protein. We conclude that BSH, in the provided doses, is safe in stable SCD patients and may induce changes in gene expression levels. We therefore propose investigation of more potent NRF2 inducers, which may elicit more robust physiological changes and offer clinical benefits to SCD patients. Trial registration: ClinicalTrials.gov NCT01715480.

  3. Belowground induction by Delia radicum or phytohormones affect aboveground herbivore communities on field-grown broccoli

    Directory of Open Access Journals (Sweden)

    Sandra Prisca Pierre

    2013-08-01

    Full Text Available Induced plant defence in response to phytophagous insects is a well described phenomenon. However, so far little is known about the effect of induced plant responses on subsequently colonizing herbivores in the field. Broccoli plants were induced in the belowground compartment using (i infestation by the root-herbivore Delia radicum, (ii root application of jasmonic acid (JA or (iii root application of salicylic acid (SA. The abundance of D. radicum and six aboveground herbivores displaying contrasting levels of host specialisation were surveyed for five weeks. Our study showed that the response of herbivores was found to differ from one another, depending on the herbivore species, its degree of specialisation and the root treatment. The abundance of the root herbivore D. radicum and particularly the number of emerging adults was decreased by both phytohormone treatments, while the number of D. radicum eggs was increased on conspecific infested plants. The root infestation exhibited moderate effects on the aboveground community. The abundance of the aphid Brevicoryne brassicae was strongly increased on D. radicum infested plants, but the other species were not impacted. Root hormone applications exhibited a strong effect on the abundance of specialist foliar herbivores. A higher number of B. brassicae and Pieris brassicae and a lower number of Plutella xylostella were found on JA treated plants. On SA treated plants we observed a decrease of the abundance of B. brassicae, Pi. rapae and P. xylostella. Surprisingly, generalist species, Mamestra brassicae and Myzus persicae were not affected by root induction treatments. Finally, root treatments had no significant effect on either glucosinolate profiles of the heads or on plant quality parameters. These results are discussed from the perspective of below- aboveground interactions and adaptations of phytophagous insects to induced plant responses according to their trophic specialisation level.

  4. Two novel bioactive glucosinolates from Broccoli (Brassica oleracea L. var. italica) florets.

    Science.gov (United States)

    Survay, Nazneen Shaik; Kumar, Brajesh; Jang, Mi; Yoon, Do-Young; Jung, Yi-Sook; Yang, Deok-Chun; Park, Se Won

    2012-09-01

    Two novel glucosinolates along with one known glucosinolate were isolated from Broccoli (Brassica oleracea L. var. italica) florets. Their structures were established mainly by 1D ((1)H and (13)C NMR), 2D NMR ((1)H-(1)H COSY, DEPT 135°, HSQC and HMBC), and Tandem MS-MS spectrometric data as 2-mercaptomethyl sulfinyl glucosinolate [(Z)-4-(methylsulfinyl)-N-(sulfooxy)-2-((2'S,3'R,4'S,5'S,6'R)-3',4',5'-trihydroxy-6'(hydroxylmethyl)-2'-mercapto tetrahydro-2H-pyran-2-yl) butane amide] 1, (Z)-1-((2S,5S)-5-hydroxytetra-hydro-2H-pyran-2-ylthio)-2-(1H-indol-3-yl) ethylidene amino sulfate 2 and a known cinnamoyl [6'-O-trans-(4″-hydroxy cinnamoyl)4-(methylsulphinyl)butyl glucosinolate] 3. Compound 1 exhibited scavenging activity against DPPH with an inhibitory concentration IC(50) of 20 mM, whereas compound 3 was a weak antioxidant when compared to the standard quercetin (5 mM) as a positive control. Both the compounds showed a significant and similar antimicrobial activity against Staphylococcus aureus with an IC(50) of <625 μg/mL when compared to antibiotic duricef. Against Salmonella typhimurium the IC(50) of 1 and 3 was determined as <625 μg/mL and <1250 μg/mL, respectively, when compared to ampicillin (IC(50) ≤ 39 μg/mL) as a positive control. Copyright © 2012 Elsevier Ltd. All rights reserved.

  5. Apoptotic role of natural isothiocyanate from broccoli (Brassica oleracea italica) in experimental chemical lung carcinogenesis.

    Science.gov (United States)

    Kalpana Deepa Priya, D; Gayathri, R; Gunassekaran, G R; Murugan, S; Sakthisekaran, D

    2013-05-01

    Sulforaphane (SFN) [1-isothiocyanato-4-(methylsulfinyl)butane] is a naturally occurring isothiocyanate found in cruciferous vegetables such as broccoli [Brassica oleracea L. var. italica Plenck. (Brassicaceae)]. Since it is among the most potent bioactive components with antioxidant and antitumor properties, it has received intense attention in the recent years for its chemopreventive properties. The present work determined the rehabilitating role in alleviating the oxidative damage caused by benzo(a)pyrene [B(a)P] to biomolecules and the apoptotic cascade mediated by orally administered isothiocyanate-SFN (9 µmol/mouse/day) against B(a)P (100 mg/kg body weight, i.p.) induced pulmonary carcinogenesis in Swiss albino mice. Oxidative damage was assessed by measuring lipid peroxidation, 8-hydroxydeoxyguanosine, hydrogen peroxide (H2O2) production, glycoprotein components, protein carbonyl levels and DNA-protein crosslinks. DNA fragmentation by agarose gel electrophoresis and caspase-3 activity by ELISA proved apoptotic induction by SFN along with the protein expression of Bcl-2, Bax and Cyt c. SFN treatment was found to decrease the H2O2 production (p < 0.001) in cancer induced animals, proving its antioxidant potential. Apoptosis was induced by increasing the release of Cyt c (p < 0.001) from mitochondria, decreasing and increasing the expression of Bcl-2 (p < 0.01) and Bax (p < 0.001), respectively. Caspase-3 activity was also enhanced (p < 0.001) which leads to DNA fragmentation in SFN treated groups. Our results reflect the rehabilitating role of SFN in B(a)P induced lung carcinogenesis.

  6. Structural basis for substrate recognition in the salicylic acid carboxyl methyltransferase family.

    Science.gov (United States)

    Zubieta, Chloe; Ross, Jeannine R; Koscheski, Paul; Yang, Yue; Pichersky, Eran; Noel, Joseph P

    2003-08-01

    Recently, a novel family of methyltransferases was identified in plants. Some members of this newly discovered and recently characterized methyltransferase family catalyze the formation of small-molecule methyl esters using S-adenosyl-L-Met (SAM) as a methyl donor and carboxylic acid-bearing substrates as methyl acceptors. These enzymes include SAMT (SAM:salicylic acid carboxyl methyltransferase), BAMT (SAM:benzoic acid carboxyl methyltransferase), and JMT (SAM:jasmonic acid carboxyl methyltransferase). Moreover, other members of this family of plant methyltransferases have been found to catalyze the N-methylation of caffeine precursors. The 3.0-A crystal structure of Clarkia breweri SAMT in complex with the substrate salicylic acid and the demethylated product S-adenosyl-L-homocysteine reveals a protein structure that possesses a helical active site capping domain and a unique dimerization interface. In addition, the chemical determinants responsible for the selection of salicylic acid demonstrate the structural basis for facile variations of substrate selectivity among functionally characterized plant carboxyl-directed and nitrogen-directed methyltransferases and a growing set of related proteins that have yet to be examined biochemically. Using the three-dimensional structure of SAMT as a guide, we examined the substrate specificity of SAMT by site-directed mutagenesis and activity assays against 12 carboxyl-containing small molecules. Moreover, the utility of structural information for the functional characterization of this large family of plant methyltransferases was demonstrated by the discovery of an Arabidopsis methyltransferase that is specific for the carboxyl-bearing phytohormone indole-3-acetic acid.

  7. Isothiocyanate metabolism, distribution, and interconversion in mice following consumption of thermally processed broccoli sprouts or purified sulforaphane.

    Science.gov (United States)

    Bricker, Gregory V; Riedl, Kenneth M; Ralston, Robin A; Tober, Kathleen L; Oberyszyn, Tatiana M; Schwartz, Steven J

    2014-10-01

    Broccoli sprouts are a rich source of glucosinolates, a group of phytochemicals that when hydrolyzed, are associated with cancer prevention. Our objectives were to investigate the metabolism, distribution, and interconversion of isothiocyanates (ITCs) in mice fed thermally processed broccoli sprout powders (BSPs) or the purified ITC sulforaphane. For 1 wk, mice were fed a control diet (n = 20) or one of four treatment diets (n = 10 each) containing nonheated BSP, 60°C mildly heated BSP, 5-min steamed BSP, or 3 mmol purified sulforaphane. Sulforaphane and erucin metabolite concentrations in skin, liver, kidney, bladder, lung, and plasma were quantified using HPLC-MS/MS. Thermal intensity of BSP processing had disparate effects on ITC metabolite concentrations upon consumption. Mild heating generally resulted in the greatest ITC metabolite concentrations in vivo, followed by the nonheated and steamed BSP diets. We observed interconversion between sulforaphane and erucin species or metabolites, and report that erucin is the favored form in liver, kidney, and bladder, even when only sulforaphane is consumed. ITC metabolites were distributed to all tissues analyzed, suggesting the potential for systemic benefits. We report for the first time tissue-dependent ratio of sulforaphane and erucin, though further investigation is warranted to assess biological activity of individual forms. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  8. Effect of time of day for harvest and postharvest treatments on the sugar metabolism of broccoli (Brassica oleracea var. italica

    Directory of Open Access Journals (Sweden)

    Joaquin Hasperue

    2014-02-01

    Full Text Available     Loss of sugars contributes to accelerate postharvest senescence of broccoli. Several treatments have been developed to delay senescence, but in many cases their effects on sugar metabolism were not analyzed. We studied the effect of harvest at different times of day (08:00, 13:00 and 18:00 h and of several postharvest treatments as heat treatment (HT, modified atmosphere (MA and 1-methylcylcopropene (1-MCP on sugar levels and activities of enzymes related to sucrose and starch degradation. Harvesting at the end of day delayed the loss of chlorophylls and caused the lowest decrement in sugars, although no differences in invertase, sucrose synthase and β-amylase activities were detected among samples. Treatments of MA and 1-MCP caused a lower loss of glucose and fructose, while HT caused a lower decrement of sucrose. Treated samples maintained higher levels of chlorophylls. The treatments reduced the activity of invertase and sucrose synthase and induced higher levels of β-amylase activity. Harvesting at the end of day and performing simultaneously a MA treatment could be a good combination to maintain the green color of the inflorescence and sugar levels during postharvest of broccoli.

  9. Effects of combined treatments of irradiation and antimicrobial coatings on reduction of food pathogens in broccoli florets

    Science.gov (United States)

    Takala, P. N.; Salmieri, S.; Vu, K. D.; Lacroix, M.

    2011-12-01

    The effect of combined treatment of antimicrobial coatings and γ-radiation on reduction of food pathogens such as Listeria monocytogenes, Escherichia coli, and Salmonella Typhimurium was evaluated in broccoli florets. Broccoli florets were inoculated with pathogenic bacteria at 10 6 CFU/g. Inoculated florets were then coated with methylcellulose-based coating containing various mixtures of antimicrobial agents: organic acids (OAs) plus lactic acid bacteria metabolites (LABs), OA plus citrus extract (CE), OA plus CE plus spice mixture (SM), and OA plus rosemary extract (RE). Coated florets were irradiated with various doses (0-3.3 kGy), and microbial analyses were used to calculate the D10 value and radiosensitive relative. The coating containing OA plus CE was the most effective formulation for increasing the sensitization of Escherichia coli by 2.4 times as compared to the control without the antimicrobial coating. For Salmonella Typhimurium, coating containing OA plus LAB was the most effective formulation, increasing radiosensitivity by 2.4 times as well. All antimicrobial coatings had almost the same effect of increasing the sensitivity of Listeria monocytogenes (from 1.31 to 1.45 times) to γ-irradiation.

  10. Effects of temperature and photoperiod on sensory quality and contents of glucosinolates, flavonols and vitamin C in broccoli florets.

    Science.gov (United States)

    Mølmann, Jørgen A B; Steindal, Anne L H; Bengtsson, Gunnar B; Seljåsen, Randi; Lea, Per; Skaret, Josefine; Johansen, Tor J

    2015-04-01

    Broccoli is grown around the world at a wide range of photoperiods and temperatures, which may influence both sensory quality and phytochemical contents. Florets produced in phytotron and at two semi-field sites (70 °N and 58 °N) were examined for effects of contrasting temperatures and photoperiods on sensory quality and contents of glucosinolates, flavonols and vitamin C. Growth conditions associated with high northern latitudes of low temperature and long photoperiods, produced bigger floral buds, and florets with sweeter taste and less colour hue than more southern conditions. The contents of vitamin C did not vary, while the response of individual glucosinolates varied with temperature and day length, and contents of quercetin and kaempferol were lower in phytotron than under semi-field conditions. Thus, our results show that contrasting temperatures and photoperiods influence the sensory quality of broccoli florets, while contents of different bioactive phytochemicals are not influenced in a unidirectional pattern. Copyright © 2014 Elsevier Ltd. All rights reserved.

  11. Gold nanoparticles synthesized by Brassica oleracea (Broccoli) acting as antimicrobial agents against human pathogenic bacteria and fungi

    Science.gov (United States)

    Piruthiviraj, Prakash; Margret, Anita; Krishnamurthy, Poornima Priyadharsani

    2016-04-01

    Production of antimicrobial agents through the synthesis of gold nanoparticles using green technology has been extensively made consistent by various researchers; yet, this study uses the flower bud's aqueous extracts of Brassica oleracea (Broccoli) as a reducing agent for chloroauric acid (1 mM). After 30 min of incubation, synthesis of gold nanoparticles (AuNps) was observed by a change in extract color from pale yellow to purple color. Synthesis of AuNps was confirmed in UV-visible spectroscopy at the range of approximately 560 nm. The SEM analysis showed the average nanoparticles size of 12-22 nm. The antimicrobial activity of AuNps was analyzed by subjecting it to human pathogenic bacteria (Gram-positive Staphylococcus aureus and Gram-negative Klebsiella pneumonia) and fungi (Aspergillus flavus, Aspergillus niger and Candida albicans) using disc diffusion method. The broccoli-synthesized AuNps showed the efficient antibacterial and antifungal activity of above-mentioned microbes. It was confirmed that AuNps have the best antimicrobial agent compared to the standard antibiotics (Gentamicin and Fluconazole). When the concentrations of AuNps were increased (10, 25, and 50 µg/ml), the sensitivity zone also increased for all the tested microbes. The synthesized AuNps are capable of rendering high antimicrobial efficacy and, hence, have a great potential in the preparation of drugs used against major bacterial and fungal diseases in humans.

  12. Differential Responses of Two Broccoli (Brassica oleracea L. var Italica Cultivars to Salinity and Nutritional Quality Improvement

    Directory of Open Access Journals (Sweden)

    Chokri Zaghdoud

    2012-01-01

    Full Text Available The comparative responses of two broccoli cultivars (Brassica oleracea var. Italica, cv. Parthenon and cv. Naxos to a 15 d exposure to different NaCl levels were investigated. Salinity led to increased concentrations of Na+ and Cl− ions in both cultivars, a disruption of the endogenous minerals levels in the shoots and roots—that varied with the cultivar and salt concentration—and decreases in the osmotic potential (Ψπ, root hydraulic conductance (L0, and stomatal conductance (Gs. The reduced biomass of Naxos at moderate NaCl indicates greater sensitivity to salinity, compared with Parthenon. Parthenon accumulated more soluble sugars, for osmotic adjustment, whereas Naxos accumulated proline, which gave the two cultivars differing nutritional characteristics. The total glucosinolates (GSLs content was not affected by salinity in Parthenon while it decreased significantly in Naxos as a consequence of the decrease in the indole GSL. However, Naxos accumulated more aliphatic GSLs under salt stress than Parthenon, which confers on this cultivar a greater nutritional value when cultivated under salinity.These results suggest that, at distinct salinity levels, each broccoli cultivar adopts a specific strategy, indicating the crucial role of the genetic background on the organoleptic and nutritional properties that each cultivar acquires.

  13. Atmospheric carbon dioxide changes photochemical activity, soluble sugars and volatile levels in broccoli (Brassica oleracea var. italica).

    Science.gov (United States)

    Krumbein, Angelika; Kläring, Hans-Peter; Schonhof, Ilona; Schreiner, Monika

    2010-03-24

    Atmospheric carbon dioxide (CO(2)) concentration is an environmental factor currently undergoing dramatic changes. The objective of the present study was to determine the effect of doubling the ambient CO(2) concentration on plant photochemistry as measured by photochemical quenching coefficient (qP), soluble sugars and volatiles in broccoli. Elevated CO(2) concentration increased qP values in leaves by up to 100% and 89% in heads, while glucose and sucrose in leaves increased by about 60%. Furthermore, in broccoli heads elevated CO(2) concentration induced approximately a 2-fold increase in concentrations of three fatty acid-derived C(7) aldehydes ((E)-2-heptenal, (E,Z)-2,4-heptadienal, (E,E)-2,4-heptadienal), two fatty acid-derived C(5) alcohols (1-penten-3-ol, (Z)-2-pentenol), and two amino acid-derived nitriles (phenyl propanenitrile, 3-methyl butanenitrile). In contrast, concentrations of the sulfur-containing compound 2-ethylthiophene and C(6) alcohol (E)-2-hexenol decreased. Finally, elevated CO(2) concentration increased soluble sugar concentrations due to enhanced photochemical activity in leaves and heads, which may account for the increased synthesis of volatiles.

  14. Differential responses of two broccoli (Brassica oleracea L. var Italica) cultivars to salinity and nutritional quality improvement.

    Science.gov (United States)

    Zaghdoud, Chokri; Alcaraz-López, Carlos; Mota-Cadenas, César; Martínez-Ballesta, María del Carmen; Moreno, Diego A; Ferchichi, Ali; Carvajal, Micaela

    2012-01-01

    The comparative responses of two broccoli cultivars (Brassica oleracea var. Italica, cv. Parthenon and cv. Naxos) to a 15 d exposure to different NaCl levels were investigated. Salinity led to increased concentrations of Na(+) and Cl(-) ions in both cultivars, a disruption of the endogenous minerals levels in the shoots and roots-that varied with the cultivar and salt concentration-and decreases in the osmotic potential (Ψ(π)), root hydraulic conductance (L(0)), and stomatal conductance (G(s)). The reduced biomass of Naxos at moderate NaCl indicates greater sensitivity to salinity, compared with Parthenon. Parthenon accumulated more soluble sugars, for osmotic adjustment, whereas Naxos accumulated proline, which gave the two cultivars differing nutritional characteristics. The total glucosinolates (GSLs) content was not affected by salinity in Parthenon while it decreased significantly in Naxos as a consequence of the decrease in the indole GSL. However, Naxos accumulated more aliphatic GSLs under salt stress than Parthenon, which confers on this cultivar a greater nutritional value when cultivated under salinity.These results suggest that, at distinct salinity levels, each broccoli cultivar adopts a specific strategy, indicating the crucial role of the genetic background on the organoleptic and nutritional properties that each cultivar acquires.

  15. UVA, UVB Light, and Methyl Jasmonate, Alone or Combined, Redirect the Biosynthesis of Glucosinolates, Phenolics, Carotenoids, and Chlorophylls in Broccoli Sprouts

    Directory of Open Access Journals (Sweden)

    Melissa Moreira-Rodríguez

    2017-11-01

    Full Text Available Broccoli sprouts contain health-promoting phytochemicals that can be enhanced by applying ultraviolet light (UV or phytohormones. The separate and combined effects of methyl jasmonate (MJ, UVA, or UVB lights on glucosinolate, phenolic, carotenoid, and chlorophyll profiles were assessed in broccoli sprouts. Seven-day-old broccoli sprouts were exposed to UVA (9.47 W/m2 or UVB (7.16 W/m2 radiation for 120 min alone or in combination with a 25 µM MJ solution, also applied to sprouts without UV supplementation. UVA + MJ and UVB + MJ treatments increased the total glucosinolate content by ~154% and ~148%, respectively. MJ induced the biosynthesis of indole glucosinolates, especially neoglucobrassicin (~538%, showing a synergistic effect with UVA stress. UVB increased the content of aliphatic and indole glucosinolates, such as glucoraphanin (~78% and 4-methoxy-glucobrassicin (~177%. UVA increased several phenolics such as gallic acid (~57% and a kaempferol glucoside (~25.4%. MJ treatment decreased most phenolic levels but greatly induced accumulation of 5-sinapoylquinic acid (~239%. MJ treatments also reduced carotenoid and chlorophyll content, while UVA increased lutein (~23%, chlorophyll b (~31%, neoxanthin (~34%, and chlorophyll a (~67%. Results indicated that UV- and/or MJ-treated broccoli sprouts redirect the carbon flux to the biosynthesis of specific glucosinolates, phenolics, carotenoids, and chlorophylls depending on the type of stress applied.

  16. The effect of the rate and method of nitrogen application on nitrogen uptake and utilization by broccoli (Brassica oleracea var. italica)

    NARCIS (Netherlands)

    Everaarts, A.P.; Willigen, de P.

    1999-01-01

    The effect of the rate and method of nitrogen application on nitrogen uptake and utilization by broccoli (Brassica oleracea var. italica) was studied in four field experiments. The methods of application were broadcast application vs band placement and split application. Maximum uptake of nitrogen

  17. Assessment of the anticancer compounds Se-methylselenocysteine and glucosinolates in Se-biofortified broccoli (Brassica oleracea L. var. italica) sprouts and florets.

    Science.gov (United States)

    Ávila, Fabricio William; Faquin, Valdemar; Yang, Yong; Ramos, Silvio Junio; Guilherme, Luiz Roberto G; Thannhauser, Theodore W; Li, Li

    2013-07-03

    Broccoli (Brassica oleracea L. var. italica) is a rich source of chemopreventive compounds. Here, we evaluated and compared the effect of selenium (Se) treatment on the accumulation of anticancer compounds Se-methylselenocysteine (SeMSCys) and glucosinolates in broccoli sprouts and florets. Total Se and SeMSCys content in sprouts increased concomitantly with increasing Se doses. Selenate was superior to selenite in inducing total Se accumulation, but selenite is equally effective as selenate in promoting SeMSCys synthesis in sprouts. Increasing sulfur doses reduced total Se and SeMSCys content in sprouts treated with selenate, but not in those with selenite. Examination of five broccoli cultivars reveals that sprouts generally have better fractional ability than florets to convert inorganic Se into SeMSCys. Distinctive glucosinolate profiles between sprouts and florets were observed, and sprouts contained approximately 6-fold more glucoraphanin than florets. In contrast to florets, glucosinolate content was not affected by Se treatment in sprouts. Thus, Se-enriched broccoli sprouts are excellent for simultaneous accumulation of chemopreventive compounds SeMSCys and glucoraphanin.

  18. Evaluation of consumer acceptance of west coast versus east coast-produced broccoli through sensory analysis of quality rating factors and nutritionally important metabolites

    Science.gov (United States)

    Production trials and germplasm evaluation of broccoli (Brassica oleracea var. italica) developed for eastern U.S. production conditions have identified lines and cultivars that are better adapted to more stressful, variable East Coast environments. As a part of this work sponsored by the USDA SCIR...

  19. Characterization of industrial broccoli discards (Brassica oleracea var. italica) for their glucosinolate, polyphenol and flavonoid contents using UPLC MS/MS and spectrophotometric methods.

    Science.gov (United States)

    Thomas, Minty; Badr, Ashraf; Desjardins, Yves; Gosselin, Andre; Angers, Paul

    2018-04-15

    The agrifood industry produces tons of waste and substandard products that are discarded at great expense. Valorization of industrial residues curbs issues related to food security and environmental problems. Broccoli (Brassica oleracea var. italica) is associated with varied beneficial health effects, but its production yields greater than 25% rejects. We aimed to characterize and quantify industrial broccoli by-products for their glucosinolate and polyphenol contents as a first step towards industrial bio-refining. Broccoli segments and rejected lots of 10 seed cultivars were analyzed using UPLC MS/MS. Variability in the contents of bioactive molecules was observed within and between the cultivars. Broccoli by-products were rich in glucosinolates (0.2-2% dry weight sample), predominantly glucoraphanin (32-64% of the total glucosinolates), whereas the polyphenolic content was less than 0.02% dry weight sample. Valorization of industrial residues facilitates the production of high value functional food ingredients along with socio-economic sustainability. Copyright © 2017 Elsevier Ltd. All rights reserved.

  20. Development of fluorescent methods for DNA methyltransferase assay

    Science.gov (United States)

    Li, Yueying; Zou, Xiaoran; Ma, Fei; Tang, Bo; Zhang, Chun-yang

    2017-03-01

    DNA methylation modified by DNA methyltransferase (MTase) plays an important role in regulating gene transcription, cell growth and proliferation. The aberrant DNA MTase activity may lead to a variety of human diseases including cancers. Therefore, accurate and sensitive detection of DNA MTase activity is crucial to biomedical research, clinical diagnostics and therapy. However, conventional DNA MTase assays often suffer from labor-intensive operations and time-consuming procedures. Alternatively, fluorescent methods have significant advantages of simplicity and high sensitivity, and have been widely applied for DNA MTase assay. In this review, we summarize the recent advances in the development of fluorescent methods for DNA MTase assay. These emerging methods include amplification-free and the amplification-assisted assays. Moreover, we discuss the challenges and future directions of this area.

  1. The role of histone methyltransferase EZH2 in myelodysplastic syndromes.

    Science.gov (United States)

    Xu, Feng; Li, Xiao

    2012-04-01

    Previous epigenetics research in myelodysplastic syndromes (MDS) mainly focused on the DNA methylation of tumor suppressor genes. Recent studies reported that around 6% of MDS patients have several EZH2 mutations including missense, frameshift and truncated mutations. Histone methyltransferase EZH2 plays a critical role in epigenetic regulation as a bridge between histone methylation/deacetylation and DNA methylation. EZH2 is frequently overexpressed and considered to be an oncogene in cancers; nevertheless, EZH2 is considered as a candidate tumor suppressor gene in MDS due to EZH2 mutations associated with poor survival. Many questions still need further discussion. Moreover, 3-deazaneplanocin can reduce EZH2 levels and H3K27 trimethylation, and synergistic effects are seen in combination with DNA demethylation agents or histone deacetylation inhibitors. All of the above give us more chances to improve epigenetic therapy in MDS. Therefore, the molecular mechanisms of EZH2 in tumorigenesis and the role of EZH2 in MDS are studied.

  2. Cloning of genes related to aliphatic glucosinolate metabolism and the mechanism of sulforaphane accumulation in broccoli sprouts under jasmonic acid treatment.

    Science.gov (United States)

    Guo, Liping; Yang, Runqiang; Gu, Zhenxin

    2016-10-01

    Cytochrome P450 79F1 (CYP79F1), cytochrome P450 83A1 (CYP83A1), UDP-glucosyltransferase 74B1 (UGT74B1), sulfotransferase 18 (ST5b) and flavin-containing monooxygenase GS-OX1 (FMOGS - OX1 ) are important enzymes in aliphatic glucosinolate biosynthesis. In this study, their full-length cDNA in broccoli was firstly cloned, then the mechanism of sulforaphane accumulation under jasmonic acid (JA) treatment was investigated. The full-length cDNA of CYP79F1, CYP83A1, UGT74B1, ST5b and FMOGS - OX1 comprised 1980, 1652, 1592, 1378 and 1623 bp respectively. The increase in aliphatic glucosinolate accumulation in broccoli sprouts treated with JA was associated with elevated expression of genes in the aliphatic glucosinolate biosynthetic pathway. Application of 100 µmol L(-1) JA increased myrosinase (MYR) activity but did not affect epithiospecifier protein (ESP) activity in broccoli sprouts, which was supported by the expression of MYR and ESP. Sulforaphane formation in 7-day-old sprouts treated with 100 µmol L(-1) JA was 3.36 and 1.30 times that in the control and 300 µmol L(-1) JA treatment respectively. JA enhanced the accumulation of aliphatic glucosinolates in broccoli sprouts via up-regulation of related gene expression. Broccoli sprouts treated with 100 µmol L(-1) JA showed higher sulforphane formation than those treated with 300 µmol L(-1) JA owing to the higher glucoraphanin content and myrosinase activity under 100 µmol L(-1) JA treatment. © 2016 Society of Chemical Industry. © 2016 Society of Chemical Industry.

  3. Chromosome doubling of microspore-derived plants from cabbage (Brassica oleracea var. capitata L. and broccoli (Brassica oleracea var. italica L.

    Directory of Open Access Journals (Sweden)

    Suxia eYuan

    2015-12-01

    Full Text Available Chromosome doubling of microspore-derived plants is an important factor in the practical application of microspore culture technology because breeding programs require a large number of genetically stable, homozygous doubled haploid plants with a high level of fertility. In the present paper, 29 populations of microspore-derived plantlets from cabbage (Brassica oleracea var. capitata and broccoli (Brassica oleracea var. italica were used to study the ploidy level and spontaneous chromosome doubling of these populations, the artificial chromosome doubling induced by colchicine, and the influence of tissue culture duration on the chromosomal ploidy of the microspore-derived regenerants. Spontaneous chromosome doubling occurred randomly and was genotype dependent. In the plant populations derived from microspores, there were haploids, diploids, and even a low frequency of polyploids and mixed-ploidy plantlets. The total spontaneous doubling in the 14 cabbage populations ranged from 0 - 76.9%, compared with 52.2 - 100% in the 15 broccoli populations. To improve the rate of chromosome doubling, an efficient and reliable artificial chromosome doubling protocol (i.e., the immersion of haploid plantlet roots in a colchicine solution was developed for cabbage and broccoli microspore-derived haploids. The optimal chromosome doubling of the haploids was obtained with a solution of 0.2% colchicine for 9 - 12 h or 0.4% colchicine for 3 - 9 h for cabbage and 0.05% colchicine for 6 - 12 h for broccoli. This protocol produced chromosome doubling in over 50% of the haploid genotypes for most of the populations derived from cabbage and broccoli. Notably, after 1 or more years in tissue culture, the chromosomes of the haploids were doubled, and most of the haploids turned into doubled haploid or mixed-ploidy plants. This is the first report indicating that tissue culture duration can change the chromosomal ploidy of microspore-derived regenerants.

  4. Plants as Biofactories: Postharvest Stress-Induced Accumulation of Phenolic Compounds and Glucosinolates in Broccoli Subjected to Wounding Stress and Exogenous Phytohormones

    Science.gov (United States)

    Villarreal-García, Daniel; Nair, Vimal; Cisneros-Zevallos, Luis; Jacobo-Velázquez, Daniel A.

    2016-01-01

    Broccoli contains high levels of bioactive molecules and is considered a functional food. In this study, postharvest treatments to enhance the concentration of glucosinolates and phenolic compounds were evaluated. Broccoli whole heads were wounded to obtain florets and wounded florets (florets cut into four even pieces) and stored for 24 h at 20 °C with or without exogenous ethylene (ET, 1000 ppm) or methyl jasmonate (MeJA, 250 ppm). Whole heads were used as a control for wounding treatments. Regarding glucosinolate accumulation, ET selectively induced the 4-hydroxylation of glucobrassicin in whole heads, resulting in ∼223% higher 4-hydroxyglucobrassicin than time 0 h samples. Additionally, glucoraphanin was increased by ∼53% in whole heads treated with ET, while neoglucobrassicin was greatly accumulated in wounded florets treated with ET or MeJA, showing increases of ∼193 and ∼286%, respectively. On the other hand, although only whole heads stored without phytohormones showed higher concentrations of phenolic compounds, which was reflected in ∼33, ∼30, and ∼46% higher levels of 1,2,2-trisinapoylgentiobose, 1,2-diferulolylgentiobiose, and 1,2-disinapoyl-2-ferulolylgentiobiose, respectively; broccoli florets stored under air control conditions showed enhanced concentrations of 3-O-caffeoylquinic acid, 1,2-disinapoylgentiobiose, and 1,2-disinapoyl-2-ferulolylgentiobiose (∼22, ∼185, and ∼65% more, respectively). Furthermore, exogenous ET and MeJA impeded individual phenolics accumulation. Results allowed the elucidation of simple and effective postharvest treatment to enhance the content of individual glucosinolates and phenolic compounds in broccoli. The stressed-broccoli tissue could be subjected to downstream processing in order to extract and purify bioactive molecules with applications in the dietary supplements, agrochemical and cosmetics markets. PMID:26904036

  5. Plants as biofactories: Postharvest Stress-Induced Accumulation of Phenolic Compounds and Glucosinolates in Broccoli Subjected to Wounding Stress and Exogenous Phytohormones

    Directory of Open Access Journals (Sweden)

    Daniel eVillarreal-García

    2016-02-01

    Full Text Available Broccoli contains high levels of bioactive molecules and is considered a functional food. In this study, postharvest treatments to enhance the concentration of glucosinolates and phenolic compounds were evaluated. Broccoli whole heads were wounded to obtain florets and wounded florets (florets cut into four even pieces and stored for 24 h at 20 ºC with or without exogenous ethylene (ET, 1000 ppm or methyl jasmonate (MeJA, 250 ppm. Whole heads were used as a control for wounding treatments. Regarding glucosinolate accumulation, ET selectively induced the 4-hydroxylation of glucobrassicin in whole heads, resulting in ~223% higher 4-hydroxyglucobrassicin than time 0 h samples. Additionally, glucoraphanin was increased by ~53% in whole heads treated with ET, while neoglucobrassicin was greatly accumulated in wounded florets treated with ET or MeJA, showing increases of ~193% and ~286%, respectively. On the other hand, although only whole heads stored without phytohormones showed higher concentrations of phenolic compounds, which was reflected in ~33%, ~30%, and 46% higher levels of 1,2,2-trisinapoylgentiobose, 1,2-diferulolylgentiobiose, and 1,2-disinapoyl-2-ferulolylgentiobiose, respectively; broccoli florets stored under air control conditions showed enhanced concentrations of 3-O-caffeoylquinic acid, 1,2-disinapoylgentiobiose, and 1,2-disinapoyl-2-ferulolylgentiobiose (~22%, ~185%, and ~65% more, respectively. However, exogenous ET and MeJA impeded individual phenolics accumulation. Results allowed the elucidation of simple and effective postharvest treatment to enhance the content of individual glucosinolates and phenolic compounds in broccoli. The stressed-broccoli tissue could be subjected to downstream processing in order to extract and purify bioactive molecules with applications in the dietary supplements, agrochemical and cosmetics markets.

  6. Chromosome Doubling of Microspore-Derived Plants from Cabbage (Brassica oleracea var. capitata L.) and Broccoli (Brassica oleracea var. italica L.).

    Science.gov (United States)

    Yuan, Suxia; Su, Yanbin; Liu, Yumei; Li, Zhansheng; Fang, Zhiyuan; Yang, Limei; Zhuang, Mu; Zhang, Yangyong; Lv, Honghao; Sun, Peitian

    2015-01-01

    Chromosome doubling of microspore-derived plants is an important factor in the practical application of microspore culture technology because breeding programs require a large number of genetically stable, homozygous doubled haploid plants with a high level of fertility. In the present paper, 29 populations of microspore-derived plantlets from cabbage (Brassica oleracea var. capitata) and broccoli (Brassica oleracea var. italica) were used to study the ploidy level and spontaneous chromosome doubling of these populations, the artificial chromosome doubling induced by colchicine, and the influence of tissue culture duration on the chromosomal ploidy of the microspore-derived regenerants. Spontaneous chromosome doubling occurred randomly and was genotype dependent. In the plant populations derived from microspores, there were haploids, diploids, and even a low frequency of polyploids and mixed-ploidy plantlets. The total spontaneous doubling in the 14 cabbage populations ranged from 0 to 76.9%, compared with 52.2 to 100% in the 15 broccoli populations. To improve the rate of chromosome doubling, an efficient and reliable artificial chromosome doubling protocol (i.e., the immersion of haploid plantlet roots in a colchicine solution) was developed for cabbage and broccoli microspore-derived haploids. The optimal chromosome doubling of the haploids was obtained with a solution of 0.2% colchicine for 9-12 h or 0.4% colchicine for 3-9 h for cabbage and 0.05% colchicine for 6-12 h for broccoli. This protocol produced chromosome doubling in over 50% of the haploid genotypes for most of the populations derived from cabbage and broccoli. Notably, after 1 or more years in tissue culture, the chromosomes of the haploids were doubled, and most of the haploids turned into doubled haploid or mixed-ploidy plants. This is the first report indicating that tissue culture duration can change the chromosomal ploidy of microspore-derived regenerants.

