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Sample records for acidic extracellular hydrolytic

  1. Isolation of Moderately Halophilic Pseudoalteromonas Producing Extracellular Hydrolytic Enzymes from Persian Gulf

    Ardakani, M. Roayaie; Poshtkouhian, A.; Amoozegar, M. A.; H. Zolgharnein

    2011-01-01

    Extracellular hydrolytic enzymes such as amylases, proteases, lipases and DNases have quite diverse potential usages in different areas such as food industry, biomedical sciences and chemical industries, also it would be of great importance to have available enzymes showing optimal activities at different values of salt concentrations and temperature. Halophiles are the most likely source of such enzymes, because not only their enzymes are salt-tolerant, but many are also thermotolerant. The ...

  2. Isolation of moderately halophilic pseudoalteromonas producing extracellular hydrolytic enzymes from persian gulf.

    Ardakani, M Roayaie; Poshtkouhian, A; Amoozegar, M A; Zolgharnein, H

    2012-03-01

    Extracellular hydrolytic enzymes such as amylases, proteases, lipases and DNases have quite diverse potential usages in different areas such as food industry, biomedical sciences and chemical industries, also it would be of great importance to have available enzymes showing optimal activities at different values of salt concentrations and temperature. Halophiles are the most likely source of such enzymes, because not only their enzymes are salt-tolerant, but many are also thermotolerant. The purpose of this study was isolation of hydrolytic extracellular enzyme producing halophilic bacteria from water and sediment of the Persian Gulf. Isolated bacteria from water and sediment were inoculated in media with concentration of 0-20% NaCl to determine the optimum salt concentration for growth, isolates were also inoculated in 4 types of solid medium containing substrates of 3 extracellular hydrolytic enzymes including amylase, Protease and Lipase, to determine the quantitative detection of enzyme production, selected strains after more accurate physiological and biochemical studies were identified regarding phylogeny and molecular characteristics using 16S rRNA technique. Isolated enzyme producing bacteria belong to Pseudoalteromonas genera. PMID:23450116

  3. Hydrolytic degradation behaviour of sucrose palmitate reinforced poly(lactic acid) nanocomposites.

    Valapa, Ravi Babu; G, Pugazhenthi; Katiyar, Vimal

    2016-08-01

    This work discusses the influence of novel biofiller, "sucrose palmitate" (SP) on the hydrolytic degradation behavior of poly(lactic acid) (PLA) nanocomposites. The influence of temperature and pH of the solution on the hydrolytic degradation behavior of PLA and PLA-SP nanocomposites was investigated. The variation in the crystallinity of PLA and PLA composites subjected to the hydrolytic degradation process is verified by XRD and DSC analysis. The morphological changes that occurred during the degradation process are observed by scanning electron microscopy (SEM). Thermo-gravimetric analysis confirms the loss of thermal stability of the neat PLA as well as composites after hydrolytic degradation process. Transparency measurements support the enhancement in opacity of both the PLA and PLA-SP nanocomposites with progress in hydrolytic degradation period. PMID:27095433

  4. Synthesis and Hydrolytic Degradation of Substituted Poly(DL-Lactic Acids

    Hideto Tsuji

    2011-08-01

    Full Text Available Non-substituted racemic poly(DL-lactic acid (PLA and substituted racemic poly(DL-lactic acids or poly(DL-2-hydroxyalkanoic acids with different side-chain lengths, i.e., poly(DL-2-hydroxybutanoic acid (PBA, poly(DL-2-hydroxyhexanoic acid (PHA, and poly(DL-2-hydroxydecanoic acid (PDA were synthesized by acid-catalyzed polycondensation of DL-lactic acid (LA, DL-2-hydroxybutanoic acid (BA, DL-2-hydroxyhexanoic acid (HA, and DL-2-hydroxydecanoic acid (DA, respectively. The hydrolytic degradation behavior was investigated in phosphate-buffered solution at 80 and 37 °C by gravimetry and gel permeation chromatography. It was found that the reactivity of monomers during polycondensation as monitored by the degree of polymerization (DP decreased in the following order: LA > DA > BA > HA. The hydrolytic degradation rate traced by DP and weight loss at 80 °C decreased in the following order: PLA > PDA > PHA > PBA and that monitored by DP at 37 °C decreased in the following order: PLA > PDA > PBA > PHA. LA and PLA had the highest reactivity during polymerization and hydrolytic degradation rate, respectively, and were followed by DA and PDA. BA, HA, PBA, and PHA had the lowest reactivity during polymerization and hydrolytic degradation rate. The findings of the present study strongly suggest that inter-chain interactions play a major role in the reactivity of non-substituted and substituted LA monomers and degradation rate of the non-substituted and substituted PLA, along with steric hindrance of the side chains as can be expected.

  5. HYDROLYTIC BREAKDOWN OF LACTOFERRICIN BY LACTIC ACID BACTERIA

    Lactoferricin is a 25 amino acid antimicrobial peptide domain that is liberated by pepsin digestion of lactoferrin in bovine milk. Along with its antibacterial properties, lactoferricin has also been reported to have immunostimulatory, antiviral, and anticarcinogenic effects. There is substantial ...

  6. Hydrolytically Degradable Hyaluronic Acid Hydrogels with Controlled Temporal Structures

    Sahoo, Sujata; CHUNG, CINDY; Khetan, Sudhir; Burdick, Jason A.

    2008-01-01

    Polysaccharides are being processed into biomaterials for numerous biological applications due to their native source in numerous tissues and biological functions. For instance, hyaluronic acid (HA) is found abundantly in the body, interacts with cells through surface receptors, and can regulate cellular behavior (e.g., proliferation, migration). HA was previously modified with reactive groups to form hydrogels that are degraded by hyaluronidases, either added exogenously or produced by cells...

  7. Hydrolytic activity of -alkoxide/acetato-bridged binuclear Cu(II) complexes towards carboxylic acid ester

    Weidong Jiang; Bin Xu; Zhen Xiang; Shengtian Huang; Fuan Liu; Ying Wang

    2013-09-01

    Two -alkoxide/acetate-bridged small molecule binuclear copper(II) complexes were synthesized, and used to promote the hydrolysis of a classic carboxylic acid ester, -nitrophenyl picolinate (PNPP). Both binuclear complexes exhibited good hydrolytic reactivity, giving rise to . 15547- and 17462-fold acceleration over background value for PNPP hydrolysis at neutral conditions, respectively. For comparing, activities of the other two mononuclear analogues were evaluated, revealing that binuclear complexes show approximately 150- and 171-fold kinetic advantage over their mononuclear analogues.

  8. Influence if acidity and concentration of aqueous uranyl nitrate solutions on the efficiency of uranium absorption by hydrolytic wood lignin

    Efficiency of uranium(VI) absorption by hydrolytic wood lignin from uranyl nitrate aqueous solutions under static conditions at room temperature, depending on solution acidity and uranium concentration, was studied using the methods of elementary analysis and IR spectroscopy. It was ascertained that hydrolytic lignin manifests a high ability to strong uranium(VI) absorption from low-acid and alkaline solutions. Interaction of uranium(VI) and hydrolytic lignin occurs both according to ion exchange mechanism and at the expense of donor-acceptor bonds formation

  9. Thermal and Hydrolytic Stability of Dithiophophoric Acids Stabilité thermique et hydrolytique des acides dithiophosphoriques

    Ivanov S. K.

    2006-11-01

    Full Text Available The thermal and hydrolytic stability of dibutyl-, diisobutyl- and diisooctyldithiophosphoric acids have been studied in the range of 40-80°C. The reaction progress was followed by the H2S evolution kinetics in volumetric equipment at constant pressure, equal to the barometric one. The kinetic parameters of the process have been determined at various acid water ratios - maximum rates of gas evolution, activation energy and the kinetic order. It has been shown that the hydrolysis proceeds mainly along the sulph-hydrate group of the acid (up to temperatures of 60°C, while at higher temperatures the ester groups and the thion sulphur are attacked too. The thermal stability of dithiophosphoric acids has been studied in the presence of stainless steel, lead and copper plates. It has been shown that stainless steel doesn't affect the process rate, while copper and especially lead increase the gas evolution and decrease the activation energy. A model describing the reaction progress at the interfacial surfaces - organic phase, water and metal surface - has been developed. Based on these data a conclusion is advanced, related to the manifacture technology of antioxidant, anticorrosion and antiwear additives of the zinc dialkyldithiophosphate type. It is pointed out that the neutralization process of dithiophosphoric acids should be carried out in a stainless steel reactor at tempeatures below 70°C. Le présent article s'efforce d'élucider l'influence de l'eau, de la température et des surfaces métalliques sur la stabilité thermique et hydrolytique des acides dibutyl-, diisobutyl- et diisooctyldithiophosphoriques, à la lumière du concept d'interaction.

  10. Scanning electron microscopic study of the hydrolytic degradation of poly(glycolic acid) suture

    This article reports the morphological observations on the surface changes of poly-(glycolic acid) sutures which have been exposed to various dosages of gamma irradiation (0, 2.5, 5.0, 10, 20 and 40 Mrad) and duration of immersion (0, 7, 14, 28, 48, 60, and 90 days) in a physiological saline buffer. The most important gross morphological characteristics of PGA suture hydrolytic degradation is the formation of surface cracks on the filaments. The regularity of the surface cracks increased with an increase in the gamma irradiation and the duration of hydrolysis. Surface cracks were not observed in irradiated sutures that had not been subjected to hydrolytic degradation. The arrangement of the surface cracks, their orientation on the filaments, and the direction of crack propagation provide very useful information for depicting the mechanism of hydrolytic degradation in this class of fibrous material. The microfibrillar model of fiber structure has been used as the basis for the proposed degradation mechanism of PGA in vitro. It is believed that hydrolysis occurs initially in the amorphous regions sandwiched between two crystalline zones, as tie-chain segments, free chain ends, and chain folds in these regions degrade into fragments. As degradation proceeds, the size of the fragments reaches the stage at which they can be dissolved into the buffer medium. This dissolution removes the fragments from the amorphous regions, and surface cracks appeared

  11. Extracellular acid proteases produced by Saccharomycopsis lipolytica.

    T. Yamada; Ogrydziak, D M

    1983-01-01

    Saccharomycopsis lipolytica CX161-1B produced at least three extracellular acid proteases during exponential growth in medium containing glycerol, Difco Proteose Peptone, and mineral salts at pH 3.4 (Difco Laboratories, Detroit, Mich.). Little extracellular acid protease activity was produced with glutamic acid as the sole nitrogen source, somewhat higher levels were obtained with peptone, and much higher levels were obtained with Difco Proteose Peptone. The relative amounts of the three prot...

  12. Effects of methyl-CD and humic acid on hydrolytic degradation of the herbicide diclofop-methyl

    CAI Xi-yun; WEN Yue-zhong; ZHONG Tian-xiang; LIU Wei-ping

    2005-01-01

    Hydrolytic degradation of the herbicide diclofop-methyl was investigated in the multi-pH deionized water, natural aquatic systems and soil suspensions. Resulting data indicated that the herbicide was stable in the acidic and nearly neutral solutions for at least 15 d. The herbicide diclofop-methyl rapidly dissipated in the natural aquatic systems and soil suspensions with half-lives less than 4 d. Methyl-CD(partially methylated β-cyclodextrin) improved its hydrolytic degradation in the pH 8 deionized water and natural aquatic systems while humic acid inhibited its hydrolytic degradation at the same conditions. But dissolved organic matter in the natural aquatic systems and soil suspensions increased its hydrolysis. Two catalysis mechanisms were introduced to describe the effects of cyclodextrin and organic matter on its hydrolytic metabolism. Though inorganic ions maybe improved its hydrolysis reaction in the natural aquatic systems, Fe2+ and Cu2+did not form complexes with the herbicide and had poor influences on its hydrolytic degradation whether cyclodextrin was added or not.

  13. Ultra-Small Fatty Acid-Stabilized Magnetite Nanocolloids Synthesized by In Situ Hydrolytic Precipitation

    Kheireddine El-Boubbou

    2015-01-01

    Full Text Available Simple, fast, large-scale, and cost-effective preparation of uniform controlled magnetic nanoparticles remains a major hurdle on the way towards magnetically targeted applications at realistic technical conditions. Herein, we present a unique one-pot approach that relies on simple basic hydrolytic in situ coprecipitation of inexpensive metal salts (Fe2+ and Fe3+ compartmentalized by stabilizing fatty acids and aided by the presence of alkylamines. The synthesis was performed at relatively low temperatures (~80°C without the use of high-boiling point solvents and elevated temperatures. This method allowed for the production of ultra-small, colloidal, and hydrophobically stabilized magnetite metal oxide nanoparticles readily dispersed in organic solvents. The results reveal that the obtained magnetite nanoparticles exhibit narrow size distributions, good monodispersities, high saturation magnetizations, and excellent colloidal stabilities. When the [fatty acid] : [Fe] ratio was varied, control over nanoparticle diameters within the range of 2–10 nm was achieved. The amount of fatty acid and alkylamine used during the reaction proved critical in governing morphology, dispersity, uniformity, and colloidal stability. Upon exchange with water-soluble polymers, the ultra-small sized particles become biologically relevant, with great promise for theranostic applications as imaging and magnetically targeted delivery vehicles.

  14. Hyaluronic acid changes of the extracellular matrix in colon carcinoma

    Arab MR.; Allahyari A.; Sargolzaie Aval F.; Rafighdoost H; Karimi M

    2007-01-01

    Background: The extracellular matrix is a complex three-dimensional network of proteins and glycosaminoglycans, which have important roles in cellular physiology and cell-cell and cell-extracellular matrix interactions. Any changes in the extracellular matrix of tumors may be implicated in cellular transformation and metastasis. The aim of the present study was to identify changes in the hyaluronic acid of the stroma of colonic carcinoma.Methods: Paraffin blocks of 30 patients with colon carc...

  15. Production of extracellular fatty acid using engineered Escherichia coli

    Liu Hui

    2012-04-01

    Full Text Available Abstract Background As an alternative for economic biodiesel production, the microbial production of extracellular fatty acid from renewable resources is receiving more concerns recently, since the separation of fatty acid from microorganism cells is normally involved in a series of energy-intensive steps. Many attempts have been made to construct fatty acid producing strains by targeting genes in the fatty acid biosynthetic pathway, while few studies focused on the cultivation process and the mass transfer kinetics. Results In this study, both strain improvements and cultivation process strategies were applied to increase extracellular fatty acid production by engineered Escherichia coli. Our results showed overexpressing ‘TesA and the deletion of fadL in E. coli BL21 (DE3 improved extracellular fatty acid production, while deletion of fadD didn’t strengthen the extracellular fatty acid production for an undetermined mechanism. Moreover, the cultivation process controls contributed greatly to extracellular fatty acid production with respect to titer, cell growth and productivity by adjusting the temperature, adding ampicillin and employing on-line extraction. Under optimal conditions, the E. coli strain (pACY-‘tesA-ΔfadL produced 4.8 g L−1 extracellular fatty acid, with the specific productivity of 0.02 g h−1 g−1dry cell mass, and the yield of 4.4% on glucose, while the ratios of cell-associated fatty acid versus extracellular fatty acid were kept below 0.5 after 15 h of cultivation. The fatty acids included C12:1, C12:0, C14:1, C14:0, C16:1, C16:0, C18:1, C18:0. The composition was dominated by C14 and C16 saturated and unsaturated fatty acids. Using the strain pACY-‘tesA, similar results appeared under the same culture conditions and the titer was also much higher than that ever reported previously, which suggested that the supposedly superior strain did not necessarily perform best for the efficient production of desired

  16. Generation of organic acids and monosaccharides by hydrolytic and oxidative transformation of food processing residues.

    Fischer, Klaus; Bipp, Hans-Peter

    2005-05-01

    Carbohydrate-rich biomass residues, i.e. sugar beet molasses, whey powder, wine yeast, potato peel sludge, spent hops, malt dust and apple marc, were tested as starting materials for the generation of marketable chemicals, e.g. aliphatic acids, sugar acids and mono-/disaccharides. Residues were oxidized or hydrolyzed under acidic or alkaline conditions applying conventional laboratory digestion methods and microwave assisted techniques. Yields and compositions of the oxidation products differed according to the oxidizing agent used. Main products of oxidation by 30% HNO(3) were acetic, glucaric, oxalic and glycolic acids. Applying H(2)O(2)/CuO in alkaline solution, the organic acid yields were remarkably lower with formic, acetic and threonic acids as main products. Gluconic acid was formed instead of glucaric acid throughout. Reaction of a 10% H(2)O(2) solution with sugar beet molasses generated formic and lactic acids mainly. Na(2)S(2)O(8) solutions were very inefficient at oxidizing the residues. Glucose, arabinose and galactose were formed during acidic hydrolysis of malt dust and apple marc. The glucose content reached 0.35 g per gram of residue. Important advantages of the microwave application were lower reaction times and reduced reagent demands. PMID:15607197

  17. New Biocompatible Polyesters Derived from α-Amino Acids: Hydrolytic Degradation Behavior

    Jeoshua Katzhendler

    2010-10-01

    Full Text Available New polymers were synthesized from α-hydroxy acids derived from the natural amino acids Ile, Leu, Phe, and Val, combined with lactic acid, glycolic acid and 6-hydroxyhexanoic acid by direct condensation. The toxicity was determined and the degradation process of these polyesters was investigated under physiological conditions by analyzing the composition of the degraded polymers and the oligomers cleaved in the buffer medium. The polymers were found to be non toxic to two cell lines. Polymers displayed a biphasic degradation behavior. In most cases, a linear relationship was found between the weight loss constant and the hydrophobicity of the polymers, Log P. Regarding the second stage of weight loss, it is apparent that polymers derived from α-hydroxy(Lisoleucine ((LHOIle and α-hydroxy(LValine ((LHOVal degraded much faster than those derived from α-hydroxy(Lleucine ((LHOLeu and α-hydroxy(Lphenylalanine ((LHOPhe, probably due to different spatial orientation of the side chains. Copolymers of 6-hydroxyhexanoic acid displayed slow degradation rates as expected, whereas the degradation profile of copolymers of lactic acid was similar to the other homopolymers. These new polyesters may serve as potential biocompatible materials for medical applications.

  18. Biocatalyzed approach for the surface functionalization of poly(L-lactic acid) films using hydrolytic enzymes.

    Pellis, Alessandro; Acero, Enrique Herrero; Weber, Hansjoerg; Obersriebnig, Michael; Breinbauer, Rolf; Srebotnik, Ewald; Guebitz, Georg M

    2015-09-01

    Poly(lactic acid) as a biodegradable thermoplastic polyester has received increasing attention. This renewable polyester has found applications in a wide range of products such as food packaging, textiles and biomedical devices. Its major drawbacks are poor toughness, slow degradation rate and lack of reactive side-chain groups. An enzymatic process for the grafting of carboxylic acids onto the surface of poly(L-lactic acid) (PLLA) films was developed using Candida antarctica lipase B as a catalyst. Enzymatic hydrolysis of the PLLA film using Humicola insolens cutinase in order to increase the number of hydroxyl and carboxylic groups on the outer polymer chains for grafting was also assessed and showed a change of water contact angle from 74.6 to 33.1° while the roughness and waviness were an order of magnitude higher in comparison to the blank. Surface functionalization was demonstrated using two different techniques, (14) C-radiochemical analysis and X-ray photoelectron spectroscopy (XPS) using (14) C-butyric acid sodium salt and 4,4,4-trifluorobutyric acid as model molecules, respectively. XPS analysis showed that 4,4,4-trifluorobutyric acid was enzymatically coupled based on an increase of the fluor content from 0.19 to 0.40%. The presented (14) C-radiochemical analyses are consistent with the XPS data indicating the potential of enzymatic functionalization in different reaction conditions. PMID:25963883

  19. Extracellular Nucleic Acids in Blood of Patients with Chronic Obstructive Pulmonary Disease

    Larissa E. Muravlyova; Vilen B. Molotov-Luchanskiy; Dmitriy A. Klyuyev; Ryszhan E. Bakirova; Ludmila A. Demidchik; Evgeniya A. Kolesnikova

    2013-01-01

    The concentrations of extracellular nucleic acids and acid-soluble precursors of nucleic acids in blood of patients with different forms and severity of chronic obstructive pulmonary disease (COPD) were evaluated. The significant increase of the content of extracellular RNA and acid-soluble precursors of nucleic acids in plasma of patients with COPD was detected. The decrease of extracellular RNA in plasma of patients with COPD worsening was diagnosed. Extracellular DNA in plasma and red bloo...

  20. Ascorbic acid: Nonradioactive extracellular space marker in canine heart

    The distribution pattern of ascorbic acid and L-[14C]ascorbic acid in myocardial tissue was compared with those of the classical radioactive extracellular space markers [3H]-inulin, [3H]sucrose, and Na82Br. A new polarographic techniques was developed for analogue registration of ascorbic acid concentration in coronary venous blood. The kinetic data of the markers were studied in an open-chest canine heart preparation during a constant tracer infusion of up to 9 min. Distribution volumes were calculated based on the mean transit time method of Zierler. The distribution volume of ascorbic acid as well as of L-[14C]ascorbic acid in myocardial tissue agreed closely with those of [3H]inulin and [3H]sucrose as well as 82Br. The obtained kinetic data confirmed that ascorbic acid exhibits the physicochemical properties of an extracellular space marker, though this compound was shown to leak slowly into myocardial cells. Favorable attributes of this indicator are its low molecular weight, high diffusibility in interstitial fluid, low binding affinity to macromolecules, and high transcapillary as well as low transplasmalemmal penetration rate. Therefore, this nonradioactive marker can be applied in a safe and simple fashion, and without untoward side effects in experimental animals as well as in patients

  1. Hyaluronic acid changes of the extracellular matrix in colon carcinoma

    Arab MR.

    2007-10-01

    Full Text Available Background: The extracellular matrix is a complex three-dimensional network of proteins and glycosaminoglycans, which have important roles in cellular physiology and cell-cell and cell-extracellular matrix interactions. Any changes in the extracellular matrix of tumors may be implicated in cellular transformation and metastasis. The aim of the present study was to identify changes in the hyaluronic acid of the stroma of colonic carcinoma.Methods: Paraffin blocks of 30 patients with colon carcinoma (10 patients at each histological grade were chosen from the pathology file of Khatam-Al-Anbia Hospital in Zahedan, Iran. Tissue sections (5-6 micrometers thick were stained with hematoxylin-eosin and the alcian blue critical electrolyte concentration histochemical technique at pH=5.8. The intensity of the staining in each section was graded as 1, 2 or 3, referring to low, moderate or severe staining reactivities, respectively. Statistical data was analyzed with nonparametric tests by SPSS (ver. 10 and histopathological reports were prepared.Results: The results of this study showed that there is a good correlation between histopathological grading and staining intensity of tumoral stroma for hyaluronic acid (p<0.005. Analysis using the Mann Whitney test revealed significant differences between staining grades 1 and 3 and grades 2 and 3 (p<0.005 and p<0.002, respectively, although there was no significant difference between staining grades 1 and 2 for hyaluronic acid.Conclusions: The difference in staining intensity of the stroma in colon carcinoma is a result of different amounts of hyaluronic acid in stroma, indicating that increased levels of hyaluronic acid are associated with the invasion and metastasis of neoplastic cells.

  2. Extracellular Nucleic Acids in Blood of Patients with Chronic Obstructive Pulmonary Disease

    Larissa E. Muravlyova

    2013-01-01

    Full Text Available The concentrations of extracellular nucleic acids and acid-soluble precursors of nucleic acids in blood of patients with different forms and severity of chronic obstructive pulmonary disease (COPD were evaluated. The significant increase of the content of extracellular RNA and acid-soluble precursors of nucleic acids in plasma of patients with COPD was detected. The decrease of extracellular RNA in plasma of patients with COPD worsening was diagnosed. Extracellular DNA in plasma and red blood cells of patients didn’t change significantly. The article examines the mechanisms of extracellular nucleic acids increase in blood of COPD patients, studies the possible role of extracellular RNA in development of coagulation disorders in COPD patients. The further research of the role of extracellular nucleic acids and their precursors in COPD progression is required

  3. Enzimas hidrolíticas extracelulares de isolados de rizóbia nativos da Amazônia Central, Amazonas, Brasil Extracellular hydrolytic enzymes in indigenous strains of rhizobia in Central Amazonia, Amazonas, Brazil

    Arlem Nascimento de Oliveira

    2006-12-01

    Full Text Available A associação rizóbia x leguminosa contribui para enriquecer o solo com nitrogênio por meio da fixação biológica. Entretanto, pouco se conhece a respeito do perfil enzimático desses microrganismos. Nesse contexto, a presente investigação propõe avaliar a produção de enzimas hidrolíticas extracelulares por isolados de rizóbia nativos da Amazônia Central. Essa triagem constitui o primeiro passo na seleção de microrganismos nativos que são potencialmente exploráveis como produtores de enzimas. Foram testados 67 isolados nativos de rizóbia para as atividades amilolítica, celulolítica, lactolítica, lipolítica, pectinolítica e proteolítica, em meio YMA modificado. A atividade ureolítica foi detectada em meio ágar-uréia. As bactérias isoladas dos nódulos de feijão caupi mostraram maior capacidade em produzir enzimas do que os isolados bacterianos de soja. De todas as enzimas hidrolíticas avaliadas, apenas a pectinase não foi detectada neste estudo. Amilase (32,8%, protease (28,4%, urease (20,9% e carboximetilcelulase (9,0% foram as enzimas mais freqüentes produzidas pelos isolados. Neste trabalho, apenas as enzimas amilase e protease variaram significativamente entre os isolados de rizóbia. Os isolados INPA R-926 e INPA R-915 exibiram os maiores índices amilolíticos (IE = 3,1 e proteolíticos (IE = 6,6, respectivamente. Este estudo revelou alguns isolados de rizóbia nativos da Amazônia Central como fontes promissoras de enzimas de importância industrial para uso biotecnológico.Legumes enrich the soil by contributing nitrogen through symbiotic biological nitrogen fixation by rhizobia bacteria. However, very little is known about the extracellular enzymatic profile of these microorganisms. In this context, the production of extracellular hydrolytic enzymes by indigenous strains of rhizobia in Central Amazonia was evaluated. This screening constitutes the first step in selecting indigenous microorganisms that are

  4. ICH GUIDANCE IN PRACTICE: DEVELOPMENT OF A VALIDATED STABILITY-INDICATING HIGH-PERFORMANCE LIQUID CHROMATOGRAPHIC ASSAY METHOD FOR FEBUXOSTAT AND DEGRADATION KINETIC STUDY IN ACID HYDROLYTIC CONDITION

    Megha V. Sheth* and Jigar J. Pandya

    2013-02-01

    Full Text Available The degradation behavior of Febuxostat was investigated under different stress degradation (hydrolytic, oxidative, photolytic and thermal conditions recommended by International Conference on Harmonization (ICH using HPLC and LCMS. Febuxostat was found to degrade significantly in acidic and alkaline conditions as well as in neutral hydrolysis. The drug was stable to dry heat, photolytic degradation and under oxidative condition. Resolution of drug and the degradation products formed under different stress studies were successfully achieved on a C-18 column utilizing Methanol- water (with 0.02%v/v TFA in the ratio of 95:5 and at the detection wavelength of 315 nm. The method was validated with respect to linearity, precision, accuracy, selectivity and specificity. The degradation kinetic of Febuxostat in acidic condition at different temperature was studied. The reaction order for Febuxostat in aqueous solvent system followed pseudo first order degradation kinetic. The catalytic rate constant and half-life at particular condition were determined. The Arrhenius plot showed the temperature dependence of Febuxostat.

  5. The Influence of Solid-State Drawing on Mechanical Properties and Hydrolytic Degradation of Melt-Spun Poly(Lactic Acid (PLA Tapes

    Fang Mai

    2015-12-01

    Full Text Available The influence of solid-state drawing on the morphology of melt-spun poly(l-lactic acid (PLLA tapes, and the accompanying changes in mechanical and degradation behaviour have been studied. Mechanical properties are found to be strongly dependent on both applied draw ratio and drawing temperature. Moduli of these highly oriented tapes are significantly increased compared to as-extruded tapes at both ambient and elevated temperatures. Interestingly, drawing leads to a significant increase in elongation to break (~3 times and toughness (~13 times compared to as-extruded tapes. Structural and morphological characterization indicates strain-induced crystallization as well as an increase in orientation of the crystalline phase at small strains. Upon further stretching, an “overdrawing” regime is observed, with decreased crystalline orientation due to the breakage of existing crystals. For fixed draw ratios, a significant increase in Young’s modulus and tensile strength is observed with increasing drawing temperature, due to a higher crystallinity and orientation obtained for tapes drawn at higher temperatures. FT-IR results indicate no crystal transformation after drawing, with the α-form being observed in all tapes. Hydrolytic degradability of PLLA was significantly reduced by solid-state drawing.

  6. Synthesis of a Stable Primary-Alkyl-Substituted Selenenyl Iodide and Its Hydrolytic Conversion to the Corresponding Selenenic Acid.

    Sase, Shohei; Kakimoto, Ryo; Kimura, Ryutaro; Goto, Kei

    2015-01-01

    A primary-alkyl-substituted selenenyl iodide was successfully synthesized through oxidative iodination of a selenol with N-iodosuccinimide by taking advantage of a cavity-shaped steric protection group. The selenenyl iodide exhibited high thermal stability and remained unchanged upon heating at 100 °C for 3 h in [D₈]toluene. The selenenyl iodide was reduced to the corresponding selenol by treatment with dithiothreitol. Hydrolysis of the selenenyl iodide under alkaline conditions afforded the corresponding selenenic acid almost quantitatively, corroborating the chemical validity of the recent proposal that hydrolysis of a selenenyl iodide to a selenenic acid is potentially involved in the catalytic mechanism of an iodothyronine deiodinase. PMID:26633336

  7. Synthesis of a Stable Primary-Alkyl-Substituted Selenenyl Iodide and Its Hydrolytic Conversion to the Corresponding Selenenic Acid

    Shohei Sase

    2015-12-01

    Full Text Available A primary-alkyl-substituted selenenyl iodide was successfully synthesized through oxidative iodination of a selenol with N-iodosuccinimide by taking advantage of a cavity-shaped steric protection group. The selenenyl iodide exhibited high thermal stability and remained unchanged upon heating at 100 °C for 3 h in [D8]toluene. The selenenyl iodide was reduced to the corresponding selenol by treatment with dithiothreitol. Hydrolysis of the selenenyl iodide under alkaline conditions afforded the corresponding selenenic acid almost quantitatively, corroborating the chemical validity of the recent proposal that hydrolysis of a selenenyl iodide to a selenenic acid is potentially involved in the catalytic mechanism of an iodothyronine deiodinase.

  8. Production of extracellular nucleic acids by genetically altered bacteria in aquatic-environment microcosms

    The factors which affect the production of extracellular DNA by genetically altered strains of Escherichia coli, Pseudomonas aeruginosa, Pseudomonas cepacia, and Bradyrhizobium japonicum in aquatic environments were investigated. Cellular nucleic acids were labeled in vivo by incubation with [3H]thymidine or [3H]adenine, and production of extracellular DNA in marine waters, artificial seawater, or minimal salts media was determined by detecting radiolabeled macromolecules in incubation filtrates. The presence or absence of the ambient microbial community had little effect on the production of extracellular DNA. Three of four organisms produced the greatest amounts of extracellular nucleic acids when incubated in low-salinity media (2% artificial seawater) rather than high-salinity media (10 to 50% artificial seawater). The greatest production of extracellular nucleic acids by P. cepacia occurred at pH 7 and 37 degree C, suggesting that extracellular-DNA production may be a normal physiologic function of the cell. Incubation of labeled P. cepacia cells in water from Bimini Harbor, Bahamas, resulted in labeling of macromolecules of the ambient microbial population. Collectively these results indicate that (i) extracellular-DNA production by genetically altered bacteria released into aquatic environments is more strongly influenced by physicochemical factors than biotic factors, (ii) extracellular-DNA production rates are usually greater for organisms released in freshwater than marine environments, and (iii) ambient microbial populations can readily utilize materials released by these organisms

  9. Production of extracellular proteolytic enzymes by Beauveria bassiana

    Józefa Chrzanowska; Maria Kołaczkowska

    2014-01-01

    The production of proteolytic enzymes by two strains of Beauveria bassiana 278, B. bassiana 446 and one strain of Ascosphera apis 496 was analysed. It was demonstrated that the strain of B. bassiana 278 proved to be the best producer of basic and acid proteases. The influence of different environmental factors such as nitrogen and carbon sources on the production of extracellular hydrolytic enzymes was assessed. In addition the acid protease from B. bassiana was partially characterized.