  7. Methyltransferase G9A regulates T cell differentiation during murine intestinal inflammation

    National Research Council Canada - National Science Library

    Antignano, Frann; Burrows, Kyle; Hughes, Michael R; Han, Jonathan M; Kron, Ken J; Penrod, Nadia M; Oudhoff, Menno J; Wang, Steven Kai Hao; Min, Paul H; Gold, Matthew J; Chenery, Alistair L; Braam, Mitchell J S; Fung, Thomas C; Rossi, Fabio M V; McNagny, Kelly M; Arrowsmith, Cheryl H; Lupien, Mathieu; Levings, Megan K; Zaph, Colby

    2014-01-01

    .... Using a murine T cell transfer model of colitis, we found that T cell-intrinsic expression of the histone lysine methyltransferase G9A was required for development of pathogenic T cells and intestinal inflammation...

  8. Molecular Basis for the Regulation of the H3K4 Methyltransferase Activity of PRDM9

    Directory of Open Access Journals (Sweden)

    Hong Wu

    2013-10-01

    Full Text Available PRDM9, a histone lysine methyltransferase, is a key determinant of the localization of meiotic recombination hot spots in humans and mice and the only vertebrate protein known to be involved in hybrid sterility. Here, we report the crystal structure of the PRDM9 methyltransferase domain in complex with a histone H3 peptide dimethylated on lysine 4 (H3K4me2 and S-adenosylhomocysteine (AdoHcy, which provides insights into the methyltransferase activity of PRDM proteins. We show that the genuine substrate of PRDM9 is histone H3 lysine 4 (H3K4 and that the enzyme possesses mono-, di-, and trimethylation activities. We also determined the crystal structure of PRDM9 in its autoinhibited state, which revealed a rearrangement of the substrate and cofactor binding sites by a concerted action of the pre-SET and post-SET domains, providing important insights into the regulatory mechanisms of histone lysine methyltransferase activity.

  9. An audit of thiopurine methyltransferase genotyping and phenotyping before intended azathioprine treatment for dermatological conditions

    DEFF Research Database (Denmark)

    Vestergaard, T; Bygum, A

    2009-01-01

    Summary Background. Determining thiopurine methyltransferase (TPMT) genotype and phenotype before azathioprine treatment predicts which patients are most likely to develop myelosuppression. Aim. To evaluate the course of azathioprine treatment in people with TPMT heterozygosity and whether this d...

  10. Presence of DNA methyltransferase activity and CpC methylation in Drosophila melanogaster.

    Science.gov (United States)

    Panikar, Chitra S; Rajpathak, Shriram N; Abhyankar, Varada; Deshmukh, Saniya; Deobagkar, Deepti D

    2015-12-01

    Drosophila melanogaster lacks DNMT1/DNMT3 based methylation machinery. Despite recent reports confirming the presence of low DNA methylation in Drosophila; little is known about the methyltransferase. Therefore, in this study, we have aimed to investigate the possible functioning of DNA methyltransferase in Drosophila. The 14 K oligo microarray slide was incubated with native cell extract from adult Drosophila to check the presence of the methyltransferase activity. After incubation under appropriate conditions, the methylated oligo sequences were identified by the binding of anti 5-methylcytosine monoclonal antibody. The antibody bound to the methylated oligos was detected using Cy3 labeled secondary antibody. Methylation sensitive restriction enzyme mediated PCR was used to assess the methylation at a few selected loci identified on the array. It could be seen that a few of the total oligos got methylated under the assay conditions. Analysis of methylated oligo sequences provides evidence for the presence of de novo methyltransferase activity and allows identification of its sequence specificity in adult Drosophila. With the help of methylation sensitive enzymes we could detect presence of CpC methylation in the selected genomic regions. This study reports presence of an active DNA methyltransferase in adult Drosophila, which exhibits sequence specificity confirmed by presence of asymmetric methylation at corresponding sites in the genomic DNA. It also provides an innovative approach to investigate methylation specificity of a native methyltransferase.

  11. Effects of broccoli stem and leaf meal on broiler performance, skin pigmentation, antioxidant function, and meat quality.

    Science.gov (United States)

    Hu, C H; Wang, D G; Pan, H Y; Zheng, W B; Zuo, A Y; Liu, J X

    2012-09-01

    Three hundred sixty 1-d-old Ross 308 male broilers were used to study the effects of broccoli stem and leaf meal (BSLM) on growth performance, skin pigmentation, antioxidant function, and meat quality. The chicks were fed 4 diets containing different levels (0, 4.0, 8.0, and 12.0%) of BSLM as partial replacement for corn and soybean meal for a period of 42 d. The results showed that dietary supplementation of BSLM had no effect (P > 0.05) on growth performance. As compared with control, dietary 4%, 8%, and 12% BSLM increased (P poultry products quality with the more skin pigmentation and the less drip loss percentage of breast meat. The more skin pigmentation mainly related to the high amount of xanthophylls in BSLM. The decreased meat drip loss fed BSLM may be caused by the antioxidative function of BSLM.

  12. EFFECT OF SOIL SULFUR FERTILIZER AND SOME FOLIAR FERTILIZERS ON GROWTH AND YIELD OF BROCCOLI IN SALINE SOIL

    Directory of Open Access Journals (Sweden)

    Ali Husain JASIM

    2015-12-01

    Full Text Available Factorial experiment was conducted in the open fields of Agricultural College, Al-Qasim Green University during the agricultural seasons of 2013/2014 and 2014/2015 to study the effect of adding two levels of agricultural sulfur (control and add 100 kg.ha-1 and four levels of nutrient spray (without spray, high-potash fertilizer, high-phosphorus fertilizer and humic acid on growth and yield of broccoli under drip irrigation and polyethylene soil mulching in saline soil (9.6 dS.m-1. Randomized complete block design with three replicates was used. The results showed that agricultural sulfur led to increase number of leaves, leaf area, leaves chlorophyll content, diameter and weight of flower head compared to control. Spraying foliar fertilizer and its interaction with sulfur fertilizer also led to increase all of parameters above (except leaves chlorophyll content significantly compared to control treatment.

  13. Variation in Broccoli Cultivar Phytochemical Content under Organic and Conventional Management Systems: Implications in Breeding for Nutrition

    Science.gov (United States)

    Renaud, Erica N. C.; Lammerts van Bueren, Edith T.; Myers, James R.; Paulo, Maria João; van Eeuwijk, Fred A.; Zhu, Ning; Juvik, John A.

    2014-01-01

    Organic agriculture requires cultivars that can adapt to organic crop management systems without the use of synthetic pesticides as well as genotypes with improved nutritional value. The aim of this study encompassing 16 experiments was to compare 23 broccoli cultivars for the content of phytochemicals associated with health promotion grown under organic and conventional management in spring and fall plantings in two broccoli growing regions in the US (Oregon and Maine). The phytochemicals quantified included: glucosinolates (glucoraphanin, glucobrassicin, neoglucobrassin), tocopherols (δ-, γ-, α-tocopherol) and carotenoids (lutein, zeaxanthin, β-carotene). For glucoraphanin (17.5%) and lutein (13%), genotype was the major source of total variation; for glucobrassicin, region (36%) and the interaction of location and season (27.5%); and for neoglucobrassicin, both genotype (36.8%) and its interactions (34.4%) with season were important. For δ- and γ- tocopherols, season played the largest role in the total variation followed by location and genotype; for total carotenoids, genotype (8.41–13.03%) was the largest source of variation and its interactions with location and season. Overall, phytochemicals were not significantly influenced by management system. We observed that the cultivars with the highest concentrations of glucoraphanin had the lowest for glucobrassicin and neoglucobrassicin. The genotypes with high concentrations of glucobrassicin and neoglucobrassicin were the same cultivars and were early maturing F1 hybrids. Cultivars highest in tocopherols and carotenoids were open pollinated or early maturing F1 hybrids. We identified distinct locations and seasons where phytochemical performance was higher for each compound. Correlations among horticulture traits and phytochemicals demonstrated that glucoraphanin was negatively correlated with the carotenoids and the carotenoids were correlated with one another. Little or no association between

  14. Two protein lysine methyltransferases methylate outer membrane protein B from Rickettsia.

    Science.gov (United States)

    Abeykoon, Amila H; Chao, Chien-Chung; Wang, Guanghui; Gucek, Marjan; Yang, David C H; Ching, Wei-Mei

    2012-12-01

    Rickettsia prowazekii, the etiologic agent of epidemic typhus, is a potential biological threat agent. Its outer membrane protein B (OmpB) is an immunodominant antigen and plays roles as protective envelope and as adhesins. The observation of the correlation between methylation of lysine residues in rickettsial OmpB and bacterial virulence has suggested the importance of an enzymatic system for the methylation of OmpB. However, no rickettsial lysine methyltransferase has been characterized. Bioinformatic analysis of genomic DNA sequences of Rickettsia identified putative lysine methyltransferases. The genes of the potential methyltransferases were synthesized, cloned, and expressed in Escherichia coli, and expressed proteins were purified by nickel-nitrilotriacetic acid (Ni-NTA) affinity chromatography. The methyltransferase activities of the purified proteins were analyzed by methyl incorporation of radioactively labeled S-adenosylmethionine into recombinant fragments of OmpB. Two putative recombinant methyltransferases (rRP789 and rRP027-028) methylated recombinant OmpB fragments. The specific activity of rRP789 is 10- to 30-fold higher than that of rRP027-028. Western blot analysis using specific antibodies against trimethyl lysine showed that both rRP789 and rRP027-028 catalyzed trimethylation of recombinant OmpB fragments. Liquid chromatography-tandem mass spectrometry (LC/MS-MS) analysis showed that rRP789 catalyzed mono-, di-, and trimethylation of lysine, while rRP027-028 catalyzed exclusively trimethylation. To our knowledge, rRP789 and rRP027-028 are the first biochemically characterized lysine methyltransferases of outer membrane proteins from Gram-negative bacteria. The production and characterization of rickettsial lysine methyltransferases provide new tools to investigate the mechanism of methylation of OmpB, effects of methylation on the structure and function of OmpB, and development of methylated OmpB-based diagnostic assays and vaccine candidates.

  15. DNA methyltransferase inhibitor CDA-II inhibits myogenic differentiation

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Zirong [State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-Sen University, Guangzhou 510060 (China); Department of Molecular Genetics and Microbiology, Shands Cancer Center, University of Florida, Gainesville, FL 32610 (United States); Jin, Guorong [State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-Sen University, Guangzhou 510060 (China); Lin, Shuibin [Department of Molecular Genetics and Microbiology, Shands Cancer Center, University of Florida, Gainesville, FL 32610 (United States); Lin, Xiumei [Department of Hematology, Guangzhou First Municipal People' s Hospital, Guangzhou 510180 (China); Gu, Yumei [Department of Molecular Genetics and Microbiology, Shands Cancer Center, University of Florida, Gainesville, FL 32610 (United States); Zhu, Yujuan; Hu, Chengbin; Zhang, Qingjiong [State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-Sen University, Guangzhou 510060 (China); Wu, Lizi [Department of Molecular Genetics and Microbiology, Shands Cancer Center, University of Florida, Gainesville, FL 32610 (United States); Shen, Huangxuan, E-mail: shenhx@mail.sysu.edu.cn [State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-Sen University, Guangzhou 510060 (China)

    2012-06-08

    Highlights: Black-Right-Pointing-Pointer CDA-II inhibits myogenic differentiation in a dose-dependent manner. Black-Right-Pointing-Pointer CDA-II repressed expression of muscle transcription factors and structural proteins. Black-Right-Pointing-Pointer CDA-II inhibited proliferation and migration of C2C12 myoblasts. -- Abstract: CDA-II (cell differentiation agent II), isolated from healthy human urine, is a DNA methyltransferase inhibitor. Previous studies indicated that CDA-II played important roles in the regulation of cell growth and certain differentiation processes. However, it has not been determined whether CDA-II affects skeletal myogenesis. In this study, we investigated effects of CDA-II treatment on skeletal muscle progenitor cell differentiation, migration and proliferation. We found that CDA-II blocked differentiation of murine myoblasts C2C12 in a dose-dependent manner. CDA-II repressed expression of muscle transcription factors, such as Myogenin and Mef2c, and structural proteins, such as myosin heavy chain (Myh3), light chain (Mylpf) and MCK. Moreover, CDA-II inhibited C1C12 cell migration and proliferation. Thus, our data provide the first evidence that CDA-II inhibits growth and differentiation of muscle progenitor cells, suggesting that the use of CDA-II might affect skeletal muscle functions.

  16. Identification and functional characterization of lysine methyltransferases of Entamoeba histolytica.

    Science.gov (United States)

    Borbolla-Vázquez, Jessica; Orozco, Esther; Medina-Gómez, Christian; Martínez-Higuera, Aarón; Javier-Reyna, Rosario; Chávez, Bibiana; Betanzos, Abigail; Rodríguez, Mario A

    2016-07-01

    Lysine methylation of histones, a posttranslational modification catalyzed by lysine methyltransferases (HKMTs), plays an important role in the epigenetic regulation of transcription. Lysine methylation of non-histone proteins also impacts the biological function of proteins. Previously it has been shown that lysine methylation of histones of Entamoeba histolytica, the protozoan parasite that infects 50 million people worldwide each year and causing up to 100,000 deaths annually, is implicated in the epigenetic machinery of this microorganism. However, the identification and characterization of HKMTs in this parasite had not yet been determined. In this work we identified four HKMTs in E. histolytica (EhHKMT1 to EhHKMT4) that are expressed by trophozoites. Enzymatic assays indicated that all of them are able to transfer methyl groups to commercial histones. EhHKMT1, EhHKMT2 and EhHKMT4 were detected in nucleus and cytoplasm of trophozoites. In addition EhHKMT2 and EhHKMT4 were located in vesicles containing ingested cells during phagocytosis, and they co-immunoprecipitated with EhADH, a protein involved in the phagocytosis of this parasite. Results suggest that E. histolytica uses its HKMTs to regulate transcription by epigenetic mechanisms, and at least two of them could also be implicated in methylation of proteins that participate in phagocytosis. © 2016 John Wiley & Sons Ltd.

  17. Cell and molecular biology of DNA methyltransferase 1.

    Science.gov (United States)

    Mohan, K Naga; Chaillet, J Richard

    2013-01-01

    The DNA cytosine methyltransferase 1 (DNMT1) is a ubiquitous nuclear enzyme that catalyzes the well-established reaction of placing methyl groups on the unmethylated cytosines in methyl-CpG:CpG base pairs in the hemimethylated DNA formed by methylated parent and unmethylated daughter strands. This activity regenerates fully methylated methyl-CpG:methyl-CpG pairs. Despite the straightforward nature of its catalytic activity, detailed biochemical, genetic, and developmental studies revealed intricate details of the central regulatory role of DNMT1 in governing the epigenetic makeup of the nuclear genome. DNMT1 mediates demethylation and also participates in seemingly wide cellular functions unrelated to maintenance DNA methylation. This review brings together mechanistic details of maintenance methylation by DNMT1, its regulation at transcriptional and posttranscriptional levels, and the seemingly unexpected functions of DNMT1 in the context of DNA methylation which is central to epigenetic changes that occur during development and the process of cell differentiation. © 2013 Elsevier Inc. All rights reserved.

  18. Theoretical insights into catalytic mechanism of protein arginine methyltransferase 1.

    Directory of Open Access Journals (Sweden)

    Ruihan Zhang

    Full Text Available Protein arginine methyltransferase 1 (PRMT1, the major arginine asymmetric dimethylation enzyme in mammals, is emerging as a potential drug target for cancer and cardiovascular disease. Understanding the catalytic mechanism of PRMT1 will facilitate inhibitor design. However, detailed mechanisms of the methyl transfer process and substrate deprotonation of PRMT1 remain unclear. In this study, we present a theoretical study on PRMT1 catalyzed arginine dimethylation by employing molecular dynamics (MD simulation and quantum mechanics/molecular mechanics (QM/MM calculation. Ternary complex models, composed of PRMT1, peptide substrate, and S-adenosyl-methionine (AdoMet as cofactor, were constructed and verified by 30-ns MD simulation. The snapshots selected from the MD trajectory were applied for the QM/MM calculation. The typical SN2-favored transition states of the first and second methyl transfers were identified from the potential energy profile. Deprotonation of substrate arginine occurs immediately after methyl transfer, and the carboxylate group of E144 acts as proton acceptor. Furthermore, natural bond orbital analysis and electrostatic potential calculation showed that E144 facilitates the charge redistribution during the reaction and reduces the energy barrier. In this study, we propose the detailed mechanism of PRMT1-catalyzed asymmetric dimethylation, which increases insight on the small-molecule effectors design, and enables further investigations into the physiological function of this family.

  19. High expression of DNA methyltransferases in primary human medulloblastoma.

    Science.gov (United States)

    Pócza, T; Krenács, T; Turányi, E; Csáthy, J; Jakab, Z; Hauser, P

    2016-01-01

    Epigenetic alterations have been implicated in cancer development. DNA methylation modulates gene expression, which is catalyzed by DNA methyltransferases (DNMTs). The objective of our study was to evaluate expression of DNMTs in medulloblastoma and analyze its correlation with clinical features. Nuclear expression of DNMT1, DNMT3A and DNMT3B was analyzed in human primary medulloblastoma of 44 patients using immunohistochemistry. Correlation of expression of DNMT levels with classical histological subtypes, novel molecular subgroups and survival of patients was analyzed. Elevated expression of DNMT1, DNMT3A and DNMT3B was observed in 63.64%, 68.18% and 72.73% of all cases, respectively. None of them showed a correlation with classical histology or survival. Concerning molecular subtypes, significantly higher expression of DNMT1 was observed in the SHH group compared to non-SHH samples (p = 0.02), but without significant difference in DNMT3A or DNMT3B levels between any subtypes. In conclusion, DNMT1, DNMT3A and DNMT3B are highly expressed in human medulloblastoma samples, suggesting that promoter hypermethylation may play a role in medulloblastoma development. Demethylation of tumor suppressor gene promoters may be considered as a possible future target in therapy of medulloblastoma.

  20. Germinal center dysregulation by histone methyltransferase EZH2 promotes lymphomagenesis

    Science.gov (United States)

    Caganova, Marieta; Carrisi, Chiara; Varano, Gabriele; Mainoldi, Federica; Zanardi, Federica; Germain, Pierre-Luc; George, Laura; Alberghini, Federica; Ferrarini, Luca; Talukder, Asoke K.; Ponzoni, Maurilio; Testa, Giuseppe; Nojima, Takuya; Doglioni, Claudio; Kitamura, Daisuke; Toellner, Kai-M.; Su, I-hsin; Casola, Stefano

    2013-01-01

    Protection against deadly pathogens requires the production of high-affinity antibodies by B cells, which are generated in germinal centers (GCs). Alteration of the GC developmental program is common in many B cell malignancies. Identification of regulators of the GC response is crucial to develop targeted therapies for GC B cell dysfunctions, including lymphomas. The histone H3 lysine 27 methyltransferase enhancer of zeste homolog 2 (EZH2) is highly expressed in GC B cells and is often constitutively activated in GC-derived non-Hodgkin lymphomas (NHLs). The function of EZH2 in GC B cells remains largely unknown. Herein, we show that Ezh2 inactivation in mouse GC B cells caused profound impairment of GC responses, memory B cell formation, and humoral immunity. EZH2 protected GC B cells against activation-induced cytidine deaminase (AID) mutagenesis, facilitated cell cycle progression, and silenced plasma cell determinant and tumor suppressor B-lymphocyte–induced maturation protein 1 (BLIMP1). EZH2 inhibition in NHL cells induced BLIMP1, which impaired tumor growth. In conclusion, EZH2 sustains AID function and prevents terminal differentiation of GC B cells, which allows antibody diversification and affinity maturation. Dysregulation of the GC reaction by constitutively active EZH2 facilitates lymphomagenesis and identifies EZH2 as a possible therapeutic target in NHL and other GC-derived B cell diseases. PMID:24200695

  1. The Role of Protein Arginine Methyltransferases in Inflammatory Responses

    Directory of Open Access Journals (Sweden)

    Ji Hye Kim

    2016-01-01

    Full Text Available Protein arginine methyltransferases (PRMTs mediate the methylation of a number of protein substrates of arginine residues and serve critical functions in many cellular responses, including cancer development, progression, and aggressiveness, T-lymphocyte activation, and hepatic gluconeogenesis. There are nine members of the PRMT family, which are divided into 4 types (types I–IV. Although most PRMTs do not require posttranslational modification (PTM to be activated, fine-tuning modifications, such as interactions between cofactor proteins, subcellular compartmentalization, and regulation of RNA, via micro-RNAs, seem to be required. Inflammation is an essential defense reaction of the body to eliminate harmful stimuli, including damaged cells, irritants, or pathogens. However, chronic inflammation can eventually cause several types of diseases, including some cancers, atherosclerosis, rheumatoid arthritis, and periodontitis. Therefore, inflammation responses should be well modulated. In this review, we briefly discuss the role of PRMTs in the control of inflammation. More specifically, we review the roles of four PRMTs (CARM1, PRMT1, PRMT5, and PRMT6 in modulating inflammation responses, particularly in terms of modulating the transcriptional factors or cofactors related to inflammation. Based on the regulatory roles known so far, we propose that PRMTs should be considered one of the target molecule groups that modulate inflammatory responses.

  2. Mapping the conformational space accessible to catechol-O-methyltransferase.

    Science.gov (United States)

    Ehler, Andreas; Benz, Jörg; Schlatter, Daniel; Rudolph, Markus G

    2014-08-01

    Methylation catalysed by catechol-O-methyltransferase (COMT) is the main pathway of catechol neurotransmitter deactivation in the prefrontal cortex. Low levels of this class of neurotransmitters are held to be causative of diseases such as schizophrenia, depression and Parkinson's disease. Inhibition of COMT may increase neurotransmitter levels, thus offering a route for treatment. Structure-based drug design hitherto seems to be based on the closed enzyme conformation. Here, a set of apo, semi-holo, holo and Michaelis form crystal structures are described that define the conformational space available to COMT and that include likely intermediates along the catalytic pathway. Domain swaps and sizeable loop movements around the active site testify to the flexibility of this enzyme, rendering COMT a difficult drug target. The low affinity of the co-substrate S-adenosylmethionine and the large conformational changes involved during catalysis highlight significant energetic investment to achieve the closed conformation. Since each conformation of COMT is a bona fide target for inhibitors, other states than the closed conformation may be promising to address. Crystallographic data for an alternative avenue of COMT inhibition, i.e. locking of the apo state by an inhibitor, are presented. The set of COMT structures may prove to be useful for the development of novel classes of inhibitors.

  3. Catechol-O-methyltransferase inhibitors in Parkinson's disease.

    Science.gov (United States)

    Müller, Thomas

    2015-02-01

    Inhibitors of catechol-O-methyltransferase (COMT) are commonly used as an adjunct to levodopa in patients with Parkinson's disease (PD) for the amelioration of wearing-off symptoms. This narrative review aims to discuss the role of COMT inhibitors on peripheral levodopa metabolism and continuous brain delivery of levodopa, and to describe their metabolic properties. Oral application of levodopa formulations with a dopa decarboxylase inhibitor (DDI) results in fluctuating levodopa plasma concentrations, predominantly due to the short half-life of levodopa and its slowing of gastric emptying. Following transport across the blood-brain barrier and its metabolic conversion to dopamine, these peripheral 'ups and downs' of levodopa are reflected in fluctuating dopamine levels in the synaptic cleft between presynaptic and postsynaptic dopaminergic neurons of the nigrostriatal system. As a result, pulsatile postsynaptic dopaminergic stimulation takes place and results in the occurrence of motor complications, such as wearing-off and dyskinesia. More continuous plasma behaviour was observed after the combination of levodopa/DDI formulations with COMT inhibitors. These compounds also weaken a levodopa/DDI-related homocysteine increase, as biomarker for an impaired methylation capacity, which is involved in an elevated oxidative stress exposure. These findings favour the concept of chronic levodopa/DDI application with concomitant inhibition of COMT and monoamine oxidase, since deamination of dopamine via this enzyme also generates free radicals. This triple combination is suggested as standard levodopa application in patients with PD who need levodopa, if they will tolerate it.

  4. Targeting histone methyltransferase EZH2 as cancer treatment.

    Science.gov (United States)

    Kondo, Yutaka

    2014-11-01

    It is widely accepted that epigenetic alterations are associated with different stages of tumour formation and progression in many cancers. Therefore, epigenetic abnormalities in cancers are emerging as important biomarkers and may have therapeutic potential. The polycomb repressive complex 2 (PRC2) is a key epigenetic regulator that catalyses trimethylation of lysine 27 on histone H3 (H3K27me3) via the histone methyltransferase, EZH2, which confers stemness and regulates differentiation during embryonic development. Given these roles of EZH2 and H3K27me3, plastic and dynamic features of cancer cells, especially cancer stem cells (CSCs), may be closely associated with this epigenetic mechanism. In addition, recent sequencing technology revealed that there are many recurrent mutations in polycomb-related genes, including EZH2, in different types of cancers. Therefore, researchers focused on targeting EZH2 as a novel cancer treatment and identified small compounds that inhibit EZH2 activity. Some of them are now under clinical trial in B-cell lymphoma. However, the underlying mechanisms by which PRC2 precisely regulate epigenetic alterations at certain genomic loci under different cellular conditions remain unclear. In this review, I focus on the recent advancements in EZH2 research, especially its dynamic regulation of epigenetic alterations in tumour cells, including the CSC population, and discuss perspectives and challenges for cancer treatment in the near future. © The Authors 2014. Published by Oxford University Press on behalf of the Japanese Biochemical Society. All rights reserved.

  5. Characterization of DNA methyltransferase and demethylase genes in Fragaria vesca.

    Science.gov (United States)

    Gu, Tingting; Ren, Shuai; Wang, Yuanhua; Han, Yuhui; Li, Yi

    2016-06-01

    DNA methylation is an epigenetic modification essential for gene regulations in plants, but understanding on how it is involved in fruit development, especially in non-climacteric fleshy fruit, is limited. The diploid woodland strawberry (Fragaria vesca) is an important model for non-climacteric fruit crops. In this study, we identified DNA methyltransferase genes and demethylase genes in Fragaria vesca and other angiosperm species. In accordance with previous studies, our phylogenetic analyses of those DNA methylation modifiers support the clustering of those genes into several classes. Our data indicate that whole-genome duplications and tandem duplications contributed to the expansion of those DNA methylation modifiers in angiosperms. We have further demonstrated that some DNA methylase and demethylase genes reach their highest expression levels in strawberry fleshy fruits when turning from white to red, suggesting that DNA methylation might undergo a dramatic change at the onset of fleshy fruit-ripening process. In addition, we have observed that expression of some DNA demethylase genes increases in response to various abiotic stresses including heat, cold, drought and salinity. Collectively, our study indicates a regulatory role of DNA methylation in the turning stage of non-climacteric fleshy fruit and responses to environment stimuli, and would facilitate functional studies of DNA methylation in the growth and development of non-climacteric fruits.

  6. Atrazine-resistant cytoplasmic male-sterile-nigra broccoli obtained by protoplast fusion between cytoplasmic male-sterile Brassica oleracea and atrazine-resistant Brassica campestris.

    Science.gov (United States)

    Christey, M C; Makaroff, C A; Earle, E D

    1991-12-01

    Protoplast fusion was used to combine the cytoplasmic traits of atrazine resistance and male sterility in Brassica oleracea var. italica (broccoli). Leaf protoplasts from broccoli with the petaloid B. nigra type of cytoplasmic male sterility were fused with hypocotyl protoplasts from an atrazine-resistant biotype of B. campestris var. oleifera cv Candle (oilseed rape). A total of 19 colonies regenerated shoots, all of which were broccolilike in phenotype, i.e., lacked trichomes. Four shoots, all from one colony, were atrazine resistant, surviving and growing in the presence of 25 μM atrazine. A leaf piece assay also confirmed that they were atrazine resistant. Molecular analysis showed that they contain chloroplasts from the atrazine-resistant B. campestris parent and mitochondria from the B. nigra parent. No recombination or rearrangement of the mitochondrial genomes in the fusion products was detected. These four plants and their progeny all showed the petaloid B. nigra type of male sterility.

  7. Influence of day length and temperature on the content of health-related compounds in broccoli (Brassica oleracea L. var. italica).

    Science.gov (United States)

    Steindal, Anne Linn Hykkerud; Mølmann, Jørgen; Bengtsson, Gunnar B; Johansen, Tor J

    2013-11-13

    Vegetables grown at different latitudes are exposed to various temperatures and day lengths, which can affect the content of health- and sensory-related compounds in broccoli florets. A 2 × 2 factorial experiment was conducted under controlled growth conditions, with contrasting temperatures (15/9 and 21/15 °C) and day lengths (12 and 24 h), to investigate the effect on glucosinolates, vitamin C, flavonols, and soluble sugars. Aliphatic glucosinolates, quercetin, and kaempferol were at their highest levels at high temperatures combined with a 12 h day. Levels of total glucosinolates, d-glucose, and d-fructose were elevated by high temperatures. Conversely, the content of vitamin C was highest with a 12 h day length combined with 15/9 °C. Our results indicate that temperature and day length influence the contents of health-related compounds in broccoli florets in a complex way, suggesting no general superiority of any of the contrasting growth conditions.

  8. Catechol-O-methyltransferase gene polymorphism and vulvar pain in women with vulvodynia.

    Science.gov (United States)

    Patanwala, Insiyyah Y; Lamvu, Georgine; Ledger, William J; Witzeman, Kathryn; Marvel, Richard; Rapkin, Andrea; Bongiovanni, Ann Marie; Feranec, Jessica; Witkin, Steven S

    2017-04-01

    The underlying causes of vulvar pain in women with vulvodynia remain poorly understood. Catechol-O-methyltransferase, an enzyme that metabolizes catecholamines, is a neuromodulator that is involved with perception and sensitivity to pain. The catechol-O-methyltransferase gene is polymorphic, and a single nucleotide polymorphism is associated with low activity and heightened pain sensitivity. The variant allele that encodes this polymorphism commonly is called the "L allele" because of its low enzyme activity as opposed to the normal H (high activity) allele. The methionine-containing catechol-O-methyltransferase protein coded by the L allele results in elevated catecholamine levels, reduced inactivation of the dopaminergic and adrenergic systems, and increased sensitivity to pain. This polymorphism not only may decrease the pain threshold in response to acute pain but also may facilitate the development of chronic pain. Therefore, the objective of our study was to assess whether a variation in the catechol-O-methyltransferase genotype is involved in increased pain sensitivity in women with vulvodynia. We conducted a prospective cohort study. Buccal swabs were collected from 167 white women with vulvodynia and 107 control subjects; the DNA was tested for a single nucleotide polymorphism at position 158 (rs4680) in the catechol-O-methyltransferase gene. Women with vulvodynia had a marginally increased, yet not significant, prevalence of the catechol-O-methyltransferase genotype that is associated with high activity of the coded protein: 32.9% in the women with vulvodynia, as opposed to 21.5% in the control subjects (odds ratio, 1.80; 95% confidence interval, 1.02-3.15). Subgrouping the cases based on pain frequency revealed that the elevated occurrence of this catechol-O-methyltransferase genotype was present in 40.6% of the subset of women who experienced pain only with sexual intercourse vs only 21.5% of control subjects (odds ratio, 2.50; 95% confidence interval

  9. Biotechnological Production of Dimethoxyflavonoids Using a Fusion Flavonoid O-Methyltransferase Possessing Both 3'- and 7-O-Methyltransferase Activities.

    Science.gov (United States)

    Lee, Danbi; Park, Hye Lin; Lee, Sang-Won; Bhoo, Seong Hee; Cho, Man-Ho

    2017-05-26

    Although they are less abundant in nature, methoxyflavonoids have distinct physicochemical and pharmacological properties compared to common nonmethylated flavonoids. Thus, enzymatic conversion and biotransformation using genetically engineered microorganisms of flavonoids have been attempted for the efficient production of methoxyflavonoids. Because of their regiospecificity, more than two flavonoid O-methyltransferases (FOMTs) and enzyme reactions are required to biosynthesize di(or poly)-methoxyflavonoids. For the one-step biotechnological production of bioactive di-O-methylflavonoids, we generated a multifunctional FOMT fusing a 3'-OMT (SlOMT3) and a 7-OMT (OsNOMT). The SlOMT3/OsNOMT fusion enzyme possessed both 3'- and 7-OMT activities to diverse flavonoid substrates, which were comparable to those of individual SlOMT3 and OsNOMT. The SlOMT3/OsNOMT enzyme also showed 3'- and 7-OMT activity for 7- or 3'-O-methylflavonoids, respectively, suggesting that the fusion enzyme can sequentially methylate flavonoids into di-O-methylflavonoids. The biotransformation of the flavonoids quercetin, luteolin, eriodictyol, and taxifolin using SlOMT3/OsNOMT-transformed Escherichia coli generated corresponding di-O-methylflavonoids, rhamnazin, velutin, 3',7-di-O-methyleriodictyol, and 3',7-di-O-methyltaxifolin, respectively. These results indicate that dimethoxyflavonoids may be efficiently produced from nonmethylated flavonoid precursors through a one-step biotransformation using the engineered E. coli harboring the SlOMT3/OsNOMT fusion gene.

  10. High frequency organogenesis in hypocotyl, cotyledon, leaf and petiole explants of broccoli (Brassica oleracea L. var. italica), an important vegetable crop.

    Science.gov (United States)

    Kumar, Pankaj; Srivastava, D K

    2015-04-01

    Broccoli (Brassica oleracea L. var. italica) is an important, nutritionally rich vegetable crop, but severely affected by environmental stresses, pests and diseases which cause massive yield and quality losses. Genetic manipulation is becoming an important method for broccoli improvement. In the present study, a reproducible and highly efficient protocol for obtaining organogenesis from hypocotyl, cotyledon, leaf and petiole explants of broccoli (Brassica oleracea L. var. italica cv. Solan green head) has been developed. Hypocotyl and cotyledon explants were used from 10 to 12 days old aseptically grown seedlings whereas leaf and petiole explants were excised from 18 to 20 days old green house grown seedlings and surface sterilized. These explants were cultured on shoot induction medium containing different concentration and combination of BAP and NAA. High efficiency shoot regeneration has been achieved in hypocotyl (83.33 %), cotyledon (90.11 %), leaf (62.96 %) and petiole (91.10 %) explants on MS medium supplemented with 3.5 mg/l BAP + 0.019 mg/l NAA 2.5 mg/l BAP + 0.5 mg/l NAA, 4.0 mg/l BAP + 0.5 mg/l NAA and 4.5 mg/l BAP + 0.019 mg/l NAA respectively. Petiole explants showed maximum shoot regeneration response as compared to other explants. MS medium supplemented with 0.10 mg/l NAA was found best for root regeneration (100 %) from in vitro developed shoots. The regenerated complete plantlets were transferred to the pots containing cocopeat and successfully acclimatized. This optimized regeneration protocol can be efficiently used for genetic transformation in broccoli. This is the first comparative report on multiple shoot induction using four different types of explants viz. hypocotyl, cotyledon, leaf and petiole.

  11. Protection against UV-light-induced skin carcinogenesis in SKH-1 high-risk mice by sulforaphane-containing broccoli sprout extracts.