  10. Production of extracellular proteolytic enzymes by Beauveria bassiana

    Józefa Chrzanowska

    2014-08-01

    Full Text Available The production of proteolytic enzymes by two strains of Beauveria bassiana 278, B. bassiana 446 and one strain of Ascosphera apis 496 was analysed. It was demonstrated that the strain of B. bassiana 278 proved to be the best producer of basic and acid proteases. The influence of different environmental factors such as nitrogen and carbon sources on the production of extracellular hydrolytic enzymes was assessed. In addition the acid protease from B. bassiana was partially characterized.

  11. Concomitant extracellular accumulation of alpha-keto acids and higher alcohols by Zygosaccharomyces rouxii.

    Van Der Sluis, Catrinus; Rahardjo, Yovita S P; Smit, Bart A; Kroon, Pieter J; Hartmans, Sybe; Ter Schure, Eelko G; Tramper, Johannes; Wijffels, Renéh

    2002-01-01

    Alpha-keto acids are key intermediates in the formation of higher alcohols, important flavor components in soy sauce, and produced by the salt-tolerant yeast Zygosaccharomyces rouxii. Unlike most of the higher alcohols, the alpha-keto acids are usually not extracellularly accumulated by Z. rouxii when it is cultivated with ammonium as the sole nitrogen source. To facilitate extracellular accumulation of the alpha-keto acids from aspartate-derived amino acid metabolism, the amino acids valine, leucine, threonine and methionine were exogenously supplied during batch and A-star cultivations of (routants of) Z. rouxii. It was shown that all alpha-keto acids from the aspartate-derived amino acid metabolism, except alpha-ketobutyrate, could be extracellularly accumulated. In addition, it appeared from the concomitant extracellular accumulation of alpha-keto acids and higher alcohols that in Z. rouxii, valine, leucine and methionine were converted via Ehrlich pathways similar to those in Saccharomyces cerevisiae. Unlike these amino acids, threonine was converted via both the Ehrlich and amino acid biosynthetic pathways in Z. rouxii. PMID:16233175

  12. Hydrolytically stable titanium-45

    Severin, Gregory; Fonslet, Jesper; Zhuravlev, Fedor

    2014-01-01

    . The high cross-section and production rates on an unenriched metal foil target contribute to make 45Ti an ideal PET radionuclide. In order to bring 45Ti to even a preclinical plat-form, the hydrolytic instability of aqueous Ti(IV) needs to be addressed. Recently, the groups of Edit Tshuva (Hebrew......, similar to ion-pairing, and eluted with isopropanol. iv. 45Ti was extracted onto a polystyrene based 1,3 diol resin (RAPP polymers) and labeling commenced on the column. Radiolabeling was slightly different in each condition, but in general the salan and dipic ligands were added to the 45Ti in pyridine...

  13. Effects of Lanthanum on Hydrolytic Enzyme Activities in Red Soil

    褚海燕; 朱建国; 谢祖彬; 李振高; 曹志洪; 曾青; 林先贵

    2002-01-01

    The effects of La on some hydrolytic enzyme activities in red soil were studied in incubation and pot culture experiments. In the incubation experiment, La slightly stimulates the activities of urease and acidic phosphatase in soil and strongly stimulates sucrase activity in soil. In the pot culture experiment, La stimulates the activities of urease, acidic phosphatase and sucrase to different degrees. The stimulative effects of rare earth elements (REE) on hydrolytic enzyme activities in soil may result in increasing yield of crops.

  14. Extracellular depolymerization of hyaluronic acid in cultured human skin fibroblasts

    The chain length of [3H]hyaluronic acid synthesized by cultivating human skin fibroblasts in the presence of [3H]glucosamine was investigated. [3H]Hyaluronic acid obtained from the matrix fraction was excluded from a Sepharose CL-2B column irrespective of the incubation period, whereas that from the medium was depolymerized into a constant chain length (Mr = 40,000). The reducing and non-reducing terminals of the depolymerized hyaluronic acid were N-acetylglucosamine and glucuronic acid, respectively. Prolonged incubation produced no oligosaccharides as shown by examination of hyaluronidase digests, suggesting the presence of a novel endo-beta-N-acetylglucosaminidase in cultured human skin fibroblasts

  15. Extracellular depolymerization of hyaluronic acid in cultured human skin fibroblasts

    Nakamura, T.; Takagaki, K.; Kubo, K.; Morikawa, A.; Tamura, S.; Endo, M. (Hirosaki Univ. School of Medicine (Japan))

    1990-10-15

    The chain length of ({sup 3}H)hyaluronic acid synthesized by cultivating human skin fibroblasts in the presence of ({sup 3}H)glucosamine was investigated. ({sup 3}H)Hyaluronic acid obtained from the matrix fraction was excluded from a Sepharose CL-2B column irrespective of the incubation period, whereas that from the medium was depolymerized into a constant chain length (Mr = 40,000). The reducing and non-reducing terminals of the depolymerized hyaluronic acid were N-acetylglucosamine and glucuronic acid, respectively. Prolonged incubation produced no oligosaccharides as shown by examination of hyaluronidase digests, suggesting the presence of a novel endo-beta-N-acetylglucosaminidase in cultured human skin fibroblasts.

  16. De novo design, synthesis and spectroscopic characterization of chiral benzimidazole-derived amino acid Zn(II) complexes: Development of tryptophan-derived specific hydrolytic DNA artificial nuclease agent

    Parveen, Shazia; Arjmand, Farukh

    2012-01-01

    Novel ternary dizinc(II) complexes 1- 3, derived from 1,2-bis(1H-benzimidazol-2-yl)ethane-1,2-diol and L-form of amino acids (viz., tryptophan, leucine and valine) were synthesized and characterized by spectroscopic (IR, 1H NMR, UV-vis, ESI-MS) and other analytical methods. To evaluate the biological preference of chiral drugs for inherently chiral target DNA, interaction of 1- 3 with calf thymus DNA in Tris-HCl buffer was studied by various biophysical techniques which reveal that all these complexes bind to CT DNA non-covalently via electrostatic interaction. The higher Kb value of L-tryptophan complex 1 suggested greater DNA binding propensity. Further, to evaluate the mode of action at the molecular level, interaction studies of complexes 1 and 2 with nucleotides (5'-GMP and 5'-TMP) were carried out by UV-vis titrations, 1H and 31P NMR which implicates the preferential selectivity of these complexes to N3 of thymine rather than N7 of guanine. Furthermore, complex 1 exhibits efficient DNA cleavage with supercoiled pBR322. The complex 1 cleaves DNA efficiently involving hydrolytic cleavage pathway. Such chiral synthetic hydrolytic nucleases with asymmetric centers are gaining considerable attention owing to their importance in biotechnology and drug design, in particular to cleave DNA with sequence selectivity different from that of the natural enzymes.

  17. Structure of an Extracellular Polysaccharide from a Strain of Lactic Acid Bacteria

    顾笑梅; 马桂荣; 吴厚铭

    2003-01-01

    A new extracellular polysaccharide (EPS-I) isolated and purified from Z222, a strain of Lactic acid bacteria has been investigated. Sugar composition analysis, methylation analysis and 1H NMR and 13C NMR spectroscopy reveal that the EPS-I is composed of a pentasaccharide repeating unit. The sequence of sugar residue was determined by using two-dlmensional NMR spectroscopy, including heteronudear multiple-bond correlation(HMBC) and nuclear overhauser effect spectroscopy (NOESY).

  18. Hydrolytic enzyme activity in landfilled refuse

    Palmisano, A.C.; Schwab, B.S.; Maruscik, D.A. (Environmental Safety Dept., Procter and Gamble Co., Ivorydale Technical Center, Cincinnati, OH (United States))

    1993-03-01

    Extracellular hydrolytic enzyme activity was assayed in 28 refuse samples excavated from 14 bore holes in Fesh Kills Landfill, Staten Island, N.Y. Esterases, proteases and amylases were present in all of the samples. Enzyme screening assays utilizing the APIZYM test system showed the incidence of enzymes in the order: Specific phosphatases>esterases>glycosyl hydrolases. Measurement of cellulase by the cellulose-azure test detected activity in two out of 28 samples. Analysis for cellulase activity using the cellulose-azure test on refuse samples from landfills in Naples, Florida, and Tucson, Arizona, also showed a limited distribution of cellulases. Mineralization of [[sup 14]C]cellulose, an independent measure of cellulase activity, ranged from <5 to 23% in a 4-week incubation, which supports a highly variable cellulolytic activity in landfilled refuse. (orig.).

  19. Biodrying of municipal solid waste with high water content by combined hydrolytic-aerobic technology

    ZHANG Dongqing; HE Pinjing; SHAO Liming; JIN Taifeng; HAN Jingyao

    2008-01-01

    The high water content of municipal solid waste (MSW) will reduce the efficiency of mechanical sorting, consequently unfavorable for beneficial utilization. In this study, a combined hydrolytic-aerobic biodrying technology was introduced to remove water from MSW. The total water removals were proved to depend on the ventilation frequency and the temporal span in the hydrolytic stage. The ventilation frequency of 6 times/d was preferable in the hydrolytic stage. The hydrolytic span should not be prolonged more than 4 d. At this optimal scenario, the final water content was 50.5% reduced from the initial water content of 72.0%, presenting a high water removal efficiency up to 78.5%. A positive correlation was observed between the organics losses and the water losses in both hydrolytic and aerobic stages (R = 0.944, p < 0.01). The evolutions of extracellular enzyme activities were shown to be consistent with the organics losses.

  20. Tyrosine Phosphatase TpbA of Pseudomonas aeruginosa Controls Extracellular DNA via Cyclic Diguanylic Acid Concentrations

    Ueda, Akihiro; Wood, Thomas K.

    2010-01-01

    Inactivating the tyrosine phosphatase TpbA of Pseudomonas aeruginosa PA14 induces biofilm formation by 150-fold via increased production of the second messenger cyclic diguanylic acid (c-di-GMP). Here, we show the tpbA mutation reduces extracellular DNA (eDNA) and that increased expression of tpbA increases eDNA; hence, eDNA is inversely proportional to c-di-GMP concentrations. Mutations in diguanylate cyclases PA0169, PA4959, and PA5487 and phosphodiesterase PA4781 corroborate this trend. Th...

  1. Compressive strength and hydrolytic stability of fly ash based geopolymers

    Nikolić Irena

    2013-01-01

    Full Text Available The process of geopolymerization involves the reaction of solid aluminosilicate materials with highly alkaline silicate solution yielding an aluminosilicate inorganic polymer named geopolymer, which may be successfully applied in civil engineering as a replacement for cement. In this paper we have investigated the influence of synthesis parameters: solid to liquid ratio, NaOH concentration and the ratio of Na2SiO3/NaOH, on the mechanical properties and hydrolytic stability of fly ash based geopolymers in distilled water, sea water and simulated acid rain. The highest value of compressive strength was obtained using 10 mol dm-3 NaOH and at the Na2SiO3/NaOH ratio of 1.5. Moreover, the results have shown that mechanical properties of fly ash based geopolymers are in correlation with their hydrolytic stability. Factors that increase the compressive strength also increase the hydrolytic stability of fly ash based geopolymers. The best hydrolytic stability of fly ash based geopolymers was shown in sea water while the lowest stability was recorded in simulated acid rain. [Projekat Ministarstva nauke Republike Srbije, br. 172054 i Nanotechnology and Functional Materials Center, funded by the European FP7 project No. 245916

  2. Science of Hyaluronic Acid Beyond Filling: Fibroblasts and Their Response to the Extracellular Matrix.

    Landau, Marina; Fagien, Steven

    2015-11-01

    Loss of viscoelasticity is one of the primarily signs of skin aging, followed by appearance of visible wrinkles. Hyaluronic acid (HA)-based fillers are widely used to fill wrinkles and compensate for volume loss. Recent clinical observations demonstrate persistence of the filling effect longer than the biological availability of the filler. Stimulation of new collagen by cross-linked HA and up-regulation of elastin have been suggested as possible explanation to this observation and have been supported experimentally. Cross-linked HA substitutes for fragmented collagen in restoring extracellular matrix required for normal activity of fibroblasts, such as collagen and elastin production. To restore extracellular matrix efficiently, serial monthly treatments are required. Boosting of facial and nonfacial skin through fibroblast activation is a new indication for HA-based products. Injectable HA has also been recently registered in Europe as agents specific for the improvement of skin quality (Restylane Skinboosters). Further explanation of the possible mechanisms supported by long-term clinical examples is presented herein. PMID:26441098

  3. Extracellular matrix assembly in extreme acidic eukaryotic biofilms and their possible implications in heavy metal adsorption

    To evaluate the importance of the extracellular matrix in relation to heavy metal binding capacity in extreme acidic environments, the extracellular polymeric substances (EPS) composition of 12 biofilms isolated from Rio Tinto (SW, Spain) was analyzed. Each biofilm was composed mainly by one or two species of eukaryotes, although other microorganisms were present. EPS ranged from 130 to 439 mg g-1 biofilm dry weight, representing between 15% and the 40% of the total biofilm dry weight (DW). Statistically significant differences (p -1 dry weight; 10% to 30% of the total biofilm dry weight. Capsular EPS ranged from 50 to 318 mg g-1 dry weight; 5% to 30% of the total biofilm dry weight. Seven of the 12 biofilms showed higher amounts of capsular than colloidal EPS (p -1 biofilm dry weight, reaching up to 16% of the total composition. In general, the heavy metal composition of the EPS extracted from the biofilms closely resembled the metal composition of the water from which the biofilms were collected

  4. Impact of Extracellular Acidity on the Activity of P-glycoprotein and the Cytotoxicity of Chemotherapeutic Drugs1

    Thews, Oliver; Gassner, Birgit; Kelleher, Debra K; Schwerdt, Gerald; Gekle, Michael

    2006-01-01

    The expression and activity of P-glycoprotein (pGP) play a role in the multidrug resistance of tumors. Because solid-growing tumors often show pronounced hypoxia or extracellular acidosis, this study attempted to analyze the impact of an acidic environment on the expression and activity of pGP and on the cytotoxicity of chemotherapeutic agents. For this, prostate carcinoma cells were exposed to an acidic extracellular environment (pH 6.6) for up to 24 hours. pGP activity was more than doubled...

  5. Impact of Extracellular Acidity on the Activity of P-glycoprotein and the Cytotoxicity of Chemotherapeutic Drugs

    Oliver Thews; Birgit Gassner; Kelleher, Debra K; Gerald Schwerd; Michael Gekle

    2006-01-01

    The expression and activity of P-glycoprotein (pGP) play a role in the multidrug resistance of tumors. Because solid-growing tumors often show pronounced hypoxia or extracellular acidosis, this study attempted to analyze the impact of an acidic environment on the expression and activity of pGP and on the cytotoxicity of chemotherapeutic agents. For this, prostate carcinoma cells were exposed to an acidic extracellular environment (pH 6.6) for up to 24 hours. pGP activity was more than doubled...

  6. Enzymatic recycling of ascorbic acid from dehydroascorbic acid by glutathione-like peptides in the extracellular loops of aminergic G-protein coupled receptors.

    Root-Bernstein, Robert; Fewins, Jenna; Rhinesmith, Tyler; Koch, Ariana; Dillon, Patrick F

    2016-07-01

    The intracellular recycling of ascorbic acid from dehydroascorbic acid by the glutathione-glutathione reductase system has been well-characterized. We propose that extracellular recycling of ascorbic acid is performed in a similar manner by cysteine-rich, glutathione-like regions of the first and second extracellular loops of some aminergic receptors including adrenergic, histaminergic, and dopaminergic receptors. Previous research in our laboratory demonstrated that ascorbic acid binds to these receptors at a site on their first or second extracellular loops, significantly enhancing ligand activity, and apparently recycling hundreds of times their own concentration of ascorbate in an enzymatic fashion. In this study, we have synthesized 25 peptides from the first and second extracellular loops of aminergic and insulin receptors and compared them directly to glutathione for their ability to prevent the oxidation of ascorbate and to regenerate ascorbate from dehydroascorbic acid. Peptide sequences that mimic glutathione in containing a cysteine and a glutamic acid-like amino acid also mimic glutathione activity in effects and in kinetics. Some (but not all) peptide sequences that contain one or more methionines instead of cysteine can significantly retard the oxidation of ascorbic acid but do not recycle it from dehydroascorbate into ascorbate. Peptides lacking both cysteines and methionines uniformly failed to alter significantly ascorbate or dehydroascorbate oxidation or reduction. We believe that this is the first proof that receptors may carry out both ligand binding and enzymatic activity extracellularly. Our results suggest the existence of a previously unknown extracellular system for recycling ascorbate. Copyright © 2016 John Wiley & Sons, Ltd. PMID:26749062

  7. Brain infection with Staphylococcus aureus leads to high extracellular levels of glutamate, aspartate, γ-aminobutyric acid, and zinc.

    Hassel, Bjørnar; Dahlberg, Daniel; Mariussen, Espen; Goverud, Ingeborg Løstegaard; Antal, Ellen-Ann; Tønjum, Tone; Maehlen, Jan

    2014-12-01

    Staphylococcal brain infections may cause mental deterioration and epileptic seizures, suggesting interference with normal neurotransmission in the brain. We injected Staphylococcus aureus into rat striatum and found an initial 76% reduction in the extracellular level of glutamate as detected by microdialysis at 2 hr after staphylococcal infection. At 8 hr after staphylococcal infection, however, the extracellular level of glutamate had increased 12-fold, and at 20 hr it had increased >30-fold. The extracellular level of aspartate and γ-aminobutyric acid (GABA) also increased greatly. Extracellular Zn(2+) , which was estimated at ∼2.6 µmol/liter in the control situation, was increased by 330% 1-2.5 hr after staphylococcal infection and by 100% at 8 and 20 hr. The increase in extracellular glutamate, aspartate, and GABA appeared to reflect the degree of tissue damage. The area of tissue damage greatly exceeded the area of staphylococcal infiltration, pointing to soluble factors being responsible for cell death. However, the N-methyl-D-aspartate receptor antagonist MK-801 ameliorated neither tissue damage nor the increase in extracellular neuroactive amino acids, suggesting the presence of neurotoxic factors other than glutamate and aspartate. In vitro staphylococci incubated with glutamine and glucose formed glutamate, so bacteria could be an additional source of infection-related glutamate. We conclude that the dramatic increase in the extracellular concentration of neuroactive amino acids and zinc could interfere with neurotransmission in the surrounding brain tissue, contributing to mental deterioration and a predisposition to epileptic seizures, which are often seen in brain abscess patients. PMID:25043715

  8. Cross Talk between Cancer and Mesenchymal Stem Cells through Extracellular Vesicles Carrying Nucleic Acids

    Lopatina, Tatiana; Gai, Chiara; Deregibus, Maria Chiara; Kholia, Sharad; Camussi, Giovanni

    2016-01-01

    Extracellular vesicles (EVs) are considered to be a novel complex mechanism of cell communication within the tumor microenvironment. EVs may act as vehicles for transcription factors and nucleic acids inducing epigenetic changes in recipient cells. Since tumor EVs may be present in patient biological fluids, it is important to investigate their function and molecular mechanisms of action. It has been shown that tumor cells release EVs, which are capable of regulating cell apoptosis, proliferation, invasion, and epithelial–mesenchymal transition, as well as to suppress activity of immune cells, to enhance angiogenesis, and to prepare a favorable microenvironment for metastasis. On the other hand, EVs derived from stromal cells, such as mesenchymal stem cells (MSCs), may influence the phenotype of tumor cells through reciprocal cross talk greatly influenced by the transcription factors and nucleic acids they carry. In particular, non-coding RNAs (ncRNAs), including microRNAs and long ncRNAs, have recently been identified as the main candidates for the phenotypic changes induced in the recipient cells by EVs. ncRNAs, which are important regulators of mRNA and protein expression, can function either as tumor suppressors or as oncogenes, depending on their targets. Herein, we have attempted to revise actual evidence reported in the literature on the role of EVs in tumor biology with particular regard to the cross talk of ncRNAs between cancer cells and MSCs.

  9. Endophytic Fungi from Frankincense Tree Improves Host Growth and Produces Extracellular Enzymes and Indole Acetic Acid.

    Abdul Latif Khan

    Full Text Available Boswellia sacra, an economically important frankincense-producing tree found in the desert woodlands of Oman, is least known for its endophytic fungal diversity and the potential of these fungi to produce extracellular enzymes and auxins. We isolated various fungal endophytes belonging to Eurotiales (11.8%, Chaetomiaceae (17.6%, Incertae sadis (29.5%, Aureobasidiaceae (17.6%, Nectriaceae (5.9% and Sporomiaceae (17.6% from the phylloplane (leaf and caulosphere (stem of the tree. Endophytes were identified using genomic DNA extraction, PCR amplification and sequencing the internal transcribed spacer regions, whereas a detailed phylogenetic analysis of the same gene fragment was made with homologous sequences. The endophytic colonization rate was significantly higher in the leaf (5.33% than the stem (0.262%. The Shannon-Weiner diversity index was H' 0.8729, while Simpson index was higher in the leaf (0.583 than in the stem (0.416. Regarding the endophytic fungi's potential for extracellular enzyme production, fluorogenic 4-methylumbelliferone standards and substrates were used to determine the presence of cellulases, phosphatases and glucosidases in the pure culture. Among fungal strains, Penicillum citrinum BSL17 showed significantly higher amounts of glucosidases (62.15±1.8 μM-1min-1mL and cellulases (62.11±1.6 μM-1min-1mL, whereas Preussia sp. BSL10 showed significantly higher secretion of glucosidases (69.4±0.79 μM-1min-1mL and phosphatases (3.46±0.31μM-1min-1mL compared to other strains. Aureobasidium sp. BSS6 and Preussia sp. BSL10 showed significantly higher potential for indole acetic acid production (tryptophan-dependent and independent pathways. Preussia sp. BSL10 was applied to the host B. sacra tree saplings, which exhibited significant improvements in plant growth parameters and accumulation of photosynthetic pigments. The current study concluded that endophytic microbial resources producing extracellular enzymes and auxin

  10. Endophytic Fungi from Frankincense Tree Improves Host Growth and Produces Extracellular Enzymes and Indole Acetic Acid.

    Khan, Abdul Latif; Al-Harrasi, Ahmed; Al-Rawahi, Ahmed; Al-Farsi, Zainab; Al-Mamari, Aza; Waqas, Muhammad; Asaf, Sajjad; Elyassi, Ali; Mabood, Fazal; Shin, Jae-Ho; Lee, In-Jung

    2016-01-01

    Boswellia sacra, an economically important frankincense-producing tree found in the desert woodlands of Oman, is least known for its endophytic fungal diversity and the potential of these fungi to produce extracellular enzymes and auxins. We isolated various fungal endophytes belonging to Eurotiales (11.8%), Chaetomiaceae (17.6%), Incertae sadis (29.5%), Aureobasidiaceae (17.6%), Nectriaceae (5.9%) and Sporomiaceae (17.6%) from the phylloplane (leaf) and caulosphere (stem) of the tree. Endophytes were identified using genomic DNA extraction, PCR amplification and sequencing the internal transcribed spacer regions, whereas a detailed phylogenetic analysis of the same gene fragment was made with homologous sequences. The endophytic colonization rate was significantly higher in the leaf (5.33%) than the stem (0.262%). The Shannon-Weiner diversity index was H' 0.8729, while Simpson index was higher in the leaf (0.583) than in the stem (0.416). Regarding the endophytic fungi's potential for extracellular enzyme production, fluorogenic 4-methylumbelliferone standards and substrates were used to determine the presence of cellulases, phosphatases and glucosidases in the pure culture. Among fungal strains, Penicillum citrinum BSL17 showed significantly higher amounts of glucosidases (62.15±1.8 μM-1min-1mL) and cellulases (62.11±1.6 μM-1min-1mL), whereas Preussia sp. BSL10 showed significantly higher secretion of glucosidases (69.4±0.79 μM-1min-1mL) and phosphatases (3.46±0.31μM-1min-1mL) compared to other strains. Aureobasidium sp. BSS6 and Preussia sp. BSL10 showed significantly higher potential for indole acetic acid production (tryptophan-dependent and independent pathways). Preussia sp. BSL10 was applied to the host B. sacra tree saplings, which exhibited significant improvements in plant growth parameters and accumulation of photosynthetic pigments. The current study concluded that endophytic microbial resources producing extracellular enzymes and auxin could

  11. Production of extracellular ferulic acid esterases by Lactobacillus strains using natural and synthetic carbon sources

    Dominik Szwajgier

    2011-09-01

    Full Text Available Background. Ferulic acid esterases (FAE, EC 3.1.1.73, also known as feruloyl esterases, cinnamic acid esterases or cinnamoyl esterases, belong to a common group of hydrolases distributed in the plant kingdom. Especially the fungal enzymes were very well characterised in the past whereas the enzyme was rarely found in the lactic acid bacteria (LAB strains. It is well known that strong antioxidants free phenolic acids can be released from the dietary fiber by the action of intestinal microflora composed among others also of Lactobacillus strains. The aim of this study was to examine four Lactobacillus strains (L. acidophilus K1, L. rhamnosus E/N, PEN, OXYfor the ability to produce extracellular FAE on different synthetic and natural carbon sources. Material and methods. The LAB strains were grown in the minimal growth media using German wheat bran, rye bran, brewers’ spent grain, isolated larchwood arabinogalactan, apple pectin, corn pectin, methyl ferulate, methyl p-coumarate, methyl syringate or methyl vanillate as the sole carbon source. FAE activity was determined using the post-cultivation supernatants, methyl ferulate and HPLC with UV detection. Results. The highest FAE activity was obtained with L. acidophilus K1 and methyl ferulate (max. 23.34 ±0.05 activity units and methyl p-coumarate (max. 14.96 ±0.47 activity units as carbon sources. L. rhamnosus E/N, OXY and PEN exhibited the limited ability to produce FAE with cinnamic acids methyl esters. Methyl syringate and methyl vanillate (MS and MV were insufficient carbon sources for FAE production. Brewers’ spent grain was the most suitable substrate for FAE production by L. acidophilus K1 (max. 2.64 ±0.06 activity units and L. rhamnosus E/N, OXY and PEN. FAE was also successfully induced by natural substrates rye bran, corn pectin (L. acidophilus K1, German wheat bran and larchwood arabinogalactan (E/N, PEN or German wheat bran and corn pectin (OXY. Conclusions. This study proved the

  12. Conserved charged amino acid residues in the extracellular region of sodium/iodide symporter are critical for iodide transport activity

    Liang Ji-An

    2010-11-01

    Full Text Available Abstract Background Sodium/iodide symporter (NIS mediates the active transport and accumulation of iodide from the blood into the thyroid gland. His-226 located in the extracellular region of NIS has been demonstrated to be critical for iodide transport in our previous study. The conserved charged amino acid residues in the extracellular region of NIS were therefore characterized in this study. Methods Fourteen charged residues (Arg-9, Glu-79, Arg-82, Lys-86, Asp-163, His-226, Arg-228, Asp-233, Asp-237, Arg-239, Arg-241, Asp-311, Asp-322, and Asp-331 were replaced by alanine. Iodide uptake abilities of mutants were evaluated by steady-state and kinetic analysis. The three-dimensional comparative protein structure of NIS was further modeled using sodium/glucose transporter as the reference protein. Results All the NIS mutants were expressed normally in the cells and targeted correctly to the plasma membrane. However, these mutants, except R9A, displayed severe defects on the iodide uptake. Further kinetic analysis revealed that mutations at conserved positively charged amino acid residues in the extracellular region of NIS led to decrease NIS-mediated iodide uptake activity by reducing the maximal rate of iodide transport, while mutations at conserved negatively charged residues led to decrease iodide transport by increasing dissociation between NIS mutants and iodide. Conclusions This is the first report characterizing thoroughly the functional significance of conserved charged amino acid residues in the extracellular region of NIS. Our data suggested that conserved charged amino acid residues, except Arg-9, in the extracellular region of NIS were critical for iodide transport.

  13. Influence of Humic Acid Complexation with Metal Ions on Extracellular Electron Transfer Activity

    Zhou, Shungui; Chen, Shanshan; Yuan, Yong; Lu, Qin

    2015-11-01

    Humic acids (HAs) can act as electron shuttles and mediate biogeochemical cycles, thereby influencing the transformation of nutrients and environmental pollutants. HAs commonly complex with metals in the environment, but few studies have focused on how these metals affect the roles of HAs in extracellular electron transfer (EET). In this study, HA-metal (HA-M) complexes (HA-Fe, HA-Cu, and HA-Al) were prepared and characterized. The electron shuttle capacities of HA-M complexes were experimentally evaluated through microbial Fe(III) reduction, biocurrent generation, and microbial azoreduction. The results show that the electron shuttle capacities of HAs were enhanced after complexation with Fe but were weakened when using Cu or Al. Density functional theory calculations were performed to explore the structural geometry of the HA-M complexes and revealed the best binding sites of the HAs to metals and the varied charge transfer rate constants (k). The EET activity of the HA-M complexes were in the order HA-Fe > HA-Cu > HA-Al. These findings have important implications for biogeochemical redox processes given the ubiquitous nature of both HAs and various metals in the environment.

  14. Extracellular alkaline-acid pH shifts evoked by iontophoresis of glutamate and aspartate in turtle cerebellum.

    Chesler, M; Rice, M E

    1991-01-01

    The effect of glutamate and aspartate iontophoresis on extracellular pH was investigated in the turtle cerebellum in vitro. Both amino acids produced a rapid alkaline transient, typically followed by a prolonged acidification. These responses could be evoked in all layers of the cerebellum. Transition from bicarbonate to N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid-buffered media amplified the pH shifts. Similar alkaline-acid transients could be evoked in the molecular layer by electrical stimulation of the parallel fibers or the ipsilateral peduncle, or by superfusion of glutamate or aspartate. However, no alkaline shifts were evoked in the granular layer by either parallel fiber or peduncle stimulation. In contrast, the iontophoretically induced alkaline shifts were largest in the granular layer. Compared with the stimulus-evoked alkalinizations, the iontophoretic alkaline shifts were relatively insensitive to Mn2+ or Cd2+. These data suggest that the activity-dependent alkalinization of brain extracellular space is generated by a bicarbonate-independent mechanism related to excitatory synaptic transmission. The results are consistent with a flux of hydrogen ions through cationic channels, but do not support a direct role for voltage-dependent Ca2+ channels. In view of the sensitivity of ion channels to changes in external pH, and the magnitude of the amino acid-induced pH shifts, these results indicate that extracellular pH could play an important modulatory role in excitatory synaptic transmission. PMID:1711651

  15. Impact of Extracellular Acidity on the Activity of P-glycoprotein and the Cytotoxicity of Chemotherapeutic Drugs

    Oliver Thews

    2006-02-01

    Full Text Available The expression and activity of P-glycoprotein (pGP play a role in the multidrug resistance of tumors. Because solid-growing tumors often show pronounced hypoxia or extracellular acidosis, this study attempted to analyze the impact of an acidic environment on the expression and activity of pGP and on the cytotoxicity of chemotherapeutic agents. For this, prostate carcinoma cells were exposed to an acidic extracellular environment (pH 6.6 for up to 24 hours. pGP activity was more than doubled after 3 to 6 hours of incubation in acidic medium, whereas cellular pGP expression remained constant, indicating that increased transport rate is the result of functional modulation. In parallel, the cytotoxic efficacy of daunorubicin showed pronounced reduction at low pH, an effect that was reversible on coincubation with a pGP inhibitor. A reduction of intracellular Ca2+ concentration by 35% under acidic conditions induced a higher transport rate of pGP, an effect comparable to that found on inhibition of protein kinase C (PKC. These data indicate that pGP activity is increased by acidic pH presumably as a result of lowered intracellular calcium levels and inhibition of PKC. These findings may explain the reduced cytotoxicity of chemotherapeutic agents in hypoxic/acidic tumors.

  16. Extracellular ATP stimulates exocytosis via localized Ca(2+) release from acidic stores in rat pancreatic beta cells.

    Xie, Li; Zhang, Ming; Zhou, Wei; Wu, Zhengxing; Ding, Jiuping; Chen, Liangyi; Xu, Tao

    2006-04-01

    Three different methods, membrane capacitance (C(m)) measurement, amperometry and FM dye labeling were used to investigate the role of extracellular ATP in insulin secretion from rat pancreatic beta cells. We found that extracellular application of ATP mobilized intracellular Ca(2+) stores and synchronously triggered vigorous exocytosis. No influence of ATP on the readily releasable pool of vesicles was observed, which argues against a direct modulation of the secretory machinery at a level downstream of Ca(2+) elevation. The stimulatory effects of ATP were greatly reduced by intracellular perfusion of BAPTA but not EGTA, suggesting a close spatial association of fusion sites with intracellular Ca(2+) releasing sites. ATP-induced Ca(2+) transients and exocytosis were not blocked by thapsigargin (TG), by a ryanodine receptor antagonist or by dissipation of pH in acidic stores by monensin alone, but they were greatly attenuated by IP(3) receptor inhibition as well as ionomycin plus monensin, suggesting involvement of IP(3)-sensitive acidic Ca(2+) stores. Taken together, our data suggest that extracellular ATP triggers exocytosis by mobilizing spatially limited acidic Ca(2+) stores through IP(3) receptors. This mechanism may explain how insulin secretion from the pancreas is coordinated through diffusible ATP that is co-released with insulin. PMID:16536741

  17. A conformation change in the extracellular domain that accompanies desensitization of acid-sensing ion channel (ASIC) 3.