    Science.gov (United States)

    Dinkova-Kostova, Albena T; Jenkins, Stephanie N; Fahey, Jed W; Ye, Lingxiang; Wehage, Scott L; Liby, Karen T; Stephenson, Katherine K; Wade, Kristina L; Talalay, Paul

    2006-08-28

    Aerobic life, UV solar radiation, genetic susceptibility, and immune status contribute collectively to the development of human skin cancers. In addition to direct DNA damage, UV radiation promotes the generation of reactive oxygen intermediates that can cause oxidative damage and inflammation, and ultimately lead to tumor formation. Treatment of murine and human keratinocytes with the isothiocyanate sulforaphane elevated phase 2 enzymes and glutathione and protected against oxidant toxicity. Topical application of sulforaphane-containing broccoli sprouts extracts induced the phase 2 response in mouse skin in vivo. Sulforaphane inhibited cytokine-dependent (gamma-interferon or lipopolysaccharide) induction of iNOS in RAW 264.7 macrophages. The UV-radiation-induced skin carcinogenesis in "initiated high-risk mice" was substantially inhibited by broccoli sprout extracts containing sulforaphane. After completion of the UV irradiation schedule (30 mJ/cm(2)/session twice a week for 20 weeks), groups of approximately 30 mice were treated topically on their backs (5 days a week for 11 weeks) with broccoli sprout extract containing either the equivalent to 0.3 micromol (low dose) or 1.0 micromol (high dose) sulforaphane, respectively. At this time point, the tumor incidence had reached 100% in the control mice. Tumor burden, incidence, and multiplicity were reduced by 50% in the animals that received the high dose of protector. Tumor incidence and multiplicity did not differ between the low dose-treated and the control groups, but the low dose treatment resulted in a substantial reduction of the overall tumor burden. Thus, topical application of sulforaphane-containing broccoli sprout extracts is a promising strategy for protecting against skin tumor formation after exposure to UV radiation.

  12. Health-promoting compounds of broccoli (Brassica oleracea L. var. italica) plants as affected by nitrogen fertilisation in projected future climatic change environments.

    Science.gov (United States)

    Zaghdoud, Chokri; Carvajal, Micaela; Moreno, Diego A; Ferchichi, Ali; Del Carmen Martínez-Ballesta, María

    2016-01-30

    The complex interactions between CO2 increase and salinity were investigated in relation to decreased N supply, in order to determine the nutritional quality of broccoli (Brassica oleracea L. var. italica) plants under these conditions. Three different decreased N fertilisation regimes (NO3(-)/NH4(+) ratios of 100:0, 50:50 and 0:100 respectively) were combined with ambient (380 ppm) and elevated (800 ppm) [CO2 ] under non-saline (0 mmol L(-1) NaCl) and saline (80 mmol L(-1) NaCl) conditions. Nutrients (minerals, soluble protein and total amino acids) and natural antioxidants (glucosinolates, phenolic acids, flavonoids and vitamin C) were determined. In NH4(+) -fed broccoli plants, a marked growth reduction was shown and a redistribution of amino acids to cope with NH4(+) toxicity resulted in higher levels of indolic glucosinolate and total phenolic compounds. However, the positive effect of the higher [CO2] - ameliorating adverse effects of salinity--was only observed when N was supplied as NO3(-). Under reduced N fertilisation, the total glucosinolates were increased by a decreased NO3(-)/NH4 (+) ratio and elevated [CO2] but were unaffected by salinity. Under future climatic challenges, such as increased salinity and elevated [CO2], a clear genotypic dependence of S metabolism was observed in broccoli plants. In addition, an influence of the form in which N was supplied on plant nutritional quality was observed; a combined NO3(-)/NH4(+) (50:50) supply allowed broccoli plants not only to deal with NH4(+) toxicity but also to modify their glucosinolate content and profile. Thus, for different modes of N fertilisation, the interaction with climatic factors must be considered in the search for an optimal balance between yield and nutritional quality. © 2015 Society of Chemical Industry.

  13. Polymer coating, germination and vigor of broccoli seeds Recobrimento polimérico, germinação e vigor de sementes de brócolos

    OpenAIRE

    Celina de Almeida; Sandra Cristina dos Santos Rocha; Luiz Fernandes Razera

    2005-01-01

    Brassica oleracea var italica occupies a special place in the internal and external market of vegetables seeds. Vegetables producers demand seeds with high degree of purity, germination and vigor, since seeds' quality is the basis for the success of the production. In this work, broccoli seeds were coated in a spouted bed, by an aqueous suspension of hidroxy-ethyl-cellulose. Effects of the operating variables: spouting air temperature, atomizing air pressure and coating suspension flow rate o...

  14. Compost enhances parasitization of Brevicoryne brassicae (L.) by Diaeretiella rapae (M’Intosh) in broccoli under different levels of crop diversification and plant competition

    OpenAIRE

    Ponti, Luigi; Altieri, Miguel A.; Gutierrez, Andrew P.

    2008-01-01

    The effects of intercropping via competition on crop biomass, pest [cabbage aphid Brevicoryne brassicae (L.)] abundance and natural enemy [the parasitoid Diaeretiella rapae (M’Intosh)] efficacy were studied in the Brassica oleracea L. var. italica system. From May to December 2004, insect populations and yield parameters were monitored in summer and fall in broccoli monoculture and polyculture systems with or without competition from Brassica spp. (mustard), or Fagopyrum esculentum Moench (bu...

  15. Modulation of the metabolism of airborne pollutants by glucoraphanin-rich and sulforaphane-rich broccoli sprout beverages in Qidong, China.

    Science.gov (United States)

    Kensler, Thomas W; Ng, Derek; Carmella, Steven G; Chen, Menglan; Jacobson, Lisa P; Muñoz, Alvaro; Egner, Patricia A; Chen, Jian Guo; Qian, Geng Sun; Chen, Tao Yang; Fahey, Jed W; Talalay, Paul; Groopman, John D; Yuan, Jian-Min; Hecht, Stephen S

    2012-01-01

    Epidemiological evidence has suggested that consumption of a diet rich in cruciferous vegetables reduces the risk of several types of cancers and chronic degenerative diseases. In particular, broccoli sprouts are a convenient and rich source of the glucosinolate, glucoraphanin, which can release the chemopreventive agent, sulforaphane, an inducer of glutathione S-transferases. Two broccoli sprout-derived beverages, one sulforaphane-rich (SFR) and the other glucoraphanin-rich (GRR), were evaluated for pharmacodynamic action in a crossover clinical trial design. Study participants were recruited from the farming community of He Zuo Township, Qidong, China, previously documented to have a high incidence of hepatocellular carcinoma with concomitant exposures to aflatoxin and more recently characterized with exposures to substantive levels of airborne pollutants. Fifty healthy participants were randomized into two treatment arms. The study protocol was as follows: a 5 days run-in period, a 7 days administration of beverage, a 5 days washout period and a 7 days administration of the opposite beverage. Urinary excretion of the mercapturic acids of acrolein, crotonaldehyde, ethylene oxide and benzene were measured both pre- and postinterventions using liquid chromatography tandem mass spectrometry. Statistically significant increases of 20-50% in the levels of excretion of glutathione-derived conjugates of acrolein, crotonaldehyde and benzene were seen in individuals receiving SFR, GRR or both compared with their preintervention baseline values. No significant differences were seen between the effects of SFR versus GRR. Intervention with broccoli sprouts may enhance detoxication of airborne pollutants and attenuate their associated health risks.

  16. Enhancement of phenolics, flavonoids and glucosinolates of Broccoli (Brassica olaracea, var. Italica as antioxidants in response to organic and bio-organic fertilizers

    Directory of Open Access Journals (Sweden)

    Abd El-Moniem M. Naguib

    2012-06-01

    Full Text Available A field experiment was carried out to study the effect of organic and bioorganic fertilizers on growth parameters, yield and the quality of two broccoli cultivars (Calabrese and Southern star. Bio-organic fertilizers gave better results for all vegetative growth parameters for Southern star cultivar compared to Calabrese cultivar. Total phenolic content (TPC, total flavonoid content (TFC and total glucosinolates content (TGsC were almost higher in Calabrese cultivar as a result of organic fertilizer treatment. The antioxidant activities of both cultivars were evaluated and Calabrese cultivar showed the higher1, 1-diphenyl-2-picrylhydrazyl DPPH. scavenging activity expressed as IC50 (its concentrations for 50% inhibition (16.56 μg/ml compared to Southern star (19.42 μg/ml. In addition, Calabrese cultivar showed the higher chelating power (75.36 μg/ml than Southern star (72.43 μg/ml at (30 μg/ml when the organic fertilizer was applied. The results indicated that there is a good margin for enhancing antioxidant compounds of broccoli for economic production using organic fertilization. This study indicated the potential application of broccoli as a potent natural source of antioxidants as nutraceuticals.

  17. Effects of the 3D-clinorotation on endogenous substances of broccoli sprout (Brassica oleracea var. italica) and its food safety

    Science.gov (United States)

    Hiraishi, K.; Tomita-Yokotani, K.; Wakabayashi, K.; Hashimoto, H.; Miyagawa, T.; Yamashita, M.

    Habitation in outer space is one of our challenges in this century We are studying on space agriculture to provide foods for space living people However careful assessment should be made on the effects of exotic environment on the endogenous production of biologically active substances and food safety of plants cultivated in space Broccoli sprout Brassica oleracea var italica is known to produce sulforaphane 4-methylsulfinybutyl isothiocyanate which is effective to function as an antioxidant and enhance immunity Because of such substance it is recognized to be good food materials Broccoli sprouts were then cultivated for 3 days under the 3D-clinorotation The amount of sulforaphane produced by this treatment showed no significant difference compared to the ground control Secondly we examined population of microorganisms adhered on the surface of sprout cultivated under the 3D-clinorotation Number of the microorganisms colony formed was statistically higher than the control Mold species was identified to penicillium sp based on the microscopic observation Poor construction of plant cell wall elements cellulose lignin etc is well known effects of microgravity Defense function of the broccoli plant cells might be weakened against microorganism We also speculate other possible causes for the high rate of contamination such as photosynthetic activity of the plant or microclimate air flow heat transport and humidity around the seedling affected by pseudo-microgravity or the 3D-clinorotation Those factors may relate to the difference in proliferation

  18. Degradation kinetics of peroxidase enzyme, phenolic content, and physical and sensorial characteristics in broccoli (Brassica oleracea L. ssp. Italica) during blanching.

    Science.gov (United States)

    Gonçalves, Elsa M; Pinheiro, Joaquina; Alegria, Carla; Abreu, Marta; Brandão, Teresa R S; Silva, Cristina L M

    2009-06-24

    The effects of water blanching treatment on peroxidase inactivation, total phenolic content, color parameters [-a*/b* and hue (h degrees*)], texture (maximum shear force), and sensory attributes (color and texture, evaluated by a trained panel) of broccoli (Brassica oleracea L. ssp. Italica) were studied at five temperatures (70, 75, 80, 85, and 90 degrees C). Experimental results showed that all studied broccoli quality parameters suffered significative changes due to blanching treatments. The vegetal total phenolic content showed a marked decline. Degradation on objective color and texture measurements and alterations in sensorial attributes were detected. Correlations between sensory and instrumental measurements have been found. Under the conditions 70 degrees C and 6.5 min or 90 degrees C and 0.4 min, 90% of the initial peroxidase activity was reduced. At these conditions, no significant alterations were detected by panelists, and a small amount of phenolic content was lost (ca. 16 and 10%, respectively). The peroxidase inactivation and phenolic content degradation were found to follow first-order reaction models. The zero-order reaction model showed a good fit to the broccoli color (-a*/b* and h degrees*), texture, and sensory parameters changes. The temperature effect was well-described by the Arrhenius law.

  19. Assessing Competence of Broccoli Consumption on Inflammatory and Antioxidant Pathways in Restraint-Induced Models: Estimation in Rat Hippocampus and Prefrontal Cortex

    Directory of Open Access Journals (Sweden)

    Leila Khalaj

    2013-01-01

    Full Text Available A growing body of evidence advocated the protective and therapeutic potential of natural compounds and phytochemicals used in diets against pathological conditions. Herein, the outcome of dietary whole broccoli consumption prior to restraint stress has been investigated in the hippocampus and prefrontal cortex of male rats, two important regions involved in the processing of responses to stressful events. Interestingly, a region-specific effect was detected regarding some of antioxidant defense system factors: nuclear factor erythroid-derived 2-related factor 2 (Nrf-2 antioxidant pathway, mitochondrial prosurvival proteins involved in mitochondrial biogenesis, and apoptotic cell death proteins. Dietary broccoli supplementation modulated the restraint-induced changes towards a consistent overall protection in the hippocampus. In the prefrontal cortex, however, despite activation of most of the protective factors, presumably as an attempt to save the system against the stress insult, some detrimental outcomes such as induced malate dehydrogenase (MDA level and cleaved form of caspase-3 were detectable. Such diversity may be attributed in one hand to the different basic levels and/or availability of defensive mechanisms within the two studied cerebral regions, and on the other hand to the probable dose-dependent and hormetic effects of whole broccoli. More experiments are essential to demonstrate these assumptions.

  20. Case Study for the Evaluation of Current Treatment Recommendations of Guanidinoacetate Methyltransferase Deficiency: Ineffectiveness of Sodium Benzoate

    NARCIS (Netherlands)

    Mercimek-Mahmutoglu, S.; Salomons, G.S.; Chan, A.

    2014-01-01

    BACKGROUND: Guanidinoacetate methyltransferase deficiency is an autosomal recessively inherited disorder of creatine biosynthesis. We report a new patient with guanidinoacetate methyltransferase deficiency and her >3-year treatment outcome. Patient This is a 6-year-old girl who was diagnosed with

  1. Catechol-O-methyltransferase association with hemoglobin A1c.

    Science.gov (United States)

    Hall, Kathryn T; Jablonski, Kathleen A; Chen, Ling; Harden, Maegan; Tolkin, Benjamin R; Kaptchuk, Ted J; Bray, George A; Ridker, Paul M; Florez, Jose C; Mukamal, Kenneth J; Chasman, Daniel I

    2016-07-01

    Catecholamines have metabolic effects on blood pressure, insulin sensitivity and blood glucose. Genetic variation in catechol-O-methyltransferase (COMT), an enzyme that degrades catecholamines, is associated with cardiometabolic risk factors and incident cardiovascular disease (CVD). Here we examined COMT effects on glycemic function and type 2 diabetes. We tested whether COMT polymorphisms were associated with baseline HbA1c in the Women's Genome Health Study (WGHS), and Meta-Analyses of Glucose and Insulin-related traits Consortium (MAGIC), and with susceptibility to type 2 diabetes in WGHS, DIAbetes Genetics Replication And Meta-analysis consortium (DIAGRAM), and the Diabetes Prevention Program (DPP). Given evidence that COMT modifies some drug responses, we examined association with type 2 diabetes and randomized metformin and aspirin treatment. COMT rs4680 high-activity G-allele was associated with lower HbA1c in WGHS (β=-0.032% [0.012], p=0.008) and borderline significant in MAGIC (β=-0.006% [0.003], p=0.07). Combined COMT per val allele effects on type 2 diabetes were significant (OR=0.98 [0.96-0.998], p=0.03) in fixed-effects analyses across WGHS, DIAGRAM, and DPP. Similar results were obtained for 2 other COMT SNPs rs4818 and rs4633. In the DPP, the rs4680 val allele was borderline associated with lower diabetes incidence among participants randomized to metformin (HR=0.81 [0.65-1.00], p=0.05). COMT rs4680 high-activity G-allele was associated with lower HbA1c and modest protection from type 2 diabetes. The directionality of COMT associations was concordant with those previously observed for cardiometabolic risk factors and CVD. Copyright © 2016 Elsevier Inc. All rights reserved.

  2. Catechol-O-methyltransferase, dopamine, and sleep-wake regulation.

    Science.gov (United States)

    Dauvilliers, Yves; Tafti, Mehdi; Landolt, Hans Peter

    2015-08-01

    Sleep and sleep disorders are complex and highly variable phenotypes regulated by many genes and environment. The catechol-O-methyltransferase (COMT) gene is an interesting candidate, being one of the major mammalian enzymes involved in the catabolism of catecholamines. The activity of COMT enzyme is genetically polymorphic due to a guanine-to-adenine transition at codon 158, resulting in a valine (Val) to methionine (Met) substitution. Individuals homozygous for the Val allele show higher COMT activity, and lower dopaminergic signaling in prefrontal cortex (PFC) than subjects homozygous for the Met allele. Since COMT has a crucial role in metabolising dopamine, it was suggested that the common functional polymorphism in the COMT gene impacts on cognitive function related to PFC, sleep-wake regulation, and potentially on sleep pathologies. The COMT Val158Met polymorphism may predict inter-individual differences in brain electroencephalography (EEG) alpha oscillations and recovery processes resulting from partial sleep loss in healthy individuals. The Val158Met polymorphism also exerts a sexual dimorphism and has a strong effect on objective daytime sleepiness in patients with narcolepsy-cataplexy. Since the COMT enzyme inactivates catecholamines, it was hypothesized that the response to stimulant drugs differs between COMT genotypes. Modafinil maintained executive functioning performance and vigilant attention throughout sleep deprivation in subjects with Val/Val genotype, but less in those with Met/Met genotype. Also, homozygous Met/Met patients with narcolepsy responded to lower doses of modafinil compared to Val/Val carriers. We review here the critical role of the common functional COMT gene polymorphism, COMT enzyme activity, and the prefrontal dopamine levels in the regulation of sleep and wakefulness in normal subjects, in narcolepsy and other sleep-related disorders, and its impact on the response to psychostimulants. Copyright © 2014 Elsevier Ltd. All

  3. Inhibitors of catechol-O-methyltransferase sensitize mice to pain

    Science.gov (United States)

    Kambur, O; Talka, R; Ansah, OB; Kontinen, VK; Pertovaara, A; Kalso, E; Männistö, PT

    2010-01-01

    BACKGROUND AND PURPOSE Catechol-O-methyltransferase (COMT) inhibitors are used in Parkinson's disease in which pain is an important symptom. COMT polymorphisms modulate pain and opioid analgesia in humans. In rats, COMT inhibitors have been shown to be pro-nociceptive in acute pain models, but also to attenuate allodynia and hyperalgesia in a model of diabetic neuropathy. Here, we have assessed the effects of acute and repeated administrations of COMT inhibitors on mechanical, thermal and carrageenan-induced nociception in male mice. EXPERIMENTAL APPROACH We used single and repeated administration of a peripherally restricted, short-acting (nitecapone) and also a centrally acting (3,5-dinitrocatechol, OR-486) COMT inhibitor. We also tested CGP 28014, an indirect inhibitor of COMT enzyme. Effects of OR-486 on thermal nociception were also studied in COMT deficient mice. Effects on spinal pathways were assessed in rats given intrathecal nitecapone. KEY RESULTS After single administration, both nitecapone and OR-486 reduced mechanical nociceptive thresholds and thermal nociceptive latencies (hot plate test) at 2 and 3 h, regardless of their brain penetration. These effects were still present after chronic treatment with COMT inhibitors for 5 days. Intraplantar injection of carrageenan reduced nociceptive latencies and both COMT inhibitors potentiated this reduction without modifying inflammation. CGP 28014 shortened paw flick latencies. OR-486 did not modify hot plate times in Comt gene deficient mice. Intrathecal nitecapone modified neither thermal nor mechanical nociception. CONCLUSIONS AND IMPLICATIONS Pro-nociceptive effects of COMT inhibitors were confirmed. The pro-nociceptive effects were primarily mediated via mechanisms acting outside the brain and spinal cord. COMT protein was required for these actions. PMID:20726980

  4. Catechol-o-methyltransferase and 3,4-({+/-})-methylenedioxymethamphetamine toxicity.

    Science.gov (United States)

    Herndon, Joseph M; Cholanians, Aram B; Lizarraga, Lucina E; Lau, Serrine S; Monks, Terrence J

    2014-05-01

    Metabolism of 3,4-(±)-methylenedioxymethamphetamine (MDMA) is necessary to elicit its neurotoxic effects. Perturbations in phase I and phase II hepatic enzymes can alter the neurotoxic profile of systemically administered MDMA. In particular, catechol-O-methyltransferase (COMT) plays a critical role in determining the fraction of MDMA that is converted to potentially neurotoxic metabolites. Thus, cytochrome P450 mediated demethylenation of MDMA, or its N-demethylated metabolite, 3,4-(±)-methylenedioxyamphetamine, give rise to the catechols, N-methyl-α-methyldopamine and α-methyldopamine, respectively. Methylation of these catechols by COMT limits their oxidation and conjugation to glutathione, a process that ultimately gives rise to neurotoxic metabolites. We therefore determined the effects of modulating COMT, a critical enzyme involved in determining the fraction of MDMA that is converted to potentially neurotoxic metabolites, on MDMA-induced toxicity. Pharmacological inhibition of COMT in the rat potentiated MDMA-induced serotonin deficits and exacerbated the acute MDMA-induced hyperthermic response. Using a genetic mouse model of COMT deficiency, in which mice lack a functional COMT gene, such mice displayed greater reductions in dopamine concentrations relative to their wild-type (WT) counterparts. Neither WT nor COMT deficient mice were susceptible to MDMA-induced decreases in serotonin concentrations. Interestingly, mice devoid of COMT were far more susceptible to the acute hyperthermic effects of MDMA, exhibiting greater increases in body temperature that ultimately resulted in death. Our findings support the view that COMT plays a pivotal role in determining the toxic response to MDMA.

  5. Structural Basis of Substrate Recognition in Thiopurine S-Methyltransferase

    Energy Technology Data Exchange (ETDEWEB)

    Peng, Yi; Feng, Qiping; Wilk, Dennis; Adjei, Araba A.; Salavaggione, Oreste E.; Weinshilboum, Richard M.; Yee, Vivien C. (Case Western); (MCCM)

    2008-09-23

    Thiopurine S-methyltransferase (TPMT) modulates the cytotoxic effects of thiopurine prodrugs such as 6-mercaptopurine by methylating them in a reaction using S-adenosyl-l-methionine as the donor. Patients with TPMT variant allozymes exhibit diminished levels of protein and/or enzyme activity and are at risk for thiopurine drug-induced toxicity. We have determined two crystal structures of murine TPMT, as a binary complex with the product S-adenosyl-l-homocysteine and as a ternary complex with S-adenosyl-l-homocysteine and the substrate 6-mercaptopurine, to 1.8 and 2.0 {angstrom} resolution, respectively. Comparison of the structures reveals that an active site loop becomes ordered upon 6-mercaptopurine binding. The positions of the two ligands are consistent with the expected S{sub N}2 reaction mechanism. Arg147 and Arg221, the only polar amino acids near 6-mercaptopurine, are highlighted as possible participants in substrate deprotonation. To probe whether these residues are important for catalysis, point mutants were prepared in the human enzyme. Substitution of Arg152 (Arg147 in murine TPMT) with glutamic acid decreases V{sub max} and increases K{sub m} for 6-mercaptopurine but not K{sub m} for S-adenosyl-l-methionine. Substitution at this position with alanine or histidine and similar substitutions of Arg226 (Arg221 in murine TPMT) result in no effect on enzyme activity. The double mutant Arg152Ala/Arg226Ala exhibits a decreased V{sub max} and increased K{sub m} for 6-mercaptopurine. These observations suggest that either Arg152 or Arg226 may participate in some fashion in the TPMT reaction, with one residue compensating when the other is altered, and that Arg152 may interact with substrate more directly than Arg226, consistent with observations in the murine TPMT crystal structure.

  6. Histone methyltransferase Setdb1 is indispensable for Meckel's cartilage development.

    Science.gov (United States)

    Yahiro, Kohei; Higashihori, Norihisa; Moriyama, Keiji

    2017-01-22

    The histone methyltransferase Setdb1 represses gene expression by catalyzing lysine 9 of histone H3 trimethylation. Given that the conventional knockout of Setdb1 is embryo-lethal at the implantation stage, its role in craniofacial development is poorly understood. Here, we investigated the role of Setdb1, using conditional knockout mice-in which Setdb1 was deleted in the Meckel's cartilage (Setdb1 CKO)-and the mouse chondrogenic cell line ATDC5-in which Setdb1 was inhibited by siRNA. Deletion of Setdb1 in Meckel's cartilage, the supportive tissue in the embryonic mandible, led to its enlargement, instead of the degeneration that normally occurs. Chondrocytes from the Meckel's cartilage of Setdb1 CKO mice showed increased size. Furthermore, at embryonic days 16.5 and 18.5, part of the perichondrium was disrupted and mineralization was observed in the Meckel's cartilage. Proliferation analysis showed that inhibition of Setdb1 caused increased proliferation in chondrocytes in the Meckel's cartilage as well as in ATDC5 cells. Quantitative RT-PCR showed decreased expression of chondrogenic genes, such as Sox9, Mmp13, Collagen II, and Aggrecan, as a result of Setdb1 inhibition in ATDC5 cells. Along with these phenomenons, SMAD-dependent BMP signaling was significantly increased by the loss of Setdb1 in both the Meckel's cartilage of Setdb1 CKO mice and ATDC5 cells. Therefore, the abnormal development of Meckel's cartilage in Setdb1 CKO mice is partly due to the enhanced SMAD-dependent BMP signaling. Overall, to our knowledge, the present study is the first to show that epigenetic regulation by Setdb1 is indispensable for the embryonic development of Meckel's cartilage. Copyright © 2016. Published by Elsevier Inc.

  7. Effects of nickel on DNA methyltransferase activity and genomic DNA methylation levels.

    Science.gov (United States)

    Lee, Y W; Broday, L; Costa, M

    1998-07-31

    Methylation of DNA plays an important role in organizing the genome into transcriptionally active and inactive zones. Nickel compounds cause chromatin condensation and DNA methylation in the transgenic gpt+ Chinese hamster cell line (G12). Here we show that nickel is an inhibitor of cytosine 5-methyltransferase activity in vivo and in vitro. In living cells, this inhibition is transient and following a recovery period after nickel treatment, Mtase activity slightly rebounds. Genomic DNA methylation levels are also somewhat decreased following nickel treatment, but with time, there is an elevation of total DNA methylation above basal levels and before any rebound of methyltransferase activity. These results suggest that nickel exposure can elevate total genomic DNA methylation levels even when DNA methyltransferase activity is depressed. These findings may explain the hypermethylation of senescence and tumor suppressor genes found during nickel carcinogenesis and support the model of a direct effect of Ni2+ on chromatin leading to de novo DNA methylation.

  8. Purification and characterization of recombinant baculovirus-expressed mouse DNA methyltransferase.

    Science.gov (United States)

    Glickman, J F; Flynn, J; Reich, N O

    1997-01-13

    DNA methylation is essential for normal embryonic development in mice. An understanding of how DNA methylation is controlled is largely dependent upon the isolation and characterization of the cellular components of the DNA methylation system. The enzyme which methylates DNA in eukaryotic cells is a C-5 cytosine DNA methyltransferase. Historically, the characterization of this enzyme has been limited by its availability and purity. Here, we present a single-step purification of 4 mg of baculovirus-expressed mouse DNA methyltransferase containing a nickel-affinity leader peptide. The recombinant DNA methyltransferase copurified with inhibitory RNA which was removed by treatment with ribonuclease A. Like its non-recombinant counterpart, the recombinant enzyme is activated by hemi-methylation. A direct steady-state kinetic comparison between the recombinant baculovirus-expressed enzyme with its MEL cell-derived counterpart is presented.

  9. Structural insights into substrate selectivity of ribosomal RNA methyltransferase RlmCD.

    Directory of Open Access Journals (Sweden)

    Yiyang Jiang

    Full Text Available RlmCD has recently been identified as the S-adenosyl methionine (SAM-dependent methyltransferase responsible for the formation of m5U at U747 and U1939 of 23S ribosomal RNA in Streptococcus pneumoniae. In this research, we determine the high-resolution crystal structures of apo-form RlmCD and its complex with SAH. Using an in-vitro methyltransferase assay, we reveal the crucial residues for its catalytic functions. Furthermore, structural comparison between RlmCD and its structural homologue RumA, which only catalyzes the m5U1939 in Escherichia coli, implicates that a unique long linker in the central domain of RlmCD is the key factor in determining its substrate selectivity. Its significance in the enzyme activity of RlmCD is further confirmed by in-vitro methyltransferase assay.

  10. Rapid and sensitive single-step radiochemical assay for catechol-O-methyltransferase

    Energy Technology Data Exchange (ETDEWEB)

    Zuercher, G.; Da Prada, M. (Hoffmann-La Roche (F.) and Co., Basel (Switzerland))

    1982-01-01

    A simple, rapid and reliable radiometric assay for the determination of catechol-O-methyltransferase activity is described. The method is based on the conversion of catechol to (/sup 3/H)guaiacol by catechol-O-methyltransferase in the presence of Mg/sup 2 +/, adenosine deaminase and S-adenosyl L-(methyl-/sup 3/H)methionine. Incubation and direct extraction of (/sup 3/H)guaiacol into organic scintillation fluid, as well as counting, are performed in the same standard scintillation vial. The assay is easy to perform and more sensitive than previous analogous procedures. The method has been applied to the assay of catechol-O-methyltransferase activity in discrete brain areas and also peripheral organs of rat and in human erythrocytes.

  11. A novel Mg(2+)-dependent O-methyltransferase in the phenylpropanoid metabolism of Mesembryanthemum crystallinum.

    Science.gov (United States)

    Ibdah, Mwafaq; Zhang, Xing-Hai; Schmidt, Jürgen; Vogt, Thomas

    2003-11-07

    Upon irradiation with elevated light intensities, the ice plant (Mesembryanthemum crystallinum) accumulates a complex pattern of methylated and glycosylated flavonol conjugates in the upper epidermal layer. Identification of a flavonol methylating activity, partial purification of the enzyme, and sequencing of the corresponding peptide fragments revealed a novel S-adenosyl-l-methionine-dependent O-methyltransferase that was specific for flavonoids and caffeoyl-CoA. Cloning and functional expression of the corresponding cDNA verified that the new methyltransferase is a multifunctional 26.6-kDa Mg(2+)-dependent enzyme, which shows a significant sequence similarity to the cluster of caffeoyl coenzyme A-methylating enzymes. Functional analysis of highly homologous members from chickweed (Stellaria longipes), Arabidopsis thaliana, and tobacco (Nicotiana tabacum) demonstrated that the enzymes from the ice plant, chickweed, and A. thaliana possess a broader substrate specificity toward o-hydroquinone-like structures than previously anticipated for Mg(2+)-dependent O-methyltransferases, and are distinctly different from the tobacco enzyme. Besides caffeoyl-CoA and flavonols, a high specificity was also observed for caffeoylglucose, a compound never before reported to be methylated by any plant O-methyltransferase. Based on phylogenetic analysis of the amino acid sequence and differences in acceptor specificities among both animal and plant O-methyltransferases, we propose that the enzymes from the Centrospermae, along with the predicted gene product from A. thaliana, form a novel subclass within the caffeoyl coenzyme A-dependent O-methyltransferases, with potential divergent functions not restricted to lignin monomer biosynthesis.

  12. Fast determination of intact glucosinolates in broccoli leaf by pressurized liquid extraction and ultra high performance liquid chromatography coupled to quadrupole time-of-flight mass spectrometry.

    Science.gov (United States)

    Ares, Ana M; Bernal, José; Nozal, María J; Turner, Charlotta; Plaza, Merichel

    2015-10-01

    In this study, we investigate for the first time the efficiency of an environmentally sustainable extraction technique (pressurized liquid extraction, PLE) in conjunction with a fast separation technique (ultra-high performance liquid chromatography, UHPLC) coupled to a selective mass spectrometry (MS) detector (quadrupole time-of-flight, qTOF) to extract, separate and quantify fifteen intact-glucosinolates (GLSs) in broccoli leaves. Firstly, we have developed and optimized by means of an experimental design an efficient extraction procedure based on PLE (using ethanol/water as a solvent), giving complete extraction within 15min; meanwhile, the average analyte recoveries were between 85% and 96% in all cases. Chromatography was performed on a UHPLC BEH Shield RP18 1.7μm 110Å (2.1×100mm) analytical column with a mobile phase composed by formic acid in water (0.5%, v/v) and formic acid in acetonitrile (0.5%, v/v) in gradient elution mode at 0.3mL/min, resulted in baseline-separated peaks and a run time of 13min. The method was fully validated in terms of selectivity, limits of detection (LOD) and quantification (LOQ), linearity, precision, and trueness; meanwhile a study of the matrix effect was also performed. A good selectivity, low LODs and LOQs, ranging from 2 to 26μg/g, wide linear ranges from LOQ to 2500μg/g, and satisfactory precision and trueness with relative standard deviation and relative error values lower than or equal to 9%, were obtained for the studied GLSs. Finally, the proposed method was successfully applied to the analysis of intact-GLSs in fifteen broccoli leaf samples from three different cultivars (Parthenon, Nubia, and Naxos). Nine intact-GLSs were detected in all the varieties, although in different concentrations, which ranged between 14 and 1136μg/g, depending on the broccoli cultivar. In addition, the highest total content of GLSs was found in broccoli leaf samples from Parthenon cultivar, being the Naxos cultivar the poorest in GLS

  13. Effectiveness of edible coatings combined with mild heat shocks on microbial spoilage and sensory quality of fresh cut broccoli (Brassica oleracea L.).

    Science.gov (United States)

    Moreira, María del R; Ponce, Alejandra; Ansorena, R; Roura, Sara I

    2011-08-01

    The use of edible coatings and mild heat shocks is proposed as postharvest treatments to prevent microbial deterioration of refrigerated broccoli. Minimally processed broccoli was coated with either chitosan or carboxymethyl-cellulose (CMC) combined or not with a previous application of a mild heat shock. The evolution of microbial populations (mesophilic, psycrotrophic, Enterobacteriaceae, molds and yeast, and lactic acid bacteria) was studied during 20 d of storage and fitted to Gompertz and logistic models. Results revealed that, at the end of the storage, chitosan coating significantly reduced all microbiological population counts, except lactic acid bacteria; while higher reduction was observed with chitosan coating combined with a heat shock treatment. A significant delay at the beginning of the exponential phase was observed for all the bacterial populations analyzed. On the other hand, CMC coating, with and without a previous thermal treatment, did not exert any antibacterial effect. Excellent agreement was found between experimental microbial counts and predicted values obtained from Gompertz and logistic models. Kinetic modeling was found to be valuable for prediction of microbiological shelf life of broccoli during storage. Results showed that the application of chitosan coating effectively maintained microbiological quality and extended shelf life of minimally processed broccoli. According to these results, the use of the edible chitosan coating alone or in combination with a heat mild shock appear to be a viable alternative for controlling microbiological growth and sensory attributes in minimally processed broccoli. The continuous consumer interest in high quality and food safety, combined with environmental concern has induced to the development and study of edible coatings that avoid the use of synthetic materials. The edible coatings, formed from generally recognized as safe materials, have the potential to reduce weight loss, respiration rate, and

  14. The RNA-methyltransferase Misu (NSun2 poises epidermal stem cells to differentiate.

    Directory of Open Access Journals (Sweden)

    Sandra Blanco

    2011-12-01

    Full Text Available Homeostasis of most adult tissues is maintained by balancing stem cell self-renewal and differentiation, but whether post-transcriptional mechanisms can regulate this process is unknown. Here, we identify that an RNA methyltransferase (Misu/Nsun2 is required to balance stem cell self-renewal and differentiation in skin. In the epidermis, this methyltransferase is found in a defined sub-population of hair follicle stem cells poised to undergo lineage commitment, and its depletion results in enhanced quiescence and aberrant stem cell differentiation. Our results reveal that post-transcriptional RNA methylation can play a previously unappreciated role in controlling stem cell fate.

  15. Cloning, expression, purification and crystallization of Schizosaccharomyces pombe Set7, a putative histone methyltransferase.

    Science.gov (United States)

    Mevius, Damiaan E H F; Shen, Yunpeng; Morishita, Masayo; di Luccio, Eric

    2016-04-01

    Dysfunction of histone-modifying enzymes affects chromatin regulation and is involved in carcinogenesis, tumour progression and other diseases. Histone methyltransferases are a family of key histone-modifying enzymes, but their structures, functions and mechanisms are incompletely understood, thus constraining drug-design efforts. Here, preliminary steps towards structure-function studies of Schizosaccharomyces pombe Set7, a putative histone methyltransferase and the first yeast full-length SET-domain-containing protein to be studied using X-ray crystallography, are reported. The methods from cloning to X-ray diffraction and phasing are discussed and the results will aid in prospective studies of histone-modifying enzymes.

  16. Radiometric assay for phenylethanolamine N-methyltransferase and catechol O-methyltransferase in a single tissue sample: application to rat hypothalamic nuclei, pineal gland, and heart

    Energy Technology Data Exchange (ETDEWEB)

    Culman, J.; Torda, T.; Weise, V.K.

    1987-08-01

    A simple and highly sensitive method for simultaneous assay of phenylethanolamine N-methyltransferase (PNMT) and catechol O-methyltransferase (COMT) is described. These enzymes are determined in a single tissue homogenate using S-(methyl-/sup 3/H) adenosyl-L-methionine as methyl donor and sequentially incubating with the substrates phenylethanolamine and epinephrine. The radioactive products of the enzymatic reactions, N-methylphenylethanolamine and metanephrine, are extracted and then separated by thin-layer chromatography. The identity of the reaction products has been established chromatographically and the conditions for both enzymatic reactions in the assay procedure have been defined. Measurement of PNMT activity in the rat pineal gland or in minute fragments of other tissues (e.g., brain nuclei) has not been possible using previously described methods. Activities of PNMT and COMT in the rat pineal gland, various hypothalamic nuclei, and the auricular and ventricular myocardia are herein reported.

  17. Genotoxicity studies of organically grown broccoli (Brassica oleracea var. italica) and its interactions with urethane, methyl methanesulfonate and 4-nitroquinoline-1-oxide genotoxicity in the wing spot test of Drosophila melanogaster.

    Science.gov (United States)

    Heres-Pulido, María Eugenia; Dueñas-García, Irma; Castañeda-Partida, Laura; Santos-Cruz, Luis Felipe; Vega-Contreras, Viridiana; Rebollar-Vega, Rosa; Gómez-Luna, Juan Carlos; Durán-Díaz, Angel

    2010-01-01

    Broccoli (Brassica oleracea var. italica) has been defined as a cancer preventive food. Nevertheless, broccoli contains potentially genotoxic compounds as well. We performed the wing spot test of Drosophila melanogaster in treatments with organically grown broccoli (OGB) and co-treatments with the promutagen urethane (URE), the direct alkylating agent methyl methanesulfonate (MMS) and the carcinogen 4-nitroquinoline-1-oxide (4-NQO) in the standard (ST) and high bioactivation (HB) crosses with inducible and high levels of cytochrome P450s (CYPs), respectively. Larvae of both crosses were chronically fed with OGB or fresh market broccoli (FMB) as a non-organically grown control, added with solvents or mutagens solutions. In both crosses, the OGB added with Tween-ethanol yielded the expected reduction in the genotoxicity spontaneous rate. OGB co-treatments did not affect the URE effect, MMS showed synergy and 4-NQO damage was modulated in both crosses. In contrast, FMB controls produced damage increase; co-treatments modulated URE genotoxicity, diminished MMS damage, and did not change the 4-NQO damage. The high dietary consumption of both types of broccoli and its protective effects in D. melanogaster are discussed. Copyright 2009 Elsevier Ltd. All rights reserved.

  18. Expression of DNA Methyltransferases in Breast Cancer Patients and to Analyze the Effect of Natural Compounds on DNA Methyltransferases and Associated Proteins

    OpenAIRE

    Mirza, Sameer; Sharma, Gayatri; Parshad, Rajinder; Gupta, Sidhartha Datta; Pandya, Pranav; Ralhan, Ranju

    2013-01-01

    Purpose The DNA methylation mediated by specific DNA methyltransferases (DNMTs), results in the epigenetic silencing of multiple genes which are implicated in human breast cancer. We hypothesized that the natural compounds modulate the expression of DNMTs and their associated proteins in the breast cancer cell lines and affect the methylation mediated gene silencing. Methods The DNMTs transcript expression was analyzed by reverse transcription-polymerase chain reaction (RT-PCR) in the tumors ...

  19. Structural and evolutionary bioinformatics of the SPOUT superfamily of methyltransferases

    Directory of Open Access Journals (Sweden)

    Purta Elzbieta

    2007-03-01

    Full Text Available Abstract Background SPOUT methyltransferases (MTases are a large class of S-adenosyl-L-methionine-dependent enzymes that exhibit an unusual alpha/beta fold with a very deep topological knot. In 2001, when no crystal structures were available for any of these proteins, Anantharaman, Koonin, and Aravind identified homology between SpoU and TrmD MTases and defined the SPOUT superfamily. Since then, multiple crystal structures of knotted MTases have been solved and numerous new homologous sequences appeared in the databases. However, no comprehensive comparative analysis of these proteins has been carried out to classify them based on structural and evolutionary criteria and to guide functional predictions. Results We carried out extensive searches of databases of protein structures and sequences to collect all members of previously identified SPOUT MTases, and to identify previously unknown homologs. Based on sequence clustering, characterization of domain architecture, structure predictions and sequence/structure comparisons, we re-defined families within the SPOUT superfamily and predicted putative active sites and biochemical functions for the so far uncharacterized members. We have also delineated the common core of SPOUT MTases and inferred a multiple sequence alignment for the conserved knot region, from which we calculated the phylogenetic tree of the superfamily. We have also studied phylogenetic distribution of different families, and used this information to infer the evolutionary history of the SPOUT superfamily. Conclusion We present the first phylogenetic tree of the SPOUT superfamily since it was defined, together with a new scheme for its classification, and discussion about conservation of sequence and structure in different families, and their functional implications. We identified four protein families as new members of the SPOUT superfamily. Three of these families are functionally uncharacterized (COG1772, COG1901, and COG4080

  20. Methyltransferase G9A Regulates Osteogenesis via Twist Gene Repression.

    Science.gov (United States)

    Higashihori, N; Lehnertz, B; Sampaio, A; Underhill, T M; Rossi, F; Richman, J M

    2017-09-01

    Here we investigate the role of epigenetic factors in controlling the timing of cranial neural crest cell differentiation. The gene coding for histone H3 lysine 9 methyltransferase G9A was conditionally deleted in neural crest cells with Wnt1-Cre. The majority of homozygous-null animals survived to birth but thereafter failed to thrive. Phenotypic analysis of postnatal animals revealed that the mutants displayed incomplete ossification and 20% shorter jaws as compared to their wild-type littermates. At E13.5, patterns of expression of the osteogenic transcription factor RUNX2 and the mesenchymal transcription factor TWIST are similar in controls and mutants; both overlap in areas of future intramembranous bone formation. At E14.5, the nonosteogenic mesenchyme expressed TWIST, whereas the ossification center had strong RUNX2 and osteopontin expression. In the mutants, TWIST protein was present in the osteogenic mesenchyme, while osteopontin was not expressed until E15.5. In addition, in mutants, small regions of TWIST-positive osteogenic mesenchyme were visible until E15.5. The delay in ossification and reduction in size of the ossification centers were correlated with an earlier decrease in proliferation. We used micromass cultures of the face to investigate the direct effects of G9A inhibition on skeletal differentiation. Addition of a small molecule inhibitor for G9A, BIX-01294, to wild-type cells upregulated Twist genes similar to what was observed in vivo. The inhibitor also caused decreases in several osteogenic markers. Chromatin immunoprecipitation analysis of primary osteogenic mesenchyme from calvaria revealed that Twist1 and Twist2 regulatory regions contain the repressive H3K9me2 marks catalyzed by G9A, which are removed when BIX-01294 is added. Our results establish a role for G9A and H3K9me2 in the regulation of Twist genes and provide novel insights into the significance of epigenetic mechanisms in controlling temporal and tissue-specific gene

  1. Genotype influences sulfur metabolism in broccoli (Brassica oleracea L.) under elevated CO2 and NaCl stress.

    Science.gov (United States)

    Rodríguez-Hernández, María del Carmen; Moreno, Diego A; Carvajal, Micaela; Martínez-Ballesta, María del Carmen

    2014-12-01

    Climatic change predicts elevated salinity in soils as well as increased carbon dioxide dioxide [CO2] in the atmosphere. The present study aims to determine the effect of combined salinity and elevated [CO2] on sulfur (S) metabolism and S-derived phytochemicals in green and purple broccoli (cv. Naxos and cv. Viola, respectively). Elevated [CO2] involved the amelioration of salt stress, especially in cv. Viola, where a lower biomass reduction by salinity was accompanied by higher sodium (Na(+)) and chloride (Cl(-)) compartmentation in the vacuole. Moreover, salinity and elevated [CO2] affected the mineral and glucosinolate contents and the activity of biosynthetic enzymes of S-derived compounds and the degradative enzyme of glucosinolate metabolism, myrosinase, as well as the related amino acids and the antioxidant glutathione (GSH). In cv. Naxos, elevated [CO2] may trigger the antioxidant response to saline stress by means of increased GSH concentration. Also, in cv. Naxos, indolic glucosinolates were more influenced by the NaCl×CO2 interaction whereas in cv. Viola the aliphatic glucosinolates were significantly increased by these conditions. Salinity and elevated [CO2] enhanced the S cellular partitioning and metabolism affecting the myrosinase-glucosinolate system. © The Author 2014. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.

  2. Selenium treatment differentially affects sulfur metabolism in high and low glucosinolate producing cultivars of broccoli (Brassica oleracea L.).

    Science.gov (United States)

    McKenzie, Marian J; Chen, Ronan K Y; Leung, Susanna; Joshi, Srishti; Rippon, Paula E; Joyce, Nigel I; McManus, Michael T

    2017-12-01

    The effect of selenium (Se) application on the sulfur (S)-rich glucosinolate (GSL)-containing plant, broccoli (Brassica oleracea L. var. italica) was examined with a view to producing germplasm with increased Se and GSL content for human health, and to understanding the influence of Se on the regulation of GSL production. Two cultivars differing in GSL content were compared. Increased Se application resulted in an increase in Se uptake in planta, but no significant change in total S or total GSL content in either cultivar. Also no significant change was observed in the activity of ATP sulfurylase (ATPS, EC 2.7.7.4) or O-acetylserine(thiol) lyase (OASTL, EC 2.5.1.47) with increased Se application. However, in the first investigation of APS kinase (APSK, EC 2.7.1.25) expression in response to Se fertilisation, an increase in transcript abundance of one variant of APS kinase 1 (BoAPSK1A) was observed in both cultivars, and an increase in BoAPSK2 transcript abundance was observed in the low GSL producing cultivar. A mechanism by which increased APSK transcription may provide a means of controlling the content of S-containing compounds, including GSLs, following Se uptake is proposed. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  3. CLONING, EXPRESSION, AND CHARACTERIZATION OF RAT S-ADENOSYL-L-METHIONINE: ARSENIC (III) METHYLTRANSFERASE (CYT19)

    Science.gov (United States)

    CLONING, EXPRESSION, AND CHARACTERIZATION OF RAT S-ADENOSYL-L-METHIONINE: ARSENIC(III) METHYLTRANSFERASE (cyt19)Stephen B. Waters1 , Felicia Walton1 , Miroslav Styblo1 , Karen Herbin-Davis2, and David J. Thomas2 1 School of Medicine, University of North Carolina at Chape...

  4. The genome-wide identification and transcriptional levels of DNA methyltransferases and demethylases in globe artichoke.

    Directory of Open Access Journals (Sweden)

    Silvia Gianoglio

    Full Text Available Changes to the cytosine methylation status of DNA, driven by the activity of C5 methyltransferases (C5-MTases and demethylases, exert an important influence over development, transposon movement, gene expression and imprinting. Three groups of C5-MTase enzymes have been identified in plants, namely MET (methyltransferase 1, CMT (chromomethyltransferases and DRM (domains rearranged methyltransferases. Here the repertoire of genes encoding C5-MTase and demethylase by the globe artichoke (Cynara cardunculus var. scolymus is described, based on sequence homology, a phylogenetic analysis and a characterization of their functional domains. A total of ten genes encoding C5-MTase (one MET, five CMTs and four DRMs and five demethylases was identified. An analysis of their predicted product's protein structure suggested an extensive level of conservation has been retained by the C5-MTases. Transcriptional profiling based on quantitative real time PCR revealed a number of differences between the genes encoding maintenance and de novo methyltransferases, sometimes in a tissue- or development-dependent manner, which implied a degree of functional specialization.

  5. The genome-wide identification and transcriptional levels of DNA methyltransferases and demethylases in globe artichoke.

    Science.gov (United States)

    Gianoglio, Silvia; Moglia, Andrea; Acquadro, Alberto; Comino, Cinzia; Portis, Ezio

    2017-01-01

    Changes to the cytosine methylation status of DNA, driven by the activity of C5 methyltransferases (C5-MTases) and demethylases, exert an important influence over development, transposon movement, gene expression and imprinting. Three groups of C5-MTase enzymes have been identified in plants, namely MET (methyltransferase 1), CMT (chromomethyltransferases) and DRM (domains rearranged methyltransferases). Here the repertoire of genes encoding C5-MTase and demethylase by the globe artichoke (Cynara cardunculus var. scolymus) is described, based on sequence homology, a phylogenetic analysis and a characterization of their functional domains. A total of ten genes encoding C5-MTase (one MET, five CMTs and four DRMs) and five demethylases was identified. An analysis of their predicted product's protein structure suggested an extensive level of conservation has been retained by the C5-MTases. Transcriptional profiling based on quantitative real time PCR revealed a number of differences between the genes encoding maintenance and de novo methyltransferases, sometimes in a tissue- or development-dependent manner, which implied a degree of functional specialization.

  6. Guanidinoacetate Methyltransferase (GAMT) Deficiency: Late Onset of Movement Disorder and Preserved Expressive Language

    Science.gov (United States)

    O'Rourke, Declan J.; Ryan, Stephanie; Salomons, Gajja; Jakobs, Cornelis; Monavari, Ahmad; King, Mary D.

    2009-01-01

    Guanidinoacetate methyltransferase (GAMT) deficiency is a disorder of creatine biosynthesis, characterized by early-onset learning disability and epilepsy in most affected children. Severe expressive language delay is a constant feature even in the mildest clinical phenotypes. We report the clinical, biochemical, imaging, and treatment data of two…

  7. The histone methyltransferase and putative oncoprotein MMSET is overexpressed in a large variety of human tumors

    DEFF Research Database (Denmark)

    Hudlebusch, Heidi Rye; Santoni-Rugiu, Eric; Simon, Ronald

    2011-01-01

    Multiple myeloma SET (Suppressor of variegation, Enhancer of zeste, and Trithorax) domain (MMSET) is a histone lysine methyltransferase deregulated in a subgroup of multiple myelomas with the t(4;14)(p16;q32) translocation and poor prognosis. With the aim of understanding, if MMSET can be involved...

  8. Guanidinoacetate methyltransferase (GAMT) deficiency : Outcomes in 48 individuals and recommendations for diagnosis, treatment and monitoring

    NARCIS (Netherlands)

    Stockler-Ipsiroglu, Sylvia; van Karnebeek, Clara; Longo, Nicola; Korenke, G. Christoph; Mercimek-Mahmutoglu, Saadet; Marquart, Iris; Barshop, Bruce; Grolik, Christiane; Schlune, Andrea; Angle, Brad; Araujo, Helena Caldeira; Coskun, Turgay; Diogo, Luisa; Geraghty, Michael; Haliloglu, Goknur; Konstantopoulou, Vassiliki; Leuzzi, Vincenzo; Levtova, Alina; MacKenzie, Jennifer; Maranda, Bruno; Mhanni, Aizeddin A.; Mitchell, Grant; Morris, Andrew; Newlove, Theresa; Renaud, Deborah; Scaglia, Fernando; Valayannopoulos, Vassili; van Spronsen, Francjan J.; Verbruggen, Krijn T.; Yuskiv, Nataliya; Nyhan, William; Schulze, Andreas

    We collected data on 48 patients from 38 families with guanidinoacetate methyltransferase (GAMT) deficiency. Global developmental delay/intellectual disability (DD/ID) with speech/language delay and behavioral problems as the most affected domains was present in 44 participants, with additional

  9. Control of substrate specificity by a single active site residue of the KsgA methyltransferase.

    Science.gov (United States)

    O'Farrell, Heather C; Musayev, Faik N; Scarsdale, J Neel; Rife, Jason P

    2012-01-10

    The KsgA methyltransferase is universally conserved and plays a key role in regulating ribosome biogenesis. KsgA has a complex reaction mechanism, transferring a total of four methyl groups onto two separate adenosine residues, A1518 and A1519, in the small subunit rRNA. This means that the active site pocket must accept both adenosine and N(6)-methyladenosine as substrates to catalyze formation of the final product N(6),N(6)-dimethyladenosine. KsgA is related to DNA adenosine methyltransferases, which transfer only a single methyl group to their target adenosine residue. We demonstrate that part of the discrimination between mono- and dimethyltransferase activity lies in a single residue in the active site, L114; this residue is part of a conserved motif, known as motif IV, which is common to a large group of S-adenosyl-L-methionine-dependent methyltransferases. Mutation of the leucine to a proline mimics the sequence found in DNA methyltransferases. The L114P mutant of KsgA shows diminished overall activity, and its ability to methylate the N(6)-methyladenosine intermediate to produce N(6),N(6)-dimethyladenosine is impaired; this is in contrast to a second active site mutation, N113A, which diminishes activity to a level comparable to L114P without affecting the methylation of N(6)-methyladenosine. We discuss the implications of this work for understanding the mechanism of KsgA's multiple catalytic steps.

  10. Catechol-O-methyltransferase gene methylation and substance use in adolescents: The TRAILS study

    NARCIS (Netherlands)

    L.J. van der Knaap (Lisette); J.M. Schäfer (Johanna); I.H.A. Franken (Ingmar); F.C. Verhulst (Frank); F.V.A. van Oort (Floor); H. Riese (Harriëtte)

    2014-01-01

    textabstractSubstance use often starts in adolescence and poses a major problem for society and individual health. The dopamine system plays a role in substance use, and catechol-O-methyltransferase (COMT) is an important enzyme that degrades dopamine. The Val108/158Met polymorphism

  11. Catechol-O-methyltransferase gene methylation and substance use in adolescents : the TRAILS study

    NARCIS (Netherlands)

    van der Knaap, L. J.; Schaefer, J. M.; Franken, I. H. A.; Verhulst, F. C.; van Oort, F. V. A.; Riese, H.

    Substance use often starts in adolescence and poses a major problem for society and individual health. The dopamine system plays a role in substance use, and catechol-O-methyltransferase (COMT) is an important enzyme that degrades dopamine. The Val(108/158)Met polymorphism modulates COMT activity

  12. Loss of the histone methyltransferase EZH2 induces resistance to multiple drugs in acute myeloid leukemia

    DEFF Research Database (Denmark)

    Göllner, Stefanie; Oellerich, Thomas; Agrawal-Singh, Shuchi

    2017-01-01

    In acute myeloid leukemia (AML), therapy resistance frequently occurs, leading to high mortality among patients. However, the mechanisms that render leukemic cells drug resistant remain largely undefined. Here, we identified loss of the histone methyltransferase EZH2 and subsequent reduction of h...

  13. Functional characterization of cinnamyl alcohol dehydrogenase and caffeic acid O-methyltransferase in Brachypodium distachyon.

    Science.gov (United States)

    Lignin is a significant recalcitrant in the conversion of plant biomass to bioethanol. Cinnamyl alcohol dehydrogenase (CAD) and caffeic acid O-methyltransferase (COMT) catalyze key steps in the pathway of lignin monomer biosynthesis. Brown midrib mutants in Zea mays and Sorghum bicolor with impaired...

  14. Polymorphisms in the catechol-o-methyltransferase gene and delirium in the elderly

    NARCIS (Netherlands)

    van Munster, Barbara C.; Baas, Frank; Tanck, Michael W.; de Rooij, Sophia E. J. A.

    2011-01-01

    Catechol-O-methyltransferase, encoded by the COMT gene, is one of the enzymes that degrade dopamine. The aim of this study was to investigate whether polymorphisms in the COMT gene were associated with delirium. Patients aged 65 years and older, acutely admitted to the medical department or to the

  15. Catechol-O-methyltransferase Val158Met and Cognitive Function in Parkinson's Disease

    NARCIS (Netherlands)

    Hoogland, Jeroen; de Bie, Rob M. A.; Williams-Gray, Caroline H.; Muslimović, Dino; Schmand, Ben; Post, Bart

    2010-01-01

    Cognitive dysfunction is one of the most incapacitating non-motor symptoms of Parkinson's disease (PD). Some cognitive deficits are thought to be related to abnormal dopamine homeostasis. The latter is influenced by catechol-O-methyltransferase (COMT), an enzyme that degrades dopamine. Previous

  16. Polymorphisms in catechol-O-methyltransferase and methylenetetrahydrofolate reductase in relation to the risk of schizophrenia.

    NARCIS (Netherlands)

    Muntjewerff, J.W.; Gellekink, H.; Heijer, M. den; Hoogendoorn, M.L.; Kahn, R.S.; Sinke, R.J.; Blom, H.J.

    2008-01-01

    BACKGROUND: Evidence is emerging for the association of aberrant homocysteine-methylation cycle and increased risk of schizophrenia. METHODS: We examined the prevalence of the catechol-O-methyltransferase (COMT) 324G>A (Val108/158Met) and methylenetetrahydrofolate reductase (MTHFR) 677C>T

  17. Association of Catechol-O-Methyltransferase (COMT) Polymorphism and Academic Achievement in a Chinese Cohort

    Science.gov (United States)

    Yeh, Ting-Kuang; Chang, Chun-Yen; Hu, Chung-Yi; Yeh, Ting-Chi; Lin, Ming-Yeh

    2009-01-01

    Catechol-O-methyltransferase (COMT) is a methylation enzyme that catalyzes the degradation pathway and inactivation of dopamine. It is accepted widely as being involved in the modulation of dopaminergic physiology and prefrontal cortex (PFC) function. The COMT Val158Met polymorphism is associated with variation in COMT activity. COMT 158Met allele…

  18. Catechol-O-methyltransferase val158met and cognitive function in Parkinson's disease

    NARCIS (Netherlands)

    Hoogland, J.; de Bie, R.M.A.; Williams-Gray, C.H.; Muslimovic, D.; Schmand, B.; Post, B.

    2010-01-01

    Cognitive dysfunction is one of the most incapacitating non-motor symptoms of Parkinson's disease (PD). Some cognitive deficits are thought to be related to abnormal dopamine homeostasis. The latter is influenced by catechol-O-methyltransferase (COMT), an enzyme that degrades dopamine. Previous

  19. Catechol-O-methyltransferase: a method for autoradiographic visualization of isozymes in cellogel

    Energy Technology Data Exchange (ETDEWEB)

    Brahe, C.; Crosti, N.; Meera Khan, P.; Serra, A.

    1984-02-01

    An electrophoretic procedure for separating the molecular forms of catechol-O-methyltransferase in cellulose acetate gel is described; the zones of enzyme activity were revealed by autoradiography. The electrophoretic patterns of the enzyme in several tissues and cell lines derived from four different species are presented.

  20. Catechol-O-methyltransferase Val158Met and the Risk of Dyskinesias in Parkinson's Disease

    NARCIS (Netherlands)

    De Lau, L.M.L.; Verbaan, D.; Marinus, J.; Heutink, P.; van Hilten, J.J.

    2012-01-01

    Background:: The A-allele of the catechol-O-methyltransferase (COMT) Val158Met polymorphism is associated with decreased enzymatic activity and higher dopamine availability. Methods:: We studied 219 patients with PD who were free of dyskinesias at baseline and underwent thorough annual examinations.

  1. Catechol-O-methyltransferase val158met and cognitive function in Parkinson's disease.

    NARCIS (Netherlands)

    Hoogland, J.; Bie, R.M. de; Williams-Gray, C.H.; Muslimovic, D.; Schmand, B.A.; Post, B.

    2010-01-01

    Cognitive dysfunction is one of the most incapacitating non-motor symptoms of Parkinson's disease (PD). Some cognitive deficits are thought to be related to abnormal dopamine homeostasis. The latter is influenced by catechol-O-methyltransferase (COMT), an enzyme that degrades dopamine. Previous

  2. Structural mechanism of S-adenosyl methionine binding to catechol O-methyltransferase.

    Directory of Open Access Journals (Sweden)

    Douglas Tsao

    Full Text Available Methyltransferases possess a homologous domain that requires both a divalent metal cation and S-adenosyl-L-methionine (SAM to catalyze its reactions. The kinetics of several methyltransferases has been well characterized; however, the details regarding their structural mechanisms have remained unclear to date. Using catechol O-methyltransferase (COMT as a model, we perform discrete molecular dynamics and computational docking simulations to elucidate the initial stages of cofactor binding. We find that COMT binds SAM via an induced-fit mechanism, where SAM adopts a different docking pose in the absence of metal and substrate in comparison to the holoenzyme. Flexible modeling of the active site side-chains is essential for observing the lowest energy state in the apoenzyme; rigid docking tools are unable to recapitulate the pose unless the appropriate side-chain conformations are given a priori. From our docking results, we hypothesize that the metal reorients SAM in a conformation suitable for donating its methyl substituent to the recipient ligand. The proposed mechanism enables a general understanding of how divalent metal cations contribute to methyltransferase function.

  3. Catechol-O-methyltransferase (COMT) gene variants and pain in chronic pancreatitis

    NARCIS (Netherlands)

    Esch, A.A.J.; Vries, E. De; Morsche, R.H.M. te; Oijen, M.G.H. van; Jansen, J.B.M.J.; Drenth, J.P.H.

    2011-01-01

    BACKGROUND: Pain is the major symptom of chronic pancreatitis. The role of genetics in pancreatic pain is unclear. Catechol-O-methyltransferase (COMT) regulates enkephalin levels and influences pain perception. The COMT gene contains functional polymorphisms that have been found to influence human

  4. Local chromatin microenvironment determines DNMT activity : from DNA methyltransferase to DNA demethylase or DNA dehydroxymethylase

    NARCIS (Netherlands)

    van der Wijst, Monique G. P.; Venkiteswaran, Muralidhar; Chen, Hui; Xu, Guo-Liang; Plosch, Torsten; Rots, Marianne G.

    2015-01-01

    Insights on active DNA demethylation disproved the original assumption that DNA methylation is a stable epigenetic modification. Interestingly, mammalian DNA methyltransferases 3A and 3B (DNMT-3A and -3B) have also been reported to induce active DNA demethylation, in addition to their well-known

  5. DNA methyltransferase and alcohol dehydrogenase: gene-nutrient interactions in relation to risk of colorectal polyps.

    NARCIS (Netherlands)

    Jung, A.Y.; Poole, E.M.; Bigler, J.; Whitton, J.; Potter, J.D.; Ulrich, C.M.

    2008-01-01

    Disturbances in DNA methylation are a characteristic of colorectal carcinogenesis. Folate-mediated one-carbon metabolism is essential for providing one-carbon groups for DNA methylation via DNA methyltransferases (DNMTs). Alcohol, a folate antagonist, could adversely affect one-carbon metabolism. In

  6. Recognition elements in rRNA for the tylosin resistance methyltransferase RlmA(II)

    DEFF Research Database (Denmark)

    Lebars, Isabelle; Husson, Clotilde; Yoshizawa, Satoko

    2007-01-01

    antibiotics. We have previously solved the solution structure of hairpin 35 in the conformation that is recognized by the RlmA(II) methyltransferase from Streptococcus pneumoniae. It was shown that while essential recognition elements are located in hairpin 35, the interactions between RlmA(II) and hairpin 35...

  7. Recognition of RNA cap in the Wesselsbron virus NS5 methyltransferase domain: implications for RNA-capping mechanisms in Flavivirus.

    Science.gov (United States)

    Bollati, Michela; Milani, Mario; Mastrangelo, Eloise; Ricagno, Stefano; Tedeschi, Gabriella; Nonnis, Simona; Decroly, Etienne; Selisko, Barbara; de Lamballerie, Xavier; Coutard, Bruno; Canard, Bruno; Bolognesi, Martino

    2009-01-09

    The mRNA-capping process starts with the conversion of a 5'-triphosphate end into a 5'-diphosphate by an RNA triphosphatase, followed by the addition of a guanosine monophosphate unit in a 5'-5' phosphodiester bond by a guanylyltransferase. Methyltransferases are involved in the third step of the process, transferring a methyl group from S-adenosyl-l-methionine to N7-guanine (cap 0) and to the ribose 2'OH group (cap 1) of the first RNA nucleotide; capping is essential for mRNA stability and proper replication. In the genus Flavivirus, N7-methyltransferase and 2'O-methyltransferase activities have been recently associated with the N-terminal domain of the viral NS5 protein. In order to further characterize the series of enzymatic reactions that support capping, we analyzed the crystal structures of Wesselsbron virus methyltransferase in complex with the S-adenosyl-l-methionine cofactor, S-adenosyl-l-homocysteine (the product of the methylation reaction), Sinefungin (a molecular analogue of the enzyme cofactor), and three different cap analogues (GpppG, (N7Me)GpppG, and (N7Me)GpppA). The structural results, together with those on other flaviviral methyltransferases, show that the capped RNA analogues all bind to an RNA high-affinity binding site. However, lack of specific interactions between the enzyme and the first nucleotide of the RNA chain suggests the requirement of a minimal number of nucleotides following the cap to strengthen protein/RNA interaction. Our data also show that, following incubation with guanosine triphosphate, Wesselsbron virus methyltransferase displays a guanosine monophosphate molecule covalently bound to residue Lys28, hinting at possible implications for the transfer of a guanine group to ppRNA. The structures of the Wesselsbron virus methyltransferase complexes obtained are discussed in the context of a model for N7-methyltransferase and 2'O-methyltransferase activities.

  8. Evolution of the Phosphatidylcholine Biosynthesis Pathways in Green Algae: Combinatorial Diversity of Methyltransferases.

    Science.gov (United States)

    Hirashima, Takashi; Toyoshima, Masakazu; Moriyama, Takashi; Sato, Naoki

    2018-01-12

    Phosphatidylcholine (PC) is one of the most common phospholipids in eukaryotes, although some green algae such as Chlamydomonas reinhardtii are known to lack PC. Recently, we detected PC in four species in the genus Chlamydomonas: C. applanata NIES-2202, C. asymmetrica NIES-2207, C. debaryana NIES-2212, and C. sphaeroides NIES-2242. To reveal the PC biosynthesis pathways in green algae and the evolutionary scenario involved in their diversity, we analyzed the PC biosynthesis genes in these four algae using draft genome sequences. Homology searches suggested that PC in these species is synthesized by phosphoethanolamine-N-methyltransferase (PEAMT) and/or phosphatidylethanolamine-N-methyltransferase (PEMT), both of which are absent in C. reinhardtii. Recombinant PEAMTs from these algae showed methyltransferase activity for phosphoethanolamine but not for monomethyl phosphoethanolamine in vitro, in contrast to land plant PEAMT, which catalyzes the three methylations from phosphoethanolamine to phosphocholine. This suggested an involvement of other methyltransferases in PC biosynthesis. Here, we characterized the putative phospholipid-N-methyltransferase (PLMT) genes of these species by genetic and phylogenetic analysis. Complementation assays using a PC biosynthesis-deficient yeast suggested that the PLMTs of these algae can synthesize PC from phosphatidylethanolamine. These results indicated that the PC biosynthesis pathways in green algae differ from those of land plants, although the enzymes involved are homologous. Phylogenetic analysis suggested that the PEAMTs and PLMTs in these algae were inherited from the common ancestor of green algae. The absence of PC biosynthesis in many Chlamydomonas species is likely a result of parallel losses of PEAMT and PLMT in this genus.

  9. Floral Benzenoid Carboxyl Methyltransferases: From in Vitro to in Planta Function

    Energy Technology Data Exchange (ETDEWEB)

    Effmert,U.; Saschenbrecker, S.; Ross, J.; Negre, F.; Fraser, C.; Noel, J.; Dudareva, N.; Piechulla, B.

    2005-01-01

    Benzenoid carboxyl methyltransferases synthesize methyl esters (e.g., methyl benzoate and methyl salicylate), which are constituents of aromas and scents of many plant species and play important roles in plant communication with the surrounding environment. Within the past five years, eleven such carboxyl methyltransferases were isolated and most of them were comprehensively investigated at the biochemical, molecular and structural level. Two types of enzymes can be distinguished according to their substrate preferences: the SAMT-type enzymes isolated from Clarkia breweri, Stephanotis floribunda, Antirrhinum majus, Hoya carnosa, and Petunia hybrida, which have a higher catalytic efficiency and preference for salicylic acid, while BAMT-type enzymes from A. majus, Arabidopsis thaliana, Arabidopsis lyrata, and Nicotiana suaveolens prefer benzoic acid. The elucidation of C. breweri SAMT's three-dimensional structure allowed a detailed modelling of the active sites of the carboxyl methyltransferases and revealed that the SAM binding pocket is highly conserved among these enzymes while the methyl acceptor binding site exhibits some variability, allowing a classification into SAMT-type and BAMT-type enzymes. The analysis of expression patterns coupled with biochemical characterization showed that these carboxyl methyltransferases are involved either in floral scent biosynthesis or in plant defense responses. While the latter can be induced by biotic or abiotic stress, the genes responsible for floral scent synthesis exhibit developmental and rhythmic expression pattern. The nature of the product and efficiency of its formation in plants depend on the availability of substrates, the catalytic efficiency of the enzyme toward benzoic acid and/or salicylic acid, and the transcriptional, translational, and post-translational regulation at the enzyme level. The biochemical properties of benzenoid carboxyl methyltransferases suggest that the genes involved in plant defenses

  10. Understanding the degradation of ascorbic acid and glutathione in relation to the levels of oxidative stress biomarkers in broccoli (Brassica oleracea L. italica cv. Bellstar) during storage and mechanical processing.

    Science.gov (United States)

    Raseetha, Siva; Leong, Sze Ying; Burritt, David John; Oey, Indrawati

    2013-06-01

    The purpose of this research was to understand the degradation of ascorbic acid and glutathione content in broccoli florets (Brassica oleracea L. italica cv. Bellstar) during prolonged storage and subsequent mechanical processing. The initial content of total ascorbic acid and glutathione in broccoli florets averaged at 5.18 ± 0.23 and 0.70 ± 0.03 μmol/g fresh weight, respectively. Results showed that the content of ascorbic acid and glutathione in broccoli degraded during storage at 23°C, for at least 4.5-fold after 6 days of storage. On each day of storage, broccoli florets were mechanically processed, but the content of total ascorbic acid and glutathione was not significantly affected. When the mechanically processed broccoli florets were further incubated for up to 6h, the amount of ascorbic acid was greatly reduced as compared to glutathione. To obtain an in-depth understanding on the degradation of ascorbic acid and glutathione, the activity of enzymes involved in plant antioxidative system via ascorbate-glutathione cycle, as a response towards oxidative stress that took place during storage was determined in this study. The content of total ascorbic acid and glutathione in broccoli florets before and after mechanical processing were found to decrease concurrently with the activity of ascorbic acid peroxidase and glutathione reductase over the experimental storage duration. Meanwhile, the effect of oxidative stress on the content of ascorbic acid and glutathione was apparent during the 6h of incubation after mechanical processing. This phenomenon was demonstrated by the level of oxidative stress biomarkers examined, in which the formation of lipid peroxides, protein carbonyls and DNA oxidised products was positively associated with the degradation of total ascorbic acid and glutathione. Copyright © 2012 Elsevier Ltd. All rights reserved.