    Cushman, Kenneth A; Marsh-Haffner, Josephine; Adelman, John P; McCleskey, Edwin W

    2007-04-01

    Acid-sensing ion channels (ASICs) are thought to trigger some forms of acid-induced pain and taste, and to contribute to stroke-induced neural damage. After activation by low extracellular pH, different ASICs undergo desensitization on time scales from 0.1 to 10 s. Consistent with a substantial conformation change, desensitization slows dramatically when temperature drops (Askwith, C.C., C.J. Benson, M.J. Welsh, and P.M. Snyder. 2001. PNAS. 98:6459-6463). The nature of this conformation change is unknown, but two studies showed that desensitization rate is altered by mutations on or near the first transmembrane domain (TM1) (Coric, T., P. Zhang, N. Todorovic, and C.M. Canessa. 2003. J. Biol. Chem. 278:45240-45247; Pfister, Y., I. Gautschi, A.-N. Takeda, M. van Bemmelen, S. Kellenberger, and L. Schild. 2006. J. Biol. Chem. 281:11787-11791). Here we show evidence of a specific conformation change associated with desensitization. When mutated from glutamate to cysteine, residue 79, which is some 20 amino acids extracellular to TM1, can be altered by cysteine-modifying reagents when the channel is closed, but not when it is desensitized; thus, desensitization appears to conceal the residue from the extracellular medium. D78 and E79 are a pair of adjacent acidic amino acids that are highly conserved in ASICs yet absent from epithelial Na(+) channels, their acid-insensitive relatives. Despite large effects on desensitization by mutations at positions 78 and 79-including a shift to 10-fold lower proton concentration with the E79A mutant-there are not significant effects on activation. PMID:17389250

  18. A Conformation Change in the Extracellular Domain that Accompanies Desensitization of Acid-sensing Ion Channel (ASIC) 3

    Cushman, Kenneth A.; Marsh-Haffner, Josephine; Adelman, John P.; McCleskey, Edwin W.

    2007-01-01

    Acid-sensing ion channels (ASICs) are thought to trigger some forms of acid-induced pain and taste, and to contribute to stroke-induced neural damage. After activation by low extracellular pH, different ASICs undergo desensitization on time scales from 0.1 to 10 s. Consistent with a substantial conformation change, desensitization slows dramatically when temperature drops (Askwith, C.C., C.J. Benson, M.J. Welsh, and P.M. Snyder. 2001. PNAS. 98:6459–6463). The nature of this conformation chang...

  19. 黑小麦麦麸戊聚糖的部分酸水解特征与甲基化分析%Partial Acid Hydrolytic Characteristics and Methylation Analysis of Pentosans from Black-grained Wheat Bran

    孙元琳; 顾小红; 陕方; 张俊明; 崔武卫

    2012-01-01

    研究了黑小麦麦麸戊聚糖(AEPH)的部分酸水解特征及其糖链的取代度和取代方式等结构信息.分别采用0.02,0.1和0.2 mol/L的三氟乙酸(Trifluoroacetic acid,TFA)对AEPH进行部分酸水解,结合甲基化分析、GC-MS和HPLC等方法对AEPH的糖链结构和相对分子质量分布进行了分析.结果表明,戊聚糖AEPH的糖残基主要由阿拉伯糖(Araf)和木糖(XylP)组成.其取代度(A/X),即Araf与Xylp的比值为0.60,并具有较高的相对分子质量(Mw=3.81×105).AEPH的主链由Xylp残基通过1→4连接形成木聚糖.主链Xylp在O2,O3或O2/O3位被Araf单取代或双取代.非取代(u)、单取代(m)和双取代(d)Xylp残基的组成比例为57.7∶22.0∶6.2.非取代与取代XylP的比值为2.1,双取代与单取代XylP比值(d/m)为0.3.Araf主要以非还原性末端T-Araf以及通过1→2和1→5连接形成低聚阿拉伯糖的形式与主链XylP相连,或与T-Xylp相连形成戊聚糖分子的支链.%The hydrolytic characteristics and the structural features of substitute degree as well as substitute patterns of pentosans ( AEPH ) from black-grained wheat bran were analyzed. AEPH was hydrolyzed with 0. 02, 0. 1, and 0. 2 mol/L trifluoroacetic acid(TFA) , respectively. The structural features and Mw of AEPH were studied with methylation methods, combined with GC-MS and HPLC instruments. The results indicated that the residues of sugar constituents were mainly Araf and Xylp, with an Ara/Xyl( A/X) ratio of 0. 60 and a high average molecular weight(Mw.) of 3. 81 ×l05. AEPH contained a xylan backbone of 1→4 linked Xylp residues which was mono- or disubstituted with Ara/ groups at position 03, 02 or at both 02 and 03 positions. The relative composition ratio of un-, mono-, and di-substituted Xylp in the arabinoxylan fraction was 57.7 : 22. 0: 6. 2, with a high proportion of unsubstituted to substituted Xylp(2. 1) , and a very low proportion of di-to monosubstituted Xylp residues(d/m, 0. 3). Ara/side groups

  20. Variation in pH Optima of Hydrolytic Enzyme Activities in Tropical Rain Forest Soils ▿

    Turner, Benjamin L.

    2010-01-01

    Extracellular enzymes synthesized by soil microbes play a central role in the biogeochemical cycling of nutrients in the environment. The pH optima of eight hydrolytic enzymes involved in the cycles of carbon, nitrogen, phosphorus, and sulfur, were assessed in a series of tropical forest soils of contrasting pH values from the Republic of Panama. Assays were conducted using 4-methylumbelliferone-linked fluorogenic substrates in modified universal buffer. Optimum pH values differed markedly am...

  1. Regulation of the arachidonic acid-stimulated respiratory burst in neutrophils by intra- cellular and extracellular calcium

    2001-01-01

    The respiratory burst is an important physiological function ofthe neutrophils in killing the bacteria invading in human body. We used chemiluminescence method to measure the exogenous arachidonic acid-stimulated respiratory burst, and measured the cytosolic free calcium concentration in neutrophils by the fluorescence method. It was found that, on one hand, the arachidonic acid-stimulated respiratory burst was enhanced by elevating the cytosolic free calcium concentration in neutrophils with a potent endomembrane Ca2+-ATPase inhibitor, Thapsgargin; on the other hand, chelating the intracellular or extracellular calcium by EGTA or BAPTA inhibited the respiratory burst. Results showed that calcium plays an important regulatory role in the signaling pathway involved in the exogenous arachidonic acid-stimulated respiratory burst of neutrophils.

  2. The effect of radiation pre-treatment on the hydrolytic degradation of cellulose

    Linters cellulose was treated with electron or γ-radiation within a wide range of dose rate and then subjected to enzymatic and acid hydrolytic degradation, respectively. At radiation of low dose rate acid hydrolytic degradation results in a strong decrease of the LODP and of the residues as compared with non-irradiated samples. Enzymatic degradation was found to be accelerated by high energy pre-irradiation. Pre-irradiation at low dose rate caused retardation of enzymatic hydrolysis as compared with non-irradiated samples. (author)

  3. Three charged amino acids in extracellular loop 1 are involved in maintaining the outer pore architecture of CFTR

    Cui, Guiying; Rahman, Kazi S.; Infield, Daniel T.; Kuang, Christopher; Prince, Chengyu Z.; McCarty, Nael A.

    2014-01-01

    The cystic fibrosis (CF) transmembrane conductance regulator (CFTR) bears six extracellular loops (ECL1–6); ECL1 is the site of several mutations associated with CF. Mutation R117H has been reported to reduce current amplitude, whereas D110H, E116K, and R117C/L/P may impair channel stability. We hypothesized that these amino acids might not be directly involved in ion conduction and permeation but may contribute to stabilizing the outer vestibule architecture in CFTR. We used cRNA injected oo...

  4. Preliminary Study on the Performance and Interaction of Recycling Hydrolytic-Aerobic Combined Process of High Concentration Starch Wastewater

    李清彪; 廖鑫凯; 吴志旺; 邓旭; 黄益丽; 卢英华; 孙道华; 洪铭媛; 王琳

    2004-01-01

    A new recycling hydrolytic-aerobic combined process was developed to treat the high concentration organic wastewater. Simulated wastewater containing 10 g·L-1 starch with a CODcr value of 10000 mg·L-1 wasused. At first, the hydrolytic degradation and aerobic degradation process were examined in two batch reactors, respectively. In the stand-alone hydrolytic process, starch in the wastewater almost disappeared after 11 h treatment, but CODCr remained as high as 5803mg·L-1 after two days. In the aerobic process, the biodegradation rate of starch was much slower during the first 11 h than that in the hydrolytic process, although the CODCr removal efficiency reached 89.6% and more than 90% starch could be degraded after 37.5 h. To determine the interaction effects of the two processes, a series of hydrolytic-aerobic combinations were examined in details. Hydrolytic process played an important role in the whole recycle combination process as it could improve the biodegradability of the high concentration starch wastewater. However, from the other experiments, the negative effect of hydrolytic acidification was found in the hydrolytic-aerobic combination, which suggested that the aerobic microorganisms needed time to adapt themselves to the acidic environment. The effect of the degrading time, which was spent in the hydrolytic and aerobic unit, and the number of circulations, with which the wastewater went through the two units were investigated. It was found that a recycle combination of 6 h hydrolytic process with 12 h aerobic process was highly effective and potentially economical, in which the final removal efficiency of CODcr and efficiency of starch degradation reached 94.1% and 98.8%, respectively.

  5. Acid potentiation of the capsaicin receptor determined by a key extracellular site.

    Jordt, S E; Tominaga, M; Julius, D

    2000-07-01

    The capsaicin (vanilloid) receptor, VR1, is a sensory neuron-specific ion channel that serves as a polymodal detector of pain-producing chemical and physical stimuli. The response of VR1 to capsaicin or noxious heat is dynamically potentiated by extracellular protons within a pH range encountered during tissue acidosis, such as that associated with arthritis, infarction, tumor growth, and other forms of injury. A molecular determinant for this important physiological activity was localized to an extracellular Glu residue (E600) in the region linking the fifth transmembrane domain with the putative pore-forming region of the channel. We suggest that this residue serves as a key regulatory site of the receptor by setting sensitivity to other noxious stimuli in response to changes in extracellular proton concentration. We also demonstrate that protons, vanilloids, and heat promote channel opening through distinct pathways, because mutations at a second site (E648) selectively abrogate proton-evoked channel activation without diminishing responses to other noxious stimuli. Our findings provide molecular evidence for stimulus-specific steps in VR1 activation and offer strategies for the development of novel analgesic agents. PMID:10859346

  6. Production of extracellular ferulic acid esterases by Lactobacillus strains using natural and synthetic carbon sources

    Dominik Szwajgier; Anna Jakubczyk

    2011-01-01

    Background. Ferulic acid esterases (FAE, EC 3.1.1.73), also known as feruloyl esterases, cinnamic acid esterases or cinnamoyl esterases, belong to a common group of hydrolases distributed in the plant kingdom. Especially the fungal enzymes were very well characterised in the past whereas the enzyme was rarely found in the lactic acid bacteria (LAB) strains. It is well known that strong antioxidants free phenolic acids can be released from the dietary fiber by the action of intestinal microflo...

  7. Yarrowia lipolytica NCIM 3589, a tropical marine yeast, degrades bromoalkanes by an initial hydrolytic dehalogenation step.

    Vatsal, Aakanksha; Zinjarde, Smita S; Kumar, Ameeta Ravi

    2015-04-01

    The widespread industrial use of organobromines which are known persistent organic pollutants has led to their accumulation in sediments and water bodies causing harm to animals and humans. While degradation of organochlorines by bacteria is well documented, information regarding degradation pathways of these recalcitrant organobromines is scarce. Hence, their fates and effects on the environment are of concern. The present study shows that a tropical marine yeast, Yarrowia lipolytica NCIM 3589 aerobically degrades bromoalkanes differing in carbon chain length and position of halogen substitution viz., 2-bromopropane (2-BP), 1-bromobutane (1-BB), 1,5 dibromopentane (1,5-DBP) and 1-bromodecane (1-BD) as seen by an increase in cell mass, release of bromide and concomitant decrease in concentration of brominated compound. The amount of bromoalkane degraded was 27.3, 21.9, 18.0 and 38.3 % with degradation rates of 0.076, 0.058, 0.046 and 0.117/day for 2-BP, 1-BB, 1,5-DBP and 1-BD, respectively. The initial product formed respectively were alcohols viz., 2-propanol, 1-butanol, 1-bromo, 5-pentanol and 1-decanol as detected by GC-MS. These were further metabolized to fatty acids viz., 2-propionic, 1-butyric and 1-decanoic acid eventually leading to carbon dioxide formation. Neither higher chain nor brominated fatty acids were detected. An inducible extracellular dehalogenase responsible for removal of bromide was detected with activities of 21.07, 18.82, 18.96 and 26.67 U/ml for 2-BP, 1-BB, 1,5-DBP and 1-BD, respectively. We report here for the first time the proposed aerobic pathway of bromoalkane degradation by an eukaryotic microbe Y. lipolytica 3589, involving an initial hydrolytic dehalogenation step. PMID:25708590

  8. Effects of acid/alkaline pretreatment and gamma-ray irradiation on extracellular polymeric substances from sewage sludge

    In order to investigate the mechanism of extracellular polymeric substances (EPS) influencing sludge characteristics, variations of extractable EPS from municipal sewage sludge by acid/alkaline pretreatment and gamma-ray irradiation were studied. The changes in constituents of EPS were analyzed by UV–vis spectra and SEM images. The effects of alkaline pretreatment and gamma-ray irradiation on the functional groups in EPS were investigated by Fourier transform infrared (FTIR) spectrometer. Results showed that the extractable EPS increased clearly with increasing irradiation dose from 0 to 15 kGy. UV–vis spectra indicated that a new absorption band from 240 nm to 300 nm existed in all irradiated samples, apart from acid condition. The results of FTIR spectroscopic analysis indicated that, irradiation influenced major functional groups in EPS, such as protein and polysaccharide, and these effects were clearer under alkaline condition. SEM images provided that after alkaline hydrolysis, gamma-ray irradiation was more effective in resulting in the sludge flocs and cells broken, compared with acid pretreatment (pH 2.50). - Highlights: • Effects of acid/alkaline pretreatment and gamma irradiation on EPS were examined. • Gamma irradiation and alkaline treatment generated remarkable synergistic effects. • The combined application could promote sludge disintegration and solubilization

  9. High extracellular Ca2+ stimulates Ca2+-activated Cl- currents in frog parathyroid cells through the mediation of arachidonic acid cascade.

    Yukio Okada

    Full Text Available Elevation of extracellular Ca(2+ concentration induces intracellular Ca(2+ signaling in parathyroid cells. The response is due to stimulation of the phospholipase C/Ca(2+ pathways, but the direct mechanism responsible for the rise of intracellular Ca(2+ concentration has remained elusive. Here, we describe the electrophysiological property associated with intracellular Ca(2+ signaling in frog parathyroid cells and show that Ca(2+-activated Cl(- channels are activated by intracellular Ca(2+ increase through an inositol 1,4,5-trisphophate (IP(3-independent pathway. High extracellular Ca(2+ induced an outwardly-rectifying conductance in a dose-dependent manner (EC(50 ∼6 mM. The conductance was composed of an instantaneous time-independent component and a slowly activating time-dependent component and displayed a deactivating inward tail current. Extracellular Ca(2+-induced and Ca(2+ dialysis-induced currents reversed at the equilibrium potential of Cl(- and were inhibited by niflumic acid (a specific blocker of Ca(2+-activated Cl(- channel. Gramicidin-perforated whole-cell recording displayed the shift of the reversal potential in extracellular Ca(2+-induced current, suggesting the change of intracellular Cl(- concentration in a few minutes. Extracellular Ca(2+-induced currents displayed a moderate dependency on guanosine triphosphate (GTP. All blockers for phospholipase C, diacylglycerol (DAG lipase, monoacylglycerol (MAG lipase and lipoxygenase inhibited extracellular Ca(2+-induced current. IP(3 dialysis failed to induce conductance increase, but 2-arachidonoylglycerol (2-AG, arachidonic acid and 12S-hydroperoxy-5Z,8Z,10E,14Z-eicosatetraenoic acid (12(S-HPETE dialysis increased the conductance identical to extracellular Ca(2+-induced conductance. These results indicate that high extracellular Ca(2+ raises intracellular Ca(2+ concentration through the DAG lipase/lipoxygenase pathway, resulting in the activation of Cl(- conductance.

  10. Activation of Plant Innate Immunity by Extracellular High Mobility Group Box 3 and Its Inhibition by Salicylic Acid.

    Choi, Hyong Woo; Manohar, Murli; Manosalva, Patricia; Tian, Miaoying; Moreau, Magali; Klessig, Daniel F

    2016-03-01

    Damage-associated molecular pattern molecules (DAMPs) signal the presence of tissue damage to induce immune responses in plants and animals. Here, we report that High Mobility Group Box 3 (HMGB3) is a novel plant DAMP. Extracellular HMGB3, through receptor-like kinases BAK1 and BKK1, induced hallmark innate immune responses, including i) MAPK activation, ii) defense-related gene expression, iii) callose deposition, and iv) enhanced resistance to Botrytis cinerea. Infection by necrotrophic B. cinerea released HMGB3 into the extracellular space (apoplast). Silencing HMGBs enhanced susceptibility to B. cinerea, while HMGB3 injection into apoplast restored resistance. Like its human counterpart, HMGB3 binds salicylic acid (SA), which results in inhibition of its DAMP activity. An SA-binding site mutant of HMGB3 retained its DAMP activity, which was no longer inhibited by SA, consistent with its reduced SA-binding activity. These results provide cross-kingdom evidence that HMGB proteins function as DAMPs and that SA is their conserved inhibitor. PMID:27007252

  11. Retinoic acid stimulation of human dermal fibroblast proliferation is dependent on suboptimal extracellular Ca2+ concentration.

    Varani, J.; Shayevitz, J.; Perry, D; Mitra, R. S.; Nickoloff, B J; Voorhees, J. J.

    1990-01-01

    Human dermal fibroblasts failed to proliferate when cultured in medium containing 0.15 mmol/l (millimolar) Ca2+ (keratinocyte growth medium [KGM]) but did when the external Ca2+ concentration was raised to 1.4 mmol/l. All-trans retinoic acid (retinoic acid) stimulated proliferation in KGM but did not further stimulate growth in Ca2(+)-supplemented KGM. The ability of retinoic acid to stimulate proliferation was inhibited in KGM prepared without Ca2+ or prepared with 0.03 mmol/l Ca2+ and in KG...

  12. The production-influencing factors of extracellular polysacchadde(EPS) from a Strain of lactic acid bacteria and EPS extraction

    CHEN Ying; SUN Liping; ZENG Yong; WANG Lei; AN Liguo

    2006-01-01

    The influencing factors of extracellular polysaccharide(EPS)produced from a strain of lactic acid bacteria(LAB L15)were studied by using the phenol-H2SO4 method.It was demonstrated that the strain produced EPS at the most amount when it was incubated for 40-48 h and when the pH value was 4 under 30℃.Glucose was the most suitable carbon source for LAB-producing EPS.The rough EPS was obtained from L15 culture after centrifugation,dialysis,deprotein,decoloration,and ethanol-precipitation.The sample was at least composed of two polysaccharides mat were completely different in molecular weight and the amount.The purified EPS was passed through the SephadexG-200 colunm and it showed that it was a sample purified by thin layer chromatography.

  13. Conversion of cheese whey into a fucose- and glucuronic acid-rich extracellular polysaccharide by Enterobacter A47.

    Antunes, Sílvia; Freitas, Filomena; Alves, Vítor D; Grandfils, Christian; Reis, Maria A M

    2015-09-20

    Cheese whey was used as the sole substrate for the production of extracellular polysaccharides (EPS) by Enterobacter A47. An EPS concentration of 6.40 g L(-1) was reached within 3.2 days of cultivation, corresponding to a volumetric productivity of 2.00 g L(-1) d(-1). The produced EPS was mainly composed of glucuronic acid (29 mol%) and fucose (29 mol%), with lower contents of glucose and galactose (21 mol% each) and a total acyl groups content of 32 wt.%. The polymer had an average molecular weight of 1.8×10(6) Da, with a polydispersity index of 1.2, and an intrinsic viscosity of 8.0 dL g(-1). EPS aqueous solutions (1.0 wt.% in 0.01 M NaCl, at pH 8.0) presented a shear thinning behavior with a viscosity of the first Newtonian plateau approaching 0.1 Pas. This novel glucuronic acid-rich polymer possesses interesting rheological properties, which, together with its high content of glucuronic acid and fucose, two bioactive sugar monomers, confers it a great potential for use in high-value applications, such as cosmetics and pharmaceuticals. PMID:26119564

  14. Effects of acid/alkaline pretreatment and gamma-ray irradiation on extracellular polymeric substances from sewage sludge

    Xie, Shuibo; Wu, Yuqi; Wang, Wentao; Wang, Jingsong; Luo, Zhiping; Li, Shiyou

    2014-04-01

    In order to investigate the mechanism of extracellular polymeric substances (EPS) influencing sludge characteristics, variations of extractable EPS from municipal sewage sludge by acid/alkaline pretreatment and gamma-ray irradiation were studied. The changes in constituents of EPS were analyzed by UV-vis spectra and SEM images. The effects of alkaline pretreatment and gamma-ray irradiation on the functional groups in EPS were investigated by Fourier transform infrared (FTIR) spectrometer. Results showed that the extractable EPS increased clearly with increasing irradiation dose from 0 to 15 kGy. UV-vis spectra indicated that a new absorption band from 240 nm to 300 nm existed in all irradiated samples, apart from acid condition. The results of FTIR spectroscopic analysis indicated that, irradiation influenced major functional groups in EPS, such as protein and polysaccharide, and these effects were clearer under alkaline condition. SEM images provided that after alkaline hydrolysis, gamma-ray irradiation was more effective in resulting in the sludge flocs and cells broken, compared with acid pretreatment (pH 2.50).

  15. OPTIMIZATION OF EXTRACELLULAR ACID PROTEASE PRODUCTION FROM ASPERGILLUS NIGER BY FACTORIAL DESIGN

    Vishalkirti Vijay Kalaskar; Narayanan Kasinathan; Volety Mallikarjuna Subrahmanyam; Josyula Venkata Rao

    2014-01-01

    The cultural conditions for acid protease production by Aspergillus niger was optimised using factorial design experiments and one factor-at-a time approach. In the production medium casein served as substrate and protease activity was measured in terms of tyrosine yield. The yield was further improved through UV mutation. Tyrosine yield amounted to 29.22 mg / g on casein substrate. Protease from this microbial strain was mesophilic. The enzyme was stable over a wide temperature range (30 to ...

  16. Expression of extracellular acid proteinase by proteolytic Candida spp. during experimental infection of oral mucosa.

    Borg, M; Rüchel, R.

    1988-01-01

    We traced an acid proteinase from Candida spp. in the initial stages of the pathogenesis of the mycosis. On infection of human buccal mucosa, proteinase antigens were detected by immuno-scanning electron microscopy on the surface of adhering blastoconidia and invading filamentous cells of C. albicans serotype A. Proteinase antigens were also present on blastoconidia of C. albicans serotype B, but were missing on filamentous cells of this serotype. Proteolytic isolates of C. tropicalis behaved...

  17. EXTRACELLULAR GAMMA-AMINOBUTYRIC-ACID IN THE SUBSTANTIA-NIGRA RETICULATA MEASURED BY MICRODIALYSIS IN AWAKE RATS - EFFECTS OF VARIOUS STIMULANTS

    TIMMERMAN, W; WESTERINK, BHC

    1995-01-01

    The gamma-aminobutyric acid (GABA)-ergic system in the substantia nigra reticulata (SNR) was challenged by local infusion of various receptor-specific agents to obtain additional information on the physiological significance of extracellular GABA levels as measured by microdialysis in awake rats. No

  18. Three charged amino acids in extracellular loop 1 are involved in maintaining the outer pore architecture of CFTR.

    Cui, Guiying; Rahman, Kazi S; Infield, Daniel T; Kuang, Christopher; Prince, Chengyu Z; McCarty, Nael A

    2014-08-01

    The cystic fibrosis (CF) transmembrane conductance regulator (CFTR) bears six extracellular loops (ECL1-6); ECL1 is the site of several mutations associated with CF. Mutation R117H has been reported to reduce current amplitude, whereas D110H, E116K, and R117C/L/P may impair channel stability. We hypothesized that these amino acids might not be directly involved in ion conduction and permeation but may contribute to stabilizing the outer vestibule architecture in CFTR. We used cRNA injected oocytes combined with electrophysiological techniques to test this hypothesis. Mutants bearing cysteine at these sites were not functionally modified by extracellular MTS reagents and were blocked by GlyH-101 similarly to WT-CFTR. These results suggest that these three residues do not contribute directly to permeation in CFTR. In contrast, mutants D110R-, E116R-, and R117A-CFTR exhibited instability of the open state and significantly shortened burst duration compared with WT-CFTR and failed to be locked into the open state by AMP-PNP (adenosine 5'-(β,γ-imido) triphosphate); charge-retaining mutants showed mainly the full open state with comparably longer open burst duration. These interactions suggest that these ECL1 residues might be involved in maintaining the outer pore architecture of CFTR. A CFTR homology model suggested that E116 interacts with R104 in both the closed and open states, D110 interacts with K892 in the fully closed state, and R117 interacts with E1126 in the open state. These interactions were confirmed experimentally. The results suggest that D110, E116, and R117 may contribute to stabilizing the architecture of the outer pore of CFTR by interactions with other charged residues. PMID:25024266

  19. Oxidation of intramyocellular lipids is dependent on mitochondrial function and the availability of extracellular fatty acids

    Corpeleijn, Eva; Hessvik, Nina P; Bakke, Siril S;

    2010-01-01

    were extended by comparing these processes in primary cultured myotubes established from healthy lean and obese type 2 diabetic (T2D) individuals, two extremes in a range of metabolic phenotypes. ICLs were prelabeled for 2 days with 100 microM [(14)C]oleic acid (OA). ICL(OX) was studied using a (14)CO......Obesity and insulin resistance are related to both enlarged intramyocellular triacylglycerol stores and accumulation of lipid intermediates. We investigated how lipid overflow can change the oxidation of intramyocellular lipids (ICL(OX)) and intramyocellular lipid storage (ICL). These experiments...... all, a lower mitochondrial mass and lower ICL(OX) were related to a higher cell-associated OA accumulation. Second, myotubes established from obese T2D individuals showed reduced ICL(OX). ICL(OX) remained lower during uncoupling (P <0.001), even with comparable mitochondrial mass, suggesting decreased...

  20. Multiple roles of the extracellular vestibule amino acid residues in the function of the rat P2X4 receptor.

    Milos B Rokic

    Full Text Available The binding of ATP to trimeric P2X receptors (P2XR causes an enlargement of the receptor extracellular vestibule, leading to opening of the cation-selective transmembrane pore, but specific roles of vestibule amino acid residues in receptor activation have not been evaluated systematically. In this study, alanine or cysteine scanning mutagenesis of V47-V61 and F324-N338 sequences of rat P2X4R revealed that V49, Y54, Q55, F324, and G325 mutants were poorly responsive to ATP and trafficking was only affected by the V49 mutation. The Y54F and Y54W mutations, but not the Y54L mutation, rescued receptor function, suggesting that an aromatic residue is important at this position. Furthermore, the Y54A and Y54C receptor function was partially rescued by ivermectin, a positive allosteric modulator of P2X4R, suggesting a rightward shift in the potency of ATP to activate P2X4R. The Q55T, Q55N, Q55E, and Q55K mutations resulted in non-responsive receptors and only the Q55E mutant was ivermectin-sensitive. The F324L, F324Y, and F324W mutations also rescued receptor function partially or completely, ivermectin action on channel gating was preserved in all mutants, and changes in ATP responsiveness correlated with the hydrophobicity and side chain volume of the substituent. The G325P mutant had a normal response to ATP, suggesting that G325 is a flexible hinge. A topological analysis revealed that the G325 and F324 residues disrupt a β-sheet upon ATP binding. These results indicate multiple roles of the extracellular vestibule amino acid residues in the P2X4R function: the V49 residue is important for receptor trafficking to plasma membrane, the Y54 and Q55 residues play a critical role in channel gating and the F324 and G325 residues are critical for vestibule widening.

  1. Hydrolytic potential of a psychrotrophic Pseudomonas isolated from refrigerated raw milk

    Ana Paula F. Corrêa

    2011-12-01

    Full Text Available The production of extracellular hydrolases by a psychrotrophic bacterium isolated from refrigerated raw milk, and identified as a Pseudomonas sp. belonging to the Pseudomonas jenssenii group, was studied. This bacterium produced proteolytic and lipolytic enzymes in all media investigated (skim milk, cheese whey, casein broth, and tryptone soy broth. High levels of α-glucosidase were produced in skim milk broth. Hydrolytic enzymes detected in skim milk broth are of particular concern, indicating that these enzymes could be produced by Pseudomonas sp. during the cold storage of raw milk, contributing to the spoilage problem in milk and dairy products.

  2. The effect of extracellular pH on radiosensitization by misonidazole and acidic or basic analogues

    Using hypoxic Chinese hamster v79 cells in vitro, misonidazole (1 mmol dm-3) gave an enhancement ratio (e.r.) of 1.6 which remained unchanged over the pHsub(e) range of 3.8-9.5. Control hypoxic survival curves in the absence of sensitizer (also) remained essentially unchanged over this pHsub(e) range. These results contrast with those seen for 0.1 mmol dm-3 Ro 03-8799 (1-(2-nitro-1-imidazolyl)-3-N-piperidino-2-propanol), a base with pKsub(a)=8.9): the ER increased from 1.4 to 2.1 as pHsub(e) increased from 5.6 to 8.4. However, with the weaker bases, Ro 03-8800 and nimorazole (morpholino derivatives with pKsub(a)=6.3 and 5.2 respectively) the e.r. remained constant over a wide pHsub(e) range. Nitroimidazoles substituted with acidic functions gave decreasing sensitization with increasing pHsub(e). For azomycin (pKsub(a)=7.2) at 1 mmol dm-3 the e.r. decreased from 1.9 at pHsub(e) 4 to 1.0 at pHsub(e) 9. The effect of the proton conductor carbonyl cyanide-3-chlorophenylhydrazone (CCCP, 10 μmol dm-3) on radiosensitization by Ro 03-8799 (0.1 mmol dm-3) and misonidazole (1.0 mmol dm-3) was also studied. At pHsub(e) 6.67 the e.r. for Ro 03-8799 was increased from 1.36 to 1.76 by the presence of CCCP, whereas at pHsub(e) 7.33 the e.r. was unchanged. In contrast the e.r. for misonidazole was unchanged at pHsub(e) 6.65 and 7.33. (author)

  3. Saturated fatty acid palmitate induces extracellular release of histone H3: A possible mechanistic basis for high-fat diet-induced inflammation and thrombosis

    Highlights: •High-fat diet feeding and palmitate induces the release of nuclear protein histone H3. •ROS production and JNK signaling mediates the release of histone H3. •Extracellular histones induces proinflammatory and procoagulant response. -- Abstract: Chronic low-grade inflammation is a key contributor to high-fat diet (HFD)-related diseases, such as type 2 diabetes, non-alcoholic steatohepatitis, and atherosclerosis. The inflammation is characterized by infiltration of inflammatory cells, particularly macrophages, into obese adipose tissue. However, the molecular mechanisms by which a HFD induces low-grade inflammation are poorly understood. Here, we show that histone H3, a major protein component of chromatin, is released into the extracellular space when mice are fed a HFD or macrophages are stimulated with the saturated fatty acid palmitate. In a murine macrophage cell line, RAW 264.7, palmitate activated reactive oxygen species (ROS) production and JNK signaling. Inhibitors of these pathways dampened palmitate-induced histone H3 release, suggesting that the extracellular release of histone H3 was mediated, in part, through ROS and JNK signaling. Extracellular histone activated endothelial cells toexpress the adhesion molecules ICAM-1 and VCAM-1 and the procoagulant molecule tissue factor, which are known to contribute to inflammatory cell recruitment and thrombosis. These results suggest the possible contribution of extracellular histone to the pathogenesis of HFD-induced inflammation and thrombosis

  4. Saturated fatty acid palmitate induces extracellular release of histone H3: A possible mechanistic basis for high-fat diet-induced inflammation and thrombosis

    Shrestha, Chandan [Department of Systems Biology in Thromboregulation, Kagoshima University Graduate School of Medical and Dental Sciences, Kagoshima (Japan); Department of Laboratory and Vascular Medicine, Kagoshima University Graduate School of Medical and Dental Sciences, Kagoshima (Japan); Ito, Takashi [Department of Systems Biology in Thromboregulation, Kagoshima University Graduate School of Medical and Dental Sciences, Kagoshima (Japan); Kawahara, Ko-ichi [Department of Biomedical Engineering, Osaka Institute of Technology, Osaka (Japan); Shrestha, Binita; Yamakuchi, Munekazu; Hashiguchi, Teruto [Department of Laboratory and Vascular Medicine, Kagoshima University Graduate School of Medical and Dental Sciences, Kagoshima (Japan); Maruyama, Ikuro, E-mail: rinken@m3.kufm.kagoshima-u.ac.jp [Department of Systems Biology in Thromboregulation, Kagoshima University Graduate School of Medical and Dental Sciences, Kagoshima (Japan)

    2013-08-09

    Highlights: •High-fat diet feeding and palmitate induces the release of nuclear protein histone H3. •ROS production and JNK signaling mediates the release of histone H3. •Extracellular histones induces proinflammatory and procoagulant response. -- Abstract: Chronic low-grade inflammation is a key contributor to high-fat diet (HFD)-related diseases, such as type 2 diabetes, non-alcoholic steatohepatitis, and atherosclerosis. The inflammation is characterized by infiltration of inflammatory cells, particularly macrophages, into obese adipose tissue. However, the molecular mechanisms by which a HFD induces low-grade inflammation are poorly understood. Here, we show that histone H3, a major protein component of chromatin, is released into the extracellular space when mice are fed a HFD or macrophages are stimulated with the saturated fatty acid palmitate. In a murine macrophage cell line, RAW 264.7, palmitate activated reactive oxygen species (ROS) production and JNK signaling. Inhibitors of these pathways dampened palmitate-induced histone H3 release, suggesting that the extracellular release of histone H3 was mediated, in part, through ROS and JNK signaling. Extracellular histone activated endothelial cells toexpress the adhesion molecules ICAM-1 and VCAM-1 and the procoagulant molecule tissue factor, which are known to contribute to inflammatory cell recruitment and thrombosis. These results suggest the possible contribution of extracellular histone to the pathogenesis of HFD-induced inflammation and thrombosis.

  5. Regulation of intracellular signaling by extracellular glycan remodeling

    Parker, Randy B.; Kohler, Jennifer J.

    2010-01-01

    The plasma membrane of eukaryotic cells is coated with carbohydrates. By virtue of their extracellular position and recognizable chemical features, cell surface glycans mediate many receptor-ligand interactions. Recently, mammalian extracellular hydrolytic enzymes have been shown to modify the structure of cell surface glycans and consequently, alter their binding properties. These cell surface glycan remodeling events can cause rapid changes in critical signal transduction phenomena. This re...

  6. Reduction of Inflammatory Responses and Enhancement of Extracellular Matrix Formation by Vanillin-Incorporated Poly(Lactic-co-Glycolic Acid) Scaffolds

    Lee, Yujung; Kwon, JeongIl; Khang, Gilson; Lee, Dongwon

    2012-01-01

    Vanillin is one of the major components of vanilla, a commonly used flavoring agent and preservative and is known to exert potent antioxidant and anti-inflammatory activities. In this work, vanillin-incorporated poly(lactic-co-glycolic acid) (PLGA) films and scaffolds were fabricated to evaluate the effects of vanillin on the inflammatory responses and extracellular matrix (ECM) formation in vitro and in vivo. The incorporation of vanillin to PLGA films induced hydrophilic nature, resulting i...

  7. Influence of Extracellular Cellulose and Colanic Acid Production on the Survival of Shiga Toxin-Producing Escherichia coli on Spinach and Lettuce after Chlorine Treatment.

    Lee, Chi-Ching; Chen, Jinru; Frank, Joseph F

    2016-04-01

    Shiga toxin-producing Escherichia coli (STEC) strains produce extracellular cellulose and colanic acid, which may influence stress tolerance. This study investigates the role of these extracellular polymers on the tolerance of STEC to chlorine treatment after attachment to lettuce and spinach. Four STEC strains, two wild-type cellulose-producing and their cellulosedeficient derivatives, were used. One strain pair produced colanic acid in addition to cellulose. Spinach and lettuce with attached cells were treated with chlorinated water (50 and 150 ppm of free chlorine). The production of the extracellular polymers by the planktonic cells had small, but significant, effects on the survival of the attached pathogen when subjected to chlorine treatment. On the lettuce surface, the colanic acid-producing, cellulose-negative mutant (49d) was most susceptible to the treatment, declining significantly (P population by 0.9 and 1.4 log units after treatment with 50 and 150 ppm of chlorine, respectively. Chlorine treatment reduced populations of cellulose-deficient cells on the intact spinach surface 1.2 log units more than the wild type when treated with 150 ppm of chlorine (P populations of cellulose-producing cells were reduced by 1.5 log units more than their mutant counterparts when the cells also produced colanic acid (P < 0.05). A greater proportion of cells attached to the spinach leaf edge were injured by chlorine treatment compared with attached to the leaf surface. These results indicate that extracellular polymers do not generally increase the ability of STEC to survive chlorine treatment and that any effects on survival are influenced by location of attachment, type of leafy green, and concentration of chlorine. PMID:27052873

  8. Silencing of acidic pathogenesis-related PR-1 genes increases extracellular beta-(1 -> 3)-glucanase activity at the onset of tobacco defence reactions

    Riviere, M.P.; Marais, A.; Ponchet, M.;

    2008-01-01

    silenced. Plants lacking extracellular PR-1s were more susceptible than wild-type plants to the oomycete Phytophthora parasitica but displayed unaffected systemic acquired resistance and developmental resistance to this pathogen. Treatment with salicylic acid up-regulates the PR-1g gene, encoding a basic...... protein of the PR-1 family, in PR-1-deficient tobacco, indicating that PR-1 expression may repress that of PR-1g. This shows that acidic PR-1s are dispensable for expression of salicylic acid-dependent acquired resistances against P. parasitica and may reveal a functional overlap in tobacco defence......The class 1 pathogenesis-related (PR) proteins are thought to be involved in plant defence responses, but their molecular functions are unknown. The function of PR-1 was investigated in tobacco by generating stable PR-1a-silenced lines in which other acidic PR-1 genes (PR-1b and PR-1c) were...

  9. Complex coordinated extracellular metabolism: Acid phosphatases activate diluted human leukocyte proteins to generate energy flow as NADPH from purine nucleotide ribose.

    Hibbs, John B; Vavrin, Zdenek; Cox, James E

    2016-08-01

    Complex metabolism is thought to occur exclusively in the crowded intracellular environment. Here we report that diluted enzymes from lysed human leukocytes produce extracellular energy. Our findings involve two pathways: the purine nucleotide catabolic pathway and the pentose phosphate pathway, which function together to generate energy as NADPH. Glucose6P fuel for NADPH production is generated from structural ribose of purine ribonucleoside monophosphates, ADP, and ADP-ribose. NADPH drives glutathione reductase to reduce an oxidized glutathione disulfide-glutathione redox couple. Acid phosphatases initiate ribose5P salvage from purine ribonucleoside monophosphates, and transaldolase controls the direction of carbon chain flow through the nonoxidative branch of the pentose phosphate pathway. These metabolic control points are regulated by pH. Biologically, this energy conserving metabolism could function in perturbed extracellular spaces. PMID:26895212

  10. Hydrolytic Amino Acids Employed as a Novel Organic Nitrogen Source for the Preparation of PGPF-Containing Bio-Organic Fertilizer for Plant Growth Promotion and Characterization of Substance Transformation during BOF Production.

    Fengge Zhang

    Full Text Available Opportunity costs seriously limit the large-scale production of bio-organic fertilizers (BOFs both in China and internationally. This study addresses the utilization of amino acids resulting from the acidic hydrolysis of pig corpses as organic nitrogen sources to increase the density of TrichodermaharzianumT-E5 (a typical plant growth-promoting fungi, PGPF. This results in a novel, economical, highly efficient and environmentally friendly BOF product. Fluorescence excitation-emission matrix (EEM spectroscopy combined with fluorescence regional integration (FRI was employed to monitor compost maturity levels, while pot experiments were utilized to test the effects of this novel BOF on plant growth. An optimization experiment, based on response surface methodologies (RSMs, showed that a maximum T-E5 population (3.72 × 108 ITS copies g-1 was obtained from a mixture of 65.17% cattle manure compost (W/W, 19.33% maggot manure (W/W, 15.50% (V/Whydrolytic amino acid solution and 4.69% (V/W inoculum at 28.7°C after a 14 day secondary solid fermentation. Spectroscopy analysis revealed that the compost transformation process involved the degradation of protein-like substances and the formation of fulvic-like and humic-like substances. FRI parameters (PI, n, PII, n, PIII, n and PV, n were used to characterize the degree of compost maturity. The BOF resulted in significantly higher increased chlorophyll content, shoot length, and shoot and root dry weights of three vegetables (cucumber, tomato and pepper by 9.9%~22.4%, 22.9%~58.5%, 31.0%~84.9%, and 24.2%~34.1%, respectively. In summary, this study presents an operational means of increasing PGPF T-E5 populations in BOF to promote plant growth with a concomitant reduction in production cost. In addition, a BOF compost maturity assessment using fluorescence EEM spectroscopy and FRI ensured its safe field application.

  11. Nanofibras de algodão obtidas sob diferentes condições de hidrólise ácida Cotton nanofibers obtained by different hydrolytic acid conditions

    Eliangela de M. Teixeira

    2010-01-01

    Full Text Available As propriedades de termo estabilidade das nanofibras de celulose estão relacionadas diretamente com a sua aplicação e em especial ao processamento de polímeros cujas temperaturas típicas de processamento encontram-se em torno de 200 °C. Neste trabalho, nanofibras de algodão comercial foram extraídas empregando-se diferentes ácidos: ácido sulfúrico, ácido clorídrico e mistura (1:1; ácido sulfúrico: ácido clorídrico. As nanofibras resultantes foram caracterizadas quanto à morfologia por microscopia de transmissão (MET, potencial zeta, análise elementar, cristalinidade por difração de raio X (DRX e estabilidade térmica em atmosfera de ar (TGA. Os resultados indicaram uma morfologia muito similar entre as nanofibras bem como a cristalinidade das mesmas, independentemente do ácido utilizado na hidrólise. A principal diferença encontrada foi em relação ao estado de agregação das mesmas e quanto à estabilidade térmica. O estado de agregação das nanofibras em suspensões seguiu a ordem HCl >H2SO4:HCl > H2SO4 devido ao aumento da carga superficial nesta ordem avaliada via medidas de potencial zeta. A incorporação de HCl ao H2SO4 para a obtenção de nanofibras de celulose revelou-se eficaz no aumento da estabilidade térmica das nanofibras relativamente a extrações empregando-se somente H2SO4 e uma melhor dispersão em relação a nanofibras obtidas com o uso somente de HCl.The thermal stability of cellulose nanofibers is related to their application and especially to polymer processing normally occurring at ca. 200 °C. In this work, nanofibers of commercial cotton were obtained by acid hydrolysis employing different acids: sulfuric, hydrochloric and a mixture (1:1; sulfuric acid: hydrochloric acid.The morphology of the nanofibers were characterized by transmission microscopy (TEM, zeta potential, elemental analysis, crystallinity by X ray diffraction (XRD and thermal stability (TGA in air atmosphere by

  12. Canceling effect leads temperature insensitivity of hydrolytic enzymes in soil

    Razavi, Bahar S.; Blagodatskaya, Evgenia; Kuzyakov, Yakov

    2015-04-01

    Extracellular enzymes are important for decomposition of many macromolecules abundant in soil such as cellulose, hemicelluloses and proteins (Allison et al., 2010; Chen et al., 2012). The temperature sensitivity of enzymes responsible for organic matter decomposition is the most crucial parameter for prediction of the effects of global warming on carbon cycle. Temperature responses of biological systems are often expressed as a Q10 functions; The Q10 describes how the rate of a chemical reaction changes with a temperature increase for 10 °C The aim of this study was to test how the canceling effect will change with variation in temperature interval, during short-term incubation. We additionally investigated, whether canceling effect occurs in a broad range of concentrations (low to high) and whether it is similar for the set of hydrolytic enzymes within broad range of temperatures. To this end, we performed soil incubation over a temperature range of 0-40°C (with 5°C steps). We determined the activities of three enzymes involved in plant residue decomposition: β-glucosidase and cellobiohydrolase, which are commonly measured as enzymes responsible for degrading cellulose (Chen et al., 2012), and xylanase, which degrades xylooligosaccharides (short xylene chain) in to xylose, thus being responsible for breaking down hemicelluloses (German et al., 2011). Michaelis-Menten kinetics measured at each temperature allowed to calculate Q10 values not only for the whole reaction rates, but specifically for maximal reaction rate (Vmax) and substrate affinity (Km). Subsequently, the canceling effect - simultaneous increase of Vmax and Km with temperature was analyzed within 10 and 5 degree of temperature increase. Three temperature ranges (below 10, between 15 and 25, and above 30 °C) clearly showed non-linear but stepwise increase of temperature sensitivity of all three enzymes and allowed to conclude for predominance of psychrophilic, mesophilic and thermophilic

  13. Tetracycline removal and effect on the formation and degradation of extracellular polymeric substances and volatile fatty acids in the process of hydrogen fermentation.

    Hou, Guangying; Hao, Xiaoyan; Zhang, Rui; Wang, Jing; Liu, Rutao; Liu, Chunguang

    2016-07-01

    Many research indicate antibiotics show adverse effect on methane fermentation, while few research focus on their effect on hydrogen fermentation. The present study aimed to gain insight of the effect of antibiotics on hydrogen fermentation with waste sludge and corn straw as substrate. For this purpose, tetracycline, as a model, was investigated with regard to tetracycline removal, hydrogen production, interaction with extracellular polymeric substances (EPSs) of substrate and volatile fatty acids (VFAs) on concentration and composition. Results show that tetracycline could be removed efficiently by hydrogen fermentation, and relative low-dose tetracycline (200mg/l) exposure affects little on hydrogen production. While tetracycline exposure could change hydrogen fermentation from butyric acid-type to propionic acid-type depending on tetracycline level. Based upon three-dimensional excitation-emission matrix fluorescence spectroscopy and UV-vis tetracycline changed the component and content of EPSs, and static quenching was the main mechanism between EPSs with tetracycline. PMID:27070285

  14. Effects of eslicarbazepine acetate on acute and chronic latrunculin A-induced seizures and extracellular amino acid levels in the mouse hippocampus

    Sierra-Paredes, Germán; Loureiro, Ana I; Wright, Lyndon C; Sierra-Marcuño, Germán; Soares-da-Silva, Patrício

    2014-01-01

    Background Latrunculin A microperfusion of the hippocampus induces acute epileptic seizures and long-term biochemical changes leading to spontaneous seizures. This study tested the effect of eslicarbazepine acetate (ESL), a novel antiepileptic drug, on latrunculin A-induced acute and chronic seizures, and changes in brain amino acid extracellular levels. Hippocampi of Swiss mice were continuously perfused with a latrunculin A solution (4 μM, 1 μl/min, 7 h/day) with continuous EEG and videotap...

  15. Doxorubicin loaded pH-sensitive micelle targeting acidic extracellular pH of human ovarian A2780 tumor in mice

    Gao, Z. G.; Lee, D H.; Kim, D I; Bae, Y. H.

    2005-01-01

    The purpose of this study was to examine the efficacy of a chemotherapeutic drug, doxorubicin (DOX), loaded in pH-sensitive micelles poly(l-histidine) (Mn:5K)-b-PEG (Mn:5K) micelles. The micelles were designed to target the acidic extracellular pH of solid tumors. Studies of pH-dependent cytotoxicity, growth rate of the tumor, pharmacokinetics and biodistribution were conducted. In vitro DOX uptake upon A2780 cells by incubating the cells in a pH 6.8 complete medium at a concentration of 20 μ...

  16. Multiple Roles of the Extracellular Vestibule Amino Acid Residues in the Function of the Rat P2X4 Receptor

    Rokic, Milos Boro; Stojilkovic, S. S.; Vávra, Vojtěch; Kuzyk, Pavlo; Tvrdoňová, Vendula; Zemková, Hana

    2013-01-01

    Roč. 8, č. 3 (2013), e59411. E-ISSN 1932-6203 R&D Projects: GA AV ČR(CZ) IAA500110910; GA ČR(CZ) GBP304/12/G069 Institutional research plan: CEZ:AV0Z50110509 Institutional support: RVO:67985823 Keywords : ATP * purinergic P2X receptor channels * transmembrane domain * extracellular vestibule * gating * ivermectin Subject RIV: ED - Physiology Impact factor: 3.534, year: 2013

  17. Screening of Marine Actinomycetes from Segara Anakan for Natural Pigment and Hydrolytic Activities

    Asnani, A.; Ryandini, D.; Suwandri

    2016-02-01

    Marine actinomycetes have become sources of great interest to natural product chemistry due to their new chemical entities and bioactive metabolites. Since April 2010, we have screened actinobacteria from five sites that represent different ecosystems of Segara Anakan lagoon. In this present study we focus on specific isolates, K-2C which covers 1) actinomycetes identification based on morphology observation and 16S rRNA gene; 2) fermentation and isolation of pigment; 3) structure determination of pigment; and 4) hydrolytic enzymes characterization; Methodologies relevant to the studies were implemented accordingly. The results indicated that K-2C was likely Streptomyces fradiae strain RSU15, and the best fermentation medium should contain starch and casein with 21 days of incubation. The isolate has extracellular as well as intracellular pigments. Isolated pigments gave purple color with λmax of 529.00 nm. The pigment was structurally characterized. Interestingly, Streptomyces K-2C was able to produce potential hydrolytic enzymes such as amylase, cellulase, protease, lipase, urease, and nitrate reductase.

  18. Oxidation of Intracellular and Extracellular Fatty Acids in Skeletal Muscle: Application of kinetic modeling, stable isotopes and liquid chromatography/electrospray ionization ion-trap tandem mass spectrometry technology

    Xu, J; Zhou, L.; Persson, X-M.; Balagopal, P.; Jensen, M D; Guo, ZK.

    2008-01-01

    Fatty acids are a major fuel for many tissues and abnormal utilization is implicated in diseases. However, tissue fatty acid oxidation has not been determined reliably in vivo. Furthermore, fatty acid oxidation has not been partitioned into intracellular and extracellular components. In this report, a one-pool model is described that enables direct quantitation of fluxes of intracellular and plasma fatty acids to mitochondria in skeletal muscle using dual stable isotopes and liquid chromatogr...

  19. Endophytic filamentous fungi from a Catharanthus roseus: Identification and its hydrolytic enzymes

    Farah Wahida Ayob

    2016-05-01

    Full Text Available This paper reported on the various filamentous fungi strains that were isolated from a wild grown Catharanthus roseus. Based on the morphological characteristics and molecular technique through a Polymerase Chain Reaction and DNA sequencing method using internal transcribed spacer (ITS, these fungi had been identified as a Colletotrichum sp., Macrophomina phaseolina, Nigrospora sphaerica and Fusarium solani. The ultrastructures of spores and hyphae were observed under a Scanning Electron Microscope. The hydrolytic enzyme test showed that all strains were positive in secreting cellulase. Colletotrichum sp. and F. solani strains also gave a positive result for amylase while only F. solani was capable to secrete protease. These fungi were putatively classified as endophytic fungi since they produced extracellular enzymes that allow them to penetrate plant cell walls and colonize with symbiotic properties.

  20. Endophytic filamentous fungi from a Catharanthus roseus: Identification and its hydrolytic enzymes.

    Ayob, Farah Wahida; Simarani, Khanom

    2016-05-01

    This paper reported on the various filamentous fungi strains that were isolated from a wild grown Catharanthus roseus. Based on the morphological characteristics and molecular technique through a Polymerase Chain Reaction and DNA sequencing method using internal transcribed spacer (ITS), these fungi had been identified as a Colletotrichum sp., Macrophomina phaseolina, Nigrospora sphaerica and Fusarium solani. The ultrastructures of spores and hyphae were observed under a Scanning Electron Microscope. The hydrolytic enzyme test showed that all strains were positive in secreting cellulase. Colletotrichum sp. and F. solani strains also gave a positive result for amylase while only F. solani was capable to secrete protease. These fungi were putatively classified as endophytic fungi since they produced extracellular enzymes that allow them to penetrate plant cell walls and colonize with symbiotic properties. PMID:27275114

  1. Halophilic Bacteria as a Source of Novel Hydrolytic Enzymes

    Encarnación Mellado

    2013-01-01

    Full Text Available Hydrolases constitute a class of enzymes widely distributed in nature from bacteria to higher eukaryotes. The halotolerance of many enzymes derived from halophilic bacteria can be exploited wherever enzymatic transformations are required to function under physical and chemical conditions, such as in the presence of organic solvents and extremes in temperature and salt content. In recent years, different screening programs have been performed in saline habitats in order to isolate and characterize novel enzymatic activities with different properties to those of conventional enzymes. Several halophilic hydrolases have been described, including amylases, lipases and proteases, and then used for biotechnological applications. Moreover, the discovery of biopolymer-degrading enzymes offers a new solution for the treatment of oilfield waste, where high temperature and salinity are typically found, while providing valuable information about heterotrophic processes in saline environments. In this work, we describe the results obtained in different screening programs specially focused on the diversity of halophiles showing hydrolytic activities in saline and hypersaline habitats, including the description of enzymes with special biochemical properties. The intracellular lipolytic enzyme LipBL, produced by the moderately halophilic bacterium Marinobacter lipolyticus, showed advantages over other lipases, being an enzyme active over a wide range of pH values and temperatures. The immobilized LipBL derivatives obtained and tested in regio- and enantioselective reactions, showed an excellent behavior in the production of free polyunsaturated fatty acids (PUFAs. On the other hand, the extremely halophilic bacterium, Salicola marasensis sp. IC10 showing lipase and protease activities, was studied for its ability to produce promising enzymes in terms of its resistance to temperature and salinity.

  2. Extracellular loop 2 of the free Fatty Acid receptor 2 mediates allosterism of a phenylacetamide ago-allosteric modulator

    Smith, Nicola J; Ward, Richard J; Stoddart, Leigh A; Hudson, Brian D; Kostenis, Evi; Ulven, Trond; Morris, Joanne C; Tränkle, Christian; Tikhonova, Irina G; Adams, David R; Milligan, Graeme

    2011-01-01

    Allosteric agonists are powerful tools for exploring the pharmacology of closely related G protein-coupled receptors that have nonselective endogenous ligands, such as the short chain fatty acids at free fatty acid receptors 2 and 3 (FFA2/GPR43 and FFA3/GPR41, respectively). We explored the molec...

  3. Evidence of hydrolytic route for anaerobic cyanide degradation.

    Fallon, R. D.

    1992-01-01

    Products observed during anaerobic cyanide transformation are consistent with a hydrolytic pathway (HCN + H2O HCONH2 + H2O HCOOH + NH3). Formate, the most frequently observed product, was generally converted to bicarbonate. Formamide was rapidly hydrolyzed to formate upon exposure to the anaerobic consortium but was not detected as an intermediate of cyanide transformation.

  4. Sponge-associated bacteria of Lakshadweep coral reefs, India: resource for extracellular hydrolytic enzymes

    Feby, A.; Nair, S.

    'Amico, S., Feller, G. and Ger- day, C. (2007) Cold-adap arctic microorganisms. Marine Biotechnology, 9(3), 293-304. [17] Lee, Y.K., Lee, J.H. and Lee, H.K. (2001) Microbial symbiosis , ti 39(4), 254-264. [18] Reiswig, H.M. (1981) Partial Carbon... esterases from Bacillus sp associated with the marine sponge Aplysina aerophoba. Applied Microbiology and Biotechnology, 67(1), 59-69. [22] Lee, O.O., Lau nikova, I., Dobretsov, S., Wu, M.C.S., Wong, P.K., Weinbauer, M. and Qian, P.Y. (2006) Shewanella...

  5. Effect of pioglitazone, quercetin and hydroxy citric acid on extracellular matrix components in experimentally induced non-alcoholic steatohepatitis

    Surapaneni Krishna Mohan

    2015-08-01

    Results:The experimental NASH rats treated with pioglitazone showed significant decrease in the levels of hyaluronic acid and significant increase in adiponectin levels when compared to experimentally induced NASH group, but did not show any effect on the levels of leptin. Contrary to these two drugs, viz. pioglitazone and hydroxy citric acid, the group treated with quercetin showedsignificant decrease in the levels of hyaluronic acid and leptin and significant decrease in adiponectin levels compared with that of experimentally induced NASH NASH group, offering maximum protection against NASH. Conclusion: Considering our findings, it could be concluded that quercetin may offer maximum protection against NASH by significantly increasing the levels of adiponectin, when compared to pioglitazone and hydroxy citric acid.

  6. Cyclic Imide Dioxime: Formation and Hydrolytic Stability

    Kang, S.O. [University of Kansas; Vukovic, Sinisa [ORNL; Custelcean, Radu [ORNL; Hay, Benjamin [ORNL

    2012-01-01

    Poly(acrylamidoximes) play an important role in the uranium extraction from seawater. The present work reports solution studies of simple analogs to address the formation and stability of two binding sites present in these polymers, open-chain amidoximes and cyclic imide dioximes, including: 1) conditions that maximize the formation of the cyclic form, 2) existence of a base-induced conversion from open-chain to cyclic form, and 3) degradation under acid and base conditions.

  7. Hydrolytic enzyme activity in rhesus monkey placenta during early gestational malaria : histochemical studies

    Nishi Saxena & P.S.R. Murthy

    2005-12-01

    Full Text Available Background & objectives: Early gestational malaria is found to be more fatal than late gestationalinfection but the pathophysiology of early gestational placenta, the maternofoetal organ responsiblefor maintenance of pregnancy, remains unexplored. Present study dealing with hydrolytic enzymes inearly gestational placenta of rhesus monkeys during Plasmodium cynomolgi infection was anticipatedto provide a better insight into the functional impairment of this organ during early gestationalmaternal malaria.Methods: Experimental monkeys (Macaca multtta at 2–2½ months of pregnancy were inoculatedwith P. cynomolgi bastianelli. After attaining first peak of parasitaemia the animals were anesthetisedand placentae were collected for histochemical studies. The snap-frozen, cryostat sections weresubjected to histochemical reactions for acid phosphatase and alkaline phosphatase.Results: The placental syncytiotrophoblast showed a loss in alkaline phosphatase activity, while thetrophoblast layers and phagocytic cells of the maternal blood showed increased acid phosphataseactivity during early gestational malarial infection. Morphological damage to the placental tissuewhenever occurred was associated with altered Alk pase activity.Interpretation & conclusion: The altered distribution of Ac pase and Alk pase in malaria infectedearly gestational placenta has been discussed in the light of placental function. It could be concludedby present studies that these malaria induced changes in hydrolytic enzyme activities in monkeyplacenta have a direct bearing on functional and morphological integrity of the placental tissue.These changes are apparently responsible for early gestational foetal death and abortions as reportedin literature

  8. Effect of amphetamine on extracellular concentrations of amino acids in striatum in neurotensin subtype 1 and 2 receptor null mice: a possible interaction between neurotensin receptors and amino acid systems for study of schizophrenia

    Li, Zhimin; Liang, Yanqi; Boules, Mona; Gordillo, Andres; Richelson, Elliott

    2010-01-01

    Neurotensin (NT) is a tridecapeptide that acts as a neuromodulator in the central nervous system mainly through two NT receptors: NTS1 and NTS2. The present study was done to determine the roles of NTS1 and NTS2 on amino acid release in striatum with the use of NTS1 or NTS2 knock-out (-/-) mice given d-amphetamine. Both NTS1-/- and NTS2-/- mice had lower extracellular concentrations of D-serine in striatum than did wild type (WT) mice. NTS2-/- but not NTS1-/- mice also had significantly lower...

  9. MODULATION OF EASTERN OYSTER HEMOCYTE ACTIVITIES BY PERKINSUS MARINUS EXTRACELLULAR PROTEINS

    The oyster pathogen Perkinsus marinusproduces many extracellular proteins (ECP) in vitro. Analysis of this ECP revealed a battery of hydrolytic enzymes. Some of these enzymes are known to modulate the activity of host defense cells. Although information on the effects of P. marin...

  10. Overexpression of OsPAP10a, A Root-Associated Acid Phosphatase, Increased Extracellular Organic Phosphorus Utilization in Rice

    Jingluan Tian; Chuang Wang; Qian Zhang; Xiaowei He; James Whelan; Huixia Shou

    2012-01-01

    Phosphorus (P) deficiency is a major limitation for plant growth and development.Among the wide set of responses to cope with low soil P,plants increase their level of intracellular and secreted acid phosphatases (APases),which helps to catalyze inorganic phosphate (Pi) hydrolysis from organophosphates.In this study we characterized the rice (Oryza sativa) purple acid phosphatase 10a (OsPAP10a).OsPAP10a belongs to group la of purple acid phosphatases (PAPs),and clusters with the principal secreted PAPs in a variety of plant species including Arabidopsis.The transcript abundance of OsPAP10a is specifically induced by Pi deficiency and is controlled by OsPHR2,the central transcription factor controlling Pi homeostasis.In gel activity assays of root and shoot protein extracts,it was revealed that OsPAP10a is a major acid phosphatase isoform induced by Pi starvation.Constitutive overexpression of OsPAP10a results in a significant increase of phosphatase activity in both shoot and root protein extracts.In vivo root 5-bromo-4-chloro-3-indolyl-phosphate (BCIP) assays and activity measurements on external media showed that OsPAP10a is a root-associated APase.Furthermore,overexpression of OsPAP10a significantly improved ATP hydrolysis and utilization compared with wild type plants.These results indicate that OsPAP10a can potentially be used for crop breeding to improve the efficiency of P use.

  11. Tailoring of the titanium surface by preparing cardiovascular endothelial extracellular matrix layer on the hyaluronic acid micro-pattern for improving biocompatibility.

    Li, Jingan; Zhang, Kun; Wu, Juejue; Zhang, Lijuan; Yang, Ping; Tu, Qiufen; Huang, Nan

    2015-04-01

    It has been proved that high molecular weight hyaluronic acid (HMW-HA, 1×10(6) Da) micro-strips on titanium (Ti) surface can elongate the human vascular endothelial cell (EC) morphology, subsequently enhance endothelial extracellular matrix (ECM) deposition in our previous work. The HMW-HA micro-strips were anticipated to possess good hemocompatibility and EC compatibility simultaneously. However, the single HMW-HA micro-strips on Ti substrate showed bad hemocompatibility. To solve this problem, a method combining HA micro-pattern and EC decellularization was developed, and the endothelial extracellular matrix layer on the HA micro-pattern (ECM/HAP) showed excellent hemocompatibility and endothelial progenitor cells (EPCs) compatibility (cell number: 14.3±0.5×10(5) cells/cm2>2.2±0.7×10(5) cells/cm2 on ECM/TiOH, 7.5±1.3×10(5) cells/cm2 on TiOH, 3.4±0.9×10(5) cells/cm2 on TiOH/HAP and 3.6±1.2×10(5) cells/cm2 on Ti). We also found that the ECM/HAP coating could significantly inhibit the excessive proliferation of smooth muscle cells (SMCs) (cck-8 absorption: 0.25±0.06coating can be used to build the bionic human endothelial ECM on the biomaterials surface, which might provide a potential and effective method for surface modification of cardiovascular devices. PMID:25684737

  12. Metabolism of [3-3H]oleanolic acid in the isolated ''Calendula officinalis'' leaf cells and transport of the synthesized glycosides, to the cell wall and the extracellular space

    It has been shown for the first time that [3-3H]oleanolic acid glycosides formed in the cytosol of ''C. officinalis'' leaf cells are transported to the extracellular space in the form of pentaglucoside VI (44%), whereas glucuronides derived from [3-3H]oleanolic acid 3-O-monoglucuronide (29%) as well as a part of glucosides (24%) were transported into the cell walls. (author). 15 refs, 2 figs, 1 tab

  13. Metabolism of [3-{sup 3}H]oleanolic acid in the isolated ``Calendula officinalis`` leaf cells and transport of the synthesized glycosides, to the cell wall and the extracellular space

    Szakiel, A.; Wasiukiewicz, I.; Janiszowska, W. [Warsaw Univ. (Poland). Katedra Biochemii

    1995-12-31

    It has been shown for the first time that [3-{sup 3}H]oleanolic acid glycosides formed in the cytosol of ``C. officinalis`` leaf cells are transported to the extracellular space in the form of pentaglucoside VI (44%), whereas glucuronides derived from [3-{sup 3}H]oleanolic acid 3-O-monoglucuronide (29%) as well as a part of glucosides (24%) were transported into the cell walls. (author). 15 refs, 2 figs, 1 tab.