  11. Protein Arginine Methyltransferase 5 as a Driver of Lymphomagenesis

    Science.gov (United States)

    Smith, Porsha Latrice

    Over the past decade, it has become clear that oncogenesis is a process driven by a wide variety of triggers including gene mutations, gene amplifications, inflammation, and immune deficiency. The growing pool of data collected from whole genome and epigenome studies of both solid and blood cancers has pointed toward dysregulation of chromatin remodelers as a unique class of cancer drivers. Next generation sequencing studies of lymphomas have identified a wide array of somatic mutations affecting enzymes that regulate epigenetic control of gene expression. Lymphoma is a type of cancer that originates in secondary lymphoid organs and manifests as an outgrowth of transformed lymphocytes, or white blood cells (WBCs) in the blood. The majority of lymphoma cases can be grouped into the Non-Hodgkins lymphoma (NHL) subset and mainly occurs in B-cells. B-cell NHL is a heterogeneous set of cancers that would benefit from new therapies to improve patient progression-free survival. Cancers such as NHL typically present with a combination of genetic and epigenetic aberrations that contribute to the malignancy program. The epigenetic modifier protein arginine methyltransferase 5 (PRMT5) is required for B-cell transformation following Epstein-Barr virus (EBV) infection, and is overexpressed in various subsets of B-cell NHL. Based on these data we hypothesized that PRMT5 is a major driver of B-cell lymphomagenesis. To explore the role of PRMT5 in the development and progression of B-cell NHL we created a small molecule inhibitors targeted to PRMT5. Using the NHL subset mantle cell lymphoma (MCL) as a model we tested the efficacy of the drug. We discovered that PRMT5 was overexpressed in MCL primary samples and cell lines as compared to normal resting B cells. Furthermore, use of the small molecule inhibitor decreased the proliferation and viability in these cells without affecting the normal B-cells. Additionally, use of inhibitors caused G2/M cell cycle and decreased the

  12. UV-B Irradiation Changes Specifically the Secondary Metabolite Profile in Broccoli Sprouts: Induced Signaling Overlaps with Defense Response to Biotic Stressors

    Science.gov (United States)

    Mewis, Inga; Schreiner, Monika; Nguyen, Chau Nhi; Krumbein, Angelika; Ulrichs, Christian; Lohse, Marc; Zrenner, Rita

    2012-01-01

    Only a few environmental factors have such a pronounced effect on plant growth and development as ultraviolet light (UV). Concerns have arisen due to increased UV-B radiation reaching the Earth’s surface as a result of stratospheric ozone depletion. Ecologically relevant low to moderate UV-B doses (0.3–1 kJ m–2 d–1) were applied to sprouts of the important vegetable crop Brassica oleracea var. italica (broccoli), and eco-physiological responses such as accumulation of non-volatile secondary metabolites were related to transcriptional responses with Agilent One-Color Gene Expression Microarray analysis using the 2×204 k format Brassica microarray. UV-B radiation effects have usually been linked to increases in phenolic compounds. As expected, the flavonoids kaempferol and quercetin accumulated in broccoli sprouts (the aerial part of the seedlings) 24 h after UV-B treatment. A new finding is the specific UV-B-mediated induction of glucosinolates (GS), especially of 4-methylsulfinylbutyl GS and 4-methoxy-indol-3-ylmethyl GS, while carotenoids and Chl levels remained unaffected. Accumulation of defensive GS metabolites was accompanied by increased expression of genes associated with salicylate and jasmonic acid signaling defense pathways and up-regulation of genes responsive to fungal and bacterial pathogens. Concomitantly, plant pre-exposure to moderate UV-B doses had negative effects on the performance of the caterpillar Pieris brassicae (L.) and on the population growth of the aphid Myzus persicae (Sulzer). Moreover, insect-specific induction of GS in broccoli sprouts was affected by UV-B pre-treatment. PMID:22773681

  13. Transcriptome and Metabolome Analyses of Glucosinolates in Two Broccoli Cultivars Following Jasmonate Treatment for the Induction of Glucosinolate Defense to Trichoplusia ni (Hübner).

    Science.gov (United States)

    Ku, Kang-Mo; Becker, Talon M; Juvik, John A

    2016-07-15

    Lepidopteran larvae growth is influenced by host plant glucosinolate (GS) concentrations, which are, in turn, influenced by the phytohormone jasmonate (JA). In order to elucidate insect resistance biomarkers to lepidopteran pests, transcriptome and metabolome analyses following JA treatments were conducted with two broccoli cultivars, Green Magic and VI-158, which have differentially induced indole GSs, neoglucobrassicin and glucobrassicin, respectively. To test these two inducible GSs on growth of cabbage looper (Trichoplusia ni), eight neonate cabbage looper larvae were placed onto each of three plants per JA treatments (0, 100, 200, 400 µM) three days after treatment. After five days of feeding, weight of larvae and their survival rate was found to decrease with increasing JA concentrations in both broccoli cultivars. JA-inducible GSs were measured by high performance liquid chromatography. Neoglucobrassicin in Green Magic and glucobrassicin in VI-158 leaves were increased in a dose-dependent manner. One or both of these glucosinolates and/or their hydrolysis products showed significant inverse correlations with larval weight and survival (five days after treatment) while being positively correlated with the number of days to pupation. This implies that these two JA-inducible glucosinolates can influence the growth and survival of cabbage looper larvae. Transcriptome profiling supported the observed changes in glucosinolate and their hydrolysis product concentrations following JA treatments. Several genes related to GS metabolism differentiate the two broccoli cultivars in their pattern of transcriptional response to JA treatments. Indicative of the corresponding change in indole GS concentrations, transcripts of the transcription factor MYB122, core structure biosynthesis genes (CYP79B2, UGT74B1, SUR1, SOT16, SOT17, and SOT18), an indole glucosinolate side chain modification gene (IGMT1), and several glucosinolate hydrolysis genes (TGG1, TGG2, and ESM1) were

  14. Transcriptome and Metabolome Analyses of Glucosinolates in Two Broccoli Cultivars Following Jasmonate Treatment for the Induction of Glucosinolate Defense to Trichoplusia ni (Hübner

    Directory of Open Access Journals (Sweden)

    Kang-Mo Ku

    2016-07-01

    Full Text Available Lepidopteran larvae growth is influenced by host plant glucosinolate (GS concentrations, which are, in turn, influenced by the phytohormone jasmonate (JA. In order to elucidate insect resistance biomarkers to lepidopteran pests, transcriptome and metabolome analyses following JA treatments were conducted with two broccoli cultivars, Green Magic and VI-158, which have differentially induced indole GSs, neoglucobrassicin and glucobrassicin, respectively. To test these two inducible GSs on growth of cabbage looper (Trichoplusia ni, eight neonate cabbage looper larvae were placed onto each of three plants per JA treatments (0, 100, 200, 400 µM three days after treatment. After five days of feeding, weight of larvae and their survival rate was found to decrease with increasing JA concentrations in both broccoli cultivars. JA-inducible GSs were measured by high performance liquid chromatography. Neoglucobrassicin in Green Magic and glucobrassicin in VI-158 leaves were increased in a dose-dependent manner. One or both of these glucosinolates and/or their hydrolysis products showed significant inverse correlations with larval weight and survival (five days after treatment while being positively correlated with the number of days to pupation. This implies that these two JA-inducible glucosinolates can influence the growth and survival of cabbage looper larvae. Transcriptome profiling supported the observed changes in glucosinolate and their hydrolysis product concentrations following JA treatments. Several genes related to GS metabolism differentiate the two broccoli cultivars in their pattern of transcriptional response to JA treatments. Indicative of the corresponding change in indole GS concentrations, transcripts of the transcription factor MYB122, core structure biosynthesis genes (CYP79B2, UGT74B1, SUR1, SOT16, SOT17, and SOT18, an indole glucosinolate side chain modification gene (IGMT1, and several glucosinolate hydrolysis genes (TGG1, TGG2, and

  15. Mouse arsenic (+3 oxidation state) methyltransferase genotype affects metabolism and tissue dosimetry of arsenicals after arsenite administration in drinking water

    Science.gov (United States)

    Arsenic (+3 oxidation state) methyltransferase (As3mt) catalyzes methylation of inorganic arsenic producing a number of methylated arsenic metabolites. Although methylation has been commonly considered a pathway for detoxification of arsenic, some highly reactive methylated ars...

  16. A conformational switch in the active site of BT_2972, a methyltransferase from an antibiotic resistant pathogen B. thetaiotaomicron.

    Directory of Open Access Journals (Sweden)

    Veerendra Kumar

    Full Text Available Methylation is one of the most common biochemical reactions involved in cellular and metabolic functions and is catalysed by the action of methyltransferases. Bacteroides thetaiotaomicron is an antibiotic-resistant bacterium that confers resistance through methylation, and as yet, there is no report on the structure of methyltransferases from this bacterium. Here, we report the crystal structure of an AdoMet-dependent methyltransferase, BT_2972 and its complex with AdoMet and AdoHcy for B. thetaiotaomicron VPI-5482 strain along with isothermal titration calorimetric assessment of the binding affinities. Comparison of the apo and complexed BT_2972 structures reveals a significant conformational change between open and closed forms of the active site that presumably regulates the association with cofactors and may aid interaction with substrate. Together, our analysis suggests that BT_2972 is a small molecule methyltransferase and might catalyze two O-methylation reaction steps involved in the ubiquinone biosynthesis pathway.

  17. Quantitative determination of dithiocarbamates in human plasma, serum, erythrocytes and urine: pharmacokinetics of broccoli sprout isothiocyanates in humans.

    Science.gov (United States)

    Ye, Lingxiang; Dinkova-Kostova, Albena T; Wade, Kristina L; Zhang, Yuesheng; Shapiro, Theresa A; Talalay, Paul

    2002-02-01

    Humans are exposed to substantial quantities of isothiocyanates and glucosinolates from vegetables. Since dietary isothiocyanates are widely regarded as potentially important chemoprotectors against cancer, reliable methods for measuring the plasma and tissue pharmacokinetics of isothiocyanates and their dithiocarbamate metabolites are essential for defining dosing regimens. Isothiocyanates (ITC) and dithiocarbamates (DTC) react quantitatively with 1,2-benzenedithiol to produce 1,3-benzodithiole-2-thione that can be quantified spectroscopically. Although this cyclocondensation reaction has been highly useful for analyzing plant material and urine samples, the determination of DTC/ITC (the total quantity of DTC and ITC components in a sample that react in the cyclocondensation reaction) in blood and tissues has been hampered by their low levels and the high concentrations of proteins that interfere with the cyclocondensation reaction. The protein content of blood and tissues was reduced by the precipitation with polyethylene glycol (PEG) or ultrafiltration, and the sensitivity of the method was increased substantially by the solid phase extraction of the cyclocondensation product. Pharmacokinetic measurements were made in four human volunteers who received single doses of about 200 micromol of broccoli sprout isothiocyanates (largely sulforaphane, with lesser amounts of iberin and erucin). Isothiocyanates were absorbed rapidly, reached peak concentrations of 0.943-2.27 micromol/l in plasma, serum and erythrocytes at 1 h after feeding and declined with first-order kinetics (half-life of 1.77+/-0.13 h). The cumulative excretion at 8 h was 58.3+/-2.8% of the dose. Clearance was 369+/-53 ml/min, indicating active renal tubular secretion. A sensitive and specific method for quantifying DTC levels in human plasma, serum, and erythrocytes has been devised. Determinations of ITC/DTC levels are important because: (i) dietary isothiocyanates are of potential value in reducing

  18. Cloning, sequencing, and expression of the uroporphyrinogen III methyltransferase cobA gene of Propionibacterium freudenreichii (shermanii).

    OpenAIRE

    Sattler, I; Roessner, C A; Stolowich, N J; Hardin, S H; Harris-Haller, L W; Yokubaitis, N T; Murooka, Y; Hashimoto, Y; Scott, A I

    1995-01-01

    We cloned, sequenced, and overexpressed cobA, the gene encoding uroporphyrinogen III methyltransferase in Propionibacterium freudenreichii, and examined the catalytic properties of the enzyme. The methyltransferase is similar in mass (27 kDa) and homologous to the one isolated from Pseudomonas denitrificans. In contrast to the much larger isoenzyme encoded by the cysG gene of Escherichia coli (52 kDa), the P. freudenreichii enzyme does not contain the additional 22-kDa peptide moiety at its N...

  19. Biochemical Studies of Mycobacterial Fatty Acid Methyltransferase: A Catalyst for the Enzymatic Production of Biodiesel.

    Science.gov (United States)

    Petronikolou, Nektaria; Nair, Satish K

    2015-11-19

    Transesterification of fatty acids yields the essential component of biodiesel, but current processes are cost-prohibitive and generate waste. Recent efforts make use of biocatalysts that are effective in diverting products from primary metabolism to yield fatty acid methyl esters in bacteria. These biotransformations require the fatty acid O-methyltransferase (FAMT) from Mycobacterium marinum (MmFAMT). Although this activity was first reported in the literature in 1970, the FAMTs have yet to be biochemically characterized. Here, we describe several crystal structures of MmFAMT, which highlight an unexpected structural conservation with methyltransferases that are involved in plant natural product metabolism. The determinants for ligand recognition are analyzed by kinetic analysis of structure-based active-site variants. These studies reveal how an architectural fold employed in plant natural product biosynthesis is used in bacterial fatty acid O-methylation. Copyright © 2015 Elsevier Ltd. All rights reserved.

  20. EmtA, a rRNA methyltransferase conferring high-level evernimicin resistance

    DEFF Research Database (Denmark)

    Mann, P. A.; Xiong, L.; Mankin, A. S.

    2001-01-01

    Enterococcus faecium strain 9631355 was isolated from animal sources on the basis of its resistance to the growth promotant avilamycin. The strain also exhibited high-level resistance to evernimicin, a drug undergoing evaluation as a therapeutic agent in humans. Ribosomes from strain 9631355...... exhibited a dramatic reduction in evernimicin binding, shown by both cell-free translation assays and direct-binding assays. The resistance determinant was cloned from strain 9631355; sequence alignments suggested it was a methyltransferase and therefore it was designated emtA for evernimicin...... methyltransferase. Evernimicin resistance was transmissible and emtA was localized to a plasmid-borne insertion element. Purified EmtA methylated 50S subunits from an evernimicin-sensitive strain 30-fold more efficiently than those from a resistant strain. Reverse transcription identified a pause site...

  1. Crystal structures of the methyltransferase and helicase from the ZIKA 1947 MR766 Uganda strain

    Energy Technology Data Exchange (ETDEWEB)

    Bukrejewska, Malgorzata; Derewenda, Urszula; Radwanska, Malwina; Engel, Daniel A.; Derewenda, Zygmunt S.

    2017-08-15

    Two nonstructural proteins encoded byZika virusstrain MR766 RNA, a methyltransferase and a helicase, were crystallized and their structures were solved and refined at 2.10 and 2.01 Å resolution, respectively. The NS5 methyltransferase contains a boundS-adenosyl-L-methionine (SAM) co-substrate. The NS3 helicase is in the apo form. Comparison with published crystal structures of the helicase in the apo, nucleotide-bound and single-stranded RNA (ssRNA)-bound states suggests that binding of ssRNA to the helicase may occur through conformational selection rather than induced fit.

  2. Super-resolution optical DNA Mapping via DNA methyltransferase-directed click chemistry

    DEFF Research Database (Denmark)

    Vranken, Charlotte; Deen, Jochem; Dirix, Lieve

    2014-01-01

    We demonstrate an approach to optical DNA mapping, which enables near single-molecule characterization of whole bacteriophage genomes. Our approach uses a DNA methyltransferase enzyme to target labelling to specific sites and copper-catalysed azide-alkyne cycloaddition to couple a fluorophore...... to the DNA. We achieve a labelling efficiency of ∼70% with an average labelling density approaching one site every 500 bp. Such labelling density bridges the gap between the output of a typical DNA sequencing experiment and the long-range information derived from traditional optical DNA mapping. We lay...... the foundations for a wider-scale adoption of DNA mapping by screening 11 methyltransferases for their ability to direct sequence-specific DNA transalkylation; the first step of the DNA labelling process and by optimizing reaction conditions for fluorophore coupling via a click reaction. Three of 11 enzymes...

  3. Crystallization and preliminary X-ray diffraction studies of a catechol-O-methyltransferase/inhibitor complex

    Energy Technology Data Exchange (ETDEWEB)

    Rodrigues, M. L. [Instituto de Tecnologia Química e Biológica (ITQB), Universidade Nova de Lisboa, Av. República, Apt. 127, 2781-901 Oeiras (Portugal); Bonifácio, M. J.; Soares-da-Silva, P. [Department of Research and Development, BIAL, 4785 S. Mamede do Coronado (Portugal); Carrondo, M. A.; Archer, M., E-mail: archer@itqb.unl.pt [Instituto de Tecnologia Química e Biológica (ITQB), Universidade Nova de Lisboa, Av. República, Apt. 127, 2781-901 Oeiras (Portugal)

    2005-01-01

    Catechol-O-methyltransferase has been co-crystallized with a novel inhibitor, which has potential therapeutic application in the Parkinson’s disease therapy. Inhibitors of the enzyme catechol-O-methyltransferase (COMT) are used as co-adjuvants in the therapy of Parkinson’s disease. A recombinant form of the soluble cytosolic COMT from rat has been co-crystallized with a new potent inhibitor, BIA 8-176 [(3,4-dihydroxy-2-nitrophenyl)phenylmethanone], by the vapour-diffusion method using PEG 6K as precipitant. Crystals diffract to 1.6 Å resolution on a synchrotron-radiation source and belong to the monoclinic space group P2{sub 1}, with unit-cell parameters a = 52.77, b = 79.63, c = 61.54 Å, β = 91.14°.

  4. Facile synthesis of nano cauliflower and nano broccoli like hierarchical superhydrophobic composite coating using PVDF/carbon soot particles via gelation technique.

    Science.gov (United States)

    Sahoo, Bichitra Nanda; Balasubramanian, Kandasubramanian

    2014-12-15

    We have elucidated a cost effective fabrication technique to produce superhydrophobic polyvinylidene fluoride (PVDF/DMF/candle soot particle and PVDF/DMF/camphor soot particle composite) porous materials. The water repellent dry composite was formed by the interaction of non-solvent (methanol) into PVDF/carbon soot particles suspension in N,N-dimethylformamide (DMF). It is seen that longer quenching time effectively changes the surface morphology of dry composites. The nano broccoli like hierarchical microstructure with micro or nano scaled roughen surface was obtained for PVDF/DMF/camphor soot particle, which reveals water contact angle of 172° with roll off angle of 2°. However, composite coating of PVDF/DMF/candle soot particle shows nano cauliflower like hierarchical, which illustrates water contact angle of 169° with roll off angle of 3°. To elucidate the enhancement of water repellent property of PVDF composites, we further divulge the evolution mechanism of nano cauliflower and nano broccoli structure. In order to evaluate the water contact angle of PVDF composites, surface diffusion of water inside the pores is investigated. Furthermore, the addition of small amount of carbon soot particles in composite not only provides the crystallization of PVDF, but also leads to dramatical amendment of surface morphology which increases the surface texture and roughness for superhydrophobicity. Copyright © 2014 Elsevier Inc. All rights reserved.

  5. Antiamnesic Effect of Broccoli (Brassica oleracea var. italica) Leaves on Amyloid Beta (Aβ)1-42-Induced Learning and Memory Impairment.

    Science.gov (United States)

    Park, Seon Kyeong; Ha, Jeong Su; Kim, Jong Min; Kang, Jin Yong; Lee, Du Sang; Guo, Tian Jiao; Lee, Uk; Kim, Dae-Ok; Heo, Ho Jin

    2016-05-04

    To examine the antiamnesic effects of broccoli (Brassica oleracea var. italica) leaves, we performed in vitro and in vivo tests on amyloid beta (Aβ)-induced neurotoxicity. The chloroform fraction from broccoli leaves (CBL) showed a remarkable neuronal cell-protective effect and an inhibition against acetylcholinesterase (AChE). The ameliorating effect of CBL on Aβ1-42-induced learning and memory impairment was evaluated by Y-maze, passive avoidance, and Morris water maze tests. The results indicated improving cognitive function in the CBL group. After the behavioral tests, antioxidant effects were detected by superoxide dismutase (SOD), oxidized glutathione (GSH)/total GSH, and malondialdehyde (MDA) assays, and inhibition against AChE was also presented in the brain. Finally, oxo-dihydroxy-octadecenoic acid (oxo-DHODE) and trihydroxy-octadecenoic acid (THODE) as main compounds were identified by quadrupole time-of-flight ultraperformance liquid chromatography (Q-TOF UPLC-MS) analysis. Therefore, our studies suggest that CBL could be used as a natural resource for ameliorating Aβ1-42-induced learning and memory impairment.

  6. Effectiveness of a spontaneous carvacrol nanoemulsion against Salmonella enterica Enteritidis and Escherichia coli O157:H7 on contaminated broccoli and radish seeds.

    Science.gov (United States)

    Landry, Kyle S; Micheli, Sean; McClements, David Julian; McLandsborough, Lynne

    2015-10-01

    The incidence of foodborne illness associated with the consumption of fresh produce has continued to increase over the past decade. Sprouts, such as mung bean, alfalfa, radish, and broccoli, are minimally processed and have been sources for foodborne illness. Currently, a 20,000 ppm calcium hypochlorite soak is recommended for the treatment of sprouting seeds. In this study, the efficacy of an antimicrobial carvacrol nanoemulsion was tested against Salmonella enterica subspecies enterica serovar Enteritidis (ATCC BAA-1045) or EGFP expressing Escherichia coli O157:H7 (ATCC 42895) contaminated sprouting seeds. Antimicrobial treatments were performed by soaking inoculated seeds in nanoemulsions (4000 or 8000 ppm) for 30 or 60 min. Following treatment, surviving cells were determined by performing plate counts and/or Most Probable Number (MPN) enumeration. Treated seeds were sprouted and tested for the presence of pathogens. Treatment successfully inactivated low levels (2 and 3 log CFU/g) of S. Enteritidis and E. coli on radish seeds when soaked for 60 min at concentrations ≥4000 (0.4%) ppm carvacrol. This treatment method was not affective on contaminated broccoli seeds. Total sprout yield was not influenced by any treatments. These results show that carvacrol nanoemulsions may be an alternative treatment method for contaminated radish seeds. Copyright © 2015 Elsevier Ltd. All rights reserved.

  7. Effects of Organic and Waste-Derived Fertilizers on Yield, Nitrogen and Glucosinolate Contents, and Sensory Quality of Broccoli (Brassica oleracea L. var. italica).

    Science.gov (United States)

    Øvsthus, Ingunn; Breland, Tor Arvid; Hagen, Sidsel Fiskaa; Brandt, Kirsten; Wold, Anne-Berit; Bengtsson, Gunnar B; Seljåsen, Randi

    2015-12-23

    Organic vegetable production attempts to pursue multiple goals concerning influence on environment, production resources, and human health. In areas with limited availability of animal manure, there is a need for considering various off-farm nutrient resources for such production. Different organic and waste-derived fertilizer materials were used for broccoli production at two latitudes (58° and 67°) in Norway during two years. The fertilizer materials were applied at two rates of total N (80 and 170 kg ha(-1)) and compared with mineral fertilizer (170 kg ha(-1)) and no fertilizer. Broccoli yield was strongly influenced by fertilizer materials (algae meal < unfertilized control < sheep manure < extruded shrimp shell < anaerobically digested food waste < mineral fertilizer). Yield, but not glucosinolate content, was linearly correlated with estimated potentially plant-available N. However, extruded shrimp shell and mineral NPK fertilizer gave higher glucosinolate contents than sheep manure and no fertilizer. Sensory attributes were less affected by fertilizer material and plant-available N.

  8. Comparative Transcriptome Analysis between Broccoli (Brassica oleracea var. italica) and Wild Cabbage (Brassica macrocarpa Guss.) in Response to Plasmodiophora brassicae during Different Infection Stages.

    Science.gov (United States)

    Zhang, Xiaoli; Liu, Yumei; Fang, Zhiyuan; Li, Zhansheng; Yang, Limei; Zhuang, Mu; Zhang, Yangyong; Lv, Honghao

    2016-01-01

    Clubroot, one of the most devastating diseases to the Brassicaceae family, is caused by the obligate biotrophic pathogen Plasmodiophora brassicae. However, studies of the molecular basis of disease resistance are still poor especially in quantitative resistance. In the present paper, two previously identified genotypes, a clubroot-resistant genotype (wild cabbage, B2013) and a clubroot-susceptible genotype (broccoli, 90196) were inoculated by P. brassicae for 0 (T0), 7 (T7), and 14 (T14) day after inoculation (DAI). Gene expression pattern analysis suggested that response changes in transcript level of two genotypes under P. brassicae infection were mainly activated at the primary stage (T7). Based on the results of DEGs functional enrichments from two infection stages, genes associated with cell wall biosynthesis, glucosinolate biosynthesis, and plant hormone signal transduction showed down-regulated at T14 compared to T7, indicating that defense responses to P. brassicae were induced earlier, and related pathways were repressed at T14. In addition, the genes related to NBS-LRR proteins, SA signal transduction, cell wall and phytoalexins biosynthesis, chitinase, Ca2+ signals and RBOH proteins were mainly up-regulated in B2013 by comparing those of 90196, indicating the pathways of response defense to clubroot were activated in the resistant genotype. This is the first report about comparative transcriptome analysis for broccoli and its wild relative during the different stages of P. brassicae infection and the results should be useful for molecular assisted screening and breeding of clubroot-resistant genotypes.

  9. Laser synthesized super-hydrophobic conducting carbon with broccoli-type morphology as a counter-electrode for dye sensitized solar cells

    Science.gov (United States)

    Gokhale, Rohan; Agarkar, Shruti; Debgupta, Joyashish; Shinde, Deodatta; Lefez, Benoit; Banerjee, Abhik; Jog, Jyoti; More, Mahendra; Hannoyer, Beatrice; Ogale, Satishchandra

    2012-10-01

    A laser photochemical process is introduced to realize superhydrophobic conducting carbon coatings with broccoli-type hierarchical morphology for use as a metal-free counter electrode in a dye sensitized solar cell. The process involves pulsed excimer laser irradiation of a thin layer of liquid haloaromatic organic solvent o-dichlorobenzene (DCB). The coating reflects a carbon nanoparticle-self assembled and process-controlled morphology that yields solar to electric power conversion efficiency of 5.1% as opposed to 6.2% obtained with the conventional Pt-based electrode.A laser photochemical process is introduced to realize superhydrophobic conducting carbon coatings with broccoli-type hierarchical morphology for use as a metal-free counter electrode in a dye sensitized solar cell. The process involves pulsed excimer laser irradiation of a thin layer of liquid haloaromatic organic solvent o-dichlorobenzene (DCB). The coating reflects a carbon nanoparticle-self assembled and process-controlled morphology that yields solar to electric power conversion efficiency of 5.1% as opposed to 6.2% obtained with the conventional Pt-based electrode. Electronic supplementary information (ESI) available: Materials and equipment details, solar cell fabrication protocol, electrolyte spreading time measurement details, XPS spectra, electronic study, film adhesion test detailed analysis and field emission results. See DOI: 10.1039/c2nr32082g

  10. Broccoli ( Brassica oleracea var. italica) sprouts and extracts rich in glucosinolates and isothiocyanates affect cholesterol metabolism and genes involved in lipid homeostasis in hamsters.

    Science.gov (United States)

    Rodríguez-Cantú, Laura N; Gutiérrez-Uribe, Janet A; Arriola-Vucovich, Jennifer; Díaz-De La Garza, Rocio I; Fahey, Jed W; Serna-Saldivar, Sergio O

    2011-02-23

    This study investigated the effects of broccoli sprouts (BS) on sterol and lipid homeostasis in Syrian hamsters with dietary-induced hypercholesterolemia. Treatments included freeze-dried BS containing 2 or 20 μmol of glucoraphanine (BSX, BS10X), glucoraphanine-rich BS extract (GRE), sulforaphane-rich BS extract (SFE), and simvastatin. Each experimental diet was offered to eight animals (male and female) for 7 weeks. Hepatic cholesterol was reduced by BS10X and SFE treatments in all animals. This correlated with a down-regulation of gene expression of sterol regulatory element-binding proteins (SREBP-1 and -2) and fatty acid synthase (FAS) caused by GRE and SFE diets. BS10X caused changes in gene expression in a gender-specific manner; additionally, it increased coprostanol excretion in females. With the same concentration of glucoraphanin, consumption of broccoli sprouts (BS10X) had more marked effects on cholesterol homeostasis than GRE; this finding reinforces the importance of the matrix effects on the bioactivity of functional ingredients.

  11. 2-D zymographic analysis of Broccoli (Brassica oleracea L. var. Italica) florets proteases: follow up of cysteine protease isotypes in the course of post-harvest senescence.

    Science.gov (United States)

    Rossano, Rocco; Larocca, Marilena; Riccio, Paolo

    2011-09-01

    Zymographic analysis of Broccoli florets (Brassica oleracea L. var. Italica) revealed the presence of acidic metallo-proteases, serine proteases and cysteine proteases. Under conditions which were denaturing for the other proteases, the study was restricted to cysteine proteases. 2-D zymography, a technique that combines IEF and zymography was used to show the presence of 11 different cysteine protease spots with molecular mass of 44 and 47-48kDa and pIs ranging between 4.1 and 4.7. pI differences could be ascribed to different degrees of phosphorylation that partly disappeared in the presence of alkaline phosphatase. Post-harvest senescence of Broccoli florets was characterized by decrease in protein and chlorophyll contents and increase of protease activity. In particular, as determined by 2-D zymography, the presence of cysteine protease clearly increased during senescence, a finding that may represent a useful tool for the control of the aging process. Copyright © 2011 Elsevier GmbH. All rights reserved.

  12. Analysis of DNA methyltransferase 3A gene mutations in patients with Philadelphia-negative myeloproliferative neoplasms

    OpenAIRE

    Neda Ketabchi; Mostafa Paridar; Javad Mohammadi-Asl; Alireza Abooali; Maria Kavianpour; Najmaldin Saki

    2017-01-01

    Context: Philadelphia (Ph)-negative myeloproliferative neoplasms (MPNs), including essential thrombocythemia (ET), polycythemia vera (PV), and primary myelofibrosis (PMF) from a group of disorders characterized by dysregulated JAK-STAT functionality, abnormal hematopoiesis, as well as increased production of proliferative cytokines. In addition to JAK2V617F mutation, additional gene alterations that are involved in epigenetic mechanisms, particularly de novo DNA methyltransferase 3A (DNMT3A),...

  13. Identification and Biochemical Characterization of Four Wood-Associated Glucuronoxylan Methyltransferases in Populus

    Science.gov (United States)

    Yuan, Youxi; Teng, Quincy; Zhong, Ruiqin; Ye, Zheng-Hua

    2014-01-01

    Wood is one of the promising bioenergy feedstocks for lignocellulosic biofuel production. Understanding how wood components are synthesized will help us design strategies for better utilization of wood for biofuel production. One of the major wood components is xylan, in which about 10% of xylosyl residues are substituted with glucuronic acid (GlcA) side chains. All the GlcA side chains of xylan in wood of Populus trichocarpa are methylated, which is different from Arabidopsis xylan in which about 60% of GlcA side chains are methylated. Genes responsible for methylation of GlcA side chains in Populus xylan have not been identified. Here, we report genetic and biochemical analyses of four DUF579 domain-containing proteins, PtrGXM1, PtrGXM2, PtrGXM3 and PtrGXM4, from Populus trichocarpa and their roles in GlcA methylation in xylan. The PtrGXM genes were found to be highly expressed in wood-forming cells and their encoded proteins were shown to be localized in the Golgi. When overexpressed in the Arabidopsis gxm1/2/3 triple mutant, PtrGXMs were able to partially complement the mutant phenotypes including defects in glucuronoxylan methyltransferase activity and GlcA methylation in xylan, indicating that PtrGXMs most likely function as glucuronoxylan methyltransferases. Direct evidence was provided by enzymatic analysis of recombinant PtrGXM proteins showing that they possessed a methyltransferase activity capable of transferring the methyl group onto GlcA-substituted xylooligomers. Kinetic analysis showed that PtrGXMs exhibited differential affinities toward the GlcA-substituted xylooligomer acceptor with PtrGXM3 and PtrGXM4 having 10 times higher Km values than PtrGXM1 and PtrGXM2. Together, these findings indicate that PtrGXMs are methyltransferases mediating GlcA methylation in Populus xylan during wood formation. PMID:24523868

  14. Functional Oligomerization of the Saccharomyces cerevisiae Isoprenylcysteine Carboxyl Methyltransferase, Ste14p

    OpenAIRE

    Griggs, Amy M.; Hahne, Kalub; Hrycyna, Christine A.

    2010-01-01

    The isoprenylcysteine carboxyl methyltransferase (Icmt) from Saccharomyces cerevisiae, also designated Ste14p, is a 26-kDa integral membrane protein that contains six transmembrane spanning segments. This protein is localized to the endoplasmic reticulum membrane where it performs the methylation step of the CAAX post-translational processing pathway. Sequence analysis reveals a putative GXXXG dimerization motif located in transmembrane 1 of Ste14p, but it is not known whether Ste14p forms or...

  15. Sequence and structural evolution of the KsgA/Dim1 methyltransferase family

    Directory of Open Access Journals (Sweden)

    Rife Jason P

    2008-10-01

    Full Text Available Abstract Background One of the 60 or so genes conserved in all domains of life is the ksgA/dim1 orthologous group. Enzymes from this family perform the same post-transcriptional nucleotide modification in ribosome biogenesis, irrespective of organism. Despite this common function, divergence has enabled some family members to adopt new and sometimes radically different functions. For example, in S. cerevisiae Dim1 performs two distinct functions in ribosome biogenesis, while human mtTFB is not only an rRNA methyltransferase in the mitochondria but also a mitochondrial transcription factor. Thus, these proteins offer an unprecedented opportunity to study evolutionary aspects of structure/function relationships, especially with respect to our recently published work on the binding mode of a KsgA family member to its 30S subunit substrate. Here we compare and contrast KsgA orthologs from bacteria, eukaryotes, and mitochondria as well as the paralogous ErmC enzyme. Results By using structure and sequence comparisons in concert with a unified ribosome binding model, we have identified regions of the orthologs that are likely related to gains of function beyond the common methyltransferase function. There are core regions common to the entire enzyme class that are associated with ribosome binding, an event required in rRNA methylation activity, and regions that are conserved in subgroups that are presumably related to non-methyltransferase functions. Conclusion The ancient protein KsgA/Dim1 has adapted to cellular roles beyond that of merely an rRNA methyltransferase. These results provide a structural foundation for analysis of multiple aspects of ribosome biogenesis and mitochondrial transcription.

  16. Catecholamine-o-methyltransferase polymorphisms are associated with postoperative pain intensity.

    LENUS (Irish Health Repository)

    Lee, Peter J

    2011-02-01

    single nucleotide polymorphisms (SNPs) in the genes for catecholamine-O-methyltransferase (COMT), μ-opioid receptor and GTP cyclohydrolase (GCH1) have been linked to acute and chronic pain states. COMT polymorphisms are associated with experimental pain sensitivity and a chronic pain state. No such association has been identified perioperatively. We carried out a prospective observational clinical trial to examine associations between these parameters and the development of postoperative pain in patients undergoing third molar (M3) extraction.

  17. A glutamate/aspartate switch controls product specificity in a protein arginine methyltransferase

    Energy Technology Data Exchange (ETDEWEB)

    Debler, Erik W.; Jain, Kanishk; Warmack, Rebeccah A.; Feng, You; Clarke, Steven G.; Blobel, Günter; Stavropoulos, Pete

    2016-02-08

    Trypanosoma brucei PRMT7 (TbPRMT7) is a protein arginine methyltransferase (PRMT) that strictly monomethylates various substrates, thus classifying it as a type III PRMT. However, the molecular basis of its unique product specificity has remained elusive. Here, we present the structure of TbPRMT7 in complex with its cofactor product S-adenosyl-L-homocysteine (AdoHcy) at 2.8 Å resolution and identify a glutamate residue critical for its monomethylation behavior. TbPRMT7 comprises the conserved methyltransferase and β-barrel domains, an N-terminal extension, and a dimerization arm. The active site at the interface of the N-terminal extension, methyltransferase, and β-barrel domains is stabilized by the dimerization arm of the neighboring protomer, providing a structural basis for dimerization as a prerequisite for catalytic activity. Mutagenesis of active-site residues highlights the importance of Glu181, the second of the two invariant glutamate residues of the double E loop that coordinate the target arginine in substrate peptides/proteins and that increase its nucleophilicity. Strikingly, mutation of Glu181 to aspartate converts TbPRMT7 into a type I PRMT, producing asymmetric dimethylarginine (ADMA). Isothermal titration calorimetry (ITC) using a histone H4 peptide showed that the Glu181Asp mutant has markedly increased affinity for monomethylated peptide with respect to the WT, suggesting that the enlarged active site can favorably accommodate monomethylated peptide and provide sufficient space for ADMA formation. In conclusion, these findings yield valuable insights into the product specificity and the catalytic mechanism of protein arginine methyltransferases and have important implications for the rational (re)design of PRMTs.

  18. Regulation of protein stability of DNA methyltransferase 1 by post-translational modifications

    OpenAIRE

    Scott, Anthony; Song, Jing; Ewing, Rob; Wang, Zhenghe

    2014-01-01

    DNA methylation is an important epigenetic mechanism that ensures correct gene expression and maintains genetic stability. DNA methyltransferase 1 (DNMT1) is the primary enzyme that maintains DNA methylation during replication. Dysregulation of DNMT1 is implicated in a variety of diseases. DNMT1 protein stability is regulated via various post-translational modifications, such as acetylation and ubiquitination, but also through protein–protein interactions. These mechanisms ensure DNMT1 is pro...