  14. Multiscale surface analyses of hydrolytic stable and degradable polyurethanes

    Špírková, Milena; Serkis, Magdalena; Poreba, Rafal; Machová, Luďka; Hodan, Jiří

    Prague : Institute of Macromolecular Chemistry AS CR, 2015. s. 57. ISBN 978-80-85009-82-8. [Functional Polymers at Bio -Material Interfaces - 79th Prague Meeting on Macromolecules. 28.06.2015-02.07.2015, Prague] R&D Projects: GA ČR(CZ) GA13-06700S Institutional support: RVO:61389013 Keywords : polyurethane elastomers * hydrolytic stability * surface analysis Subject RIV: CD - Macromolecular Chemistry

  15. Hydrolytic studies in physiologically simulated conditions II. Degradable aliphatic polyurethanes

    Poreba, Rafal; Machová, Luďka; Serkis, Magdalena; Hodan, Jiří; Kredatusová, Jana; Špírková, Milena

    Košice: Ústav materiálového výskumu SAV, 2015. s. 6-7. ISBN 978-80-89782-02-04. [Funkčné kompozitné materiály. 19.05.2015, Košice] R&D Projects: GA ČR(CZ) GA13-06700S Institutional support: RVO:61389013 Keywords : polyurethanes * hydrolytic degradation * film Subject RIV: CD - Macromolecular Chemistry

  16. Hydrolytic studies in physiologically simulated conditions I. Stable aliphatic polyurethanes

    Špírková, Milena; Poreba, Rafal; Serkis, Magdalena; Hodan, Jiří; Kredatusová, Jana

    Košice : Ústav materiálového výskumu SAV, 2015. s. 4-5. ISBN 978-80-89782-02-04. [Funkčné kompozitné materiály. 19.05.2015, Košice] R&D Projects: GA ČR(CZ) GA13-06700S Institutional support: RVO:61389013 Keywords : polyurethanes * hydrolytic stability * film Subject RIV: CD - Macromolecular Chemistry

  17. General characterization of noncommercial microbial lipases in hydrolytic and synthetic reactions.

    Otero, C; Berrendero, M A; Cardenas, F; Alvarez, E; Elson, S W

    2005-03-01

    Fourteen noncommercial preparations of microbial lipases were investigated with respect to their catalytic activity for hydrolysis and synthesis of ester bonds. Six of the lipases were derived from microorganisms that have not previously been described as lipase producers, and another four were characterized for the first time. The synthetic reactions were carried out in two solvents of different polarities (n-heptane and acetone) using a series of fatty acids and primary and secondary alcohols with different chain lengths. Under the culture conditions employed, Pseudomonas cepacia produced more active enzyme than the other microorganisms. The lipase preparations produced using Ovadendron sulphureo-ochraceum, Monascus mucoroides, Monascus sp., Fusarium oxysporum, Penicillium chrysogenum, Rhodotorula araucariae, Pseudomonas cepacia, Streptomyces halstedii, and Streptomyces sp.were the most efficient catalysts for hydrolysis at lipid-water interfaces. Enzyme preparations from P. cepacia, Streptomyces sp., S. halstedii, and R. araucariae were good biocatalysts for esterification in the polar medium (acetone). When the lipase preparations with the greatest activity for hydrolytic reactions were excluded, regression analysis of the data for the hydrolytic and synthetic activities of the remaining lipase preparations yielded high multiple correlation coefficients for these reactions in both n-heptane and acetone (R = 0.82 and 0.91, respectively). PMID:15767695

  18. A method to measure hydrolytic activity of adenosinetriphosphatases (ATPases.

    Gianluca Bartolommei

    Full Text Available The detection of small amounts (nanomoles of inorganic phosphate has a great interest in biochemistry. In particular, phosphate detection is useful to evaluate the rate of hydrolysis of phosphatases, that are enzymes able to remove phosphate from their substrate by hydrolytic cleavage. The hydrolysis rate is correlated to enzyme activity, an extremely important functional parameter. Among phosphatases there are the cation transporting adenosinetriphosphatases (ATPases, that produce inorganic phosphate by cleavage of the γ-phosphate of ATP. These membrane transporters have many fundamental physiological roles and are emerging as potential drug targets. ATPase hydrolytic activity is measured to test enzyme functionality, but it also provides useful information on possible inhibitory effects of molecules that interfere with the hydrolytic process. We have optimized a molybdenum-based protocol that makes use of potassium antimony (III oxide tartrate (originally employed for phosphate detection in environmental analysis to allow its use with phosphatase enzymes. In particular, the method was successfully applied to native and recombinant ATPases to demonstrate its reliability, validity, sensitivity and versatility. Our method introduces significant improvements to well-established experimental assays, which are currently employed for ATPase activity measurements. Therefore, it may be valuable in biochemical and biomedical investigations of ATPase enzymes, in combination with more specific tests, as well as in high throughput drug screening.

  19. Loss of force induced by high extracellular [K+] in rat muscle: effect of temperature, lactic acid and beta2-agonist.

    Pedersen, Thomas Holm; Clausen, Torben; Nielsen, Ole Baekgaard

    2003-08-15

    Loss of K+ from active muscles, leading to increased [K+]o, has been proposed to cause muscle fatigue by reducing excitability. Since exercise increases muscle temperature, we investigated the influence of temperature on muscle [K+]o sensitivity. Intact rat soleus or extensor digitorum longus (EDL) muscles were mounted on force transducers and stimulated electrically to evoke short isometric tetani at regular intervals. In each experiment, control force at 4 mM K+ was initially determined at every temperature used. In soleus muscles at 20 degrees C, 9 mM K+ reduced force to 33 +/- 5 % of control force. Increasing the temperature to 30 degrees C restored force to 89 +/- 5 % of control force. Likewise, at 30 degrees C 11 mM K+ reduced force to 16 +/- 4 % and increasing the temperature to 35 degrees C restored force to 35 +/- 5 %. Similar results were obtained using EDL. The force recovery induced by elevating temperature, reflecting reduced [K+]o sensitivity, was associated with improved excitability assessed from compound action potentials. Force recovery induced by a temperature elevation from 20 to 30 degrees C was associated with hyperpolarization (5 mV), reduced [Na+]i and a 93 % increase in Na+-K+ pump activity. The force recovery was blocked by ouabain. Since intensive exercise leads to lactic acidosis and increased plasma catecholamines, the effect of these two factors was also investigated. At 11 mM K+, force was completely restored by combining temperature elevation (30 to 35 degrees C), L-lactic acid (10 mM) and the beta2-agonist salbutamol (10-5 M). We suggest an exercise scenario where the depressing action of exercise-induced hyperkalaemia is counteracted by elevated muscle temperature, lactic acidosis and catecholamines. PMID:12813152

  20. Electrostatic and potential cation-pi forces may guide the interaction of extracellular loop III with Na+ and bile acids for human apical Na+-dependent bile acid transporter.

    Banerjee, Antara; Hussainzada, Naissan; Khandelwal, Akash; Swaan, Peter W

    2008-03-01

    The hASBT (human apical Na(+)-dependent bile acid transporter) constitutes a key target of anti-hypercholesterolaemic therapies and pro-drug approaches; physiologically, hASBT actively reclaims bile acids along the terminal ileum via Na(+) co-transport. Previously, TM (transmembrane segment) 7 was identified as part of the putative substrate permeation pathway using SCAM (substitute cysteine accessibility mutagenesis). In the present study, SCAM was extended through EL3 (extracellular loop 3; residues Arg(254)-Val(286)) that leads into TM7 from the exofacial matrix. Activity of most EL3 mutants was significantly hampered upon cysteine substitution, whereas ten (out of 31) were functionally inactive (<10% activity). Since only E282C lacked plasma membrane expression, EL3 amino acids predominantly fulfill critical functional roles during transport. Oppositely charged membrane-impermeant MTS (methanethiosulfonate) reagents {MTSET [(2-trimethylammonium) ethyl MTS] and MTSES [(2-sulfonatoethyl) MTS]} produced mostly similar inhibition profiles wherein only middle and descending loop segments (residues Thr(267)-Val(286)) displayed significant MTS sensitivity. The presence of bile acid substrate significantly reduced the rates of MTS modification for all MTS-sensitive mutants, suggesting a functional association between EL3 residues and bile acids. Activity assessments at equilibrative [Na(+)] revealed numerous Na(+)-sensitive residues, possibly performing auxiliary functions during transport such as transduction of protein conformational changes during translocation. Integration of these data suggests ligand interaction points along EL3 via electrostatic interactions with Arg(256), Glu(261) and probably Glu(282) and a potential cation-pi interaction with Phe(278). We conclude that EL3 amino acids are essential for hASBT activity, probably as primary substrate interaction points using long-range electrostatic attractive forces. PMID:18028035

  1. Ultrasound-assisted extraction and characterization of hydrolytic and oxidative enzymes produced by solid state fermentation.

    Szabo, Orsolya Erzsebet; Csiszar, Emilia; Toth, Karolina; Szakacs, George; Koczka, Bela

    2015-01-01

    Ligninolytic and hydrolytic enzymes were produced with six selected fungi on flax substrate by solid state fermentation (SSF). The extracellular enzyme production of the organisms in two SSF media was evaluated by measuring the soluble protein concentration and the filter paper, endoxylanase, 1,4-β-d-glucosidase, 1,4-β-d-endoglucanase, polygalacturonase, lignin peroxidase, manganese peroxidase and laccase activities of the clear culture solutions produced by conventional extraction from the SSF materials. The SSF material of the best enzyme producer (Trichoderma virens TUB F-498) was further investigated to enhance the enzyme recovery by low frequency ultrasound treatment. Performance of both the original and ultrasound macerated crude enzyme mixtures was evaluated in degradation of the colored lignin-containing and waxy materials of raw linen fabric. Results proved that sonication (at 40%, 60% and 80% amplitudes, for 60min) did not result in reduction in the filter paper, lignin peroxidase and laccase activities of the crude enzyme solution, but has a significant positive effect on the efficiency of enzyme extraction from the SSF material. Depending on the parameters of sonication, the enzyme activities in the extracts obtained can be increased up to 129-413% of the original activities measured in the control extracts recovered by a common magnetic stirrer. Sonication also has an effect on both the enzymatic removal of the lignin-containing color materials and hydrophobic surface layer from the raw linen. PMID:25043555

  2. Advances in hexitol and ethylene glycol production by one-pot hydrolytic hydrogenation and hydrogenolysis of cellulose

    In this review, recent advances in the one-pot hydrolytic hydrogenation and hydrogenolysis of cellulose to value-added polyols, including hexitols (sorbitol, mannitol, and isosorbide) and 1,2-alkanediols (ethylene glycol and 1,2-propylene glycol), are summarized. Methods for the generation of H+ in the first step of cellulose hydrolysis to form intermediate sugars, such as the use of soluble acids (mineral acids and heteropoly acids) and H+ produced in situ from functional supports and H2 dissociation, are classified and analyzed, considering its combination with active metals for the subsequent hydrogenation or hydrogenolysis of sugars to polyols. The interaction of non-noble metals such as nickel, bimetals, and tungsten with support materials in the catalytic conversion of intermediate sugars to hexitols and ethylene glycol is reviewed. The corresponding reaction pathways and mechanisms are discussed, including the conversion process using basic supports and solution conditions. Major challenges and promising routes are also suggested for the future development of the chemocatalytic conversion of cellulose. - Highlights: • Advances in the one-pot hydrolytic hydrogenation/hydrogenolysis of cellulose are summarized. • The interaction of non-noble metals with support materials for cellulose conversion is reviewed. • Method for the generation of in situ H+ and effects of the acidic groups on supports are discussed. • Incomplete identification of intermediates/products causes mechanism complications. • Efficient conversion, separation and purification are other concerns for cellulose degrading

  3. Comparison of hydrolytic and non-hydrolytic atomic layer deposition chemistries: Interfacial electronic properties at alumina-silicon interfaces

    Marstell, Roderick J.; Strandwitz, Nicholas C.

    2015-11-01

    We report the differences in the passivation and electronic properties of aluminum oxide (Al2O3) deposited on silicon via traditional hydrolytic atomic layer deposition (ALD) and non-hydrolytic (NH) ALD chemistries. Traditional films were grown using trimethylaluminum (TMA) and water and NHALD films grown using TMA and isopropanol at 300 °C. Hydrolytically grown ALD films contain a smaller amount of fixed charge than NHALD films (oxide fixed charge Qf Traditional = -8.1 × 1011 cm-2 and Qf NHALD = -3.6 × 1012 cm-2), and a larger degree of chemical passivation than NHALD films (density of interface trap states, Dit Traditional = 5.4 × 1011 eV-1 cm-2 and Dit NHALD = 2.9 × 1012 eV-1 cm-2). Oxides grown with both chemistries were found to have a band gap of 7.1 eV. The conduction band offset was 3.21 eV for traditionally grown films and 3.38 eV for NHALD. The increased Dit for NHALD films may stem from carbon impurities in the oxide layer that are at and near the silicon surface, as evidenced by both the larger trap state time constant (τTraditional = 2.2 × 10-9 s and τNHALD = 1.7 × 10-7 s) and the larger carbon concentration. We have shown that the use of alcohol-based oxygen sources in NHALD chemistry can significantly affect the resulting interfacial electronic behavior presenting an additional parameter for understanding and controlling interfacial electronic properties at semiconductor-dielectric interfaces.

  4. Comparison of hydrolytic and non-hydrolytic atomic layer deposition chemistries: Interfacial electronic properties at alumina-silicon interfaces

    We report the differences in the passivation and electronic properties of aluminum oxide (Al2O3) deposited on silicon via traditional hydrolytic atomic layer deposition (ALD) and non-hydrolytic (NH) ALD chemistries. Traditional films were grown using trimethylaluminum (TMA) and water and NHALD films grown using TMA and isopropanol at 300 °C. Hydrolytically grown ALD films contain a smaller amount of fixed charge than NHALD films (oxide fixed charge Qf Traditional = −8.1 × 1011 cm−2 and Qf NHALD = −3.6 × 1012 cm−2), and a larger degree of chemical passivation than NHALD films (density of interface trap states, Dit Traditional = 5.4 × 1011 eV−1 cm−2 and Dit NHALD = 2.9 × 1012 eV−1 cm−2). Oxides grown with both chemistries were found to have a band gap of 7.1 eV. The conduction band offset was 3.21 eV for traditionally grown films and 3.38 eV for NHALD. The increased Dit for NHALD films may stem from carbon impurities in the oxide layer that are at and near the silicon surface, as evidenced by both the larger trap state time constant (τTraditional = 2.2 × 10−9 s and τNHALD = 1.7 × 10−7 s) and the larger carbon concentration. We have shown that the use of alcohol-based oxygen sources in NHALD chemistry can significantly affect the resulting interfacial electronic behavior presenting an additional parameter for understanding and controlling interfacial electronic properties at semiconductor-dielectric interfaces

  5. An amino acid residue in the second extracellular loop determines the agonist-dependent tolerance property of the human D3 dopamine receptor.

    Gil-Mast, Sara; Kortagere, Sandhya; Kota, Kokila; Kuzhikandathil, Eldo V

    2013-06-19

    The D3 dopamine receptor is a therapeutic target for treating various nervous system disorders such as schizophrenia, Parkinson's disease, depression, and addictive behaviors. The crystal structure of the D3 receptor bound to an antagonist was recently described; however, the structural features that contribute to agonist-induced conformational changes and signaling properties are not well understood. We have previously described the conformation-dependent tolerance and slow response termination (SRT) signaling properties of the D3 receptor and identified the C147 residue in the second intracellular loop (IL2) of the D3 receptor as important for the tolerance property. Interestingly, while IL2 and the C147 residue, in particular, were important for dopamine- and quinpirole-induced tolerance, this residue did not affect the severe tolerance induced by the high affinity, D3 receptor-selective agonist, PD128907. Here, we used D2/D3 receptor chimeras and site-specific D3 receptor mutants to identify another residue, D187, in the second extracellular loop (EC2) of the human D3 receptor that mediates the tolerance property induced by PD128907, quinpirole, pramipexole, and dopamine. Molecular dynamics simulations confirmed the distinct conformation adopted by D3 receptor during tolerance and suggested that in the tolerant D3 receptor the D187 residue in EC2 forms a salt bridge with the H354 residue in EC3. Indeed, site-directed mutation of the H354 residue resulted in loss of PD1287907-induced tolerance. The mapping of specific amino acid residues that contribute to agonist-dependent conformation changes and D3 receptor signaling properties refines the agonist-bound D3 receptor pharmacophore model which will help develop novel D3 receptor agonists. PMID:23477444

  6. Exercise pre‑conditioning alleviates brain damage via excitatory amino acid transporter 2 and extracellular signal‑regulated kinase 1/2 following ischemic stroke in rats.

    Wang, Xiao; Zhang, Min; Feng, Rui; Li, Wen-Bin; Ren, Shi-Qing; Zhang, Feng

    2015-02-01

    Previous studies have reported that physical exercise may exert a neuroprotective effect in humans as well as animals. However, the detailed mechanisms underlying the neuroprotective effect of exercise has remained to be elucidated. The aim of the present study was to explore the possible signaling pathways involved in the protective effect of pre‑ischemic treadmill training for ischemic stroke in rats. A total of 36 male Sprague‑Dawley rats were divided at random into three groups as follows (n=12 for each): Sham surgery group; middle cerebral artery occlusion (MCAO) group; and exercise with MCAO group. Following treadmill training for three weeks, the middle cerebral artery was occluded for 90 min in order to induce ischemic stroke, followed by reperfusion. Following 24 h post‑reperfusion, six rats from each group were assessed for neurological deficits and then sacrificed to calculate the infarct volume. The remaining rats (n=6 for each group) were sacrificed and the expression levels of excitatory amino acid transporter 2 (EAAT‑2) and extracellular signal‑regulated kinase 1/2 (ERK1/2) were detected using western blot analysis. The results of the present study demonstrated that rats that underwent pre‑ischemic exercise intervention had a significantly decreased brain infarct volume and neurological deficits; in addition, the pre‑ischemic exercise group showed decreased overexpression of phosphorylated ERK1/2 and increased expression of EAAT‑2 following ischemic stroke. In conclusion, treadmill training exercise prior to ischemic stroke alleviated brain damage in rats via regulation of EAAT‑2 and ERK1/2. PMID:25370789

  7. Tumor extracellular acidity-activated nanoparticles as drug delivery systems%肿瘤酸度活化的纳米药物载体

    李洪军; 王均

    2014-01-01

    pH响应性的纳米粒(nanoparticles,NPs)作为药物输送载体用于肿瘤治疗倍受关注。实体瘤的弱酸性肿瘤微环境(pHe约为6.8)为抗肿瘤药物纳米载体的设计提供了新思路和发展契机。相对于靶向基团介导的肿瘤靶向策略,对pHe响应的纳米载体更具有普适性。本文概述了pHe活化纳米药物载体在肿瘤治疗中的设计和应用现状,重点介绍pHe触发表面电荷反转的纳米颗粒用于抗肿瘤药物和小干扰RNA(siRNA)输送的研究进展,并展望这种新型载体在增强肿瘤疗效方面的应用潜力。%pH-responsive nanoparticles (NPs) are currently under intense development as carriers of drug delivery systems for cancer therapy. Of these NPs, those that are designed to target the slightly acidic extracellular pH environment (pHe~6.8) of solid tu-mors offer a new paradigm of tumor-targeted drug delivery. Compared with conventional, specific, surface-targeting approaches, the pHe-targeting strategy is considered to be more general because of the common occurrence of an acidic microenvironment in solid tu-mors. This review mainly focuses on the design and applications of pHe-activated NPs, particularly on pHe-activated surface-charge re-versal NPs, for drug and siRNA delivery to tumors. The novel development of NPs described in this review has high potential for achieving stronger therapeutic effects in cancer treatment.

  8. Preliminary Study on the Performance and Interaction of Recycling Hydrolytic-Aerobic Combined Process of High Concentration Starch Wastewater%高浓度淀粉废水水解--好氧循环一体化处理工艺的初步研究

    李清彪; 廖鑫凯; 吴志旺; 邓旭; 黄益丽; 卢英华; 孙道华; 洪铭嫒; 王琳

    2004-01-01

    A new recycling hydrolytic-aerobic combined process was developed to treat the high concentration used. At first, the hydrolytic degradation and aerobic degradation process were examined in two batch reactors,respectively. In the stand-alone hydrolytic process, starch in the wastewater almost disappeared after 11 h treatment,but CODCr remained as high as 5803mg. L-1 after two days. In the aerobic process, the biodegradation rate of starch was much slower during the first 11 h than that in the hydrolytic process, although the CODCr removal efficiency reached 89.6% and more than 90% starch could be degraded after 37.5 h. To determine the interaction effects of the two processes, a series of hydrolytic-aerobic combinations were examined in details. Hydrolytic process played an important role in the whole recycle combination process as it could improve the biodegradability of the high concentration starch wastewater. However, from the other experiments, the negative effect of hydrolytic acidification was found in the hydrolytic-aerobic combination, which suggested that the aerobic microorganisms needed time to adapt themselves to the acidic environment. The effect of the degrading time, which was spent in the hydrolytic and aerobic unit, and the number of circulations, with which the wastewater went through the two units were investigated. It was found that a recycle combination of 6 h hydrolytic process with 12 h aerobic process was highly effective and potentially economical, in which the final removal efficiency of CODCr and efficiency of starch degradation reached 94.1% and 98.8%, respectively.

  9. Extracellular Acidification Alters Lysosomal Trafficking in Human Breast Cancer Cells

    Kristine Glunde; Sandra E. Guggino; Meiyappan Solaiyappan; Pathak, Arvind P.; Yoshitaka Ichikawa; Bhujwalla, Zaver M.

    2003-01-01

    Cancer cells invade by secreting degradative enzymes, which are sequestered in lysosomal vesicles. In this study, the impact of an acidic extracellular environment on lysosome size, number, and distance from the nucleus in human mammary epithelial cells (HMECs) and breast cancer cells of different degrees of malignancy was characterized because the physiological microenvironment of tumors is frequently characterized by extracellular acidity. An acidic extracellular pH (pHe) resulted in a dist...

  10. Comparison of hydrolytic and non-hydrolytic atomic layer deposition chemistries: Interfacial electronic properties at alumina-silicon interfaces

    Marstell, Roderick J.; Strandwitz, Nicholas C., E-mail: strand@lehigh.edu [Department of Materials Science and Engineering and Center for Advanced Materials and Nanotechnology, Lehigh University, Bethlehem, Pennsylvania 18015 (United States)

    2015-11-14

    We report the differences in the passivation and electronic properties of aluminum oxide (Al{sub 2}O{sub 3}) deposited on silicon via traditional hydrolytic atomic layer deposition (ALD) and non-hydrolytic (NH) ALD chemistries. Traditional films were grown using trimethylaluminum (TMA) and water and NHALD films grown using TMA and isopropanol at 300 °C. Hydrolytically grown ALD films contain a smaller amount of fixed charge than NHALD films (oxide fixed charge Q{sub f} {sub Traditional} = −8.1 × 10{sup 11 }cm{sup −2} and Q{sub f} {sub NHALD} = −3.6 × 10{sup 12 }cm{sup −2}), and a larger degree of chemical passivation than NHALD films (density of interface trap states, D{sub it} {sub Traditional} = 5.4 × 10{sup 11 }eV{sup −1 }cm{sup −2} and D{sub it} {sub NHALD} = 2.9 × 10{sup 12 }eV{sup −1 }cm{sup −2}). Oxides grown with both chemistries were found to have a band gap of 7.1 eV. The conduction band offset was 3.21 eV for traditionally grown films and 3.38 eV for NHALD. The increased D{sub it} for NHALD films may stem from carbon impurities in the oxide layer that are at and near the silicon surface, as evidenced by both the larger trap state time constant (τ{sub Traditional} = 2.2 × 10{sup −9} s and τ{sub NHALD} = 1.7 × 10{sup −7} s) and the larger carbon concentration. We have shown that the use of alcohol-based oxygen sources in NHALD chemistry can significantly affect the resulting interfacial electronic behavior presenting an additional parameter for understanding and controlling interfacial electronic properties at semiconductor-dielectric interfaces.

  11. SAMHD1, the Aicardi-Goutières syndrome gene and retroviral restriction factor, is a phosphorolytic ribonuclease rather than a hydrolytic ribonuclease.

    Ryoo, Jeongmin; Hwang, Sung-Yeon; Choi, Jongsu; Oh, Changhoon; Ahn, Kwangseog

    2016-09-01

    SAMHD1 plays diverse roles in innate immunity, autoimmune diseases and HIV restriction, but the mechanisms involved are still unclear. SAMHD1 has been reported to have both dNTPase and RNase activities. However, whether SAMHD1 possesses RNase activity remains highly controversial. Here, we found that, unlike conventional hydrolytic exoribonucleases, SAMHD1 requires inorganic phosphate to degrade RNA substrates and produces nucleotide diphosphates rather than nucleoside monophosphates, which indicated that SAMHD1 is a phosphorolytic but not hydrolytic 3'-5' exoribonuclease. Furthermore, SAMHD1 preferentially cleaved single-stranded RNAs comprising A20 or U20, whereas neither C20 nor G20 was susceptible to SAMHD1-mediated degradation. Our findings will facilitate more advanced studies into the role of the SAMHD1 RNase function in the cellular pathogenesis implicated in nucleic acid-triggered inflammatory responses and the anti-retroviral function of SAMHD1. PMID:27387229

  12. Stability-indicating spectrophotometric methods for determination of the anticoagulant drug apixaban in the presence of its hydrolytic degradation product

    Tantawy, Mahmoud A.; El-Ragehy, Nariman A.; Hassan, Nagiba Y.; Abdelkawy, Mohamed

    2016-04-01

    Apixaban (a novel anticoagulant agent) was subjected to a stress stability study including acid, alkali, oxidative, photolytic, and thermal degradation. The drug was found to be only liable to acidic and alkaline hydrolysis. The degradation product was then isolated and identified by IR and GC-mass spectrometry. Four spectrophotometric methods, namely; first derivative (D1), derivative ratio (DR), ratio difference (RD) and mean centering of ratio spectra (MCR), have been suggested for the determination of apixaban in presence of its hydrolytic degradation product. The proposed methods do not require any preliminary separation step. The accuracy, precision and linearity ranges of the proposed methods were determined, and the methods were validated as per ICH guidelines and the specificity was assessed by analyzing synthetic mixtures containing different percentages of the degradation product with the drug. The developed methods were successfully applied for the determination of apixaban in bulk powder and its tablet dosage form.

  13. Bioglass dissolution: a comparison between SBF solution and hydrolytic etching

    The evaluation of chemical dissolution phenomena of the glasses in general is important because it is related to the bioactivity. This paper aims a comparative study of the bioglass dissolution between in vitro bioactivity test in SBF (Simulated Body Fluid) solution and the chemical durability test, in glasses on SiO2-Na2O-CaO with 6wt% de P2O5 system. The glasses were obtained by melting at 1500°C/2h and annealed at 500°C/2h followed by natural cooling. To in vitro test, the samples were immersed in SBF solution in different periods (1, 3, 7 and 14 days) at 7,25 pH and 37 deg C. For the hydrolytic resistance test were performed in a Soxhlet column. The physic-chemical and morphological characterization of the samples before and after both tests was realized through XRD, DRIFT and SEM. The samples presented a difference in kinetics dissolution for both tests that allow an improved understanding of the glasses bioactivity mechanism. (author)

  14. Hydrolytic catalysis and structural stabilization in a designed metalloprotein

    Zastrow, Melissa L.; Peacock, Anna F. A.; Stuckey, Jeanne A.; Pecoraro, Vincent L.

    2012-02-01

    Metal ions are an important part of many natural proteins, providing structural, catalytic and electron transfer functions. Reproducing these functions in a designed protein is the ultimate challenge to our understanding of them. Here, we present an artificial metallohydrolase, which has been shown by X-ray crystallography to contain two different metal ions—a Zn(II) ion, which is important for catalytic activity, and a Hg(II) ion, which provides structural stability. This metallohydrolase displays catalytic activity that compares well with several characteristic reactions of natural enzymes. It catalyses p-nitrophenyl acetate (pNPA) hydrolysis with an efficiency only ~100-fold less than that of human carbonic anhydrase (CA)II and at least 550-fold better than comparable synthetic complexes. Similarly, CO2 hydration occurs with an efficiency within ~500-fold of CAII. Although histidine residues in the absence of Zn(II) exhibit pNPA hydrolysis, miniscule apopeptide activity is observed for CO2 hydration. The kinetic and structural analysis of this first de novo designed hydrolytic metalloenzyme reveals necessary design features for future metalloenzymes containing one or more metals.

  15. High throughput screening of hydrolytic enzymes from termites using a natural substrate derived from sugarcane bagasse

    Lucena Severino A; Lima Leile S; Cordeiro Luís SA; Sant'Anna Celso; Constantino Reginaldo; Azambuja Patricia; Souza Wanderley de; Garcia Eloi S; Genta Fernando A

    2011-01-01

    Abstract Background The description of new hydrolytic enzymes is an important step in the development of techniques which use lignocellulosic materials as a starting point for fuel production. Sugarcane bagasse, which is subjected to pre-treatment, hydrolysis and fermentation for the production of ethanol in several test refineries, is the most promising source of raw material for the production of second generation renewable fuels in Brazil. One problem when screening hydrolytic activities i...

  16. High throughput screening of hydrolytic enzymes from termites using a natural substrate derived from sugarcane bagasse

    Lucena, Severino A; Lima, Leile S; Cordeiro, Luís SA; Sant'Anna, Celso; Constantino, Reginaldo; de Azambuja, Patricia; de Souza, Wanderley; Eloi S. Garcia; Fernando A Genta

    2011-01-01

    Background The description of new hydrolytic enzymes is an important step in the development of techniques which use lignocellulosic materials as a starting point for fuel production. Sugarcane bagasse, which is subjected to pre-treatment, hydrolysis and fermentation for the production of ethanol in several test refineries, is the most promising source of raw material for the production of second generation renewable fuels in Brazil. One problem when screening hydrolytic activities is that th...

  17. Treatment of anthraquinone dye wastewater by hydrolytic acidification-aerobic process

    YANG Jian; WU Min; Li Dan

    2004-01-01

    Experiment on microbial degradation with two kinds of biological process, hydrolytic acidification-aerobic process and aerobic process was conducted to treat the anthraquinone dye wastewater with CODCr concentration of 400 mg/L and chroma 800. The experimental result demonstrated that the hydrolytic-aerobic process could raise the biodegradability of anthraquinone dye wastewater effectively. The effluent CODCr can reach 120-170 mg/L and chroma 150 which is superior to that from simple aerobic process.

  18. Silicate Esters of Paclitaxel and Docetaxel: Synthesis, Hydrophobicity, Hydrolytic Stability, Cytotoxicity, and Prodrug Potential

    Wohl, Adam R.; Michel, Andrew R.; Kalscheuer, Stephen; Macosko, Christopher W.; Panyam, Jayanth; Hoye, Thomas R.

    2014-01-01

    We report here the synthesis and selected properties of various silicate ester derivatives (tetraalkoxysilanes) of the taxanes paclitaxel (PTX) and docetaxel (DTX) [i.e., PTX-OSi(OR)3 and DTX-OSi(OR)3]. Both the hydrophobicity and hydrolytic lability of these silicates can be (independently) controlled by choice of the alkyl group (R). The synthesis, structural characterization, hydrolytic reactivity, and in vitro cytotoxicity against the MDA-MB-231 breast cancer cell line of most of these de...

  19. MODIFICATION OF POLYAMIDE 6 WITH POLYAMINOAMIDE-g-POLY(ETHYLENE GLYCOL) VIA HYDROLYTIC POLYMERIZATION

    Yao-chi Liu; Wei Xu; Yuan-qin Xiong; Fan Zhang; Wei-jian Xu

    2009-01-01

    To enhance the impact strength of polyamide 6, hydrolytic polymerization modification by the polyaminoamide-g-poly(ethylene glycol) (PAAEG) derivatives with poly(ethylene glycol) (PEG) molecular weight of 400-10000 was studied. Amide groups of polyaminoamide segments were postulated to form hydrogen bonding with polyamide 6, and hydroxy groups of PAAEG units were expected to react with carboxylic acid groups of polyamide 6 forming copolymers during the polymerization. The improved compatibility in amorphous regions of blends has been confirmed by differential scanning calorimetry (DSC) and scanning electron microscopy (SEM) of fracture surfaces. The effects of PAAEG on the water absorption and notch sensitivity of blends were investigated, using water uptake measurement and mechanical testings, respectively. For comparison, pure polyamide 6 and the blend of PEG/polyamide 6 were also investigated. The addition of PAAEG retarded the crystallization of polyamide 6, but did not make remarkable influences on its crystalline structure. As a consequence of the strong interactions between the dispersed phases and polyamide 6 matrices, PAAEG was a more suitable additive for improving the notched impact strength of polyarnide 6 than PEG.

  20. The pulmonary extracellular lining.

    George, G; Hook, G E

    1984-01-01

    The extracellular lining of the lungs is reviewed. The pulmonary extracellular lining is a complex mixture of phospholipids, proteins and carbohydrates which is absolutely essential for the maintenance of normal pulmonary functions such as gas exchange. Without the lining the lungs would collapse. Alterations in the pulmonary extracellular lining may underlie some disease conditions induced by toxic agents, especially those which interfere with the formation of pulmonary surfactant. The extra...

  1. Hydrolytic and radiolytic degradation of TBP in TBP.30% V/V-dodecane/UO2(NO3)2.HNO3.H2O systems

    The hydrolytic and radiolytic degradation of TBP is investigated in systems of TBP 30% V/V-dodecane/H2O . HNO3 . UO2(NO3)2by gas chromatographic determination of HDBP. No direct relation between the concentration of HDBP formed and the quantity of HNO3 extracted by the organic phase is observed in the studies of hydrolysis of TBP. The HDBP concentration is seen to increase non-linearly with the concentration of HNO3 extracted by the organic phase. Radiolytic studies show that for doses greater than 1 Wh/l, the concentration of HDBP formed increases with the dose absorbed by the system. Whith doses smaller than 1 Wh/l and acid concentration greater than 2 M, two distinct patterns of behavior are observed. The concentration of HDBP as a function of the radiation dose absorbed by the system presents a minimum for uranyl nitrate concentrations smaller than 0.9 M; for uranyl nitrate concentrations greater than 1.3 M the concentration of radiolytic HDBP cannot be calculated because the concentration of the hydrolytic HDBP determined is greater than the sum of the experimental concentrations of hydrolytic and radiolytic HDBP. It is known that the dose absorbed by the process solutions during the reprocessing of light water reactor fuel elements is smaller than one Wh/l. Thus, dose rates between zero and one Wh/l should be studied for this system. (Author)

  2. Conditions of Mytilus edulis extracellular body fluids and shell composition in a pH-treatment experiment: Acid-base status, trace elements and δ11B

    Heinemann, Agnes; Fietzke, Jan; Melzner, Frank; BöHm, Florian; Thomsen, JöRn; Garbe-SchöNberg, Dieter; Eisenhauer, Anton

    2012-01-01

    Mytilus edulis were cultured for 3 months under six different seawater pCO2 levels ranging from 380 to 4000 μatm. Specimen were taken from Kiel Fjord (Western Baltic Sea, Germany) which is a habitat with high and variable seawater pCO2 and related shifts in carbonate system speciation (e.g., low pH and low CaCO3 saturation state). Hemolymph (HL) and extrapallial fluid (EPF) samples were analyzed for pH and total dissolved inorganic carbon (CT) to calculate pCO2 and [HCO3-]. A second experiment was conducted for 2 months with three different pCO2 levels (380, 1400 and 4000 μatm). Boron isotopes (δ11B) were investigated by LA-MC-ICP-MS (Laser Ablation-Multicollector-Inductively Coupled Plasma-Mass Spectrometry) in shell portions precipitated during experimental treatment time. Additionally, elemental ratios (B/Ca, Mg/Ca and Sr/Ca) in the EPF of specimen from the second experiment were measured via ICP-OES (Inductively Coupled Plasma-Optical Emission Spectrometry). Extracellular pH was not significantly different in HL and EPF but systematically lower than ambient water pH. This is due to high extracellular pCO2 values, a prerequisite for metabolic CO2 excretion. No accumulation of extracellular [HCO3-] was measured. Elemental ratios (B/Ca, Mg/Ca and Sr/Ca) in the EPF increased slightly with pH which is in accordance with increasing growth and calcification rates at higher seawater pH values. Boron isotope ratios were highly variable between different individuals but also within single shells. This corresponds to a high individual variability in fluid B/Ca ratios and may be due to high boron concentrations in the organic parts of the shell. The mean δ11B value shows no trend with pH but appears to represent internal pH (EPF) rather than ambient water pH.

  3. Comparative studies on the influence of some radioprotectors on the postirradiation hydrolytic function of subcellular fractions of pancreas

    Comparative studies on the influence of the two radioprotectors Fenchlorfos and cysteamine on the hydroproteolytic activity of pancreatic subcellular fractions of rats were carried out. The animals were irradiated with the single dose of 800 R of X-rays. Biochemical estimation of lipase, β-glucuronidase, acid phosphatase and catheptic (caseonolytic) activity were done 24 hours after exposition. The two radioprotective substances have a lot of common in their action against hydrolytic activity. They induce the decrease of some enzymes of subcellular fractions we checked both in control as well as in irradiated groups. The ambivalent influence of Trichlorfon on the lipase activity in granules and supernatant fraction seems to be of interest. (orig.)

  4. Non-hydrolytic Sol-gel Synthesis of Tin Sulfides

    Kaur, Rajvinder

    The non-hydrolytic sol-gel (NHSG) process is an effective low temperature route well known for preparing homogeneous metal oxides. Thermodynamically as well as kinetically favored products, which cannot be prepared with the traditional solid-state routes, can be produced using NHSG. This project is focused on the exploration of NHSG synthesis of binary tin sulfides. In the past few years, metal sulfides have been the subject of significant interest. Much effort has been devoted to understand these materials because of their potential applications in electronic, optical, and superconductor devices.4 Among these materials, tin sulfides are materials of technological importance, which are being explored as semiconductors, anode materials for Li ion batteries, photoconductors, photocatalysts and absorber layer materials in photovoltaic solar cell devices. All of these applications depend upon features like homogeneity, oxidation state, high surface area and purity of the materials. These properties can be difficult to achieve by employing traditional synthetic routes, which require high temperatures due to slow diffusion, limiting the products to thermodynamically stable phases and prohibiting control over properties like particle size and surface area. A variety of low temperature methods are being explored due to the increased demand for such advanced materials. This project is focused on exploring the NHSG approach to synthesize binary tin sulfides, with the main goal of establishing conditions for the targeted synthesis of different tin sulfide polymorphs with controlled particle size. Being non-oxide materials, tin sulfides can be air sensitive, which requires special attention in handling. All reactions were carried out in absence of oxygen. This project explores the reaction of tin halides with thioethers in a dry solvent medium, leading to the formation of tin sulfides. There are a number of synthetic parameters that can be varied for the NHSG approach. A

  5. Extracellular ionic locks determine variation in constitutive activity and ligand potency between species orthologs of the free fatty acid receptors FFA2 and FFA3

    Hudson, Brian D; Tikhonova, Irina G; Pandey, Sunil K;

    2012-01-01

    ) is non-selective. Although C2 was confirmed to be selective for human FFA2 over FFA3, this ligand was not selective between the mouse orthologs. Moreover, although C3 was indeed not selective between the human orthologs, it displayed clear selectivity for mouse FFA3 over mouse FFA2. This altered...... selectivity to C2 and C3 resulted from broad differences in SCFAs potency at the mouse orthologs. In studies to define the molecular basis for these observations, marked variation in ligand-independent constitutive activity was identified using a [(35)S]GTPγS assay. The orthologs with higher potency for the...... SCFAs, human FFA2 and mouse FFA3, displayed high constitutive activity in this assay, whereas the orthologs with lower potency for the agonist ligands, mouse FFA2 and human FFA3, did not. Sequence alignments of the second extracellular loop identified single negatively charged residues in FFA2 and FFA3...

  6. Alternative methods for characterization of extracellular vesicles

    Fatemeh eMomen-Heravi

    2012-09-01

    Full Text Available Extracellular vesicles are nano-sized vesicles released by all cells in vitro as well as in vivo. Their role has been implicated mainly in cell-cell communication, but also in disease biomarkers and more recently in gene delivery. They represent a snapshot of the cell status at the moment of release and carry bioreactive macromolecules such as nucleic acids, proteins and lipids. A major limitation in this emerging new field is the availability/awareness of techniques to isolate and properly characterize Extracellular vesicles. The lack of gold standards makes comparing different studies very difficult and may potentially hinder some Extracellular vesicles -specific evidence. Characterization of Extracellular vesicles has also recently seen many advances with the use of Nanoparticle Tracking Analysis (NTA, flow cytometry, cryo-EM instruments and proteomic technologies. In this review, we discuss the latest developments in translational technologies involving characterization methods including the facts in their support and the challenges they face.

  7. A biodegradation study of forest biomass by Aspergillus niger F7: correlation between enzymatic activity, hydrolytic percentage and biodegradation index

    Nivedita Sharma

    2012-06-01

    Full Text Available Aspergillus niger F7 isolated from soil was found to be the potent producer of cellulase and xylanase. The residue of forest species Toona ciliata, Celtris australis, Cedrus deodara and Pinus roxburghii was selected as substrate for biodegradation study due to its easy availability and wide use in industry. It was subjected to alkali (sodium hydroxide treatment for enhancing its degradation. Biodegradation of forest waste by hydrolytic enzymes (cellulase and xylanase secreted by A. niger under solid state fermentation (SSF was explored. SSF of pretreated forest biomass was found to be superior over untreated forest biomass. Highest extracellular enzyme activity of 2201±23.91 U/g by A. niger was shown in pretreated C. australis wood resulting in 6.72±0.20 percent hydrolysis and 6.99±0.23 biodegradation index (BI. The lowest BI of 1.40±0.08 was observed in untreated saw dust of C. deodara having the least enzyme activity of 238±1.36 U/g of dry matter. Biodegradation of forest biomass under SSF was increased many folds when moistening agent i.e. tap water had been replaced with modified basal salt media (BSM. In BSM mediated degradation of forest waste with A. niger, extracellular enzyme activity was increased up to 4089±67.11 U/g of dry matter in turn resulting in higher BI of 15.4±0.41 and percent hydrolysis of 19.38±0.81 in pretreated C. australis wood. A. niger exhibited higher enzyme activity on pretreated biomass when moistened with modified BSM in this study. Statistically a positive correlation has been drawn between these three factors i.e. enzyme activity, BI and percent hydrolysis of forest biomass thus proving their direct relationship with each other.

  8. A novel method to control hydrolytic degradation of nanocomposite biocompatible materials via imparting superhydrophobicity

    Khakbaz, Mobina; Hejazi, Iman; Seyfi, Javad; Jafari, Seyed-Hassan; Khonakdar, Hossein Ali; Davachi, Seyed Mohammad

    2015-12-01

    Acceleration of hydrolytic degradation of biomedical materials is not always desirable. For instance, terpolymers based on L-lactide, glycolide and trimethylene carbonate exhibit very fast hydrolytic degradation due to their amorphous structure, hydrophilicity, and high water absorption capability. Therefore, an attempt was made in the current study to impede the hydrolytic degradation for these materials through imparting superhydrophobicity to their surfaces. The used terpolymer has been shown to have promising potential applications as bio-absorbable surgical sutures and other biomedical materials, and thus, its applicability could be further extended upon impeding its hydrolytic degradation. Moreover, a novel method including combined use of non-solvent and nanoparticles was utilized to achieve superhydrophobicity. Very diverse wettability results were obtained which were attributed to the obtained various morphologies according to scanning electron microscopy results. More importantly, a unique hierarchical morphology was found to be responsible for the observed water repellent behavior. X-ray photoelectron spectroscopy results revealed co-existence of nanosilica particles and terpolymer chains on the surface's top layer. Finally, it was found that the superhydrophobic sample exhibited a significantly impeded hydrolytic degradation as compared with the hydrophilic pure terpolymer which was attributed to the formation of air pockets on the surface's top layer.

  9. Determination of the Proportion of Total Soil Extracellular Acid Phosphomonoesterase (E.C. 3.1.3.2 Activity Represented by Roots in the Soil of Different Forest Ecosystems

    Klement Rejsek

    2012-01-01

    Full Text Available The aim of this study is to present a new method for determining the root-derived extracellular acid phosphomonoesterase (EAPM activity fraction within the total EAPM activity of soil. EAPM activity was determined for roots, organic and mineral soil. Samples were collected using paired PVC cylinders, inserted to a depth of 15 cm, within seven selected forest stands. Root-derived EAPM formed between 4 and18% of the total EAPM activity of soil from forests of differing maturity. A new approach, presented in this work, enables separation of root-derived EAPM activity from total soil EAPM. Separation of root-derived EAPM from soil provides a better understanding of its role in P-cycling in terrestrial ecosystems. The method presented in this work is a first step towards the separation of root- and microbe-derived EAPM in soils, which are thought to possess different kinetic properties and different sensitivity to environmental change.

  10. Microbial respiration and kinetics of extracellular enzymes activities through rhizosphere and detritusphere at agricultural site

    Löppmann, Sebastian; Blagodatskaya, Evgenia; Kuzyakov, Yakov

    2014-05-01

    detritivore communities in the soil. The kinetics (Km and Vmax) of four extracellular hydrolytic enzymes responsible for C- and phosphorous-cycle (β-glucosidase, β-xylosidase, β-cellobiohydrolase and acid phosphatase), microbial biomass, basal respiration (BR) and substrate-induced respiration (SIR) were measured in rhizosphere, detritusphere and control from 0 - 10 and 10 - 20 cm. The metabolic quotient (qCO2) was calculated as specific indicator for efficiency of microbial substrate utilization. We observed clear differences in enzymes activities at low and high concentrations of substrate. At substrate saturation enzyme activity rates of were significantly higher in rooted plots compared to litter amended plots, whereas at lower concentration no treatment effect could be found. The BR, SIR and qCO2 values were significantly higher at 0 - 10 cm of the planted treatment compared to litter and control plots, revealing a significantly higher respiration at lower efficiency of microbial substrate utilization in the rhizosphere. The Michaelis-Menten constant (Km) decreased with depth, especially for β-glucosidase, acid phosphatase and β-xylosidase, indicating higher substrate affinity of microorganisms in deeper soil and therefore different enzyme systems functioning. The substrate affinity factor (Vmax/Km) increased 2-fold with depth for various enzymes, reflecting a switch of predominantly occurring microbial strategies. Vmax/Km ratio indicated relative domination of zymogenous microbial communities (r-strategists) in 0 - 10 cm depth as compared with 10 - 20 cm depth where the K-strategists dominated.

  11. PRODUCTION OF EXTRACELLULAR ENZYMES BY HALOPHILIC BACTERIA ISOLATED FROM SOLAR SALTERNS

    Jhuma Biswas

    2013-12-01

    Full Text Available During the course of survey of halophilic microorganisms, a total of sixteen bacterial isolates were obtained from coastal solar salterns of Orissa and West Bengal, India. Morphological, physiological and biochemical characteristics of these isolates indicate that majority of them belong to the genus Halomonas, however, members belonging to Cobetia and Halococcus were not uncommon. These isolates were screened for the production of extracellular enzymes such as amylase, glutaminase, asparaginase, xylanase, cellulase, gelatinase, inulinase, caseinase, pectinase, urease and lipase. Among these hydrolytic enzymes, glutamine and asparagine hydrolytic activities were predominant, although lipid and casein degrading activities were not inferior. However, amylase and gelatinase production were rare. None of these halophiles was able to degrade cellulose, inulin, pectin and xylan and only one isolate was capable of hydrolyzing urea

  12. AN INTEGRATED, ANIMATED MODEL OF THE (NA, K-ATPase HYDROLYTIC CYCLE

    F.A. Leone

    2006-07-01

    Full Text Available The  (Na,  K-ATPase,  or  sodium  pump,  is  the  principal,  active  transport  system  that  establishes  sodium  and potassium  gradients  across  the  plasma  membranes  of  all  animal  cells.  Such  gradients  are  critical  to  sustaining important cellular functions like osmotic equilibrium, cell volume and pH homeostasis, among many others (Ann Rev Physiol 65: 817, 2003; Physiol 19: 377, 2004. This transport protein is a heterodimer that consists of a 110-kDa  -subunit  and  a  55-kDa,  glycosylated  -subunit.  A  group  of  seven  small  proteins,  known  as  FXYD  proteins  from  the sequence  of  a  conserved  motif  has  been  identified  recently,  and  one  of  these,  FXYD2,  constitutes  the  (Na,  K-ATPase  -subunit.  Our  model  is  based  on  conformational  changes  occurring  between  the  E1  and  E2  forms  of  the enzyme, which initiates its hydrolytic cycle at a high ATP/ADP ratio. While all steps are reversible, the model does not include  the reverse  reactions that can  take  place under appropriate conditions. The  E1 state  corresponds to that of the SERCA, recently crystallized (Science 304; 1672, 2004; Nature 430: 529, 2004. The animation was developed in Macromedia  Flash  8.0® and  illustrates  the  principle  of  an  alternating-access  model  of  an  ion  pump.  The  protein  is embedded  in  the  membrane  with  the  extracellular  face  uppermost  and  the  cytoplasmic  face  at  the  bottom.  Access from  the  cytoplasmic  or  extracellular  faces  to  the  cation-binding  sites,  located  in  the  transmembrane  moiety,  are controlled  by  two  gates  (moving  horizontal  bars,  and  conformations  showing  the  two  gates  closed  correspond  to states with occluded Na+ and K+ sites. Changes in cation-binding site structure entail

  13. A review on hydrolytic enzymes in the treatment of wastewater with high oil and grease content.

    Cammarota, M C; Freire, D M G

    2006-11-01

    Wastewater from dairies and slaughterhouses contains high levels of fats and proteins that present low biodegradability. A large number of pretreatment systems are employed to remove oil and grease (O&G) to prevent a host of problems that may otherwise arise in the biological process, and reduce the efficiency of the treatment station. Problems caused by excessive O&G include a reduction in the cell-aqueous phase transfer rates, a sedimentation hindrance due to the development of filamentous microorganisms, development and flotation of sludge with poor activity, clogging and the emergence of unpleasant odors. Therefore the application of a pretreatment to hydrolyze and dissolve lipids may improve the biological degradation of fatty wastewaters, accelerating the process and improving time efficiency. However thus far, only a few studies describing the degradation of fats and oils by alkaline/acid/enzymatic hydrolysis have been reported; the treatment of effluents from several origins is a new and promising application for lipases. Among the strains that produce the hydrolytic enzymes studied, the fungus Penicillium restrictum is a particularly promising one. When cultivated in low-cost solid medium composed of agro-industrial waste, P. restrictum produces a pool of hydrolases capable of degrading the most complex organic compounds. This degradation enables a considerable increase in organic matter removal efficiency to be realized, which results in the attainment of a high-quality effluent in the subsequent biological treatment stage. Consequently, there is presently a wide variety of ongoing scientific investigation in the field of developing enzymatic hydrolysis processes to precede traditional biological treatment. PMID:16621527

  14. A review on hydrolytic enzymes in the treatment of wastewater with high oil and grease content

    Cammarota, M.C. [Federal University of Rio de Janeiro (Brazil). School of Chemistry; Freire, D.M.G. [Federal University of Rio de Janeiro (Brazil). Institute of Chemistry

    2006-11-15

    Wastewater from dairies and slaughterhouses contains high levels of fats and proteins that present low biodegradability. A large number of pretreatment systems are employed to remove oil and grease (OandG) to prevent a host of problems that may otherwise arise in the biological process, and reduce the efficiency of the treatment station. Problems caused by excessive OandG include a reduction in the cell-aqueous phase transfer rates, a sedimentation hindrance due to the development of filamentous microorganisms, development and flotation of sludge with poor activity, clogging and the emergence of unpleasant odors. Therefore the application of a pretreatment to hydrolyze and dissolve lipids may improve the biological degradation of fatty wastewaters, accelerating the process and improving time efficiency. However thus far, only a few studies describing the degradation of fats and oils by alkaline/acid/enzymatic hydrolysis have been reported; the treatment of effluents from several origins is a new and promising application for lipases. Among the strains that produce the hydrolytic enzymes studied, the fungus Penicillium restrictum is a particularly promising one. When cultivated in low-cost solid medium composed of agro-industrial waste, P. restrictum produces a pool of hydrolases capable of degrading the most complex organic compounds. This degradation enables a considerable increase in organic matter removal efficiency to be realized, which results in the attainment of a high-quality effluent in the subsequent biological treatment stage. Consequently, there is presently a wide variety of ongoing scientific investigation in the field of developing enzymatic hydrolysis processes to precede traditional biological treatment. (author)

  15. Interesterification of butter fat by partially purified extracellular lipases from Pseudomonas putida, Aspergillus niger and Rhizopus oryzae.

    Pabai, F; Kermasha, S; Morin, A

    1995-11-01

    Three extracellular lipases were produced by batch fermentation of Pseudomonas putida ATCC 795, Aspergillus niger CBS 131.52 and Rhizopus oryzae ATCC 34612 during the late phase of growth, at 72, 96 and 96 h, respectively. The lipases were partially purified by (NH4)2SO4 fractionation. The lipase of P. putida was optimal at pH 8.0 whereas those from A. niger and R. oryzae were optimal at pH 7.5. The A. niger lipase had the lowest V max value (0.51×10(-3) U/min) and R. oryzae the highest (1.86×10(-3) U/min). The K m values for P. putida, A. niger and R. oryzae lipases were 1.18, 0.97, and 0.98 mg/ml, respectively. Native PAGE of the partially-purified lipase extracts showed two to four major bands. The interesterification of butter fat by A. niger lipase decreased the water activity as well as the hydrolytic activity. The A. niger lipase had the highest interesterification yield value (26%) and the R. oryzae lipase the lowest (4%). In addition, A. niger lipase exhibited the highest decrease (17%) in long-chain hypercholesterolemic fatty acids (C12:0, C14:0 and C16:0) at the sn-2-position; the P. putida lipase demonstrated the least favourable changes in specificity at the same position. PMID:24415019

  16. The effects of extracellular pH and hydroxycinnamic acids influence the intracellular pH of Brettanomyces bruxellensis DSM 7001

    Campolongo, Simona; Siegumfeldt, Henrik; Aabo, Thomas Ask;

    2014-01-01

    Brettanomyces bruxellensis yeast produces ethyl phenols, which cause wine spoilage and severe economic losses. Ethyl phenols are produced by a class of polyphenols that are present in grapes and musts, called hydrocinnamic acids, which are capable of inhibiting yeasts and bacteria. The viability...

  17. γ-Aminobutyric Acid Type A (GABAA) Receptor Subunits Play a Direct Structural Role in Synaptic Contact Formation via Their N-terminal Extracellular Domains.

    Brown, Laura E; Nicholson, Martin W; Arama, Jessica E; Mercer, Audrey; Thomson, Alex M; Jovanovic, Jasmina N

    2016-07-01

    The establishment of cell-cell contacts between presynaptic GABAergic neurons and their postsynaptic targets initiates the process of GABAergic synapse formation. GABAA receptors (GABAARs), the main postsynaptic receptors for GABA, have been recently demonstrated to act as synaptogenic proteins that can single-handedly induce the formation and functional maturation of inhibitory synapses. To establish how the subunit composition of GABAARs influences their ability to induce synaptogenesis, a co-culture model system incorporating GABAergic medium spiny neurons and the HEK293 cells, stably expressing different combinations of receptor subunits, was developed. Analyses of HEK293 cell innervation by medium spiny neuron axons using immunocytochemistry, activity-dependent labeling, and electrophysiology have indicated that the γ2 subunit is required for the formation of active synapses and that its effects are influenced by the type of α/β subunits incorporated into the functional receptor. To further characterize this process, the large N-terminal extracellular domains (ECDs) of α1, α2, β2, and γ2 subunits were purified using the baculovirus/Sf9 cell system. When these proteins were applied to the co-cultures of MSNs and α1/β2/γ2-expressing HEK293 cells, the α1, β2, or γ2 ECD each caused a significant reduction in contact formation, in contrast to the α2 ECD, which had no effect. Together, our experiments indicate that the structural role of GABAARs in synaptic contact formation is determined by their subunit composition, with the N-terminal ECDs of each of the subunits directly participating in interactions between the presynaptic and postsynaptic elements, suggesting the these interactions are multivalent and specific. PMID:27129275

  18. Extracellular calcium sensing and extracellular calcium signaling

    Brown, E. M.; MacLeod, R. J.; O'Malley, B. W. (Principal Investigator)

    2001-01-01

    The cloning of a G protein-coupled extracellular Ca(2+) (Ca(o)(2+))-sensing receptor (CaR) has elucidated the molecular basis for many of the previously recognized effects of Ca(o)(2+) on tissues that maintain systemic Ca(o)(2+) homeostasis, especially parathyroid chief cells and several cells in the kidney. The availability of the cloned CaR enabled the development of DNA and antibody probes for identifying the CaR's mRNA and protein, respectively, within these and other tissues. It also permitted the identification of human diseases resulting from inactivating or activating mutations of the CaR gene and the subsequent generation of mice with targeted disruption of the CaR gene. The characteristic alterations in parathyroid and renal function in these patients and in the mice with "knockout" of the CaR gene have provided valuable information on the CaR's physiological roles in these tissues participating in mineral ion homeostasis. Nevertheless, relatively little is known about how the CaR regulates other tissues involved in systemic Ca(o)(2+) homeostasis, particularly bone and intestine. Moreover, there is evidence that additional Ca(o)(2+) sensors may exist in bone cells that mediate some or even all of the known effects of Ca(o)(2+) on these cells. Even more remains to be learned about the CaR's function in the rapidly growing list of cells that express it but are uninvolved in systemic Ca(o)(2+) metabolism. Available data suggest that the receptor serves numerous roles outside of systemic mineral ion homeostasis, ranging from the regulation of hormonal secretion and the activities of various ion channels to the longer term control of gene expression, programmed cell death (apoptosis), and cellular proliferation. In some cases, the CaR on these "nonhomeostatic" cells responds to local changes in Ca(o)(2+) taking place within compartments of the extracellular fluid (ECF) that communicate with the outside environment (e.g., the gastrointestinal tract). In others

  19. Extracellular hemicellulolytic enzymes from the maize endophyte Acremonium zeae.

    Bischoff, Kenneth M; Wicklow, Donald T; Jordan, Douglas B; de Rezende, Sebastiao T; Liu, Siqing; Hughes, Stephen R; Rich, Joseph O

    2009-05-01

    Microorganisms that colonize plants require a number of hydrolytic enzymes to help degrade the cell wall. The maize endophyte Acremonium zeae was surveyed for production of extracellular enzymes that hydrolyze cellulose and hemicellulose. The most prominent enzyme activity in cell-free culture medium from A. zeae NRRL 6415 was xylanase, with a specific activity of 60 U/mg from cultures grown on crude corn fiber. Zymogram analysis following SDS-PAGE indicated six functional xylanase polypeptides of the following masses: 51, 44, 34, 29, 23, and 20 kDa. Xylosidase (0.39 U/mg), arabinofuranosidase (1.2 U/mg), endoglucanase (2.3 U/mg), cellobiohydrolase (1.3 U/mg), and beta-glucosidase (0.85 U/mg) activities were also detected. Although apparently possessing a full complement of hemicellulolytic activities, cell-free culture supernatants prepared from A. zeae required an exogenously added xylosidase to release more than 90% of the xylose and 80% of the arabinose from corn cob and wheat arabinoxylans. The hydrolytic enzymes from A. zeae may be suitable for application in the bioconversion of lignocellulosic biomass into fermentable sugars. PMID:19184610

  20. Identification of hydrolytic and isomeric N-oxide degradants of vilazodone by on line LC-ESI-MS/MS and APCI-MS.

    Kalariya, Pradipbhai D; Talluri, M V N Kumar; Patel, Prinesh N; Srinivas, R

    2015-01-01

    The present study reports the degradation behavior of a new antidepressant drug, vilazodone, under various stress conditions as per International Conference on Harmonization guidelines (ICH, Q1A(R2). The investigation involved monitoring decomposition of the drug under hydrolytic (acidic, basic and neutral), oxidative, photolytic and thermal stress conditions and identifying degradation products. A rapid, precise, accurate and robust ultra high performance liquid chromatography (UPLC) method has been developed on a Waters CSH Phenyl-Hexyl column (100 mm × 2.1 mm, 1.7 μm) using gradient elution of 10mM ammonium acetate buffer (pH 5.0) and acetonitrile as mobile phase. The drug was found to be degraded in hydrolytic (acidic and basic) and oxidative conditions, whereas it was stable under neutral hydrolytic, photolytic and thermal stress conditions. The method was extended to quadrupole time-of-flight mass spectrometry (QTOF-MS) for the structural characterization of degradation products. It has been observed that isomeric N-oxide degradation products were formed under oxidative stress condition. The exact location of N-oxidation in the drug was investigated using atmospheric pressure chemical ionization (APCI) due to the formation of characteristic fragment ions. These fragment ions resulted from Meisenheimer rearrangement owing to thermal energy activation at the vaporizer of APCI source. All degradation products were comprehensively characterized by UPLC-ESI-MS/MS and UPLC-APCI-MS experiments. The most probable mechanisms for the formation of degradation products have also been proposed. The method was validated in terms of specificity, linearity, accuracy, precision, and robustness as per ICH guidelines. PMID:25459935

  1. All-trans retinoic acid regulates the expression of the extracellular matrix protein fibulin-1 in the guinea pig sclera and human scleral fibroblasts

    Li, Chuanxu; McFadden, Sally A.; Morgan, Ian; Cui, Dongmei; Hu, Jianmin; Wan, Wenjuan; Zeng, Junwen

    2010-01-01

    Purpose Fibulin-1 (FBLN1) mRNA is expressed in human sclera and is an important adhesion modulatory protein that can affect cell–matrix interactions and tissue remodeling. Scleral remodeling is influenced by all-trans retinoic acid (RA). Our purpose was to confirm the presence of fibulin-1 protein in guinea pig sclera and investigate the effect of RA on the expression of fibulin-1 in guinea pig sclera in vivo and in cultured human scleral fibroblasts (HSFs). Methods Confocal fluorescence micr...

  2. Mesoporous titanosilicates by templated non-hydrolytic sol–gel reactions

    Škoda, D.; Stýskalík, A.; Moravec, Z.; Bezdička, Petr; Barnes, C.E.; Pinkas, J.

    2015-01-01

    Roč. 74, č. 3 (2015), s. 810-822. ISSN 0928-0707 Institutional support: RVO:61388980 Keywords : Acetamide elimination * Catalysis * Epoxidation * Mesoporous * Non-hydrolytic sol-gel * Templating Subject RIV: CA - Inorganic Chemistry Impact factor: 1.532, year: 2014

  3. Persistence of pyrazosulfuron-ethyl and halosulfuron-methyl in aqueous solutions: Comparing hydrolytic dissipation

    Pyrazosulfuron-ethyl and halosulfuron-methyl are two new highly active sulfonylurea herbicides that have been widely used for weed control in many crops. Chemical hydrolysis is a primary process to determine the environmental fates of this group of pesticides. The hydrolytic dissipation of two herbi...

  4. Hydrolytic stability of polycarbonate-based polyurethane elastomers tested in physiologically simulated conditions

    Serkis, Magdalena; Špírková, Milena; Poreba, Rafal; Hodan, Jiří; Kredatusová, Jana; Kubies, Dana

    2015-01-01

    Roč. 119, September (2015), s. 23-34. ISSN 0141-3910 R&D Projects: GA ČR(CZ) GA13-06700S Institutional support: RVO:61389013 Keywords : polyurethane * elastomer * hydrolytic stability Subject RIV: CD - Macromolecular Chemistry Impact factor: 3.163, year: 2014

  5. Xyloglucan endotransglycosylases (XETS) from nasturtium seeds do not posses hydrolytic activity

    Stratilová, E.; Ait-Mohand, F.; Řehulka, Pavel; Garajová, S.; Flodrová, D.; Řehulková, Helena; Farkaš, V.

    Bratislava, 2008. s. 72. [Bratislava Symposium on Saccharides /11./. 21.09.2008-26.09.2008, Smolenice] R&D Projects: GA MŠk 1M06030 Institutional research plan: CEZ:AV0Z40310501 Keywords : xyloglucan endotransglycosylase * hydrolytic activity Subject RIV: CB - Analytical Chemistry, Separation

  6. The effect of D,L-lactide-based linker on the hydrolytic stability of polyurethane films

    Špírková, Milena; Serkis, Magdalena; Poreba, Rafal; Kredatusová, Jana; Machová, Luďka; Hodan, Jiří

    Belgrade: Materials Research Society of Serbia , 2015 - (Uskoković, D.; Radmilović, V.). s. 15 [Annual Conference YUCOMAT 2015 /17./. 31.08.2015-04.09.2015, Herceg Novi] R&D Projects: GA ČR(CZ) GA13-06700S Institutional support: RVO:61389013 Keywords : polyurethanes * hydrolytic stability * surface analysis Subject RIV: CD - Macromolecular Chemistry

  7. Single amino acid insertions in extracellular loop 2 of Bombyx mori ABCC2 disrupt its receptor function for Bacillus thuringiensis Cry1Ab and Cry1Ac but not Cry1Aa toxins.