  19. Mutations in the DNA methyltransferase gene DNMT3A cause an overgrowth syndrome with intellectual disability

    DEFF Research Database (Denmark)

    Tatton-Brown, Katrina; Seal, Sheila; Ruark, Elise

    2014-01-01

    Overgrowth disorders are a heterogeneous group of conditions characterized by increased growth parameters and other variable clinical features such as intellectual disability and facial dysmorphism. To identify new causes of human overgrowth, we performed exome sequencing in ten proband...... and histone binding. Similar mutations were not present in 1,000 UK population controls (13/152 cases versus 0/1,000 controls; P intellectual disability and greater height. DNMT3A encodes a DNA methyltransferase essential for establishing...

  20. Ultrasound assisted extraction of phenolic acids from broccoli vegetable and using sonochemistry for preparation of MOF-5 nanocubes: Comparative study based on micro-dilution broth and plate count method for synergism antibacterial effect.

    Science.gov (United States)

    Pezeshkpour, Vahid; Khosravani, Seyed Abdolmajid; Ghaedi, Mehrorang; Dashtian, Kheibar; Zare, Fahimeh; Sharifi, Asghar; Jannesar, Ramin; Zoladl, Mohammad

    2018-01-01

    The aim of this work was comparison study of dilution and plating method for evaluation of the synergism effect of metal-organic framework nanocubes (MOF-5-NCs) and broccoli extract (Brassica oleracea) on antibacterial activity of standard and clinical Pseudomonas aeruginosa strains. For this purpose, sonochemical synthesis of MOF-5-NCs was performed and it was characterized using XRD, FT-IR, FESEM and EDS techniques. Maceration extraction (ME) and ultrasound assisted extraction (UAE) methods in three different solvents were prepared and applicability of their extracts were compared in some cases such as radical scavenging and antioxidant activity. The HPLC/UV analysis was applied for separation, identification and evaluation of phenolic acids in prepared broccoli extracts. Then, antimicrobial activity of MOF-5NCs and broccoli extract against gram-negative bacteria, Pseudomonas aeruginosa was evaluated by detection of minimal inhibition concentration (MIC), minimal bactericidal concentration (MBC) and zone of inhibition (ZOI). The results of in vitro assays showed that dilution method due to flase estimation of 4% viability percentage which is not logic by consideration of MBC well could not be able to estimate MBC. Therefore, plate count method was performed for precise calculation of MBC. MIC of broccoli extract and MOF-5-NCs on Pseudomonas aeruginosa strains were 7.81mgmL -1 and 3.13mgmL -1 , respectively. Copyright © 2017 Elsevier B.V. All rights reserved.

  1. Lack of involvement of known DNA methyltransferases in familial hydatidiform mole implies the involvement of other factors in establishment of imprinting in the human female germline

    OpenAIRE

    Picton H M; Huntriss J; Hodge D; Judson H; de Vos M; Hayward B E; Sheridan E; Bonthron DT

    2003-01-01

    Abstract Background Differential methylation of the two alleles is a hallmark of imprinted genes. Correspondingly, loss of DNA methyltransferase function results in aberrant imprinting and abnormal post-fertilization development. In the mouse, mutations of the oocyte-specific isoform of the DNA methyltransferase Dnmt1 (Dnmt1o) and of the methyltransferase-like Dnmt3L gene result in specific failures of imprint establishment or maintenance, at multiple loci. We have previously shown in humans ...

  2. Near-isogenic lines for measuring phenotypic effects of DIMBOA-Glc methyltransferase activity in maize.

    Science.gov (United States)

    Mijares, Valeria; Meihls, Lisa N; Jander, Georg; Tzin, Vered

    2013-10-01

    Three O-methyltransferases (BX10a, b, c) catalyze the conversion of 2,4-dihydroxy-7-methoxy-1,4-benzoxazin-3-one glucoside (DIM BOA-Glc) to 2-hydroxy-4,7-dimethoxy-1,4-benzoxazin-3-one glucoside (HDMBOA -Glc) in maize (Zea mays). Variation in benzoxazinoid accumulation and resistance to Rhopalosiphum maidis (corn leaf aphid) was attributed to a natural CACTA family transposon insertion that inactivates Bx10c. Whereas maize inbred line B73 has this transposon insertion, line CM L277 does not. To characterize the phenotypic effects of DIM BOA-Glc methyltransferase activity, we created near-isogenic lines derived from B73 and CM L277 that do or do not contain the transposon insertion. Bx10c inactivation causes high DIM BOA -Glc, low HDMBOA-Glc, and decreased aphid reproduction relative to near-isogenic lines that have a functional Bx10c gene. These results confirm the importance of this locus in maize aphid resistance. The availability of Bx10c near-isogenic lines will facilitate further research on the function of different benzoxazinoids and DIM BOA-Glc methyltransferase activity in maize defense against herbivores and pathogens.

  3. Independent Recruitment of an O-Methyltransferase for Syringyl Lignin Biosynthesis in Selaginella moellendorffii[W

    Science.gov (United States)

    Weng, Jing-Ke; Akiyama, Takuya; Ralph, John; Chapple, Clint

    2011-01-01

    Syringyl lignin, an important component of the secondary cell wall, has traditionally been considered to be a hallmark of angiosperms because ferns and gymnosperms in general lack lignin of this type. Interestingly, syringyl lignin was also detected in Selaginella, a genus that represents an extant lineage of the most basal of the vascular plants, the lycophytes. In angiosperms, syringyl lignin biosynthesis requires the activity of ferulate 5-hydroxylase (F5H), a cytochrome P450-dependent monooxygenase, and caffeic acid/5-hydroxyferulic acid O-methyltransferase (COMT). Together, these two enzymes divert metabolic flux from the biosynthesis of guaiacyl lignin, a lignin type common to all vascular plants, toward syringyl lignin. Selaginella has independently evolved an alternative lignin biosynthetic pathway in which syringyl subunits are directly derived from the precursors of p-hydroxyphenyl lignin, through the action of a dual specificity phenylpropanoid meta-hydroxylase, Sm F5H. Here, we report the characterization of an O-methyltransferase from Selaginella moellendorffii, COMT, the coding sequence of which is clustered together with F5H at the adjacent genomic locus. COMT is a bifunctional phenylpropanoid O-methyltransferase that can methylate phenylpropanoid meta-hydroxyls at both the 3- and 5-position and function in concert with F5H in syringyl lignin biosynthesis in S. moellendorffii. Phylogenetic analysis reveals that Sm COMT, like F5H, evolved independently from its angiosperm counterparts. PMID:21742988

  4. Identification of a Pseudomonas aeruginosa PAO1 DNA Methyltransferase, Its Targets, and Physiological Roles

    Directory of Open Access Journals (Sweden)

    Sebastian Doberenz

    2017-02-01

    Full Text Available DNA methylation is widespread among prokaryotes, and most DNA methylation reactions are catalyzed by adenine DNA methyltransferases, which are part of restriction-modification (R-M systems. R-M systems are known for their role in the defense against foreign DNA; however, DNA methyltransferases also play functional roles in gene regulation. In this study, we used single-molecule real-time (SMRT sequencing to uncover the genome-wide DNA methylation pattern in the opportunistic pathogen Pseudomonas aeruginosa PAO1. We identified a conserved sequence motif targeted by an adenine methyltransferase of a type I R-M system and quantified the presence of N6-methyladenine using liquid chromatography-tandem mass spectrometry (LC-MS/MS. Changes in the PAO1 methylation status were dependent on growth conditions and affected P. aeruginosa pathogenicity in a Galleria mellonella infection model. Furthermore, we found that methylated motifs in promoter regions led to shifts in sense and antisense gene expression, emphasizing the role of enzymatic DNA methylation as an epigenetic control of phenotypic traits in P. aeruginosa. Since the DNA methylation enzymes are not encoded in the core genome, our findings illustrate how the acquisition of accessory genes can shape the global P. aeruginosa transcriptome and thus may facilitate adaptation to new and challenging habitats.

  5. Transcriptome profiling of Set5 and Set1 methyltransferases: Tools for visualization of gene expression

    Directory of Open Access Journals (Sweden)

    Glòria Mas Martín

    2014-12-01

    Full Text Available Cells regulate transcription by coordinating the activities of multiple histone modifying complexes. We recently identified the yeast histone H4 methyltransferase Set5 and discovered functional overlap with the histone H3 methyltransferase Set1 in gene expression. Specifically, using next-generation RNA sequencing (RNA-Seq, we found that Set5 and Set1 function synergistically to regulate specific transcriptional programs at subtelomeres and transposable elements. Here we provide a comprehensive description of the methodology and analysis tools corresponding to the data deposited in NCBI's Gene Expression Omnibus (GEO under the accession number GSE52086. This data complements the experimental methods described in Mas Martín G et al. (2014 and provides the means to explore the cooperative functions of histone H3 and H4 methyltransferases in the regulation of transcription. Furthermore, a fully annotated R code is included to enable researchers to use the following computational tools: comparison of significant differential expression (SDE profiles; gene ontology enrichment of SDE; and enrichment of SDE relative to chromosomal features, such as centromeres, telomeres, and transposable elements. Overall, we present a bioinformatics platform that can be generally implemented for similar analyses with different datasets and in different organisms.

  6. Kinetic analysis of Yersinia pestis DNA adenine methyltransferase activity using a hemimethylated molecular break light oligonucleotide.

    Directory of Open Access Journals (Sweden)

    Robert J Wood

    Full Text Available BACKGROUND: DNA adenine methylation plays an important role in several critical bacterial processes including mismatch repair, the timing of DNA replication and the transcriptional control of gene expression. The dependence of bacterial virulence on DNA adenine methyltransferase (Dam has led to the proposal that selective Dam inhibitors might function as broad spectrum antibiotics. METHODOLOGY/PRINCIPAL FINDINGS: Herein we report the expression and purification of Yersinia pestis Dam and the development of a continuous fluorescence based assay for DNA adenine methyltransferase activity that is suitable for determining the kinetic parameters of the enzyme and for high throughput screening against potential Dam inhibitors. The assay utilised a hemimethylated break light oligonucleotide substrate containing a GATC methylation site. When this substrate was fully methylated by Dam, it became a substrate for the restriction enzyme DpnI, resulting in separation of fluorophore (fluorescein and quencher (dabcyl and therefore an increase in fluorescence. The assays were monitored in real time using a fluorescence microplate reader in 96 well format and were used for the kinetic characterisation of Yersinia pestis Dam, its substrates and the known Dam inhibitor, S-adenosylhomocysteine. The assay has been validated for high throughput screening, giving a Z-factor of 0.71+/-0.07 indicating that it is a sensitive assay for the identification of inhibitors. CONCLUSIONS/SIGNIFICANCE: The assay is therefore suitable for high throughput screening for inhibitors of DNA adenine methyltransferases and the kinetic characterisation of the inhibition.

  7. miR-29 Represses the Activities of DNA Methyltransferases and DNA Demethylases

    Directory of Open Access Journals (Sweden)

    Izuho Hatada

    2013-07-01

    Full Text Available Members of the microRNA-29 (miR-29 family directly target the DNA methyltransferases, DNMT3A and DNMT3B. Disturbances in the expression levels of miR-29 have been linked to tumorigenesis and tumor aggressiveness. Members of the miR-29 family are currently thought to repress DNA methylation and suppress tumorigenesis by protecting against de novo methylation. Here, we report that members of the miR-29 family repress the activities of DNA methyltransferases and DNA demethylases, which have opposing roles in control of DNA methylation status. Members of the miR-29 family directly inhibited DNA methyltransferases and two major factors involved in DNA demethylation, namely tet methylcytosine dioxygenase 1 (TET1 and thymine DNA glycosylase (TDG. Overexpression of miR-29 upregulated the global DNA methylation level in some cancer cells and downregulated DNA methylation in other cancer cells, suggesting that miR-29 suppresses tumorigenesis by protecting against changes in the existing DNA methylation status rather than by preventing de novo methylation of DNA.

  8. Structural insights into mechanisms of the small RNA methyltransferase HEN1

    Energy Technology Data Exchange (ETDEWEB)

    Huang, Ying; Ji, Lijuan; Huang, Qichen; Vassylyev, Dmitry G.; Chen, Xuemei; Ma, Jin-Biao; (UAB); (UCR)

    2010-02-22

    RNA silencing is a conserved regulatory mechanism in fungi, plants and animals that regulates gene expression and defence against viruses and transgenes. Small silencing RNAs of {approx}20-30 nucleotides and their associated effector proteins, the Argonaute family proteins, are the central components in RNA silencing. A subset of small RNAs, such as microRNAs and small interfering RNAs (siRNAs) in plants, Piwi-interacting RNAs in animals and siRNAs in Drosophila, requires an additional crucial step for their maturation; that is, 2'-O-methylation on the 3' terminal nucleotide. A conserved S-adenosyl-L-methionine-dependent RNA methyltransferase, HUA ENHANCER 1 (HEN1), and its homologues are responsible for this specific modification. Here we report the 3.1 {angstrom} crystal structure of full-length HEN1 from Arabidopsis in complex with a 22-nucleotide small RNA duplex and cofactor product S-adenosyl-L-homocysteine. Highly cooperative recognition of the small RNA substrate by multiple RNA binding domains and the methyltransferase domain in HEN1 measures the length of the RNA duplex and determines the substrate specificity. Metal ion coordination by both 2' and 3' hydroxyls on the 3'-terminal nucleotide and four invariant residues in the active site of the methyltransferase domain suggests a novel Mg{sup 2+}-dependent 2'-O-methylation mechanism.

  9. MtrA of the sodium ion pumping methyltransferase binds cobalamin in a unique mode.

    Science.gov (United States)

    Wagner, Tristan; Ermler, Ulrich; Shima, Seigo

    2016-06-21

    In the three domains of life, vitamin B12 (cobalamin) is primarily used in methyltransferase and isomerase reactions. The methyltransferase complex MtrA-H of methanogenic archaea has a key function in energy conservation by catalysing the methyl transfer from methyl-tetrahydromethanopterin to coenzyme M and its coupling with sodium-ion translocation. The cobalamin-binding subunit MtrA is not homologous to any known B12-binding proteins and is proposed as the motor of the sodium-ion pump. Here, we present crystal structures of the soluble domain of the membrane-associated MtrA from Methanocaldococcus jannaschii and the cytoplasmic MtrA homologue/cobalamin complex from Methanothermus fervidus. The MtrA fold corresponds to the Rossmann-type α/β fold, which is also found in many cobalamin-containing proteins. Surprisingly, the cobalamin-binding site of MtrA differed greatly from all the other cobalamin-binding sites. Nevertheless, the hydrogen-bond linkage at the lower axial-ligand site of cobalt was equivalently constructed to that found in other methyltransferases and mutases. A distinct polypeptide segment fixed through the hydrogen-bond linkage in the relaxed Co(III) state might be involved in propagating the energy released upon corrinoid demethylation to the sodium-translocation site by a conformational change.

  10. Identification of des-methyl-DIF-1 methyltransferase in Dictyostelium purpureum.

    Science.gov (United States)

    Motohashi, Kazunori A; Morita, Naoki; Kato, Atsushi; Saito, Tamao

    2012-01-01

    The signalling molecule 1-(3,5-dichloro-2,6-dihydroxy-4-methoxyphenyl) hexan-1-one (DIF-1) is required for differentiation and pattern formation in Dictyostelium discoideum development. DIF-1 is synthesized by three enzymes, a hybrid polyketide synthase, a flavin-dependent halogenase, and a des-methyl-DIF-1 methyltransferase. The genome data on the related species D. purpureum are now public. Using this genome information, des-methyl-DIF-1 methyltransferase of D. purpureum was identified, and was named Dp dmtA. Overexpression of Dp dmtA complemented the defects in basal disc formation and lower cup formation in a dmtA knock-out mutant of D. discoideum. This indicates that Dp dmtA has the same function as D. discoideum dmtA and compensates for loss of the dmtA gene in the D. discoideum dmtA mutant. The materials released in the medium by D. purpureum contained stalk-inducing activity with the same retention time as that of DIF-1 in HPLC fractionation. This indicates that the stalk-inducing signal of DIF-1 and des-methyl-DIF-1 methyltransferase are conserved in D. purpureum.

  11. Mechanistic insights into small RNA recognition and modification by the HEN1 methyltransferase.

    Science.gov (United States)

    Plotnikova, Alexandra; Baranauskė, Simona; Osipenko, Aleksandr; Klimašauskas, Saulius; Vilkaitis, Giedrius

    2013-07-15

    The HEN1 methyltransferase from Arabidopsis thaliana modifies the 3'-terminal nucleotides of small regulatory RNAs. Although it is one of the best characterized members of the 2'-O-methyltransferase family, many aspects of its interactions with the cofactor and substrate RNA remained unresolved. To better understand the substrate interactions and contributions of individual steps during HEN1 catalysis, we studied the binding and methylation kinetics of the enzyme using a series of unmethylated, hemimethylated and doubly methylated miRNA and siRNA substrates. The present study shows that HEN1 specifically binds double-stranded unmethylated or hemimethylated miR173/miR173* substrates with a subnanomolar affinity in a cofactor-dependent manner. Kinetic studies under single turnover and pre-steady state conditions in combination with isotope partitioning analysis showed that the binary HEN1-miRNA/miRNA* complex is catalytically competent; however, successive methylation of the two strands in a RNA duplex occurs in a non-processive (distributive) manner. We also find that the observed moderate methylation strand preference is largely exerted at the RNA-binding step and is fairly independent of the nature of the 3'-terminal nucleobase, but shows some dependency on proximal nucleotide mispairs. The results of the present study thus provide novel insights into the mechanism of RNA recognition and modification by a representative small RNA 2'-O-methyltransferase.

  12. The activity of a yeast Family 16 methyltransferase, Efm2, is affected by a conserved tryptophan and its N-terminal region.

    Science.gov (United States)

    Hamey, Joshua J; Hart-Smith, Gene; Erce, Melissa A; Wilkins, Marc R

    2016-12-01

    The Family 16 methyltransferases are a group of eukaryotic nonhistone protein methyltransferases. Sixteen of these have recently been described in yeast and human, but little is known about their sequence and structural features. Here we investigate one of these methyltransferases, Saccharomyces cerevisiae elongation factor methyltransferase 2 (Efm2), by site-directed mutagenesis and truncation. We show that an active site-associated tryptophan, invariant in Family 16 methyltransferases and at position 222 in Efm2, is important for methyltransferase activity. A second highly conserved tryptophan, at position 318 in Efm2, is likely involved in S-adenosyl methionine binding but is of lesser consequence for catalysis. By truncation analysis, we show that the N-terminal 50-200 amino acids of Efm2 are critical for its methyltransferase activity. As N-terminal regions are variable among Family 16 methyltransferases, this suggests a possible role in determining substrate specificity. This is consistent with recently solved structures that show the core of Family 16 methyltransferases to be near-identical but the N termini to be structurally quite different. Finally, we show that Efm2 can exist as an oligomer but that its N terminus is not necessary for oligomerisation to occur.

  13. Mapping of Post-translational Modifications of Transition Proteins, TP1 and TP2, and Identification of Protein Arginine Methyltransferase 4 and Lysine Methyltransferase 7 as Methyltransferase for TP2*

    Science.gov (United States)

    Gupta, Nikhil; Madapura, M. Pradeepa; Bhat, U. Anayat; Rao, M. R. Satyanarayana

    2015-01-01

    In a unique global chromatin remodeling process during mammalian spermiogenesis, 90% of the nucleosomal histones are replaced by testis-specific transition proteins, TP1, TP2, and TP4. These proteins are further substituted by sperm-specific protamines, P1 and P2, to form a highly condensed sperm chromatin. In spermatozoa, a small proportion of chromatin, which ranges from 1 to 10% in mammals, retains the nucleosomal architecture and is implicated to play a role in transgenerational inheritance. However, there is still no mechanistic understanding of the interaction of chromatin machinery with histones and transition proteins, which facilitate this selective histone replacement from chromatin. Here, we report the identification of 16 and 19 novel post-translational modifications on rat endogenous transition proteins, TP1 and TP2, respectively, by mass spectrometry. By in vitro assays and mutational analysis, we demonstrate that protein arginine methyltransferase PRMT4 (CARM1) methylates TP2 at Arg71, Arg75, and Arg92 residues, and lysine methyltransferase KMT7 (Set9) methylates TP2 at Lys88 and Lys91 residues. Further studies with modification-specific antibodies that recognize TP2K88me1 and TP2R92me1 modifications showed that they appear in elongating to condensing spermatids and predominantly associated with the chromatin-bound TP2. This work establishes the repertoire of post-translational modifications that occur on TP1 and TP2, which may play a significant role in various chromatin-templated events during spermiogenesis and in the establishment of the sperm epigenome. PMID:25818198

  14. Mapping of Post-translational Modifications of Transition Proteins, TP1 and TP2, and Identification of Protein Arginine Methyltransferase 4 and Lysine Methyltransferase 7 as Methyltransferase for TP2.

    Science.gov (United States)

    Gupta, Nikhil; Madapura, M Pradeepa; Bhat, U Anayat; Rao, M R Satyanarayana

    2015-05-08

    In a unique global chromatin remodeling process during mammalian spermiogenesis, 90% of the nucleosomal histones are replaced by testis-specific transition proteins, TP1, TP2, and TP4. These proteins are further substituted by sperm-specific protamines, P1 and P2, to form a highly condensed sperm chromatin. In spermatozoa, a small proportion of chromatin, which ranges from 1 to 10% in mammals, retains the nucleosomal architecture and is implicated to play a role in transgenerational inheritance. However, there is still no mechanistic understanding of the interaction of chromatin machinery with histones and transition proteins, which facilitate this selective histone replacement from chromatin. Here, we report the identification of 16 and 19 novel post-translational modifications on rat endogenous transition proteins, TP1 and TP2, respectively, by mass spectrometry. By in vitro assays and mutational analysis, we demonstrate that protein arginine methyltransferase PRMT4 (CARM1) methylates TP2 at Arg(71), Arg(75), and Arg(92) residues, and lysine methyltransferase KMT7 (Set9) methylates TP2 at Lys(88) and Lys(91) residues. Further studies with modification-specific antibodies that recognize TP2K88me1 and TP2R92me1 modifications showed that they appear in elongating to condensing spermatids and predominantly associated with the chromatin-bound TP2. This work establishes the repertoire of post-translational modifications that occur on TP1 and TP2, which may play a significant role in various chromatin-templated events during spermiogenesis and in the establishment of the sperm epigenome. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  15. The use of biodegradable mulch for tomato and broccoli production: Crop yield and quality, mulch deterioration, and growers' perceptions

    Science.gov (United States)

    Cowan, Jeremy Scott

    Biodegradable mulch may offer the benefits of polyethylene mulch for crop production with the added benefit of biodegradability. Four studies were carried out in Mount Vernon, WA to evaluate biodegradable mulch for tomato (Solanum lycopersicum L.) and broccoli (Brassica oleracea var. italica) production. The first study compared four biodegradable mulch treatments: BioAgri, BioTelo, WeedGuardPlus (cellulose product), and SB-PLA-10/11/12 (experimental, non-woven fabric), to polyethylene mulch and bare ground in high tunnels and in the open field for tomato yield and fruit quality over three growing seasons. Biodegradable plastic films produced yields and fruit quality comparable to polyethylene. Moreover, high tunnels increased total and marketable fruit weight five and eight times, respectively, compared to the open field. The second study quantified relationships among visual assessment parameters and mulch mechanical properties. Visual assessments and mechanical property tests of polyethylene, BioAgri, BioTelo, WeedGuardPlus, and SB-PLA-10/11/12, were made over three growing seasons. Regression analyses found the strongest relationship overall (r2 = 0.41) to be between the percent of initial breaking force in the machine direction and log 10 of percent visual deterioration. However, evaluating mulch products individually and increasing sample frequency are recommended for future research. The third study evaluated three biodegradable mulch products, BioAgri, Crown 1, and SB-PLA-11, after soil-incorporation. The average area of recovered mulch fragments decreased for all mulch products over time. The number of mulch fragments initially increased for all mulch products, with the greatest number of Crown 1 and BioAgri fragments recovered 132 and 299 days after incorporation, respectively. At 397 days after soil-incorporation, the total area of recovered fragments of Crown 1 and BioAgri was 0% and 34% of the theoretical maximum area, respectively. The fourth study

  16. The catechol-O-methyltransferase inhibitory potential of Z-vallesiachotamine by in silicoand in vitro approaches

    Directory of Open Access Journals (Sweden)

    Carolina dos Santos Passos

    Full Text Available AbstractZ-Vallesiachotamine is a monoterpene indole alkaloid that has a β-N-acrylate group in its structure. This class of compounds has already been described in different Psychotriaspecies. Our research group observed that E/Z-vallesiachotamine exhibits a multifunctional feature, being able to inhibit targets related to neurodegeneration, such as monoamine oxidase A, sirtuins 1 and 2, and butyrylcholinesterase enzymes. Aiming at better characterizing the multifunctional profile of this compound, its effect on cathecol-O-methyltransferase activity was investigated. The cathecol-O-methyltransferase activity was evaluated in vitro by a fluorescence-based method, using S-(5′-adenosyl-l-methionine as methyl donor and aesculetin as substrate. The assay optimization was performed varying the concentrations of methyl donor (S-(5′-adenosyl-l-methionine and enzyme. It was observed that the highest concentrations of both factors (2.25 U of the enzyme and 100 µM of S-(5′-adenosyl-l-methionine afforded the more reproducible results. The in vitro assay demonstrated that Z-vallesiachotamine was able to inhibit the cathecol-O-methyltransferase activity with an IC50 close to 200 µM. Molecular docking studies indicated that Z-vallesiachotamine can bind the catechol pocket of catechol-O-methyltransferase enzyme. The present work demonstrated for the first time the inhibitory properties of Z-vallesiachotamine on cathecol-O-methyltransferase enzyme, affording additional evidence regarding its multifunctional effects in targets related to neurodegenerative diseases.

  17. Intrinsic resistance to aminoglycosides in Enterococcus faecium is conferred by the 16S rRNA m5C1404-specific methyltransferase EfmM

    DEFF Research Database (Denmark)

    Galimand, Marc; Schmitt, Emmanuelle; Panvert, Michel

    2011-01-01

    structure determination of EfmM at 2.28 Å resolution reveals an N-terminal domain connected to a central methyltransferase domain that is linked by a flexible lysine-rich region to two C-terminal subdomains. Mutagenesis of the methyltransferase domain established that two cysteines at specific tertiary...

  18. Insights into the structure, function and evolution of the radical-SAM 23S rRNA methyltransferase Cfr that confers antibiotic resistance in bacteria

    DEFF Research Database (Denmark)

    Karminska, K. H.; Purta, E.; Hansen, L .H.

    2010-01-01

    The Cfr methyltransferase confers combined resistance to five classes of antibiotics that bind to the peptidyl tranferase center of bacterial ribosomes by catalyzing methylation of the C-8 position of 23S rRNA nucleotide A2503. The same nucleotide is targeted by the housekeeping methyltransferase...

  19. Crystal structure of the Escherichia coli 23S rRNA:m5C methyltransferase RlmI (YccW) reveals evolutionary links between RNA modification enzymes

    DEFF Research Database (Denmark)

    Sunita, S; Tkaczuk, Karolina L; Purta, Elzbieta

    2008-01-01

    Methylation is the most common RNA modification in the three domains of life. Transfer of the methyl group from S-adenosyl-l-methionine (AdoMet) to specific atoms of RNA nucleotides is catalyzed by methyltransferase (MTase) enzymes. The rRNA MTase RlmI (rRNA large subunit methyltransferase gene I...

  20. Effects of Trichoderma harzianum Rifai over Plasmodiophora brassicae Woronin in broccoli, in Escagüey, municipality of Rangel, Mérida state

    Directory of Open Access Journals (Sweden)

    Mirna Labrador Morales

    2014-04-01

    Full Text Available The effectiveness of Trichoderma harzianum in suppressing clubroot of brassicas, which is caused by Plasmodiophora brassicae, was tested on broccoli (Brassica oleracea var. italica Plenck, in field conditions, in Escagüey, municipality of Rangel, Merida State, Venezuela. The experiment showed that the applications of a biopesticide based on this antagonist had a significant effect on the percentage of health plants and the crop yields, in dependence of the dose used. In addition, the relation benefits/cost was also favorable. These results showed that, for these particular conditions, P. brassicae is an adequate biological alternative to control the clubroot of brassicas, no aggressive to environment and human beings, useful for the transition phase toward a sustainable agriculture, without chemical pesticides.

  1. Effects of Trichoderma harzianum Rifai over Plasmodiophora brassicae Woronin in broccoli, in Escagüey, municipality of Rangel, Mérida State

    Directory of Open Access Journals (Sweden)

    Mirna Labrador Morales

    2016-03-01

    Full Text Available The effectiveness of Trichoderma harzianum in suppressing clubroot of brassicas, which is caused by Plasmodiophora brassicae, was tested on broccoli (Brassica oleracea var. italica Plenck, in field conditions, in Escagüey, municipality of Rangel, Merida State, Venezuela. The experiment showed that the applications of a biopesticide based on this antagonist had a significant effect on the percentage of health plants and the crop yields, in dependence of the dose used. In addition, the relation benefits/cost was also favorable. These results showed that, for these particular conditions, P. brassicae is an adequate biological alternative to control the clubroot of brassicas, no aggressive to environment and human beings, useful for the transition phase toward a sustainable agriculture, without chemical pesticides.

  2. Impact of Synergistic Association of ZnO-Nanorods and Symbiotic Fungus Piriformospora indica DSM 11827 on Brassica oleracea var. botrytis (Broccoli).

    Science.gov (United States)

    Singhal, Uma; Khanuja, Manika; Prasad, Ram; Varma, Ajit

    2017-01-01

    In the present work, novel nanotool called 'nano-embedded fungus' formed by impact of synergistic association of ZnO-nanorods and fungus Piriformospora indica DSM 11827, for growth of Brassica oleracea var. botrytis (Broccoli) is reported. ZnO-nanorods were synthesized by mechanical assisted thermal decomposition process and characterized by scanning electron microscopy (SEM) for morphology, X-ray diffraction for structural studies and UV-vis absorption spectroscopy for band gap determination. Nanoembedded fungus is prepared by optimizing ZnO-nanorods concentration (500 ppm) which resulted in the increased biomass of P. indica, as confirmed by dry weight method, spore count, spread plate and microscopy techniques viz. SEM and confocal microscopy. Enhancement in B. oleracea var. botrytis is reported on treatment with nanoembedded fungus. According to the authors, this is the first holistic study focusing on the impact of ZnO-nanorods in the enhancement of fungal symbiont for enhanced biomass productivity of B. oleracea plant.

  3. Vigor de sementes de brócolos submetidas a coberturas biodegradáveis e micronutrientes Vigor of broccoli seeds submitted to biodegradable coatings and micronutrients

    Directory of Open Access Journals (Sweden)

    Juliana A. Batista

    2005-07-01

    Full Text Available O desenvolvimento e a aplicação de filmes ou coberturas biodegradáveis na horticultura são técnicas praticadas há vários anos, que visam promover melhoria na qualidade do produto. no presente trabalho desenvolveu-se e caracterizou-se biofilmes de pectina, gelatina e ácidos graxos e verificou-se sua eficiência como cobertura para sementes de brócolos (Brassica oleracea L. var. italica. Os filmes foram caracterizados quanto à solubilidade em água. A contagem e a observação de plantas de brócolos, a partir da germinação de sementes cobertas e não cobertas foi realizada em média a cada três dias, aos 27 dias fêz-se a quantificação da matéria fresca e seca. Os filmes de pectina e ácido esteárico foram 100% solúveis em água, enquanto os elaborados com pectina e gelatina (1/1 foram apenas 18%. A aplicação das coberturas filmogênicas nas sementes de brócolos não afetou a emergência das plantas, demonstrando assim sua potencialidade para uso comercial. Os fertilizantes molibdato de sódio (0,2 mg L-1 e ácido bórico (1mg L-1, aplicados juntamente com os biofilmes, também não afetaram a emergência e o desenvolvimento das plantas.The development and application of biodegradable coatings and films have been used for many years in order to improve quality of the coated or packed product. The development and characterization of pectin and pectin/gelatin-based biofilms with fatty acids was evaluated and verified their efficiency as coatings to broccoli seeds (Brassica oleracea L. var. italica. The solubility in water of the films was determined. The counting and visual observations of the germinated broccoli seeds were done each three days and fresh and dried weight of the plants were determined at the end of the experiment. Pectin-based films with stearic acid were 100% soluble in water and composite films of pectin and gelatin were 18%. The application of the biodegradable coatings on broccoli seeds had no effect on the

  4. Proposed Method for Estimating Health-Promoting Glucosinolates and Hydrolysis Products in Broccoli (Brassica oleracea var. italica) Using Relative Transcript Abundance.

    Science.gov (United States)

    Becker, Talon M; Jeffery, Elizabeth H; Juvik, John A

    2017-01-18

    Due to the importance of glucosinolates and their hydrolysis products in human nutrition and plant defense, optimizing the content of these compounds is a frequent breeding objective for Brassica crops. Toward this goal, we investigated the feasibility of using models built from relative transcript abundance data for the prediction of glucosinolate and hydrolysis product concentrations in broccoli. We report that predictive models explaining at least 50% of the variation for a number of glucosinolates and their hydrolysis products can be built for prediction within the same season, but prediction accuracy decreased when using models built from one season's data for prediction of an opposing season. This method of phytochemical profile prediction could potentially allow for lower phytochemical phenotyping costs and larger breeding populations. This, in turn, could improve selection efficiency for phase II induction potential, a type of chemopreventive bioactivity, by allowing for the quick and relatively cheap content estimation of phytochemicals known to influence the trait.

  5. Characterization of biomasses, concentrates, and permeates of dried powder of Kombucha fermentation of spinach (Amaranthus sp.) and broccoli (Brassica oleracea) with membrane microfiltration and freeze drying techniques for natural sources of folic acid

    Science.gov (United States)

    Nugraha, Tutun; Susilowati, Agustine; Aspiyanto, Lotulung, Puspa Dewi; Maryati, Yati

    2017-11-01

    Fermentation of spinach (Amaranthus sp) and Broccoli (Brassica oleracea) using Kombucha Culture has been shown to produce biomass that has the potential to become natural sources of folic acid. To produce the materials, following the fermentation, the biomass was filtered using membrane microfiltration (0.15 µm) at a pressure of 40 psia, at room temperature, yielding the concentrate and the permeate fractions. Following this step, freeze drying process was done on the biomass feeds, as well as on the concentrate and permeate fractions. For the freeze drying stage, the samples were frozen, and the condenser was kept at -50°C for 40 hours, while the pressure in the chamber was set at 200 Pa. Freeze drying results showed that the final products, have differences in compositions, as well as differences in the dominat monomers of folates. After water content was driven out, freeze drying increased the concentrations of folic acid in the dried products, and was found to be the highest in the concentrate fractions. Freeze drying has been shown to be capable of protecting the folates from heat and oxidative damages that typicaly occur with other types of drying. The final freeze dried concentrates of fermentation of spinach and broccoli were found to contain folic acid at 2531.88 µg/mL and 1626.94 µg/mL, total solids at 87.23% and 88.65 %, total sugar at 22.66 µg/mL and 25.13 µg/mL, total reducing sugar at 34.46 mg/mL and 15.22 mg/mL, as well as disolved protein concentrations at 0.93 mg/mL and 1.45 mg/mL. Liquid Chromatography Mass Spectometry (LC-MS) identification of the folates in the freeze dried concentrates of fermented spinach and broccoli was done using folic acid and glutamic acid standard solutions as the reference materials. The results showed the presence of folic acid and showed that the dominant monomers of molecules of folates with molecular weights of 441.44 Da. and 441.54 Da. for spinach and broccoli respectively. Moreover, the monomers of glutamic

  6. Arabidopsis DNA methyltransferase AtDNMT2 associates with histone deacetylase AtHD2s activity

    Energy Technology Data Exchange (ETDEWEB)

    Song, Yuan [Key Laboratory of Arid and Grassland Agroecology, Ministry of Education, School of Life Science, Lanzhou University, Lanzhou 730000 (China); Southern Crop Protection and Food Research Centre, Agriculture and Agri-Food Canada, 1391 Sandford Street, London, ON, Canada N5V4T3 (Canada); Wu, Keqiang [Institute of Plant Biology, National Taiwan University, Taipei 106, Taiwan (China); Dhaubhadel, Sangeeta [Southern Crop Protection and Food Research Centre, Agriculture and Agri-Food Canada, 1391 Sandford Street, London, ON, Canada N5V4T3 (Canada); An, Lizhe, E-mail: lizhean@lzu.edu.cn [Key Laboratory of Arid and Grassland Agroecology, Ministry of Education, School of Life Science, Lanzhou University, Lanzhou 730000 (China); Tian, Lining, E-mail: tianl@agr.gc.ca [Southern Crop Protection and Food Research Centre, Agriculture and Agri-Food Canada, 1391 Sandford Street, London, ON, Canada N5V4T3 (Canada)

    2010-05-28

    DNA methyltransferase2 (DNMT2) is always deemed to be enigmatic, because it contains highly conserved DNA methyltransferase motifs but lacks the DNA methylation catalytic capability. Here we show that Arabidopsis DNA methyltransferase2 (AtDNMT2) is localized in nucleus and associates with histone deacetylation. Bimolecular fluorescence complementation and pull-down assays show AtDNMT2 interacts with type-2 histone deacetylases (AtHD2s), a unique type of histone deacetylase family in plants. Through analyzing the expression of AtDNMT2: ss-glucuronidase (GUS) fusion protein, we demonstrate that AtDNMT2 has the ability to repress gene expression at transcription level. Meanwhile, the expression of AtDNMT2 gene is altered in athd2c mutant plants. We propose that AtDNMT2 possibly involves in the activity of histone deacetylation and plant epigenetic regulatory network.