    Tanaka, Shiho; Miyamoto, Kazuhisa; Noda, Hiroaki; Endo, Haruka; Kikuta, Shingo; Sato, Ryoichi

    2016-04-01

    In a previous report, seven Cry1Ab-resistant strains were identified in the silkworm, Bombyx mori; these strains were shown to have a tyrosine insertion at position 234 in extracellular loop 2 of the ABC transporter C2 (BmABCC2). This insertion was confirmed to destroy the receptor function of BmABCC2 and confer the strains resistance against Cry1Ab and Cry1Ac. However, these strains were susceptible to Cry1Aa. In this report, we examined the mechanisms of the loss of receptor function of the transporter by expressing mutations in Sf9 cells. After replacement of one or two of the five amino acid residues in loop 2 of the susceptible BmABCC2 gene [BmABCC2_S] with alanine, cells still showed susceptibility, retaining the receptor function. Five mutants with single amino acid insertions at position 234 in BmABCC2 were also generated, resulting in loop 2 having six amino acids, which corresponds to replacing the tyrosine insertion in the resistant BmABCC2 gene [BmABCC2_R(+(234)Y)] with another amino acid. All five mutants exhibited loss of function against Cry1Ab and Cry1Ac. These results suggest that the amino acid sequence in loop 2 is less important than the loop size (five vs. six amino acids) or loop structure for Cry1Ab and Cry1Ac activity. Several domain-swapped mutant toxins were then generated among Cry1Aa, Cry1Ab, and Cry1Ac, which are composed of three domains. Swapped mutants containing domain II of Cry1Ab or Cry1Ac did not kill Sf9 cells expressing BmABCC2_R(+(234)Y), suggesting that domain II of the Cry toxin is related to the interaction with the receptor function of BmABCC2. This also suggests that different reactions against Bt-toxins in some B. mori strains, that is, Cry1Ab resistance or Cry1Aa susceptibility, are attributable to structural differences in domain II of Cry1A toxins. PMID:26928903

  8. Activity of selected hydrolytic enzymes in Allium sativum L. anthers.

    Winiarczyk, Krystyna; Gębura, Joanna

    2016-05-01

    The aim of the study was to determine enzymatic activity in sterile Allium sativum anthers in the final stages of male gametophyte development (the stages of tetrads and free microspores). The analysed enzymes were shown to occur in the form of numerous isoforms. In the tetrad stage, esterase activity was predominant, which was manifested by the greater number of isoforms of the enzyme. In turn, in the microspore stage, higher numbers of isoforms of acid phosphatases and proteases were detected. The development of sterile pollen grains in garlic is associated with a high level of protease and acid phosphatase activity and lower level of esterase activities in the anther locule. Probably this is the first description of the enzymes activity (ACPH, EST, PRO) in the consecutives stages of cell wall formation which is considered to be one of the causes of male sterility in flowering plant. PMID:26901781

  9. Porous Materials for Hydrolytic Dehydrogenation of Ammonia Borane

    Tetsuo Umegaki; Qiang Xu; Yoshiyuki Kojima

    2015-01-01

    Hydrogen storage is still one of the most significant issues hindering the development of a “hydrogen energy economy”. Ammonia borane is notable for its high hydrogen densities. For the material, one of the main challenges is to release efficiently the maximum amount of the stored hydrogen. Hydrolysis reaction is a promising process by which hydrogen can be easily generated from this compound. High purity hydrogen from this compound can be evolved in the presence of solid acid or metal based ...

  10. Cultural conditions on the production of extracellular enzymes by Trichoderma isolates from tobacco rhizosphere

    K.L.N Mallikharjuna Rao

    2016-03-01

    Full Text Available Abstract Twelve isolates of Trichoderma spp. isolated from tobacco rhizosphere were evaluated for their ability to produce chitinase and β-1,3-glucanase extracellular hydrolytic enzymes. Isolates ThJt1 and TvHt2, out of 12 isolates, produced maximum activities of chitinase and β-1,3-glucanase, respectively. In vitro production of chitinase and β-1,3-glucanase by isolates ThJt1 and TvHt2 was tested under different cultural conditions. The enzyme activities were significantly influenced by acidic pH and the optimum temperature was 30 °C. The chitin and cell walls of Sclerotium rolfsii, as carbon sources, supported the maximum and significantly higher chitinase activity by both isolates. The chitinase activity of isolate ThJt1 was suppressed significantly by fructose (80.28%, followed by glucose (77.42%, whereas the β-1,3-glucanase activity of ThJt1 and both enzymes of isolate TvHt2 were significantly suppressed by fructose, followed by sucrose. Ammonium nitrate as nitrogen source supported the maximum activity of chitinase in both isolates, whereas urea was a poor nitrogen source. Production of both enzymes by the isolates was significantly influenced by the cultural conditions. Thus, the isolates ThJt1 and TvHt2 showed higher levels of chitinase and β-1,3-glucanase activities and were capable of hydrolyzing the mycelium of S. rolfsii infecting tobacco. These organisms can be used therefore for assessment of their synergism in biomass production and biocontrol efficacy and for their field biocontrol ability against S. rolfsii and Pythium aphanidermatum infecting tobacco.

  11. Cultural conditions on the production of extracellular enzymes by Trichoderma isolates from tobacco rhizosphere.

    Mallikharjuna Rao, K L N; Siva Raju, K; Ravisankar, H

    2016-01-01

    Twelve isolates of Trichoderma spp. isolated from tobacco rhizosphere were evaluated for their ability to produce chitinase and β-1,3-glucanase extracellular hydrolytic enzymes. Isolates ThJt1 and TvHt2, out of 12 isolates, produced maximum activities of chitinase and β-1,3-glucanase, respectively. In vitro production of chitinase and β-1,3-glucanase by isolates ThJt1 and TvHt2 was tested under different cultural conditions. The enzyme activities were significantly influenced by acidic pH and the optimum temperature was 30°C. The chitin and cell walls of Sclerotium rolfsii, as carbon sources, supported the maximum and significantly higher chitinase activity by both isolates. The chitinase activity of isolate ThJt1 was suppressed significantly by fructose (80.28%), followed by glucose (77.42%), whereas the β-1,3-glucanase activity of ThJt1 and both enzymes of isolate TvHt2 were significantly suppressed by fructose, followed by sucrose. Ammonium nitrate as nitrogen source supported the maximum activity of chitinase in both isolates, whereas urea was a poor nitrogen source. Production of both enzymes by the isolates was significantly influenced by the cultural conditions. Thus, the isolates ThJt1 and TvHt2 showed higher levels of chitinase and β-1,3-glucanase activities and were capable of hydrolyzing the mycelium of S. rolfsii infecting tobacco. These organisms can be used therefore for assessment of their synergism in biomass production and biocontrol efficacy and for their field biocontrol ability against S. rolfsii and Pythium aphanidermatum infecting tobacco. PMID:26887223

  12. Extracellular Vesicle (EV) Array

    Jørgensen, Malene; Bæk, Rikke; Pedersen, Shona;

    2013-01-01

    Exosomes are one of the several types of cell-derived vesicles with a diameter of 30-100 nm. These extracellular vesicles are recognized as potential markers of human diseases such as cancer. However, their use in diagnostic tests requires an objective and high-throughput method to define...

  13. Bacterial extracellular lignin peroxidase

    Crawford, Donald L.; Ramachandra, Muralidhara

    1993-01-01

    A newly discovered lignin peroxidase enzyme is provided. The enzyme is obtained from a bacterial source and is capable of degrading the lignin portion of lignocellulose in the presence of hydrogen peroxide. The enzyme is extracellular, oxidative, inducible by lignin, larch wood xylan, or related substrates and capable of attacking certain lignin substructure chemical bonds that are not degradable by fungal lignin peroxidases.

  14. Extracellular Acidification Alters Lysosomal Trafficking in Human Breast Cancer Cells1

    Glunde, Kristine; Sandra E. Guggino; Solaiyappan, Meiyappan; Pathak, Arvind P.; Ichikawa, Yoshitaka; Bhujwalla, Zaver M.

    2003-01-01

    Cancer cells invade by secreting degradative enzymes, which are sequestered in lysosomal vesicles. In this study, the impact of an acidic extracellular environment on lysosome size, number, and distance from the nucleus in human mammary epithelial cells (HMECs) and breast cancer cells of different degrees of malignancy was characterized because the physiological microenvironment of tumors is frequently characterized by extracellular acidity. An acidic extracellular pH (pHe) resulted in a dist...

  15. Influence of pH on Morphology and Structure during Hydrolytic Degradation of the Segmented GL-b-[GL-co-TMC-co-CL]-b-GL Copolymer

    Yolanda Márquez

    2015-09-01

    Full Text Available Hydrolytic degradation in media having a continuous variation of pH from 2 to 12 was studied for a copolymer having two polyglycolide hard blocks and a middle soft segment constituted by glycolide, trimethylene carbonate, and ɛ-caprolactone units. The last units were susceptible to cross-linking reactions by γ irradiation that led to an increase of the molecular weight of the sample. Nevertheless, the susceptibility to hydrolytic degradation was enhanced with respect to non-irradiated samples and consequently such samples were selected to analyze the degradation process through weight loss measurements and the evaluation of changes on molecular weight, morphology, and SAXS patterns. Results reflected the different hydrolytic mechanisms that took place in acid and basic media and the different solubilization of the degradation products. Thus, degradation was faster and solubilization higher in the basic media. In this case, fibers showed a high surface erosion and the formation of both longitudinal and deep circumferential cracks that contrasted with the peeling process detected at intermediate pHs (from 6 to 8 and the absence of longitudinal cracks at low pHs. SAXS measurements indicated that degradation was initiated through the hydrolysis of the irregular molecular folds placed on the amorphous interlamellar domains but also affected lamellar crystals at the last stages. Subsequent heating processes performed with degraded samples were fundamental to reveal the changes in microstructure that occurred during degradation and even the initial lamellar arrangement. In particular, the presence of interfibrillar domains and the disposition of lamellar domains at different levels along the fiber axis for a determined cross-section were evidenced.

  16. Hydrolytic charge-reversal of PEGylated polyplexes enhances intracellular un-packaging and activity of siRNA.

    Werfel, Thomas A; Swain, Corban; Nelson, Christopher E; Kilchrist, Kameron V; Evans, Brian C; Miteva, Martina; Duvall, Craig L

    2016-04-01

    Hydrolytically degrading nano-polyplexes (HDG-NPs) that reverse charge through conversion of tertiary amines to carboxylic acids were investigated to improve intracellular un-packaging of siRNA and target gene silencing compared to a non-degradable analog (non-HDG-NPs). Both NP types comprised reversible addition-fragmentation chain-transfer (RAFT) synthesized diblock copolymers of a poly(ethylene glycol) (PEG) corona-forming block and a cationic block for nucleic acid packaging that incorporated butyl methacrylate (BMA) and either dimethylaminoethyl methacrylate (DMAEMA, non-HDG-NPs) or dimethylaminoethyl acrylate (DMAEA, HDG-NPs). HDG-NPs decreased significantly in size and released significantly more siRNA (∼40%) than non-HDG-NPs after 24 h in aqueous solution. While both HDG-NPs and non-HDG-NPs had comparable uptake and cytotoxicity up to 150 nM siRNA doses, HDG-NPs achieved significantly higher target gene silencing of the model gene luciferase in vitro. High resolution FRET confocal microscopy was used to monitor the intracellular un-packaging of siRNA. Non-HDG-NPs had significantly higher FRET efficiency than HDG-NPs, indicating that siRNA delivered from HDG-NPs was more fully un-packaged and therefore had improved intracellular bioavailability. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 917-927, 2016. PMID:26691570

  17. Fulvic acid promotes extracellular anti-cancer mediators from RAW 264.7 cells, causing to cancer cell death in vitro.

    Jayasooriya, Rajapaksha Gedara Prasad Tharanga; Dilshara, Matharage Gayani; Kang, Chang-Hee; Lee, Seungheon; Choi, Yung Hyun; Jeong, Yong Kee; Kim, Gi-Young

    2016-07-01

    Fulvic acid (FA) is known to promote electrochemical balance as a donor or a receptor possessing many biomedical functions. Nevertheless, the effect of FA on the anti-cancer activity has not been elucidated. In the current study, we first isolated FA from humus and investigated whether FA regulates immune-stimulating functions, such as production of nitric oxide (NO), in RAW 264.7 cells. Our data showed that FA slightly enhances cell viability in a dose-dependent manner and secretion of NO from RAW 264.7 cells. It upregulated the protein and mRNA expression of inducible NO synthesis (iNOS). In addition, FA enhanced the DNA-binding activity of nuclear factor-κB (NF-κB) in RAW 264.7 cells; the NF-κB inhibitor, pyrrolidine dithiocarbamate (PDTC) effectively attenuated the expression of FA-stimulated iNOS, suggesting that FA stimulates NF-κB to promote iNOS and NO production. Finally, FA-stimulated culture media (FA-CM) from RAW 264.7 cells were collected and MCA-102 fibrosarcoma cells were cultured in this media. The FA-CM augmented MCA-102 fibrosarcoma cell apoptosis; however, an NO inhibitor N(G)-monomethyl-l-arginine (NMMA) slightly inhibited the FA-CM-mediated MCA-102 fibrosarcoma cell apoptosis, which was accompanied by low levels of NO. In the present study, we found that FA induces the generation of NO and iNOS in RAW 264.7 cells by inducing NF-κB activation; however, NO did not significantly stimulate MCA-102 fibrosarcoma cell apoptosis in the current study. In addition, FA-CM enhanced cell death in various human cancer cells such as Hep3B, LNCaP, and HL60. Taken together, FA most likely stimulates immune-modulating molecules such as NO and induces cancer cell apoptosis. PMID:27177083

  18. All-trans retinoic acid regulates the expression of the extracellular matrix protein fibulin-1 in the guinea pig sclera and human scleral fibroblasts

    Li, Chuanxu; McFadden, Sally A.; Morgan, Ian; Cui, Dongmei; Hu, Jianmin; Wan, Wenjuan

    2010-01-01

    Purpose Fibulin-1 (FBLN1) mRNA is expressed in human sclera and is an important adhesion modulatory protein that can affect cell–matrix interactions and tissue remodeling. Scleral remodeling is influenced by all-trans retinoic acid (RA). Our purpose was to confirm the presence of fibulin-1 protein in guinea pig sclera and investigate the effect of RA on the expression of fibulin-1 in guinea pig sclera in vivo and in cultured human scleral fibroblasts (HSFs). Methods Confocal fluorescence microscopy was used to study fibulin-1 and aggrecan expression and localization in sclera from control guinea pigs and in animals given RA by daily gavage from 4 to 8 days of age. The effects of RA (from 10−9 to 10−5 M) on fibulin-1 expression in HSFs were observed by immunohistochemistry and assayed by real-time PCR and western blot analysis. Results Fibulin-1 protein expression was detected by confocal fluorescence microscopy in guinea pig sclera and in cultured HSFs. Upregulation of fibulin-1 in scleral tissue was observed after feeding with RA. In vitro, the level of Fbln1 mRNA was increased after treatment of HSFs with RA (at concentrations of 10−8 to 10−6 M; p<0.001), with a maximum effect at 10−7 M. Fibulin-1 protein levels were significantly increased after treatment of HSFs with 10−7 M of RA for 24 or 48 h (p<0.05). Conclusions Fibulin-1 protein was expressed in guinea pig sclera and cultured HSFs. Expression was regulated by RA, a molecule known to be involved in the regulation of eye growth. Further studies on the role of fibulin-1 in the regulation of eye growth, including during the development of myopia, are therefore warranted. PMID:20405022

  19. Porous Materials for Hydrolytic Dehydrogenation of Ammonia Borane

    Tetsuo Umegaki

    2015-07-01

    Full Text Available Hydrogen storage is still one of the most significant issues hindering the development of a “hydrogen energy economy”. Ammonia borane is notable for its high hydrogen densities. For the material, one of the main challenges is to release efficiently the maximum amount of the stored hydrogen. Hydrolysis reaction is a promising process by which hydrogen can be easily generated from this compound. High purity hydrogen from this compound can be evolved in the presence of solid acid or metal based catalyst. The reaction performance depends on the morphology and/or structure of these materials. In this review, we survey the research on nanostructured materials, especially porous materials for hydrogen generation from hydrolysis of ammonia borane.

  20. Volatile fatty acids platform from thermally hydrolysed secondary sewage sludge enhanced through recovered micronutrients from digested sludge.

    Kumi, Philemon J; Henley, Adam; Shana, Achame; Wilson, Victoria; Esteves, Sandra R

    2016-09-01

    The extracellular polymeric substances and microbial cytoplasmic contents seem to hold inorganic ions and organic products, such as proteins and carbohydrates that are of critical importance for the metabolism of hydrolytic and acidogenic anaerobic microorganisms. The addition of soluble microbially recovered nutrients from thermally treated digestate sludge, for the fermentation of thermally hydrolysed waste activated sludge, resulted in higher volatile fatty acids yields (VFAs). The yield of VFAs obtained from the recovered microbial nutrients was 27% higher than the no micronutrients control, and comparable to the yield obtained using a micronutrients commercial recipe. In addition, the use of a low pH resulting from a high sucrose dose to select spore forming acidogenic bacteria was effective for VFA production, and yielded 20% higher VFAs than without the pH shock and this associated with the addition of recovered microbial nutrients would overcome the need to thermally pre-treat the inoculum. PMID:27206055

  1. Hydrolytic study of the copolymer Poly pyrrole/ Polyethyleneglycol and Poly pyrrole synthesized by plasma

    In this work the study about the hydrolytic compatibility of semiconductor polymers, copolymer Poly pyrrole/ Polyethyleneglycol (PPy/PEG) and Poly pyrrole (PPy) for their possible use as biomaterials. The polymers were synthesized by plasma between 10 and 100 W, with discharges of splendor RF to 13.5 MHz with resistive coupling. The hydrolytic affinity was evaluated calculating the contact angle with solutions of NaCl, NaCl-MgSO4 and Krebs-Ringer. The results show a hydrophilicity increment due to the increase of the surface ruggedness with the synthesis energy. On the contrary, the crystallinity diminishes when increasing the power in PPy and it stays approximately constant in PPy/PEG. The electric conductivity presents a growth from 2 to 4 magnitude orders in function of the water content in the polymers. (Author)

  2. Shape and size controlled synthesis of uniform iron oxide nanocrystals through new non-hydrolytic routes.

    Li, Wenlu; Lee, Seung Soo; Wu, Jiewei; Hinton, Carl H; Fortner, John D

    2016-08-12

    New, non-hydrolytic routes to synthesize highly crystalline iron oxide nanocrystals (8-40 nm, magnetite) are described in this report whereby particle size and morphology were precisely controlled through reactant (precursor, e.g. (FeO(OH)) ratios, co-surfactant and organic additive, and/or reaction time. Particle size, with high monodispersivity (materials synthesized/purified in organic solvents are broadly water dispersible through a variety of phase (aqueous) transfer method(s). PMID:27354334

  3. Biosynthesis of hydrolytic enzymes of UV-mutants of Bacillus subtilis B-3 strain

    The studies were carried out on 12 UV-mutants of Bacillus subtilis B-3 which synthesize from 18 to 105% more hydrolytic enzymes (proteinases, amylases, cellulases and lipases) than the wild strain. After 5 years' storage of the mutants, a 26-fold increase in amylase biosynthesis was observed for a G7 UV-mutant. This mutant showed also other phenotypic differences. The increased ability of mutant G7 to synthesize amylases was controlled by the maltose-substrate induction. (author)

  4. Cellulose Surface Degradation by a Lytic Polysaccharide Monooxygenase and Its Effect on Cellulase Hydrolytic Efficiency*

    Eibinger, Manuel; Ganner, Thomas; Bubner, Patricia; Rošker, Stephanie; Kracher, Daniel; Haltrich, Dietmar; Ludwig, Roland; Plank, Harald; Nidetzky, Bernd

    2014-01-01

    Lytic polysaccharide monooxygenase (LPMO) represents a unique principle of oxidative degradation of recalcitrant insoluble polysaccharides. Used in combination with hydrolytic enzymes, LPMO appears to constitute a significant factor of the efficiency of enzymatic biomass depolymerization. LPMO activity on different cellulose substrates has been shown from the slow release of oxidized oligosaccharides into solution, but an immediate and direct demonstration of the enzyme action on the cellulos...

  5. Optimization of the hydrolytic-acidogenic anaerobic digestion stage (55 degrees C) of sewage sludge: influence of pH and solid content.

    Ponsá, Sergio; Ferrer, Ivet; Vázquez, Felícitas; Font, Xavier

    2008-08-01

    In conventional single-stage anaerobic digestion processes, hydrolysis is regarded as the rate-limiting step in the degradation of complex organic compounds, such as sewage sludge. Two-stage systems have been proposed to enhance this process. However, so far it is not clear which are the best conditions for a two-stage anaerobic digestion process of sewage sludge, in terms of temperature and hydraulic retention time of each stage. The aim of this work was to determine the optimal conditions for the hydrolytic-acidogenic stage treating real sludge with a high concentration of total solids (40-50gL(-1)) and volatile solids (25-30gL(-1)), named high concentration sludge. The variables considered for this first stage were: hydraulic retention time (1-4 days) and temperature (55 and 65 degrees C). Maximum volatile fatty acids generation was obtained at 4 days and 3 days hydraulic retention time for 55 degrees C and 65 degrees C, respectively. Consequently, 4 days hydraulic retention time and temperature of 55 degrees C were set as the working conditions for the hydrolytic-acidogenic stage treating high concentration sludge. The results obtained when operating with high concentration sludge were compared with a low concentration sludge consisting of 17-28gL(-1) total solids and 13-21gL(-1) volatile solids. The effect of decreasing the influent sludge pH, when working at the optimal conditions established, was also evaluated. PMID:18687452

  6. Durability of switchable QR code carriers under hydrolytic and photolytic conditions

    Following a guest diffusion approach, the surface of a shape memory poly(ester urethane) (PEU) was either black or blue colored. Bowtie-shaped quick response (QR) code carriers were then obtained from laser engraving and cutting, before thermo-mechanical functionalization (programming) was applied to stabilize the PEU in a thermo-responsive (switchable) state. The stability of the dye within the polymer surface and long-term functionality of the polymer were investigated against UVA and hydrolytic ageing. Spectrophotometric investigations verified UVA ageing-related color shifts from black to yellow-brownish and blue to petrol-greenish whereas hydrolytically aged samples changed from black to greenish and blue to light blue. In the case of UVA ageing, color changes were accompanied by dye decolorization, whereas hydrolytic ageing led to contrast declines due to dye diffusion. The Michelson contrast could be identified as an effective tool to follow ageing-related contrast changes between surface-dyed and laser-ablated (undyed) polymer regions. As soon as the Michelson contrast fell below a crucial value of 0.1 due to ageing, the QR code was no longer decipherable with a scanning device. Remarkably, the PEU information carrier base material could even then be adequately fixed and recovered. Hence, the surface contrast turned out to be the decisive parameter for QR code carrier applicability. (paper)

  7. Identification and characterization of hydrolytic enzymes from the midgut of the cotton bollworm, Helicoverpa armigera Hübner (Lepidoptera: Noctuidae)

    Özgür, Ebru; Yücel, Meral; Öktem, Hüseyin Avni

    2009-01-01

    Midgut hydrolytic enzymes of Helicoverpa armigera Hübner (Lepidoptera: Noctuidae) were identified and partially characterized. Km, Vmax, optimum pH, and specific activity were determined for proteolytic enzymes and a-amylases. All hydrolytic enzyme activity had an optimum pH value in the alkaline pH range. We observed major serine protease activity, together with minor cysteine-like activity, the former being significantly inhibited by soybean trypsin inhibitor (SBTI) and aprotinin. Moreover,...

  8. RAPD analysis of genetic diversity and qualitative assessment of hydrolytic activities in a collection of Bacillus sp. isolate

    Berić Tanja; Urdaci Maria C.; Stanković S.; Knežević-Vukčević Jelena

    2009-01-01

    Genetic diversity and production of hydrolytic enzymes of 205 Bacillus isolates from different geographical and ecological niches in Serbia were studied. Combining RAPD analysis and 16S DNA sequencing, we determined 13 different groups of RAPD profiles within four (five) species: B. subtilis, B. cereus/B. thuringiensis, B. pumilus, and B. firmus. Screening for production of hydrolytic enzymes showed that there was no correlation of enzyme production with species. Most of the isolates from all...

  9. Botryosphaeriales fungi produce extracellular enzymes with biotechnological potential.

    Esteves, Ana Cristina; Saraiva, Márcia; Correia, António; Alves, Artur

    2014-05-01

    Phytopathogenic fungi are known for producing an arsenal of extracellular enzymes whose involvement in the infection mechanism has been suggested. However, these enzymes are largely unknown and their biotechnological potential also remains poorly understood. In this study, the production and thermostability of extracellular enzymes produced by phytopathogenic Botryosphaeriaceae was investigated. Hydrolytic and oxidative activities were detected and quantified at different temperatures. Most strains (70%; 37/53) were able to produce simultaneously cellulases, laccases, xylanases, pectinases, pectin lyases, amylases, lipases, and proteases. Surprisingly for mesophilic filamentous fungi, several enzymes proved to be thermostable: cellulases from Neofusicoccum mediterraneum CAA 001 and from Dothiorella prunicola CBS 124723, lipases from Diplodia pinea (CAA 015 and CBS 109726), and proteases from Melanops tulasnei CBS 116806 were more active at 70 °C than at any of the other temperatures tested. In addition, lipases produced by Diplodia pinea were found to be significantly more active than any other known lipase from Botryosphaeriales. The thermal activity profile and the wide array of activities secreted by these fungi make them optimal producers of biotechnologically relevant enzymes that may be applied in the food and the health industries (proteases), the pulp-and-paper and biofuel industries (cellulases), or even in the detergent industry (lipases, proteases, amylases, and cellulases). PMID:24802941

  10. Extracellular Enzyme Activity assay as indicator of soil microbial functional diversity and activity

    Hendriksen, Niels Bohse; Winding, Anne

    2012-01-01

    Extracellular Enzyme Activity assay as indicator of soil microbial functional diversity and activity Niels Bohse Hendriksen, Anne Winding. Department of Environmental Science, Aarhus University, 4000 Roskilde, Denmark Soil enzymes originate from a variety of organisms, notably fungi and bacteria...... and especially hydrolytic extracellular enzymes are of pivotal importance for decomposition of organic substrates and biogeochemical cycling. Their activity reflects the functional diversity and activity of the microorganisms involved in decomposition processes which are essential processes for soil...... functioning and soil ecosystem services. The soil enzyme activity has been measured by the use of fluorogenic model substrates e.g. methylumbelliferyl (MUF) substrates for a number of enzymes involved in the degradation of polysaccharides as cellulose, hemicellulose and chitin, while degradation of proteins...

  11. Extracellular enzyme activity assay as indicator of soil microbial functional diversity and activity

    Hendriksen, Niels Bohse; Winding, Anne

    2012-01-01

    Extracellular enzyme activity assay as indicator of soil microbial functional diversity and activity Niels Bohse Hendriksen, Anne Winding. Department of Environmental Science, Aarhus University, 4000 Roskilde, Denmark Soils provide numerous essential ecosystem services such as carbon cycling......, recycling of nutrients and waste, soil remediation, plant growth support and regulation of above ground biodiversity, resilience, and soil suppressiveness. As such, soil ecosystem services are beneficial and vital for human life and at the same time threatened by anthropogenic activities. Increasing...... of soil microbial functions is still needed. In soil, enzymes originate from a variety of organisms, notably fungi and bacteria and especially hydrolytic extracellular enzymes are of pivotal importance for decomposition of organic substrates and biogeochemical cycling. Their activity will reflect the...

  12. Influence of Wall Teichoic Acid on Lysozyme Resistance in Staphylococcus aureus▿

    Bera, Agnieszka; Biswas, Raja; Herbert, Silvia; Kulauzovic, Emir; Weidenmaier, Christopher; Peschel, Andreas; Götz, Friedrich

    2006-01-01

    Staphylococcus aureus peptidoglycan (PG) is completely resistant to the hydrolytic activity of lysozyme. Here we show that modifications in PG by O acetylation, wall teichoic acid, and a high degree of cross-linking contribute to this resistance.

  13. Fragmentation of extracellular matrix by hypochlorous acid

    Woods, Alan A; Davies, Michael Jonathan

    2003-01-01

    /chloramide decomposition, with copper and iron ions being effective catalysts, and decreased by compounds which scavenge chloramines/chloramides, or species derived from them. The effect of such matrix modifications on cellular behaviour is poorly understood, though it is known that changes in matrix materials can have...

  14. Streptococcus mutans-derived extracellular matrix in cariogenic oral biofilms

    Klein, Marlise I.; Hwang, Geelsu; Santos, Paulo H. S.; Campanella, Osvaldo H.; Koo, Hyun

    2015-01-01

    Biofilms are highly structured microbial communities that are enmeshed in a self-produced extracellular matrix. Within the complex oral microbiome, Streptococcus mutans is a major producer of extracellular polymeric substances including exopolysaccharides (EPS), eDNA, and lipoteichoic acid (LTA). EPS produced by S. mutans-derived exoenzymes promote local accumulation of microbes on the teeth, while forming a spatially heterogeneous and diffusion-limiting matrix that protects embedded bacteria...

  15. Streptococcus mutans-derived extracellular matrix in cariogenic oral biofilms

    Marlise eKlein; Geelsu eHwang; Paulo eSantos; Osvaldo eCampanella; Hyun eKoo

    2015-01-01

    Biofilms are highly structured microbial communities that are enmeshed in a self-produced extracellular matrix. Within the complex oral microbiome, Streptococcus mutans is a major producer of extracellular polymeric substances including exopolysaccharides (EPS), eDNA and lipoteichoic acid (LTA). EPS produced by S. mutans-derived exoenzymes promote local accumulation of microbes on the teeth, while forming a spatially heterogeneous and diffusion-limiting matrix that protects embedded bacteria....

  16. Extracellular slime associated with Proteus mirabilis during swarming.

    Stahl, S. J.; Stewart, K R; Williams, F. D.

    1983-01-01

    Light microscopy, transmission electron microscopy, and scanning electron microscopy were used to visualize the extracellular slime of Proteus mirabilis swarm cells. Slime was observed with phase-contrast microscopy after fixation in hot sulfuric acid-sodium borate. Ruthenium red was used to stain slime for transmission electron microscopy. Copious quantities of extracellular slime were observed surrounding swarm cells; the slime appeared to provide a matrix through which the cells could migr...

  17. Extracellular Control of Limb Regeneration

    Calve, S.; Simon, H.-G.

    Adult newts possess the ability to completely regenerate organs and appendages. Immediately after limb loss, the extracellular matrix (ECM) undergoes dramatic changes that may provide mechanical and biochemical cues to guide the formation of the blastema, which is comprised of uncommitted stem-like cells that proliferate to replace the lost structure. Skeletal muscle is a known reservoir for blastema cells but the mechanism by which it contributes progenitor cells is still unclear. To create physiologically relevant culture conditions for the testing of primary newt muscle cells in vitro, the spatio-temporal distribution of ECM components and the mechanical properties of newt muscle were analyzed. Tenascin-C and hyaluronic acid (HA) were found to be dramatically upregulated in the amputated limb and were co-expressed around regenerating skeletal muscle. The transverse stiffness of muscle measured in situ was used as a guide to generate silicone-based substrates of physiological stiffness. Culturing newt muscle cells under different conditions revealed that the cells are sensitive to both matrix coating and substrate stiffness: Myoblasts on HA-coated soft substrates display a rounded morphology and become more elongated as the stiffness of the substrate increases. Coating of soft substrates with matrigel or fibronectin enhanced cell spreading and eventual cell fusion.

  18. Replacement of traditional seawater-soluble pigments by starch and hydrolytic enzymes in polishing antifouling coatings

    Olsen, Søren Martin; Pedersen, L. T.; Dam-Johansen, Kim;

    2010-01-01

    The use of starch and hydrolytic enzymes as replacement for traditional polishing pigments (e.g., Cu2O and ZnO) in antifouling coatings has been investigated. The enzymes facilitate a slow conversion of water-insoluble starch into water-soluble glucose, and dissolution of glucose causes the...... starch-enzyme coatings tested; however, polishing is only detected for two out of four binder systems investigated. Suitable polishing rates of 7-10 mu m/month, based on the enzymatic starch-degradation, have been measured. Controls containing only starch (no enzyme) did not polish....