  7. Epigenetic changes of Arabidopsis genome associated with altered DNA methyltransferase and demethylase expressions after gamma irradiation

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Ji Eun; Cho, Eun Ju; Kim, Ji Hong; Chung, Byung Yeoup; Kim, Jin Hong [Korea Atomic Energy Research Institute, Daejeon (Korea, Republic of)

    2012-05-15

    DNA methylation at carbon 5 of cytosines is a hall mark of epigenetic inactivation and heterochromatin in both plants and mammals. In Arabidopsis, DNA methylation has two roles that protect the genome from selfish DNA elements and regulate gene expression. Plant genome has three types of DNA methyltransferase, METHYLTRANSFERASE 1 (MET1), DOMAINREARRANGED METHYLASE (DRM) and CHROMOMETHYLASE 3 (CMT3) that are capable of methylating CG, CHG (where H is A, T, or C) and CHH sites, respectively. MET1 is a maintenance DNA methyltransferase that controls CG methylation. Two members of the DRM family, DRM1 and DRM2, are responsible for de novo methylation of CG, CHG, and CHH sites but show a preference for CHH sites. Finally, CMT3 principally carries out CHG methylation and is involved in both de novo methylation and maintenance. Alternatively, active DNA demethylation may occur through the glycosylase activity by removing the methylcytosines from DNA. It may have essential roles in preventing transcriptional silencing of transgenes and endogenous genes and in activating the expression of imprinted genes. DNA demetylation in Arabidopsis is mediated by the DEMETER (DME) family of bifunctional DNA glycosylase. Three targets of DME are MEA (MEDEA), FWA (FLOWERING WAGENINGEN), and FIS2 (FERTILIZATION INDEPENDENT SEED 2). The DME family contains DEMETER-LIKE 2 (DML2), DML3, and REPRESSOR OF SILENING 1 (ROS1). DNA demetylation by ROS1, DML2, and DML3 protect the hypermethylation of specific genome loci. ROS1 is necessary to suppress the promoter methylation and the silencing of endogenous genes. In contrast, the function of DML2 and DML3 has not been reported. Several recent studies have suggested that epigenetic alterations such as change in DNA methylation and histone modification should be caused in plant genomes upon exposure to ionizing radiation. However, there is a lack of data exploring the underlying mechanisms. Therefore, the present study aims to characterize and

  8. Water balance and N-metabolism in broccoli (Brassica oleracea L. var. Italica) plants depending on nitrogen source under salt stress and elevated CO2.

    Science.gov (United States)

    Zaghdoud, Chokri; Carvajal, Micaela; Ferchichi, Ali; Del Carmen Martínez-Ballesta, María

    2016-11-15

    Elevated [CO2] and salinity in the soils are considered part of the effects of future environmental conditions in arid and semi-arid areas. While it is known that soil salinization decreases plant growth, an increased atmospheric [CO2] may ameliorate the negative effects of salt stress. However, there is a lack of information about the form in which inorganic nitrogen source may influence plant performance under both conditions. Single factor responses and the interactive effects of two [CO2] (380 and 800ppm), three different NO3(-)/NH4(+) ratios in the nutrient solution (100/0, 50/50 and 0/100, with a total N concentration of 3.5mM) and two NaCl concentrations (0 and 80mM) on growth, leaf gas exchange parameters in relation to root hydraulic conductance and N-assimilating enzymes of broccoli (Brassica oleracea L. var. Italica) plants were determined. The results showed that a reduced NO3(-) or co-provision of NO3(-) and NH4(+) could be an optimal source of inorganic N for broccoli plants. In addition, elevated [CO2] ameliorated the effect of salt exposure on the plant growth through an enhanced rate of photosynthesis, even at low N-concentration. However, NO3(-) or NO3(-)/NH4(+) co-provision display differential plant response to salt stress regarding water balance, which was associated to N metabolism. The results may contribute to our understanding of N-fertilization modes under increasing atmospheric [CO2] to cope with salt stress, where variations in N nutrition significantly influenced plant response. Copyright © 2016 Elsevier B.V. All rights reserved.

  9. Dietary Intake of Sulforaphane-Rich Broccoli Sprout Extracts during Juvenile and Adolescence Can Prevent Phencyclidine-Induced Cognitive Deficits at Adulthood.

    Directory of Open Access Journals (Sweden)

    Yumi Shirai

    Full Text Available Oxidative stress and inflammation play a role in cognitive impairment, which is a core symptom of schizophrenia. Furthermore, a hallmark of the pathophysiology of this disease is the dysfunction of cortical inhibitory γ-aminobutyric acid (GABA neurons expressing parvalbumin (PV, which is also involved in cognitive impairment. Sulforaphane (SFN, an isothiocyanate derived from broccoli, is a potent activator of the transcription factor Nrf2, which plays a central role in the inducible expressions of many cytoprotective genes in response to oxidative stress. Keap1 is a cytoplasmic protein that is essential for the regulation of Nrf2 activity. Here, we found that pretreatment with SFN attenuated cognitive deficits, the increase in 8-oxo-dG-positive cells, and the decrease in PV-positive cells in the medial prefrontal cortex and hippocampus after repeated administration of phencyclidine (PCP. Furthermore, PCP-induced cognitive deficits were improved by the subsequent subchronic administration of SFN. Interestingly, the dietary intake of glucoraphanin (a glucosinolate precursor of SFN during the juvenile and adolescence prevented the onset of PCP-induced cognitive deficits as well as the increase in 8-oxo-dG-positive cells and the decrease in PV-positive cells in the brain at adulthood. Moreover, the NRF2 gene and the KEAP1 gene had an epistatic effect on cognitive impairment (e.g., working memory and processing speed in patients with schizophrenia. These findings suggest that SFN may have prophylactic and therapeutic effects on cognitive impairment in schizophrenia. Therefore, the dietary intake of SFN-rich broccoli sprouts during the juvenile and adolescence may prevent the onset of psychosis at adulthood.

  10. Preferentially enhancing anti-cancer isothiocyanates over glucosinolates in broccoli sprouts: How NaCl and salicylic acid affect their formation.

    Science.gov (United States)

    Esfandiari, Azadeh; Saei, Ali; McKenzie, Marian J; Matich, Adam J; Babalar, Mesbah; Hunter, Donald A

    2017-06-01

    Broccoli (Brassica oleracea L. var. italica) sprouts contain glucosinolates (GLs) that when hydrolysed yield health promoting isothiocyanates such as sulforaphane (SF). SF content can be increased by salt (NaCl) stress, although high salt concentrations negatively impact plant growth. Salicylic acid (SA) treatments can attenuate the negative effects of salt on growth. To test whether sprout isothiocyanate content could be elevated without sprout growth being compromised, broccoli seed were germinated and grown for seven days in salt (0, 80 and 160 mM) alone and in combination with 100 μM SA. Increasing concentrations of salt lowered transcript accumulation of GL biosynthetic genes which was reflected in lowered content of Gluconapin, 4-methoxyglucobrassicin and neoglucobrassicin glucosinolates. Other glucosinolates such as glucoraphanin did not alter significantly. Salt (160 mM) increased transcript abundance of the GL hydrolytic gene MYROSINASE (BoMYO) and its cofactor EPITHIOSPECIFIER MODIFIER1 (BoESM1) whose encoded product directs MYROSINASE to produce isothiocyanate rather than nitrile forms. SF content was increased 6-fold by the 160 mM salt treatment, but the salt treatment reduced percentage seed germination, slowed seed germination, and reduced sprout hypocotyl elongation. This growth inhibition was prevented if 100 μM SA was included with the salt treatment. These findings suggest that the increase in SF production by salt occurs in part because of increased transcript abundance of genes in the hydrolytic pathway, which occurs independently of the negative impact of salt on sprout growth. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  11. Active edible coating and γ-irradiation as cold combined treatments to assure the safety of broccoli florets (Brassica oleracea L.).

    Science.gov (United States)

    Ben-Fadhel, Yosra; Saltaji, Sabrina; Khlifi, Mohamed Ali; Salmieri, Stephane; Dang Vu, Khanh; Lacroix, Monique

    2017-01-16

    The antimicrobial activity of essential oils (EOs), organic acid (OA) salts and natamycin, a natural antifungal produced during fermentation by the bacterium Streptomyces natalensis, was assessed against four pathogens (Escherichia coli O157:H7, Listeria monocytogenes, Salmonella Typhimurium and Aspergillus niger). The Minimum Inhibitory Concentration (MIC) of each antimicrobial (AM) was assessed to determine their efficiency on tested microbial species in order to select the most efficient. Then, the interactions between different antimicrobial compounds showing the lowest MIC were determined by the checkerboard method. The most effective antimicrobial formulation showing synergistic or additive effects was then encapsulated in an alginate matrix to protect the antimicrobial efficiency during storage. The effectiveness of the formulation was then evaluated in situ using broccoli as a food model. A combined treatment of active coating and γ-irradiation (0.4 and 0.8kGy) was also done in order to evaluate the possible synergistic effect between treatments. The results of this study allowed the selection of 4 EOs, one OA salt and the natamycin as an antifungal agent exhibiting lower MIC values. The interactive antimicrobial effects between them showed that an antimicrobial formulation composed of 300ppm of lemongrass EO, 5000ppm of sodium diacetate and 80ppm of natamycin resulted in an additive effect against A. niger, E. coli and S. Typhimurium and showing synergistic effect against L. monocytogenes. Finally, in situ analyses showed a synergistic antimicrobial activity between active coating and γ-irradiation and allowed the extension of the shelf-life of ready-to-eat (RTE) broccoli during storage at 4°C. Copyright © 2016 Elsevier B.V. All rights reserved.

  12. Crystal structure of phosphoethanolamine methyltransferase from Plasmodium falciparum in complex with amodiaquine

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Soon Goo; Alpert, Tara D.; Jez, Joseph M. (WU)

    2012-07-17

    Phosphoethanolamine N-methyltransferase (PMT) is essential for phospholipid biogenesis in the malarial parasite Plasmodium falciparum. PfPMT catalyzes the triple methylation of phosphoethanolamine to produce phosphocholine, which is then used for phosphatidylcholine synthesis. Here we describe the 2.0 {angstrom} resolution X-ray crystal structure of PfPMT in complex with amodiaquine. To better characterize inhibition of PfPMT by amodiaquine, we determined the IC{sub 50} values of a series of aminoquinolines using a direct radiochemical assay. Both structural and functional analyses provide a possible approach for the development of new small molecule inhibitors of PfPMT.

  13. A fluorescence resonance energy transfer-based method for histone methyltransferases

    DEFF Research Database (Denmark)

    Devkota, Kanchan; Lohse, Brian; Nyby Jakobsen, Camilla

    2015-01-01

    A simple dye–quencher fluorescence resonance energy transfer (FRET)-based assay for methyltransferases was developed and used to determine kinetic parameters and inhibitory activity at EHMT1 and EHMT2. Peptides mimicking the truncated histone H3 tail were functionalized in each end with a dye...... and a quencher, respectively. When lysine-9 residues in the peptides were methylated, they were protected from cleavage by endoproteinase–EndoLysC, whereas unmethylated peptides were cleaved, resulting in an increase in fluorescent intensity....

  14. Turning a Substrate Peptide into a Potent Inhibitor for the Histone Methyltransferase SETD8

    Energy Technology Data Exchange (ETDEWEB)

    Judge, Russell A.; Zhu, Haizhong; Upadhyay, Anup K.; Bodelle, Pierre M.; Hutchins, Charles W.; Torrent, Maricel; Marin, Violeta L.; Yu, Wenyu; Vedadi, Masoud; Li, Fengling; Brown, Peter J.; Pappano, William N.; Sun, Chaohong; Petros, Andrew M.

    2016-12-08

    SETD8 is a histone H4–K20 methyltransferase that plays an essential role in the maintenance of genomic integrity during mitosis and in DNA damage repair, making it an intriguing target for cancer research. While some small molecule inhibitors for SETD8 have been reported, the structural binding modes for these inhibitors have not been revealed. Using the complex structure of the substrate peptide bound to SETD8 as a starting point, different natural and unnatural amino acid substitutions were tested, and a potent (Ki 50 nM, IC50 0.33 μM) and selective norleucine containing peptide inhibitor has been obtained.

  15. A rat brain cytosolic N-methyltransferase(s) activity converting phosphorylethanolamine into phosphorylcholine.

    Science.gov (United States)

    Andriamampandry, C; Massarelli, R; Freysz, L; Kanfer, J N

    1990-09-14

    It had been previously speculated upon but never proved that the methylation of phosphorylethanolamine could contribute to the production of choline containing compounds. However, experimental evidence obtained with neuronal cultures was interpreted as showing that the stepwise methylation of phosphobases may be an important route for this biosynthesis. We demonstrate that cytosolic fraction from rat brain possesses a N-methyltransferase activity capable of methylating phosphorylethanolamine and its mono- and dimethyl-derivatives into phosphorylcholine. The level of activity detectable in rat liver cytosol is only 18% of that found in the brain cytosol.

  16. Synthesis and optimization of N-heterocyclic pyridinones as catechol-O-methyltransferase (COMT) inhibitors.

    Science.gov (United States)

    Zhao, Zhijian; Harrison, Scott T; Schubert, Jeffrey W; Sanders, John M; Polsky-Fisher, Stacey; Zhang, Nanyan Rena; McLoughlin, Debra; Gibson, Christopher R; Robinson, Ronald G; Sachs, Nancy A; Kandebo, Monika; Yao, Lihang; Smith, Sean M; Hutson, Pete H; Wolkenberg, Scott E; Barrow, James C

    2016-06-15

    A series of N-heterocyclic pyridinone catechol-O-methyltransferase (COMT) inhibitors were synthesized. Physicochemical properties, including ligand lipophilic efficiency (LLE) and clogP, were used to guide compound design and attempt to improve inhibitor pharmacokinetics. Incorporation of heterocyclic central rings provided improvements in physicochemical parameters but did not significantly reduce in vitro or in vivo clearance. Nevertheless, compound 11 was identified as a potent inhibitor with sufficient in vivo exposure to significantly affect the dopamine metabolites homovanillic acid (HVA) and dihydroxyphenylacetic acid (DOPAC), and indicate central COMT inhibition. Copyright © 2016 Elsevier Ltd. All rights reserved.

  17. DNA Methyltransferase Protein Synthesis Is Reduced in CXXC Finger Protein 1–Deficient Embryonic Stem Cells

    OpenAIRE

    Butler, Jill S; Palam, Lakshmi R.; Tate, Courtney M.; Sanford, Jeremy R; Wek, Ronald C.; Skalnik, David G.

    2009-01-01

    CXXC finger protein 1 (CFP1) binds to unmethylated CpG dinucleotides and is required for embryogenesis. CFP1 is also a component of the Setd1A and Setd1B histone H3K4 methyltransferase complexes. Murine embryonic stem (ES) cells lacking CFP1 fail to differentiate, and exhibit a 70% reduction in global genomic cytosine methylation and a 50% reduction in DNA methyltransferase (DNMT1) protein and activity. This study investigated the underlying mechanism for reduced DNMT1 expression in CFP1-defi...

  18. Identification of the methyltransferase targeting C2499 in Deinococcus radiodurans 23S ribosomal RNA

    DEFF Research Database (Denmark)

    Nielsen, Julie Mundus; Flyvbjerg, Karen Freund; Kirpekar, Finn

    2016-01-01

    and in vivo. We also inactivated the DR_0049 gene in D. radiodurans through insertion of a chloramphenicol resistance cassette. This resulted in complete absence of the cytidine 2499 methylation, which all together demonstrates that DR_0049 encodes the methyltransferase producing m(5)C2499 in D. radiodurans...... 2499 reported. Using homology search, we identified the open reading frame DR_0049 as the primary candidate gene for the methyltransferase that modifies cytidine 2499. Mass spectrometric analysis demonstrated that recombinantly expressed DR0049 protein methylates E. coli cytidine 2499 both in vitro...

  19. Association between TPMT*3C and decreased thiopurine S-methyltransferase activity in patients with neuromyelitis optica spectrum disorders in China.

    Science.gov (United States)

    Gong, Xiaoqing; Mei, Shenghui; Li, Xindi; Li, Xingang; Zhou, Heng; Liu, Yonghong; Zhou, Anna; Yang, Li; Zhao, Zhigang; Zhang, Xinghu

    2017-12-01

    Thiopurines are effective drugs in treating neuromyelitis optica spectrum disorders and other diseases. Thiopurines' toxicity is mainly imputed to thiopurine S-methyltransferase activity. In Chinese population, the most common and important variation of thiopurine S-methyltransferase is TPMT*3C (rs1142345). This study aims to reveal the association between thiopurine S-methyltransferase activity and genetic polymorphisms of thiopurine S-methyltransferase in patients with neuromyelitis optica spectrum disorders in China. A liquid chromatography tandem mass/mass method was used to evaluate the thiopurine S-methyltransferase activity by using 6-mercapthioprine as the substrate in human erythrocyte haemolysate via 1 h incubation at 37 °C to form its methylated product 6-methylmercaptopurine. The amount of 6-methylmercaptopurine was adjusted by haematocrit and normalized to 8 × 10 8 erythrocytes. The selected polymorphisms of thiopurine S-methyltransferase were identified using MassARRAY system (Sequenom) and multiple SNaPshot technique. In 69 patients with neuromyelitis optica spectrum disorders, thiopurine S-methyltransferase activity was 80.29-154.53 (127.51 ± 16.83) pmol/h/8 × 10 8 erythrocytes. TPMT*3C (rs1142345) was associated with lower thiopurine S-methyltransferase activity (BETA = -25.37, P = 0.011). Other selected variants were not associated with thiopurine S-methyltransferase activity. TPMT*3C affects TPMT activity in Chinese patients with neuromyelitis optica spectrum disorders. Further studies are warranted to confirm the results. TPRs = thiopurines; NMOSD = neuromyelitis optica spectrum disorders; TPMT = thiopurine S-methyltransferase; LC-MS/MS = liquid chromatography tandem mass/mass; 6-MMP = 6-methylmercaptopurine; IS = internal standard; SNP = single nucleotide polymorphism; MAF = minor allele frequency; HWE = Hardy-Weinberg equilibrium; BETA = regression coefficients; UTR-3 = untranslated region 3.

  20. Identification of a methyltransferase catalyzing the final step of methyl anthranilate synthesis in cultivated strawberry.

    Science.gov (United States)

    Pillet, Jeremy; Chambers, Alan H; Barbey, Christopher; Bao, Zhilong; Plotto, Anne; Bai, Jinhe; Schwieterman, Michael; Johnson, Timothy; Harrison, Benjamin; Whitaker, Vance M; Colquhoun, Thomas A; Folta, Kevin M

    2017-08-31

    Methyl anthranilate (MA) contributes an attractive fruity note to the complex flavor and aroma of strawberry (Fragaria spp.), yet it is rare in modern cultivars. The genetic basis for its biosynthesis has not been elucidated. Understanding the specific genes required for its synthesis could allow  the development of gene/allele-specific molecular markers to speed breeding of flavorful strawberries. Ripe fruits from individuals in an F1 population resulting from a cross between a MA producer and a non-producer were examined using a bulk-segregant transcriptome approach. MA producer and non-producer transcriptomes were compared, revealing five candidate transcripts that strictly co-segregated with MA production. One candidate encodes an annotated methyltransferase. MA levels are lower when this transcript is suppressed with RNAi, and bacterial cultures expressing the protein produced MA in the presence of anthranilic acid. Frozen fruit powders reconstituted with anthranilic acid and a methyl donor produced MA only if the transcript was detected in the fruit powder. A DNA-based molecular marker was developed that segregates with the MA-producing gene variant. These analyses indicate that the methyltransferase, now noted ANTHRANILIC ACID METHYL TRANSFERASE (FanAAMT), mediates the ultimate step of MA production in cultivated strawberry. Identification of this gene and its associated molecular marker may hasten breeding efforts to introduce this important volatile into modern cultivars.

  1. Catalysis in glycine N-methyltransferase: testing the electrostatic stabilization and compression hypothesis.

    Science.gov (United States)

    Soriano, Alejandro; Castillo, Raquel; Christov, Christo; Andrés, Juan; Moliner, Vicente; Tuñón, Iñaki

    2006-12-19

    Glycine N-methyltransferase (GNMT) is an S-adenosyl-l-methionine dependent enzyme that catalyzes glycine transformation to sarcosine. Here, we present a hybrid quantum mechanics/molecular mechanics (QM/MM) computational study of the reaction compared to the counterpart process in water. The process takes place through an SN2 mechanism in both media with a transition state in which the transferring methyl group is placed in between the donor (SAM) and the acceptor (the amine group of glycine). Comparative analysis of structural, electrostatic, and electronic characteristics of the in-solution and enzymatic transition states allows us to get a deeper insight into the origins of the enzyme's catalytic power. We found that the enzyme is able to stabilize the substrate in its more active basic form by means of a positively charged residue (Arg175) placed in the active site. However, the maximum stabilization is attained for the transition state. In this case, the enzyme is able to form stronger hydrogen bonds with the positively charged amine group. Finally, we show that in agreement with previous computational studies on other methyltransferases, there is no computational evidence for the compression hypothesis, as was formulated by Schowen (Hegazi, M. F., Borchardt, R. T., and Schowen, R. L. (1979) J. Am. Chem. Soc. 101, 4359-4365).

  2. Characterization of a Bvg-regulated fatty acid methyl-transferase in Bordetella pertussis.

    Science.gov (United States)

    Rivera-Millot, Alex; Lesne, Elodie; Solans, Luis; Coutte, Loic; Bertrand-Michel, Justine; Froguel, Philippe; Dhennin, Véronique; Hot, David; Locht, Camille; Antoine, Rudy; Jacob-Dubuisson, Françoise

    2017-01-01

    The whooping cough agent Bordetella pertussis controls the expression of its large virulence regulon in a coordinated manner through the two-component signal transduction system BvgAS. In addition to the genes coding for bona fide virulence factors, the Bvg regulon comprises genes of unknown function. In this work, we characterized a new Bvg-activated gene called BP2936. Homologs of BP2936 are found in other pathogenic Bordetellae and in several other species, including plant pathogens and environmental bacteria. We showed that the gene product of BP2936 is a membrane-associated methyl-transferase of free fatty acids. We thus propose to name it FmtB, for fatty acid methyl-transferase of Bordetella. The role of this protein was tested in cellular and animal models of infection, but the loss of BP2936 did not appear to affect host-pathogen interactions in those assays. The high level of conservation of BP2936 among B. pertussis isolates nevertheless argues that it probably plays a role in the life cycle of this pathogen.

  3. Identification of the methyltransferase targeting C2499 in Deinococcus radiodurans 23S ribosomal RNA.

    Science.gov (United States)

    Mundus, Julie; Flyvbjerg, Karen Freund; Kirpekar, Finn

    2016-01-01

    The bacterium Deinococcus radiodurans-like all other organisms-introduces nucleotide modifications into its ribosomal RNA. We have previously found that the bacterium contains a Carbon-5 methylation on cytidine 2499 of its 23S ribosomal RNA, which is so far the only modified version of cytidine 2499 reported. Using homology search, we identified the open reading frame DR_0049 as the primary candidate gene for the methyltransferase that modifies cytidine 2499. Mass spectrometric analysis demonstrated that recombinantly expressed DR0049 protein methylates E. coli cytidine 2499 both in vitro and in vivo. We also inactivated the DR_0049 gene in D. radiodurans through insertion of a chloramphenicol resistance cassette. This resulted in complete absence of the cytidine 2499 methylation, which all together demonstrates that DR_0049 encodes the methyltransferase producing m(5)C2499 in D. radiodurans 23S rRNA. Growth experiments disclosed that inactivation of DR_0049 is associated with a severe growth defect, but available ribosome structures show that cytidine 2499 is positioned very similar in D. radiodurans harbouring the modification and E. coli without the modification. Hence there is no obvious structure-based explanation for the requirement for the C2499 posttranscriptional modification in D. radiodurans.

  4. DNA methyltransferase 1 (Dnmt1) mutation affects Snrpn imprinting in the mouse male germ line.

    Science.gov (United States)

    Saferali, Aabida; Moussette, Sanny; Chan, Donovan; Trasler, Jacquetta; Chen, Taiping; Rozen, Rima; Naumova, Anna K

    2012-09-01

    DNA methylation and DNA methyltransferases are essential for spermatogenesis. Mutations in the DNA methyltransferase Dnmt1 gene exert a paternal effect on epigenetic states and phenotypes of offspring, suggesting that DNMT1 is important for the epigenetic remodeling of the genome that takes place during spermatogenesis. However, the specific role of DNMT1 in spermatogenesis and the establishment of genomic imprints in the male germ line remains elusive. To further characterize the effect of DNMT1 deficiency on the resetting of methylation imprints during spermatogenesis, we analyzed the methylation profiles of imprinted regions in the spermatozoa of mice that were heterozygous for a Dnmt1 loss-of-function mutation. The mutation did not affect the H19 or IG differentially methylated regions (DMRs) that are usually highly methylated but led to a partial hypermethylation of the Snrpn DMR, a region that should normally be unmethylated in mature spermatozoa. This defect does not appear in mouse models with mutations in Dnmt3a and Mthfr genes and, therefore, it is specific for the Dnmt1 gene and is suggestive of a role of DNMT1 in imprint resetting or maintenance in the male germ line.

  5. The murine catecholamine methyltransferase mTOMT is essential for mechanotransduction by cochlear hair cells

    Science.gov (United States)

    Cunningham, Christopher L; Wu, Zizhen; Jafari, Aria; Zhao, Bo; Schrode, Kat; Harkins-Perry, Sarah; Lauer, Amanda; Müller, Ulrich

    2017-01-01

    Hair cells of the cochlea are mechanosensors for the perception of sound. Mutations in the LRTOMT gene, which encodes a protein with homology to the catecholamine methyltransferase COMT that is linked to schizophrenia, cause deafness. Here, we show that Tomt/Comt2, the murine ortholog of LRTOMT, has an unexpected function in the regulation of mechanotransduction by hair cells. The role of mTOMT in hair cells is independent of mTOMT methyltransferase function and mCOMT cannot substitute for mTOMT function. Instead, mTOMT binds to putative components of the mechanotransduction channel in hair cells and is essential for the transport of some of these components into the mechanically sensitive stereocilia of hair cells. Our studies thus suggest functional diversification between mCOMT and mTOMT, where mTOMT is critical for the assembly of the mechanotransduction machinery of hair cells. Defects in this process are likely mechanistically linked to deafness caused by mutations in LRTOMT/Tomt. DOI: http://dx.doi.org/10.7554/eLife.24318.001 PMID:28504928

  6. The motility of a human parasite, Toxoplasma gondii, is regulated by a novel lysine methyltransferase.

    Directory of Open Access Journals (Sweden)

    Aoife T Heaslip

    2011-09-01

    Full Text Available Protozoa in the phylum Apicomplexa are a large group of obligate intracellular parasites. Toxoplasma gondii and other apicomplexan parasites, such as Plasmodium falciparum, cause diseases by reiterating their lytic cycle, comprising host cell invasion, parasite replication, and parasite egress. The successful completion of the lytic cycle requires that the parasite senses changes in its environment and switches between the non-motile (for intracellular replication and motile (for invasion and egress states appropriately. Although the signaling pathway that regulates the motile state switch is critical to the pathogenesis of the diseases caused by these parasites, it is not well understood. Here we report a previously unknown mechanism of regulating the motility activation in Toxoplasma, mediated by a protein lysine methyltransferase, AKMT (for Apical complex lysine (K methyltransferase. AKMT depletion greatly inhibits activation of motility, compromises parasite invasion and egress, and thus severely impairs the lytic cycle. Interestingly, AKMT redistributes from the apical complex to the parasite body rapidly in the presence of egress-stimulating signals that increase [Ca²⁺] in the parasite cytoplasm, suggesting that AKMT regulation of parasite motility might be accomplished by the precise temporal control of its localization in response to environmental changes.

  7. Histone methyltransferase 1 regulates the encystation process in the parasite Giardia lamblia.

    Science.gov (United States)

    Salusso, Agostina; Zlocowski, Natacha; Mayol, Gonzalo F; Zamponi, Nahuel; Rópolo, Andrea S

    2017-08-01

    In eukaryotes, histone lysine methylation is associated with either active or repressed chromatin states, depending on the status of methylation. Even when the amino-terminus of Giardia lamblia histones diverges from other organisms, these regions contain lysine residues that are potential targets for methylation. When we examined the role of the histone methyltransferase 1 (HMT1) in the regulation of the encystation process by giardial histone methyltransferase 1 (GlHMT1) overexpression or downregulation, we observed an increase or a decrease in cyst production, respectively, compared to wild-type trophozoites. A time-lapse analysis of encystation showed that overexpression of GlHMT1 induced an earlier and faster process than in wild-type cells together with an upregulation of mRNA expression of cyst wall proteins. Subcellular localization studies indicated that GlHMT1-hemaglutinin was mainly associated with the nuclear and perinuclear region in both growing and encysting parasites, in agreement with bioinformatics analyses showing that GlHMT-1 possesses nuclear localization signals in addition to the classical SU(var)3-9, Enhancer-of-Zeste, Trithorax (SET), and post-SET domains. Altogether, these findings suggest that the function of HMT1 is critical for the success and timing of the encystation process, and reinforce the idea that epigenetic marks are critical for cyst formation in G. lamblia. © 2017 Federation of European Biochemical Societies.

  8. The murine catecholamine methyltransferase mTOMT is essential for mechanotransduction by cochlear hair cells.

    Science.gov (United States)

    Cunningham, Christopher L; Wu, Zizhen; Jafari, Aria; Zhao, Bo; Schrode, Kat; Harkins-Perry, Sarah; Lauer, Amanda; Müller, Ulrich

    2017-05-15

    Hair cells of the cochlea are mechanosensors for the perception of sound. Mutations in the LRTOMT gene, which encodes a protein with homology to the catecholamine methyltransferase COMT that is linked to schizophrenia, cause deafness. Here, we show that Tomt/Comt2, the murine ortholog of LRTOMT, has an unexpected function in the regulation of mechanotransduction by hair cells. The role of mTOMT in hair cells is independent of mTOMT methyltransferase function and mCOMT cannot substitute for mTOMT function. Instead, mTOMT binds to putative components of the mechanotransduction channel in hair cells and is essential for the transport of some of these components into the mechanically sensitive stereocilia of hair cells. Our studies thus suggest functional diversification between mCOMT and mTOMT, where mTOMT is critical for the assembly of the mechanotransduction machinery of hair cells. Defects in this process are likely mechanistically linked to deafness caused by mutations in LRTOMT/Tomt.

  9. Identification and expression profiling of DNA methyltransferases during development and stress conditions in Solanaceae.

    Science.gov (United States)

    Kumar, Rahul; Chauhan, Pankaj Kumar; Khurana, Ashima

    2016-09-01

    DNA methyltransferase (DMTase) enzymes contribute to plant development and stress responses by de novo establishment and subsequent maintenance of DNA methylation during replication. However, the molecular mechanism underlying this activity remains obscure, especially in crop species. Using DMTase homolog complement in six Solanaceae species, we demonstrated here that their number remained conserved in Solanum lineage, whereas it was expanded in both pepper and Nicotiana benthamiana. Non-synonymous vs synonymous (Ka/Ks) substitution ratio revealed that most of the Solanaceous DMTase homologs undergo purifying selection. The genomic sequences of tomato DMT homologs in its wild relative, Solanum pennellii, remained highly conserved in their exons and methyltransferase domains. Structure analysis further revealed highly similar folding of DMTase homologs and conservation in the residues participating in protein-protein interaction in Solanum lineage, whereas a considerable diversification was observed of pepper homologs. Transcript profiling of DMTases highlighted both similar and distinct expression patterns of tomato homologs in other species during fruit development and stress responses. Overall, our analysis provides a strong basis for in-depth exploration of both conserved as well as distinct functions of tomato DMTase homologs in other economically important Solanaceae species.

  10. Regulation of DNA replication and chromosomal polyploidy by the MLL-WDR5-RBBP5 methyltransferases

    Directory of Open Access Journals (Sweden)

    Fei Lu

    2016-10-01

    Full Text Available DNA replication licensing occurs on chromatin, but how the chromatin template is regulated for replication remains mostly unclear. Here, we have analyzed the requirement of histone methyltransferases for a specific type of replication: the DNA re-replication induced by the downregulation of either Geminin, an inhibitor of replication licensing protein CDT1, or the CRL4CDT2 ubiquitin E3 ligase. We found that siRNA-mediated reduction of essential components of the MLL-WDR5-RBBP5 methyltransferase complexes including WDR5 or RBBP5, which transfer methyl groups to histone H3 at K4 (H3K4, suppressed DNA re-replication and chromosomal polyploidy. Reduction of WDR5/RBBP5 also prevented the activation of H2AX checkpoint caused by re-replication, but not by ultraviolet or X-ray irradiation; and the components of MLL complexes co-localized with the origin recognition complex (ORC and MCM2-7 replicative helicase complexes at replication origins to control the levels of methylated H3K4. Downregulation of WDR5 or RBBP5 reduced the methylated H3K4 and suppressed the recruitment of MCM2-7 complexes onto replication origins. Our studies indicate that the MLL complexes and H3K4 methylation are required for DNA replication but not for DNA damage repair.

  11. Structure of the arginine methyltransferase PRMT5-MEP50 reveals a mechanism for substrate specificity.

    Directory of Open Access Journals (Sweden)

    Meng-Chiao Ho

    Full Text Available The arginine methyltransferase PRMT5-MEP50 is required for embryogenesis and is misregulated in many cancers. PRMT5 targets a wide variety of substrates, including histone proteins involved in specifying an epigenetic code. However, the mechanism by which PRMT5 utilizes MEP50 to discriminate substrates and to specifically methylate target arginines is unclear. To test a model in which MEP50 is critical for substrate recognition and orientation, we determined the crystal structure of Xenopus laevis PRMT5-MEP50 complexed with S-adenosylhomocysteine (SAH. PRMT5-MEP50 forms an unusual tetramer of heterodimers with substantial surface negative charge. MEP50 is required for PRMT5-catalyzed histone H2A and H4 methyltransferase activity and binds substrates independently. The PRMT5 catalytic site is oriented towards the cross-dimer paired MEP50. Histone peptide arrays and solution assays demonstrate that PRMT5-MEP50 activity is inhibited by substrate phosphorylation and enhanced by substrate acetylation. Electron microscopy and reconstruction showed substrate centered on MEP50. These data support a mechanism in which MEP50 binds substrate and stimulates PRMT5 activity modulated by substrate post-translational modifications.

  12. The Histone Methyltransferase Activity of MLL1 Is Dispensable for Hematopoiesis and Leukemogenesis

    Directory of Open Access Journals (Sweden)

    Bibhu P. Mishra

    2014-05-01

    Full Text Available Despite correlations between histone methyltransferase (HMT activity and gene regulation, direct evidence that HMT activity is responsible for gene activation is sparse. We address the role of the HMT activity for MLL1, a histone H3 lysine 4 (H3K4 methyltransferase critical for maintaining hematopoietic stem cells (HSCs. Here, we show that the SET domain, and thus HMT activity of MLL1, is dispensable for maintaining HSCs and supporting leukemogenesis driven by the MLL-AF9 fusion oncoprotein. Upon Mll1 deletion, histone H4 lysine 16 (H4K16 acetylation is selectively depleted at MLL1 target genes in conjunction with reduced transcription. Surprisingly, inhibition of SIRT1 is sufficient to prevent the loss of H4K16 acetylation and the reduction in MLL1 target gene expression. Thus, recruited MOF activity, and not the intrinsic HMT activity of MLL1, is central for the maintenance of HSC target genes. In addition, this work reveals a role for SIRT1 in opposing MLL1 function.

  13. Comparative analysis of DNA methyltransferase gene family in fungi: a focus on Basidiomycota

    Directory of Open Access Journals (Sweden)

    Ruirui Huang

    2016-10-01

    Full Text Available DNA methylation plays a crucial role in the regulation of gene expression in eukaryotes. Mushrooms belonging to the phylum Basidiomycota are highly valued for both nutritional and pharmaceutical uses. A growing number of studies have demonstrated the significance of DNA methylation in the development of plants and animals. However, our understanding of DNA methylation in mushrooms is limited. In this study, we identified and conducted comprehensive analyses on DNA methyltransferases (DNMtases in representative species from Basidiomycota and Ascomycota, and obtained new insights into their classification and characterization in fungi. Our results revealed that DNMtases in basidiomycetes can be divided into two classes, the Dnmt1 class and the newly defined Rad8 class. We also demonstrated that the fusion event between the characteristic domains of the DNMtases family and Snf2 family in the Rad8 class is fungi-specific, possibly indicating a functional novelty of Rad8 DNMtases in fungi. Additionally, expression profiles of DNMtases in the edible mushroom Pleurotus ostreatus revealed diverse expression patterns in various organs and developmental stages. For example, DNMtase genes displayed higher expression levels in dikaryons than in monokaryons. Consistent with the expression profiles, we found that dikaryons are more susceptible to the DNA methyltransferase inhibitor 5-azacytidine. Taken together, our findings pinpoint an important role of DNA methylation during the growth of mushrooms and provide a foundation for understanding of DNMtases in basidiomycetes.

  14. Molecular cloning and functional identification of sterol C24-methyltransferase gene from Tripterygium wilfordii

    Directory of Open Access Journals (Sweden)

    Hongyu Guan

    2017-09-01

    Full Text Available Sterol C24-methyltransferase (SMT plays multiple important roles in plant growth and development. SMT1, which belongs to the family of transferases and transforms cycloartenol into 24-methylene cycloartenol, is involved in the biosynthesis of 24-methyl sterols. Here, we report the cloning and characterization of a cDNA encoding a sterol C24-methyltransferase from Tripterygium wilfordii (TwSMT1. TwSMT1 (GenBank access number KU885950 is a 1530 bp cDNA with a 1041 bp open reading frame predicted to encode a 346-amino acid, 38.62 kDa protein. The polypeptide encoded by the SMT1 cDNA was expressed and purified as a recombinant protein from Escherichia coli (E. coli and showed SMT activity. The expression of TwSMT1 was highly up-regulated in T. wilfordii cell suspension cultures treated with methyl jasmonate (MeJA. Tissue expression pattern analysis showed higher expression in the phellem layer compared to the other four organs (leaf, stem, xylem and phloem, which is about ten times that of the lowest expression in leaf. The results are meaningful for the study of sterol biosynthesis of T. wilfordii and will further lay the foundations for the research in regulating both the content of other main compounds and growth and development of T. wilfordii.

  15. Colorimetric activity measurement of a recombinant putrescine N-methyltransferase from Datura stramonium.

    Science.gov (United States)

    Biastoff, Stefan; Teuber, Michael; Zhou, Zhaohui Sunny; Dräger, Birgit

    2006-10-01

    Putrescine N-methyltransferase (PMT, EC 2.1.1.53) catalyses the S-adenosyl- L-methionine (SAM or AdoMet)-dependent methylation of putrescine to N-methylputrescine within the biosynthetic pathways of calystegines, nicotine, and tropane alkaloids in medicinal plants and produces S-adenosyl- L-homocysteine (SAH or AdoHcy). Determination of PMT activity was time-consuming and hardly reproducible in the past because it required tedious separation steps after chemical derivatisation or radioactive labelling of N-methylputrescine. A convenient and accurate enzyme-coupled colorimetric assay is based on the conversion of SAH to homocysteine by 5'-methylthioadenosine/ S-adenosylhomocysteine nucleosidase (MTAN/SAHN, EC 3.2.2.9) and S-ribosylhomocysteine lyase (LuxS, EC 4.4.1.21). Homocysteine is quantified by 5,5'-dithiobis-2-nitrobenzoic acid. Putrescine was shown not to interfere with MTAN or LuxS. The colorimetric assay was validated by HPLC analysis. K(m) values determined by the assay, 108 microM for putrescine and 42 microM for SAM, are lower than the previously reported values, due to alleviation of PMT inhibition by SAH. DTNB:5,5'-dithiobis-2-nitrobenzoic acid LuxS: S-ribosylhomocysteine lyase MTAN:5'-methylthioadenosine nucleosidase PMT:putrescine N-methyltransferase SAH: S-adenosyl- L-homocysteine SAM: S-adenosyl- L-methionine TNB:2-nitro-5-thiobenzoic acid.

  16. Structural insights into methyltransferase KsgA function in 30S ribosomal subunit biogenesis.

    Science.gov (United States)

    Boehringer, Daniel; O'Farrell, Heather C; Rife, Jason P; Ban, Nenad

    2012-03-23

    The assembly of the ribosomal subunits is facilitated by ribosome biogenesis factors. The universally conserved methyltransferase KsgA modifies two adjacent adenosine residues in the 3'-terminal helix 45 of the 16 S ribosomal RNA (rRNA). KsgA recognizes its substrate adenosine residues only in the context of a near mature 30S subunit and is required for the efficient processing of the rRNA termini during ribosome biogenesis. Here, we present the cryo-EM structure of KsgA bound to a nonmethylated 30S ribosomal subunit. The structure reveals that KsgA binds to the 30S platform with the catalytic N-terminal domain interacting with substrate adenosine residues in helix 45 and the C-terminal domain making extensive contacts to helix 27 and helix 24. KsgA excludes the penultimate rRNA helix 44 from adopting its position in the mature 30S subunit, blocking the formation of the decoding site and subunit joining. We suggest that the activation of methyltransferase activity and subsequent dissociation of KsgA control conformational changes in helix 44 required for final rRNA processing and translation initiation.

  17. Structural Insights into Methyltransferase KsgA Function in 30S Ribosomal Subunit Biogenesis*

    Science.gov (United States)

    Boehringer, Daniel; O'Farrell, Heather C.; Rife, Jason P.; Ban, Nenad

    2012-01-01

    The assembly of the ribosomal subunits is facilitated by ribosome biogenesis factors. The universally conserved methyltransferase KsgA modifies two adjacent adenosine residues in the 3′-terminal helix 45 of the 16 S ribosomal RNA (rRNA). KsgA recognizes its substrate adenosine residues only in the context of a near mature 30S subunit and is required for the efficient processing of the rRNA termini during ribosome biogenesis. Here, we present the cryo-EM structure of KsgA bound to a nonmethylated 30S ribosomal subunit. The structure reveals that KsgA binds to the 30S platform with the catalytic N-terminal domain interacting with substrate adenosine residues in helix 45 and the C-terminal domain making extensive contacts to helix 27 and helix 24. KsgA excludes the penultimate rRNA helix 44 from adopting its position in the mature 30S subunit, blocking the formation of the decoding site and subunit joining. We suggest that the activation of methyltransferase activity and subsequent dissociation of KsgA control conformational changes in helix 44 required for final rRNA processing and translation initiation. PMID:22308031

  18. Inhibition of histone methyltransferase EZH2 ameliorates early acute renal allograft rejection in rats.

    Science.gov (United States)

    Li, Long; Zhang, Yi; Xu, Ming; Rong, Ruiming; Wang, Jina; Zhu, Tongyu

    2016-10-26

    Although histone methyltransferases EZH2 has been proved to have significant regulatory effect on the immune rejection after hematopoietic stem cell transplantation, its role in solid-organ transplantation remains uncovered. In this study, we investigate whether histone methylation regulation can impact renal allograft rejection in rat models. Allogeneic rat renal transplantation model (Wistar to Lewis) was established, and the recipients were administrated with EZH2 inhibitor DZNep after transplantation. Renal allografts and peripheral blood were collected on day 5 after transplantation for histological examination and mechanism investigation. We found that inhibition of EZH2 by DZNep after transplantation significantly ameliorated acute rejection (AR), with decreased histological injury and reduced inflammatory infiltration in renal allografts. Attenuation of AR was due to the prohibited activation of alloreactive T cells, the subsequent impaired production of inflammatory cytokines, and also the elevated apoptosis of alloreactive T cells in both renal allografts and periphery. However, inhibition of EZH2 did not increase the regulatory T cells during the AR. Disruption of EZH2 by DZNep suppressed the immune responses of alloreactive T cells and ameliorated AR of renal allografts. This suggests a therapeutic potential of targeting histone methyltransferases EZH2 in treating allograft rejection after solid organ transplantation.

  19. Folate deficiency and aberrant expression of DNA methyltransferase 1 were associated with cervical cancerization.

    Science.gov (United States)

    Wang, Jin-tao; Ding, Ling; Jiang, Shi-Wen; Hao, Junxia; Zhao, Wei-min; Zhou, Quin; Yang, Zuo-kai; Zhang, Li

    2014-01-01

    DNA methyltransferase 1 (DNMT1) plays a significant role in maintaining DNA methylation. Aberrant DNA methylation is a recognized feature of human cancers and folate is directly involved in DNA methylation via one-carbon metabolism. Previous reports also have suggested that folate deficiency was associated with many cancers. The aim of the present study was to evaluate the effect of folate deficiency and aberrant expression of DNA methyltransferase 1 (DNMT1) on cervical cancerization. The expression of DNMT1 protein and mRNA and levels of serum folate were detected in 238 women with a diagnosis of normal cervix (NC,n = 53), cervical intraepithelial neoplasia (CIN I, n = 52; CIN II/III, n = 53), and squamous cell carcinoma of the cervix (SCC; n = 80). In addition, the expression of DNMT1 protein and mRNA was measured in cervical cancer cells (Caski and C33A) treated by different concentration of folate. Serum folate levels decreased and expression levels of DNMT1 protein and mRNA increased gradually with progressive severity of the cervix lesions (Pcancerization. Folate supplement and recovery of aberrant DNA methylation status may offer a new strategy for prevention and therapy of cancers.

  20. Recovery of biological active catechol-O-methyltransferase isoforms from Q-sepharose.

    Science.gov (United States)

    Correia, F F; Santos, F M; Pedro, A Q; Bonifácio, M J; Queiroz, J A; Passarinha, L A

    2014-01-01

    The development of new catechol-O-methyltransferase inhibitors has led to an improvement in the treatment of Parkinson's disease. However, despite the fact that the soluble isoform has been extensively investigated, few studies have been published concerning membrane isoform chromatographic recovery and bioactivity levels. In this work, chromatographic profiles of both catechol-O-methyltransferase isoforms were compared using quaternary amine as a ligand to evaluate its activity levels and recovery rates. Results show that both proteins required different conditions for adsorption; the soluble isoform adsorption was performed at low ionic strength, while the membrane isoform required increasing linear salt gradient. However, the application of 0.5% Triton X-100 promoted membrane isoform adsorption even at low ionic strength. Indeed, chromatographic conditions of both isoforms became similar when detergents were applied. The developed methods also appear to be highly effective in bioactivity recovery, presenting rates of 107% for soluble protein and 67 and 91% for membrane isoform without and with detergents, respectively. The chromatographic strategies with and without detergents resulted in a 4.3- and sevenfold purification, respectively, corresponding to specific activity values of 331 and 496 nmol/h/mg. Thus, the use of Q-sepharose as anion exchanger was effective in the recovery of both enzymes, which is a requirement for further kinetic and pharmacological trials. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  1. Human catechol-O-methyltransferase: Cloning and expression of the membrane-associated form

    Energy Technology Data Exchange (ETDEWEB)

    Bertocci, B.; Miggiano, V.; Da Prada, M.; Dembic, Z.; Lahm, H.W.; Malherbe, P. (F. Hoffmann-La Roche Ltd., Basel (Switzerland))

    1991-02-15

    A cDNA clone for human catechol-O-methyltransferase was isolated from a human hepatoma cell line (Hep G2) cDNA library by hybridization screening with a porcine cDNA probe. The cDNA clone was sequenced and found to have an insert of 1226 nucleotides. The deduced primary structure of hCOMT is composed of 271 amino acid residues with the predicted molecular mass of 30 kDa. At its N terminus it has a hydrophobic segment of 21 amino acid residues that may be responsible for insertion of hCOMT into the endoplasmic reticulum membrane. The primary structure of hCOMT exhibits high homology to the porcine partial cDNA sequence (93%). The deduced amino acid sequence contains two tryptic peptide sequences (T-22, T-33) found in porcine liver catechol-O-methyltransferase (CEMT). The coding region of hCOMT cDNA was placed under the control of the cytomegalovirus promoter to transfect human kidney 293 cells. The recombinant hCOMT was shown by immunoblot analysis to be mainly associated with the membrane fraction. RNA blot analysis revealed one COMT mRNA transcript of 1.4 kilobases in Hep G2 poly(A){sup +} RNA.

  2. Improved radioenzymatic assay for plasma norepinephrine using purified phenylethanolamine n-methyltransferase

    Energy Technology Data Exchange (ETDEWEB)

    Bowsher, R.R.; Henry, D.P.

    1986-03-01

    Radioenzymatic assays have been developed for catecholamines using either catechol O-methyltransferase (COMT) or phenylethanolamine N-methyltransferase (PNMT). Assays using PNMT are specific for norepinephrine (NE) and require minimal manipulative effort but until now have been less sensitive than the more complex procedures using COMT. The authors report an improved purification scheme for bovine PNMT which has permitted development of an NE assay with dramatically improved sensitivity (0.5 pg), specificity and reproducibility (C.V. < 5%). PNMT was purified by sequential pH 5.0 treatment and dialysis and by column chromatographic procedures using DEAE-Sephacel, Sepharcryl S-200 and Phenyl-Boronate Agarose. Recovery of PNMT through the purification scheme was 50%, while blank recovery was <.001%. NE can be directly quantified in 25 ul of human plasma and an 80 tube assay can be completed within 4 h. The capillary to venous plasma NE gradient was examined in 8 normotensive male subjects. Capillary plasma (NE (211.2 +/- 61.3 pg/ml)) was lower than venous plasma NE (366.6 +/- 92.5 pg/ml) in all subjects (p < 0.005). This difference suggests that capillary (NE) may be a unique indicator of sympathetic nervous system activity in vivo. In conclusion, purification of PNMT has facilitated development of an improved radioenzymatic for NE with significantly improved sensitivity.

  3. Structural Characterization of the Mitomycin 7-O-Methyltransferase MmcR

    Science.gov (United States)

    Singh, Shanteri; Chang, Aram; Goff, Randal D.; Bingman, Craig A.; Grüschow, Sabine; Sherman, David H.; Phillips, George N.; Thorson, Jon S.

    2011-01-01

    Mitomycins are quinone-containing antibiotics, widely used as anti-tumor drugs in chemotherapy. Mitomycin-7-O-methyltransferase (MmcR), a key tailoring enzyme involved in the biosynthesis of mitomycin in Streptomyces lavendulae, catalyzes the 7-O-methylation of both C9β- and C9α-configured 7-hydroxymitomycins. We have determined the crystal structures of the MmcR–S-adenosylhomocysteine (SAH) binary complex and MmcR–SAH-Mitomycin A (MMA) ternary complex at resolutions of 1.9 and 2.3 Å, respectively. The study revealed MmcR to adopt a common SAM-dependent O-MTase fold and the presence of a structurally-conserved active site general acid-base pair is consistent with a proton assisted methyltransfer common to most methyltransferases. Given the importance of C7 alkylation to modulate mitomycin redox potential this study may also present a template toward the future engineering of catalysts to generate uniquely bioactive mitomycins. PMID:21538548

  4. Targeting epigenetic regulators for cancer therapy: modulation of bromodomain proteins, methyltransferases, demethylases, and microRNAs.

    Science.gov (United States)

    Gelato, Kathy A; Shaikhibrahim, Zaki; Ocker, Matthias; Haendler, Bernard

    2016-07-01

    Histone deacetylases (HDACs) and DNA methyltransferases (DNMTs) were the first epigenetic targets to be successfully addressed for cancer treatment, but more recently additional families of epigenetic modulators have been the subject of intense research. Potent inhibitors have been identified in several instances and have proven to be invaluable tools for studying these proteins in normal physiology and in disease. Some have now progressed to clinical studies in hematological and solid tumors, and encouraging early results have been reported. This article reviews recent advances regarding the roles of new epigenetic players beyond HDACs and DNMTs in cancer, and discusses the impact of selective chemical probes on unravelling their function. The emerging field of non-coding RNAs (ncRNAs) and ongoing clinical studies with epigenetic drugs and microRNAs (miRNAs) are also addressed. The roles of different epigenetic factors in numerous cancers have been unraveled recently, leading to the initiation of clinical studies. With inhibitors of BET bromodomain proteins, the histone methyltransferases EZH2 and DOT1L, and the histone demethylase LSD1 progressing through clinical trials, and the recognition of the importance of ncRNAs as potential biomarkers and therapeutics, this bears the hope that novel epigenetic therapies will be approved soon.

  5. Structure and Mechanism of the Rebeccamycin Sugar 4'-O-Methyltransferase RebM

    Energy Technology Data Exchange (ETDEWEB)

    Singh, Shanteri; McCoy, Jason G.; Zhang, Changsheng; Bingman, Craig A.; Phillips, Jr., George N.; Thorson, Jon S. (UW)

    2008-12-12

    The 2.65-{angstrom} crystal structure of the rebeccamycin 4'-O-methyltransferase RebM in complex with S-adenosyl-l-homocysteine revealed RebM to adopt a typical S-adenosylmethionine-binding fold of small molecule O-methyltransferases (O-MTases) and display a weak dimerization domain unique to MTases. Using this structure as a basis, the RebM substrate binding model implicated a predominance of nonspecific hydrophobic interactions consistent with the reported ability of RebM to methylate a wide range of indolocarbazole surrogates. This model also illuminated the three putative RebM catalytic residues (His{sup 140/141} and Asp{sup 166}) subsequently found to be highly conserved among sequence-related natural product O-MTases from GC-rich bacteria. Interrogation of these residues via site-directed mutagenesis in RebM demonstrated His{sup 140} and Asp{sup 166} to be most important for catalysis. This study reveals RebM to be a member of the general acid/base-dependent O-MTases and, as the first crystal structure for a sugar O-MTase, may also present a template toward the future engineering of natural product MTases for combinatorial applications.

  6. Protein substrates of the arginine methyltransferase Hmt1 identified by proteome arrays.

    Science.gov (United States)

    Low, Jason K K; Im, Hogune; Erce, Melissa A; Hart-Smith, Gene; Snyder, Michael P; Wilkins, Marc R

    2016-02-01

    Arginine methylation on nonhistone proteins is associated with a number of cellular processes including RNA splicing, protein localization, and the formation of protein complexes. In this manuscript, Saccharomyces cerevisiae proteome arrays carrying 4228 proteins were used with an antimethylarginine antibody to first identify 88 putatively arginine-methylated proteins. By treating the arrays with recombinant arginine methyltransferase Hmt1, 42 proteins were found to be possible substrates of this enzyme. Analysis of the putative arginine-methylated proteins revealed that they were predominantly nuclear or nucleolar in localization, consistent with the localization of Hmt1. Many are involved in known methylarginine-associated functions, such as RNA processing and ribonucleoprotein complex biogenesis, yet others are of newer classes, namely RNA/DNA helicases and tRNA-associated proteins. Using ex vivo methylation and MS/MS, a set of 12 proteins (Brr1, Dia4, Hts1, Mpp10, Mrd1, Nug1, Prp43, Rpa43, Rrp43, Spp381, Utp4, and Npl3), including the RNA helicase Prp43 and tRNA ligases Dia4 and Hts1, were all validated as Hmt1 substrates. Interestingly, the majority of these also had human orthologs, or family members, that have been documented elsewhere to carry arginine methylation. These results confirm arginine methylation as a widespread modification and Hmt1 as the major arginine methyltransferase in the S. cerevisiae cell. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  7. Novel Function of Lysine Methyltransferase G9a in the Regulation of Sox2 Protein Stability.

    Directory of Open Access Journals (Sweden)

    Jae-Young Lee

    Full Text Available G9a is a lysine methyltransferase (KMTase for histone H3 lysine 9 that plays critical roles in a number of biological processes. Emerging evidence suggests that aberrant expression of G9a contributes to tumor metastasis and maintenance of a malignant phenotype in cancer by inducing epigenetic silencing of tumor suppressor genes. Here, we show that G9a regulates Sox2 protein stability in breast cancer cells. When G9a lysine methyltransferase activity was chemically inhibited in the ER(+ breast cancer cell line MCF7, Sox2 protein levels were decreased. In addition, ectopic overexpression of G9a induced accumulation of Sox2. Changes in cell migration, invasion, and mammosphere formation by MCF7 cells were correlated with the activity or expression level of G9a. Ectopic expression of G9a also increased Sox2 protein levels in another ER(+ breast cancer cell line, ZR-75-1, whereas it did not affect Sox2 expression in MDA-MB-231 cells, an ER(- breast cancer cell line, or in glioblastoma cell lines. Furthermore, treatment of mouse embryonic stem cells with a KMT inhibitor, BIX-01294, resulted in a rapid reduction in Sox2 protein expression despite increased Sox2 transcript levels. This finding suggests that G9a has a novel function in the regulation of Sox2 protein stability in a cell type-dependent manner.

  8. Discovery of Potent and Selective Inhibitors for G9a-Like Protein (GLP) Lysine Methyltransferase

    Energy Technology Data Exchange (ETDEWEB)

    Xiong, Yan; Li, Fengling; Babault, Nicolas; Dong, Aiping; Zeng, Hong; Wu, Hong; Chen, Xin; Arrowsmith, Cheryl H.; Brown, Peter J.; Liu, Jing; Vedadi, Masoud; Jin, Jian

    2017-02-14

    G9a-like protein (GLP) and G9a are highly homologous protein lysine methyltransferases (PKMTs) sharing approximately 80% sequence identity in their catalytic domains. GLP and G9a form a heterodimer complex and catalyze mono- and dimethylation of histone H3 lysine 9 and nonhistone substrates. Although they are closely related, GLP and G9a possess distinct physiological and pathophysiological functions. Thus, GLP or G9a selective small-molecule inhibitors are useful tools to dissect their distinct biological functions. We previously reported potent and selective G9a/GLP dual inhibitors including UNC0638 and UNC0642. Here we report the discovery of potent and selective GLP inhibitors including 4 (MS0124) and 18 (MS012), which are >30-fold and 140-fold selective for GLP over G9a and other methyltransferases, respectively. The cocrystal structures of GLP and G9a in the complex with either 4 or 18 displayed virtually identical binding modes and interactions, highlighting the challenges in structure-based design of selective inhibitors for either enzyme.

  9. Structural Basis for the Methylation of G1405 in 16S rRNA by Aminoglycoside Resistance Methyltransferase Sgm from an Antibiotic Producer: a Diversity of Active Sites in m7G Methyltransferases

    Energy Technology Data Exchange (ETDEWEB)

    Husain, N.; Tkaczuk, K; Tulsidas, S; Kaminska, K; Cubrilo, S; Maravic -Vlahovicek, G; Bujnicki, J; Sivaraman, J

    2010-01-01

    Sgm (Sisomicin-gentamicin methyltransferase) from antibiotic-producing bacterium Micromonospora zionensis is an enzyme that confers resistance to aminoglycosides like gentamicin and sisomicin by specifically methylating G1405 in bacterial 16S rRNA. Sgm belongs to the aminoglycoside resistance methyltransferase (Arm) family of enzymes that have been recently found to spread by horizontal gene transfer among disease-causing bacteria. Structural characterization of Arm enzymes is the key to understand their mechanism of action and to develop inhibitors that would block their activity. Here we report the structure of Sgm in complex with cofactors S-adenosylmethionine (AdoMet) and S-adenosylhomocysteine (AdoHcy) at 2.0 and 2.1 {angstrom} resolution, respectively, and results of mutagenesis and rRNA footprinting, and protein-substrate docking. We propose the mechanism of methylation of G1405 by Sgm and compare it with other m{sup 7}G methyltransferases, revealing a surprising diversity of active sites and binding modes for the same basic reaction of RNA modification. This analysis can serve as a stepping stone towards developing drugs that would specifically block the activity of Arm methyltransferases and thereby re-sensitize pathogenic bacteria to aminoglycoside antibiotics.

  10. Conversion of nicotinic acid to trigonelline is catalyzed by N-methyltransferase belonged to motif B′ methyltransferase family in Coffea arabica

    Energy Technology Data Exchange (ETDEWEB)

    Mizuno, Kouichi, E-mail: koumno@akita-pu.ac.jp [Faculty of Bioresource Sciences, Akita Prefectural University, Akita City, Akita 010-0195 (Japan); Matsuzaki, Masahiro [Faculty of Bioresource Sciences, Akita Prefectural University, Akita City, Akita 010-0195 (Japan); Kanazawa, Shiho [Graduate School of Humanities and Sciences, Ochanomizu University, Otsuka, Bunkyo-ku, Tokyo 112-8610 (Japan); Tokiwano, Tetsuo; Yoshizawa, Yuko [Faculty of Bioresource Sciences, Akita Prefectural University, Akita City, Akita 010-0195 (Japan); Kato, Misako [Graduate School of Humanities and Sciences, Ochanomizu University, Otsuka, Bunkyo-ku, Tokyo 112-8610 (Japan)

    2014-10-03

    Graphical abstract: Trigonelline synthase catalyzes the conversion of nicotinic acid to trigonelline. We isolated and characterized trigonelline synthase gene(s) from Coffea arabica. - Highlights: • Trigonelline is a major compound in coffee been same as caffeine is. • We isolated and characterized trigonelline synthase gene. • Coffee trigonelline synthases are highly homologous with coffee caffeine synthases. • This study contributes the fully understanding of pyridine alkaloid metabolism. - Abstract: Trigonelline (N-methylnicotinate), a member of the pyridine alkaloids, accumulates in coffee beans along with caffeine. The biosynthetic pathway of trigonelline is not fully elucidated. While it is quite likely that the production of trigonelline from nicotinate is catalyzed by N-methyltransferase, as is caffeine synthase (CS), the enzyme(s) and gene(s) involved in N-methylation have not yet been characterized. It should be noted that, similar to caffeine, trigonelline accumulation is initiated during the development of coffee fruits. Interestingly, the expression profiles for two genes homologous to caffeine synthases were similar to the accumulation profile of trigonelline. We presumed that these two CS-homologous genes encoded trigonelline synthases. These genes were then expressed in Escherichiacoli, and the resulting recombinant enzymes that were obtained were characterized. Consequently, using the N-methyltransferase assay with S-adenosyl[methyl-{sup 14}C]methionine, it was confirmed that these recombinant enzymes catalyzed the conversion of nicotinate to trigonelline, coffee trigonelline synthases (termed CTgS1 and CTgS2) were highly identical (over 95% identity) to each other. The sequence homology between the CTgSs and coffee CCS1 was 82%. The pH-dependent activity curve of CTgS1 and CTgS2 revealed optimum activity at pH 7.5. Nicotinate was the specific methyl acceptor for CTgSs, and no activity was detected with any other nicotinate derivatives, or

  11. Correlation Does Not Imply Causation: Histone Methyltransferases, but Not Histone Methylation, SET the Stage for Enhancer Activation.

    Science.gov (United States)

    Pollex, Tim; Furlong, Eileen E M

    2017-05-18

    Although H3K4me1 is a pervasive "mark" of enhancers, its functional requirement for enhancer activity remains unclear. In this issue of Molecular Cell, Dorighi et al. (2017) show that in some contexts, the methyltransferase complex, rather than the H3K4me1 mark, is required for gene expression. Copyright © 2017 Elsevier Inc. All rights reserved.

  12. Characterization of cytosine methylated regions and 5-cytosine DNA methyltransferase (Ehmeth) in the protozoan parasite Entamoeba histolytica

    Science.gov (United States)

    Fisher, Ohad; Siman-Tov, Rama; Ankri, Serge

    2004-01-01

    The DNA methylation status of the protozoan parasite Entamoeba histolytica was heretofore unknown. In the present study, we developed a new technique, based on the affinity of methylated DNA to 5-methylcytosine antibodies, to identify methylated DNA in this parasite. Ribosomal DNA and ribosomal DNA circles were isolated by this method and we confirmed the validity of our approach by sodium bisulfite sequencing. We also report the identification and the characterization of a gene, Ehmeth, encoding a DNA methyltransferase strongly homologous to the human DNA methyltransferase 2 (Dnmt2). Immunofluorescence microscopy using an antibody raised against a recombinant Ehmeth showed that Ehmeth is concentrated in the nuclei of trophozoites. The recombinant Ehmeth has a weak but significant methyltransferase activity when E.histolytica genomic DNA is used as substrate. 5-Azacytidine (5-AzaC), an inhibitor of DNA methyltransferase, was used to study in vivo the role of DNA methylation in E.histolytica. Genomic DNA of trophozoites grown with 5-AzaC (23 µM) was undermethylated and the ability of 5-AzaC-treated trophozoites to kill mammalian cells or to cause liver abscess in hamsters was strongly impaired. PMID:14715927

  13. Downregulation of DNA (cytosine-5-)methyltransferase is a late event in NGF-induced PC12 cell differentiation.

    Science.gov (United States)

    Deng, J; Szyf, M

    1999-07-23

    DNA methylation patterns are a critical component of the epigenetic machinery that controls the expression of genetic programs in vertebrates. DNA methyltransferase gene (dnmt1) encodes the enzyme catalyzing the methylation of DNA during replication. We tested the hypothesis that the expression of dnmt1 is regulated with the developmental state of neuronal cells. We show that DNA methyltransferase (Dnmt1) activity is sharply reduced 4 days after induction of differentiation of PC12 cells with NGF. Similarly, the adult brain expresses reduced levels of Dnmt1 activity. We propose that the level of Dnmt1 is downregulated to adjust the activity of the DNA methyltransferase to a different role in mature post-mitotic neurons. Both the abundance of dnmt1 mRNA as well as the Dnmt1 polypeptide are downregulated. Downregulation of dnmt1 parallels other indicators of withdrawal from the cell cycle such as induction of p21, and downregulation of the S phase maker PCNA (proliferating cell nuclear antigen). The temporal pattern of downregulation of dnmt1 in nerve growth factor (NGF)-induced PC12 cells is different from myotube differentiation where downregulation of DNA methyltransferase and demethylation is an early event and was proposed to play a causal role in differentiation. We propose that NGF differentiation of PC12 cells represents a different paradigm of involvement of DNA methylation in terminal differentiation. Copyright 1999 Elsevier Science B.V.

  14. The Cfr rRNA methyltransferase confers resistance to Phenicols, Lincosamides, Oxazolidinones, Pleuromutilins, and Streptogramin A antibiotics

    DEFF Research Database (Denmark)

    Long, K. S.; Poehlsgaard, Jacob; Kehrenberg, C.

    2006-01-01

    A novel multidrug resistance phenotype mediated by the Cfr rRNA methyltransferase is observed in Staphylococcus aureus and Escherichia coli. The cfr gene has previously been identified as a phenicol and lincosamide resistance gene on plasmids isolated from Staphylococcus spp. of animal origin...

  15. Polymorphisms in O-methyltransferase genes are associated with stover cell wall digestibility in European maize (Zea mays L.)

    DEFF Research Database (Denmark)

    Brenner, Everton A; Zein, Imad; Chen, Yongsheng

    2010-01-01

    Background OMT (O-methyltransferase) genes are involved in lignin biosynthesis, which relates to stover cell wall digestibility. Reduced lignin content is an important determinant of both forage quality and ethanol conversion efficiency of maize stover. Results Variation in genomic sequences codi...

  16. Identification of the methyltransferase targeting C2499 in Deinococcus radiodurans 23S ribosomal RNA

    DEFF Research Database (Denmark)

    Nielsen, Julie Mundus; Flyvbjerg, Karen Freund; Kirpekar, Finn

    2016-01-01

    2499 reported. Using homology search, we identified the open reading frame DR_0049 as the primary candidate gene for the methyltransferase that modifies cytidine 2499. Mass spectrometric analysis demonstrated that recombinantly expressed DR0049 protein methylates E. coli cytidine 2499 both in vitro...

  17. Thiopurine Methyltransferase Enzyme Activity Determination before Treatment of Inflammatory Bowel Disease with Azathioprine: Effect on Cost and Adverse Events

    Directory of Open Access Journals (Sweden)

    Farzana A Sayani

    2005-01-01

    Full Text Available BACKGROUND: Azathioprine (AZA, used to treat inflammatory bowel disease (IBD, is metabolized by thiopurine methyltransferase (TPMT. The accumulation of individual metabolites varies because humans display genetic polymorphism for TPMT expression. Deficiencies in TPMT result in accumulation of toxic metabolites, followed by neutropenia and hepatic inflammation. Concern over acute toxicity frequently leads to under dosing and frequent monitoring tests and visits.

  18. Vagus nerve contributes to the development of steatohepatitis and obesity in phosphatidylethanolamine N-methyltransferase deficient mice

    NARCIS (Netherlands)

    Gao, Xia; van der Veen, Jelske N.; Zhu, Linfu; Chaba, Todd; Ordonez, Marta; Lingrell, Susanne; Koonen, Debby P. Y.; Dyck, Jason R. B.; Gomez-Munoz, Antonio; Vance, Dennis E.; Jacobs, Rene L.

    BACKGROUND & AIMS: Phosphatidylethanolamine N-methyltransferase (PEMT), a liver enriched enzyme, is responsible for approximately one third of hepatic phosphatidylcholine biosynthesis. When fed a high-fat diet (HFD), Pemt(-/-) mice are protected from HF-induced obesity; however, they develop

  19. Thirteen New Patients with Guanidinoacetate Methyltransferase Deficiency and Functional Characterization of Nineteen Novel Missense Variants in the GAMT Gene

    NARCIS (Netherlands)

    Mahmutoglu, S.; Ndika, J.D.T.; Kanhai, W.; de Villemeur, T.B.; Cheillan, D.; Christensen, E.; Dorison, N.; Hannig, V.; Hendriks, Y.M.C.; Hofstede, F.C.; Lion-Francois, L.; Lund, A.M.; Mundy, H.; Pitelet, G.; Raspall-Chaure, M.; Scott-Schwoerer, J.A.; Szakszon, K.; Valayannopoulos, V.; Williams, M.; Salomons, G.S.

    2014-01-01

    Guanidinoacetate methyltransferase deficiency (GAMT-D) is an autosomal recessively inherited disorder of creatine biosynthesis. Creatine deficiency on cranial proton magnetic resonance spectroscopy, and elevated guanidinoacetate levels in body fluids are the biomarkers of GAMT-D. In 74 patients, 50

  20. Catechol-O-methyltransferase gene and obsessive-compulsive symptoms in patients with recent-onset schizophrenia: Preliminary results

    NARCIS (Netherlands)

    Zinkstok, Janneke; van Nimwegen, Lonneke; van Amelsvoort, Therese; de Haan, Lieuwe; Yusuf, Maryan Abdulkadir; Baas, Frank; Linszen, Don

    2008-01-01

    The catechol-O-methyltransferase (COMT) gene is a candidate gene for schizophrenia because of its role in the breakdown of dopamine in the prefrontal cortex. The COMT gene contains a functional polymorphism changing enzyme activity that has been associated with some neuropsychiatric