  19. Thermogravimetry for optimization of chloride pyro hydrolytic separations in zirconia-magnesia matrix

    A fast and accurate method for chloride ion determination in zirconia-magnesia matrix was studied the method consists in the pyro hydrolysis of the oxides at 900o C, using a quartz apparatus, during thirty minutes and further determination of the chloride ion collected by means of either ion-selective electrode or ion chromatography. The thermogravimetric curves (TG curves) of the metal oxides and oxychlorides provide all the information about the chloride ion evolution temperature and the influence of pyro hydrolytic accelerators (U3 O8) on ion evolution. (author)

  20. Novel Materials through Non-Hydrolytic Sol-Gel Processing: Negative Thermal Expansion Oxides and Beyond

    Cora Lind

    2010-04-01

    Full Text Available Low temperature methods have been applied to the synthesis of many advanced materials. Non-hydrolytic sol-gel (NHSG processes offer an elegant route to stable and metastable phases at low temperatures. Excellent atomic level homogeneity gives access to polymorphs that are difficult or impossible to obtain by other methods. The NHSG approach is most commonly applied to the preparation of metal oxides, but can be easily extended to metal sulfides. Exploration of experimental variables allows control over product stoichiometry and crystal structure. This paper reviews the application of NHSG chemistry to the synthesis of negative thermal expansion oxides and selected metal sulfides.

  1. Catalytic efficiency of hydrolytic enzymes is related to microbial community structure and is constrained by resource availability

    Tischer, Alexander; Hamer, Ute; Blagodatskaya, Evgenia

    2015-04-01

    The turnover of nutrients bound to organic matter is mediated by extracellular enzymes (EHEs) produced by soil microorganisms. We linked catalytic properties of extracellular enzymes to a set of environmental and microbial variables in soils of a land-use sequence ranging from undisturbed natural forest and pastures of different ages to secondary succession in the Andes of southern Ecuador. The sensitivity of substrate affinity constant (Km) and maximal rate (Vmax) - of six EHEs (β-cellobiohydrolase, β-glucosidase, N-acetylglucosaminidase, α-glucosidase, xylanase, acid phosphomonoesterase) to changing environmental conditions was approximated by Michaelis-Menten kinetics. Microbial communities adapted to environmental changes, demonstrated high flexibility of extracellular enzyme systems (determined by Km) and selected for the enzymes with higher catalytic efficiency as compared with pure cultures. Our study showed that high turnover rates of starch, hemicellulose, and monophosphoesters was associated with Gram(-)-bacteria Land-use associated declines in microbial biomass and soil N strongly reduced the turnover of monoester-bound organic P in soils during long-term pasture-use and secondary succession. Our data further suggest an increase in turnover of microbial biomass leads to an increase of DOC concentrations, as a consequence of N-acetylglucosaminidase activity.

  2. In vitro differential activity of phospholipases and acid proteinases of clinical isolates of Candida

    Aurean D'Eça Júnior

    2011-06-01

    Full Text Available INTRODUCTION: Candida yeasts are commensals; however, if the balance of normal flora is disrupted or the immune defenses are compromised, Candida species can cause disease manifestations. Several attributes contribute to the virulence and pathogenicity of Candida, including the production of extracellular hydrolytic enzymes, particularly phospholipase and proteinase. This study aimed to investigate the in vitro activity of phospholipases and acid proteinases in clinical isolates of Candida spp. METHODS: Eighty-two isolates from hospitalized patients collected from various sites of origin were analyzed. Phospholipase production was performed in egg yolk medium and the production of proteinase was verified in a medium containing bovine serum albumin. The study was performed in triplicate. RESULTS: Fifty-six (68.3% of isolates tested were phospholipase positive and 16 (44.4% were positive for proteinase activity. C. tropicalis was the species with the highest number of positive isolates for phospholipase (91.7%. Statistically significant differences were observed in relation to production of phospholipases among species (p<0,0001 and among the strains from different sites of origin (p=0.014. Regarding the production of acid protease, the isolates of C. parapsilosis tested presented a larger number of producers (69.2%. Among the species analyzed, the percentage of protease producing isolates did not differ statistically (χ2=1.9 p=0.5901 (χ2=1.9 p=0.5901. CONCLUSIONS: The majority of C. non-albicans and all C. albicans isolates were great producers of hydrolytic enzymes and, consequently, might be able to cause infection under favorable conditions.

  3. HYDROLYTIC DEGRADATION BEHAVIOR OF PLLA NANOCOMPOSITES REINFORCED WITH MODIFIED CELLULOSE NANOCRYSTALS

    Everton Luiz de Paula

    2015-09-01

    Full Text Available Bionanocomposites derived from poly(L-Lactide (PLLA were reinforced with chemically modified cellulose nanocrystals (m-CNCs. The effects of these modified cellulose nanoparticles on the mechanical and hydrolytic degradation behavior of polylactide were studied. The m-CNCs were prepared by a method in which hydrolysis of cellulose chains is performed simultaneously with the esterification of hydroxyl groups to produce modified nanocrystals with ester groups. FTIR, elemental analysis, TEM, XRD and contact angle measurements were used to confirm and characterize the chemical modifications of the m-CNCs. These bionanocomposites gave considerably better mechanical properties than neat PLLA based on an approximately 100% increase in tensile strength. Due to the hydrophobic properties of the esterified nanocrystals incorporated into a polymer matrix, it was also demonstrated that a small amount of m-CNCs could lead to a remarkable decrease in the hydrolytic degradation rate of the biopolymer. In addition, the m-CNCs considerably delay the degradation of the nanocomposite by providing a physical barrier that prevents the permeation of water, which thus hinders the overall absorption of water into the matrix. The results obtained in this study show the nanocrystals can be used to reinforce polylactides and fine-tune their degradation rates in moist or physiological environments.

  4. Unfolded protein response is required for Aspergillus oryzae growth under conditions inducing secretory hydrolytic enzyme production.

    Tanaka, Mizuki; Shintani, Takahiro; Gomi, Katsuya

    2015-12-01

    Unfolded protein response (UPR) is an intracellular signaling pathway for adaptation to endoplasmic reticulum (ER) stress. In yeast UPR, Ire1 cleaves the unconventional intron of HAC1 mRNA, and the functional Hac1 protein translated from the spliced HAC1 mRNA induces the expression of ER chaperone genes and ER-associated degradation genes for the refolding or degradation of unfolded proteins. In this study, we constructed an ireA (IRE1 ortholog) conditionally expressing strain of Aspergillus oryzae, a filamentous fungus producing a large amount of amylolytic enzymes, and examined the contribution of UPR to ER stress adaptation under physiological conditions. Repression of ireA completely blocked A. oryzae growth under conditions inducing the production of hydrolytic enzymes, such as amylases and proteases. This growth defect was restored by the introduction of unconventional intronless hacA (hacA-i). Furthermore, UPR was observed to be induced by amylolytic gene expression, and the disruption of the transcriptional activator for amylolytic genes resulted in partial growth restoration of the ireA-repressing strain. In addition, a homokaryotic ireA disruption mutant was successfully generated using the strain harboring hacA-i as a parental host. These results indicated that UPR is required for A. oryzae growth to alleviate ER stress induced by excessive production of hydrolytic enzymes. PMID:26496881

  5. High throughput screening of hydrolytic enzymes from termites using a natural substrate derived from sugarcane bagasse

    Lucena Severino A

    2011-11-01

    Full Text Available Abstract Background The description of new hydrolytic enzymes is an important step in the development of techniques which use lignocellulosic materials as a starting point for fuel production. Sugarcane bagasse, which is subjected to pre-treatment, hydrolysis and fermentation for the production of ethanol in several test refineries, is the most promising source of raw material for the production of second generation renewable fuels in Brazil. One problem when screening hydrolytic activities is that the activity against commercial substrates, such as carboxymethylcellulose, does not always correspond to the activity against the natural lignocellulosic material. Besides that, the macroscopic characteristics of the raw material, such as insolubility and heterogeneity, hinder its use for high throughput screenings. Results In this paper, we present the preparation of a colloidal suspension of particles obtained from sugarcane bagasse, with minimal chemical change in the lignocellulosic material, and demonstrate its use for high throughput assays of hydrolases using Brazilian termites as the screened organisms. Conclusions Important differences between the use of the natural substrate and commercial cellulase substrates, such as carboxymethylcellulose or crystalline cellulose, were observed. This suggests that wood feeding termites, in contrast to litter feeding termites, might not be the best source for enzymes that degrade sugarcane biomass.

  6. Selective optical response of hydrolytically stable stratified Si rugate mirrors to liquid infiltration.

    Jalkanen, Tero; Torres-Costa, Vicente; Mäkilä, Ermei; Kaasalainen, Martti; Koda, Ryo; Sakka, Tetsuo; Ogata, Yukio H; Salonen, Jarno

    2014-02-26

    Stratified optical filters with distinct spectral features and layered surface chemistry were prepared on silicon substrates with stepwise anodic porosification and thermal carbonization. The use of differing parameters for successive carbonization treatments enabled the production of hydrolytically stable porous silicon-based layered optical structures where the adsorption of water to the lower layer is inhibited. This enables selective shifting of reflectance bands by means of liquid infiltration. The merit of using thermal carbonization for creating layered functionality was demonstrated by comparing the hydrolytic stability resulting from this approach to other surface chemistries available for Si. The functionality of the stratified optical structures was demonstrated under water and ethanol infiltration, and changes in the adsorption properties after 9 months of storage were evaluated. The changes observed in the structure were explained using simulations based on the transfer matrix method and the Bruggeman effective medium approximation. Scanning electron microscopy was used for imaging the morphology of the porous structure. Finally, the adaptability of the method for preparing complex structures was demonstrated by stacking superimposed rugate structures with several reflective bands. PMID:24450851

  7. A fluorescence-based hydrolytic enzyme activity assay for quantifying toxic effects of Roundup® to Daphnia magna

    Ørsted, Michael; Roslev, Peter

    2015-01-01

    Daphnia magna is a widely used model organism for aquatic toxicity testing. In the present study, we investigated the hydrolytic enzyme activity of D. magna after exposure to toxicant stress. In vivo enzyme activity was quantified using 15 fluorogenic enzyme probes based on 4-methylumbelliferyl...... that the fluorescence based hydrolytic enzyme activity assay (FLEA assay) can be used as an index of D. magna stress. Combining enzyme activity with fluorescence measurements may be applied as a simple and quantitative supplement for toxicity testing with D. magna....... or 7-amino-4-methylcoumarin. Probing D. magna enzyme activity was evaluated using short-term exposure (24-48 h) to the reference chemical K2Cr2O7, or the herbicide formulation Roundup®. Toxicant induced changes in hydrolytic enzyme activity were compared to changes in mobility (ISO 6341). The results...

  8. Benthic oxygen uptake, hydrolytic potentials and microbial biomass at the Arctic continental slope

    Boetius, Antje; Damm, Ellen

    1998-02-01

    Oxygen (O 2) uptake and microbial activity in sediments of the eastern Arctic continental slope were investigated in both ice-covered and ice-free areas of the Laptev Sea. Total O 2 flux ( J) decreased markedly from 2 mmol m -2 d -1 at the shelf edge (50 m) to 0.07 mmol m -2 d -1 at the bottom of the slope (3500 m), matched by the more than tenfold decline in chlorophyll pigments (CPE), protein and dissolved amino acids (DFAA). Furthermore, concentrations of these labile organic compounds were strongly correlated with extracellular enzyme potentials (EEA) in the sediments as well as with microbial biomass. The concentrations of labile substances and total microbial biomass (TMB) as well as the rates of O 2 uptake and EEA were independent of the distribution of TOC, probably due to the dominance of non-labile terrigenous compounds. Differences in O 2 uptake and microbial EEA between ice-covered and ice-free transects were relatively small. Values of O 2 uptake, CPE, EEA and TMB at the Laptev Sea slope were considerably lower than at temperate continental slopes but nevertheless higher than in the central Arctic deep-sea basin. Considering newly published data on primary productivity in the central Arctic, our results indicate that the benthic respiratory demand at the Laptev Sea slope and in the Arctic basin could be satisfied by the vertical flux of POC and does not necessarily depend on lateral advection of POC from the shelf seas as previously anticipated.

  9. Towards early detection of the hydrolytic degradation of poly(bisphenol A)carbonate by hyphenated liquid chromatography and comprehensive two-dimensional liquid chromatography

    Coulier, L.; Kaal, E.R.; Hankemeier, Th.

    2006-01-01

    The hydrolytic degradation of poly(bisphenol A)carbonate (PC) has been characterized by various liquid chromatography techniques. Size exclusion chromatography (SEC) showed a significant decrease in molecular mass as a result of hydrolytic degradation, while 'liquid chromatography at critical condit

  10. Inhibitors of the Hydrolytic Enzyme Dimethylarginine Dimethylaminohydrolase (DDAH): Discovery, Synthesis and Development.

    Murphy, Rhys B; Tommasi, Sara; Lewis, Benjamin C; Mangoni, Arduino A

    2016-01-01

    Dimethylarginine dimethylaminohydrolase (DDAH) is a highly conserved hydrolytic enzyme found in numerous species, including bacteria, rodents, and humans. In humans, the DDAH-1 isoform is known to metabolize endogenous asymmetric dimethylarginine (ADMA) and monomethyl arginine (l-NMMA), with ADMA proposed to be a putative marker of cardiovascular disease. Current literature reports identify the DDAH family of enzymes as a potential therapeutic target in the regulation of nitric oxide (NO) production, mediated via its biochemical interaction with the nitric oxide synthase (NOS) family of enzymes. Increased DDAH expression and NO production have been linked to multiple pathological conditions, specifically, cancer, neurodegenerative disorders, and septic shock. As such, the discovery, chemical synthesis, and development of DDAH inhibitors as potential drug candidates represent a growing field of interest. This review article summarizes the current knowledge on DDAH inhibition and the derived pharmacokinetic parameters of the main DDAH inhibitors reported in the literature. Furthermore, current methods of development and chemical synthetic pathways are discussed. PMID:27187323

  11. Metagenomics as a Tool for Enzyme Discovery: Hydrolytic Enzymes from Marine-Related Metagenomes.

    Popovic, Ana; Tchigvintsev, Anatoly; Tran, Hai; Chernikova, Tatyana N; Golyshina, Olga V; Yakimov, Michail M; Golyshin, Peter N; Yakunin, Alexander F

    2015-01-01

    This chapter discusses metagenomics and its application for enzyme discovery, with a focus on hydrolytic enzymes from marine metagenomic libraries. With less than one percent of culturable microorganisms in the environment, metagenomics, or the collective study of community genetics, has opened up a rich pool of uncharacterized metabolic pathways, enzymes, and adaptations. This great untapped pool of genes provides the particularly exciting potential to mine for new biochemical activities or novel enzymes with activities tailored to peculiar sets of environmental conditions. Metagenomes also represent a huge reservoir of novel enzymes for applications in biocatalysis, biofuels, and bioremediation. Here we present the results of enzyme discovery for four enzyme activities, of particular industrial or environmental interest, including esterase/lipase, glycosyl hydrolase, protease and dehalogenase. PMID:26621459

  12. Industrially important hydrolytic enzyme diversity explored in stove ash bacterial isolates.

    Kiran, Tabbassum; Asad, Wajeeha; Siddiqui, Shahla; Ajaz, Munazza; Rasool, Sheikh Ajaz

    2015-11-01

    Extreme environments merit special attention and significance because of the possible existence of thermophilic microorganisms in such ecological niches. Keeping this in mind indigenous stove ash samples were explored for extremophilic bacteria in term of their biodiversity. Accordingly, this study reports 37 bacterial isolates from the local wood run oven (Tandoor) ash samples. All the isolated strains belong to genus Bacillus on the bases of morpho-cultural and biochemical considerations. The average temperature tolerance profile was >45°C thereby, indicating towards the thermophilic nature of the isolated strains. The Bacillus isolates were screened for 10 different hydrolytic enzymes (cellulase, xylanase, amylase, pectinase, caseinase, keratinase, lipase, esterase, dextranase and β-galactosidase) by plate screening method using the medium incorporated with specific substrate(s). It was found that keratinase was produced by all the isolates while, 36 (97.2%) isolates showed caseinase and esterase production. Amylase was produced by 35(94.6%) isolates and 34 (91.8%) isolates were able to degrade Tween-80 and xylan as substrate for lipase and xylanase respectively. The enzyme, β-galactosidase was produced by 31 (89.1%) of the isolates. Cellulase and dextranase were produced by 26 (70.2%) and 22 (59.4%) isolates respectively. None of the isolates could (under the existing conditions) produce pectin-hydrolyzing enzyme. According to the Tukey's post hoc test, significant difference was found between the mean enzyme index of all the (screened) enzymes. Thus, the isolated bacterial strains with diverse hydrolytic potential may be of great value and relevance for the existing (national) industrial setups. PMID:26639497

  13. Acute acidification of stratum corneum membrane domains using polyhydroxyl acids improves lipid processing and inhibits degradation of corneodesmosomes.

    Hachem, Jean-Pierre; Roelandt, Truus; Schürer, Nanna; Pu, Xu; Fluhr, Joachim; Giddelo, Christina; Man, Mao-Qiang; Crumrine, Debra; Roseeuw, Diane; Feingold, Kenneth R; Mauro, Theodora; Elias, Peter M

    2010-02-01

    Neutralization of the normally acidic stratum corneum (SC) has deleterious consequences for permeability barrier homeostasis and SC integrity/cohesion attributable to serine proteases (SPs) activation leading to deactivation/degradation of lipid-processing enzymes and corneodesmosomes (CD). As an elevated pH compromises SC structure and function, we asked here whether SC hyperacidification would improve the structure and function. We lowered the pH of mouse SC using two polyhydroxyl acids (PHA), lactobionic acid (LBA), or gluconolactone (GL). Applications of the PHA reduced the pH at all levels of SC of hairless mouse, with further selective acidification of SC membrane domains, as shown by fluorescence lifetime imaging. Hyperacidification improved permeability barrier homeostasis, attributable to increased activities of two key membrane-localized, ceramide-generating hydrolytic enzymes (beta-glucocerebrosidase and acidic sphingomyelinase), which correlated with accelerated extracellular maturation of SC lamellar membranes. Hyperacidification generated "supernormal" SC integrity/cohesion, attributable to an SP-dependent decreased degradation of desmoglein-1 (DSG1) and the induction of DSG3 expression in lower SC. As SC hyperacidification improves the structure and function, even of normal epidermis, these studies lay the groundwork for an assessment of the potential utility of SC acidification as a therapeutic strategy for inflammatory dermatoses, characterized by abnormalities in barrier function, cohesion, and surface pH. PMID:19741713

  14. COMPLEX HYDROLYTIC PROCESSING OF PENTOSAN PLANT BIOMASS FOR PRODUCTION FURFURAL AND PROTEIN-CONTAINING FEED ADDITIVE (REVIEW

    Валерий Станиславович Болтовский

    2014-10-01

    Full Text Available The review is devoted to methods of complex hydrolytic and microbial processing for the production of furfural from pentose hydrolysates and protein-containing feed additives, said process comprising bioconversion the said pentose hydrolysates and protein-containing feed additives by solid-phase fermentation of lignocellulose.

  15. The influence of morphology on the (hydrolytic degradation of as-polymerized and hot-drawn poly (L-lactide))

    Joziasse, CAP; Grijpma, DW; Cordewener, FW; Bos, RRM; Pennings, AJ

    1998-01-01

    The influence of morphology on the hydrolytic degradation behavior of poly(L-lactide) has been studied. High molecular weight and highly crystalline as-polymerized poly(L-lactide) was obtained in high yields through melt polymerization. Poly(L-lactide) fiber with a draw ratio of 5.6 was obtained by

  16. Humic Acid-Like and Fulvic Acid-Like Inhibition on the Hydrolysis of Cellulose and Tributyrin

    Fernandes, T.V.; Lier, van J.B.; Zeeman, Grietje

    2015-01-01

    Enzymatic hydrolysis of complex wastes is a critical step for efficient biogas production in anaerobic digesters. Inhibition of this hydrolytic step was studied by addition of humic acid-like (HAL) and fulvic acid-like (FAL) substances, extracted from maize silage and fresh cow manure, to batch t

  17. Extracellular proteases of Halobacillus blutaparonensis strain M9, a new moderately halophilic bacterium

    Anderson F. Santos

    2013-12-01

    Full Text Available Halophilic microorganisms are source of potential hydrolytic enzymes to be used in industrial and/or biotechnological processes. In the present study, we have investigated the ability of the moderately halophilic bacterium Halobacillus blutaparonensis (strain M9, a novel species described by our group, to release proteolytic enzymes. This bacterial strain abundantly proliferated in Luria-Bertani broth supplemented with 2.5% NaCl as well as secreted proteases to the extracellular environment. The production of proteases occurred in bacterial cells grown under different concentration of salt, ranging from 0.5% to 10% NaCl, in a similar way. The proteases secreted by H. blutaparonensis presented the following properties: (i molecular masses ranging from 30 to 80 kDa, (ii better hydrolytic activities under neutral-alkaline pH range, (iii expression modulated according to the culture age, (iv susceptibility to phenylmethylsulphonyl fluoride, classifying them as serine-type proteases, (v specific cleavage over the chymotrypsin substrate, and (vi enzymatic stability in the presence of salt (up to 20% NaCl and organic solvents (e.g., ether, isooctane and cyclohexane. The proteases described herein are promising for industrial practices due to its haloalkaline properties.

  18. CHARACTERIZATION OF EXTRACELLULAR GABA IN THE SUBSTANTIA-NIGRA-RETICULATA BY MEANS OF BRAIN MICRODIALYSIS

    TIMMERMAN, W; ZWAVELING, J; WESTERINK, BHC

    1992-01-01

    Brain microdialysis was used to characterize extracellular gamma-aminobutyric acid (GABA) in the substantia nigra reticulata (SNR) of freely moving rats. The extracellular GABA in the SNR was characterized using acutely implanted probes (4-8 h after surgery; day 1) and chronically implanted probes (

  19. Crystallization and preliminary X-ray crystallographic study of the wild type and two mutants of the CP1 hydrolytic domain from Aquifex aeolicus leucyl-tRNA synthetase

    The wild-type editing CP1 domain of A. aeolicus leucyl-tRNA synthetase and two mutant CP1 domains have been overexpressed, purified and crystallized. X-ray diffraction data have been collected to 1.8 Å, which has enabled determination of the structures by molecular replacement. The editing or hydrolytic CP1 domain of leucyl-tRNA synthetase (LeuRS) hydrolyses several misactivated amino acids. The CP1 domain of Aquifex aeolicus LeuRS was expressed, purified and crystallized by the hanging-drop vapour-diffusion method using ammonium sulfate as precipitant. Crystals belong to space group P212121, with unit-cell parameters a = 38.8, b = 98.4, c = 116.7 Å. Crystals diffract to beyond 1.8 Å resolution and contain two monomers in the asymmetric unit. Two CP1 mutants in which a conserved threonine residue essential for the fidelity of the hydrolytic pathway is mutated to alanine or glutamic acid have also been expressed and crystallized. Crystals of the two CP1 mutants are isomorphs of the wild type and diffract to beyond 1.9 Å resolution. All structures were solved by molecular-replacement techniques

  20. Crystallization and preliminary X-ray crystallographic study of the wild type and two mutants of the CP1 hydrolytic domain from Aquifex aeolicus leucyl-tRNA synthetase

    Cura, Vincent; Olieric, Natacha; Guichard, Alexandre [Département de Biologie et Génomique Structurales, UMR 7104, Institut de Génétique et de Biologie Moléculaire et Cellulaire, CNRS/INSERM/ULP Strasbourg, 1 Rue Laurent Fries, 67404 Illkirch (France); Wang, En-Duo [State Key Laboratory of Molecular Biology, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, The Chinese Academy of Sciences, 320 Yue Yang Road, Shanghai 200031 (China); Moras, Dino [Département de Biologie et Génomique Structurales, UMR 7104, Institut de Génétique et de Biologie Moléculaire et Cellulaire, CNRS/INSERM/ULP Strasbourg, 1 Rue Laurent Fries, 67404 Illkirch (France); Eriani, Gilbert [Architecture et Réactivité de l’ARN, UPR 9002, Institut de Biologie Moléculaire et Cellulaire du CNRS, 15 Rue René Descartes, 67084 Strasbourg (France); Cavarelli, Jean, E-mail: cava@igbmc.u-strasbg.fr [Département de Biologie et Génomique Structurales, UMR 7104, Institut de Génétique et de Biologie Moléculaire et Cellulaire, CNRS/INSERM/ULP Strasbourg, 1 Rue Laurent Fries, 67404 Illkirch (France)

    2005-10-01

    The wild-type editing CP1 domain of A. aeolicus leucyl-tRNA synthetase and two mutant CP1 domains have been overexpressed, purified and crystallized. X-ray diffraction data have been collected to 1.8 Å, which has enabled determination of the structures by molecular replacement. The editing or hydrolytic CP1 domain of leucyl-tRNA synthetase (LeuRS) hydrolyses several misactivated amino acids. The CP1 domain of Aquifex aeolicus LeuRS was expressed, purified and crystallized by the hanging-drop vapour-diffusion method using ammonium sulfate as precipitant. Crystals belong to space group P2{sub 1}2{sub 1}2{sub 1}, with unit-cell parameters a = 38.8, b = 98.4, c = 116.7 Å. Crystals diffract to beyond 1.8 Å resolution and contain two monomers in the asymmetric unit. Two CP1 mutants in which a conserved threonine residue essential for the fidelity of the hydrolytic pathway is mutated to alanine or glutamic acid have also been expressed and crystallized. Crystals of the two CP1 mutants are isomorphs of the wild type and diffract to beyond 1.9 Å resolution. All structures were solved by molecular-replacement techniques.

  1. Extracellular enzyme activities in a tropical mountain rainforest region of southern Ecuador affected by low soil P status and land-use change

    Tischer, Alexander; Blagodatskaya, Evgenia; Ute, Hamer

    2014-05-01

    Little is known about the enzymatic response of microorganisms in soils having a low P status and being subjected to global change phenomena, such as forest disturbance and land-use change. Along a land-use sequence (natural forest - young pasture - old pasture - abandoned pasture - shrubland) in the Andes of southern Ecuador mineral topsoils of Cambisols / Umbrisols were investigated. We tested whether the activities of the six hydrolytic enzymes (cellobiohydrolase, β-glucosidase, N-acetylglucosaminidase, α-glucosidase, xylanase, acid phosphomonoesterase) were affected by nutrient status and land-use induced alterations in soil pH (pHH2O from 3.7 to 5.2), resource quantity and quality (e.g. a SOC:N:P ratio from 182:13:1 to 1050:38:1) and microbial community structure (as monitored by phospholipid fatty acids). Microbial production of acid phosphatase responded to the low P status of the sites by a higher investment in the acquisition of P compared to C. We determined three major drivers of enzyme activities: 1.) Microbial demand for P regulated the production of acid phosphatase, provided that N and C were available. At the natural forest site the two-fold higher specific activity of acid phosphatase pointed to a high microbial P-demand, whereas the production of acid phosphatase was constrained by the availability of N and DOC after pasture abandonment. 2.) Microbial biomass that was controlled by pH and resource availability (total soil N (organic and inorganic N), organic P (Bray-fraction)) was the main driver for cellobiohydrolase, β-glucosidase and N-acetylglucosaminidase activities. 3.) Substrate induction due to increased litter inputs of herbaceous plant species seemed to regulate α-glucosidase and xylanase activities during secondary succession. In contrast, alterations in the abundance of microbial groups affected the variation in extracellular enzyme activities only marginally. At the level of broadly defined microbial groups (PLFA), our results

  2. Electrospun polyvinyl alcohol-collagen-hydroxyapatite nanofibers: a biomimetic extracellular matrix for osteoblastic cells

    Song, Wei; Markel, David C.; Wang, Sunxi; Shi, Tong; Mao, Guangzhao; Ren, Weiping

    2012-03-01

    The failure of prosthesis after total joint replacement is due to the lack of early implant osseointegration. In this study polyvinyl alcohol-collagen-hydroxyapatite (PVA-Col-HA) electrospun nanofibrous meshes were fabricated as a biomimetic bone-like extracellular matrix for the modification of orthopedic prosthetic surfaces. In order to reinforce the PVA nanofibers, HA nanorods and Type I collagen were incorporated into the nanofibers. We investigated the morphology, biodegradability, mechanical properties and biocompatibility of the prepared nanofibers. Our results showed these inorganic-organic blended nanofibers to be degradable in vitro. The encapsulated nano-HA and collagen interacted with the PVA content, reinforcing the hydrolytic resistance and mechanical properties of nanofibers that provided longer lasting stability. The encapsulated nano-HA and collagen also enhanced the adhesion and proliferation of murine bone cells (MC3T3) in vitro. We propose the PVA-Col-HA nanofibers might be promising modifying materials on implant surfaces for orthopedic applications.

  3. Electrospun polyvinyl alcohol–collagen–hydroxyapatite nanofibers: a biomimetic extracellular matrix for osteoblastic cells

    The failure of prosthesis after total joint replacement is due to the lack of early implant osseointegration. In this study polyvinyl alcohol–collagen–hydroxyapatite (PVA-Col-HA) electrospun nanofibrous meshes were fabricated as a biomimetic bone-like extracellular matrix for the modification of orthopedic prosthetic surfaces. In order to reinforce the PVA nanofibers, HA nanorods and Type I collagen were incorporated into the nanofibers. We investigated the morphology, biodegradability, mechanical properties and biocompatibility of the prepared nanofibers. Our results showed these inorganic–organic blended nanofibers to be degradable in vitro. The encapsulated nano-HA and collagen interacted with the PVA content, reinforcing the hydrolytic resistance and mechanical properties of nanofibers that provided longer lasting stability. The encapsulated nano-HA and collagen also enhanced the adhesion and proliferation of murine bone cells (MC3T3) in vitro. We propose the PVA-Col-HA nanofibers might be promising modifying materials on implant surfaces for orthopedic applications. (paper)

  4. Microbial decomposition in aquatic environments: combined process of extracellular enzyme activity and substrate uptake

    The aim of this study was to define a model for the coupling between extracellular enzyme activity and substrate uptake by bacterial populations in natural waters. The balance between uptake of leucine and extracellular hydrolytic production of leucine from a peptide model substrate was investigated in a combined fluorescence-radiotracer experiment with [3H] leucine as a marker for the leucine pool and L-leucine-4-methyl-7-coumarinylamide (Leu-MCA) as a marker for the pool of dissolved peptide substrates. Results show that at low concentrations of the model substrate the input and uptake processes of leucine are nearly balanced, whereas at high concentrations of the model substrate much more leucine is liberated than taken up. In addition, samples from one polluted and one less polluted station in the Kiel Fjord were investigated for their extracellular enzymatic and uptake properties in an annual cycle. Calculated on an annual average basis, turnover rates were ca. nine times higher than hydrolysis rates at the polluted station and ca., five times higher at the less polluted station. From the described model, this would mean that the relative fraction of polymers within the total dissolved organic carbon pool (with regard to the substrate combination dissolved protein-leucine) is about twice that at the polluted than at the less polluted station

  5. Extracellular proteins limit the dispersal of biogenic nanoparticles

    Moreau, J.W.; Weber, P.K.; Martin, M.C.; Gilbert, B.; Hutcheon, I.D.; Banfield, J.F.

    2007-01-01

    High-spatial-resolution secondary ion microprobe spectrometry, synchrotron radiation-based Fourier-transform infrared spectroscopy, and polyacrylamide gel analysis demonstrated the intimate association of proteins with spheroidal aggregates of biogenic zinc sulfide nanocrystals, an example of extracellular biomineralization. Experiments involving synthetic zinc sulfide nanoparticles and representative amino acids indicated a driving role for cysteine in rapid nanoparticle aggregation. These findings suggest that microbially derived extracellular proteins can limit the dispersal of nanoparticulate metal-bearing phases, such as the mineral products of bioremediation, that may otherwise be transported away from their source by subsurface fluid flow.

  6. Neutrophil extracellular traps in sheep mastitis.

    Pisanu, Salvatore; Cubeddu, Tiziana; Pagnozzi, Daniela; Rocca, Stefano; Cacciotto, Carla; Alberti, Alberto; Marogna, Gavino; Uzzau, Sergio; Addis, Maria Filippa

    2015-01-01

    Neutrophil extracellular traps (NETs) are structures composed of DNA, histones, and antimicrobial proteins that are released extracellularly by neutrophils and other immune cells as a means for trapping and killing invading pathogens. Here, we describe NET formation in milk and in mammary alveoli of mastitic sheep, and provide a dataset of proteins found in association to these structures. Nucleic acid staining, immunomicroscopy and fluorescent in-situ hybridization of mastitic mammary tissue from sheep infected with Streptococcus uberis demonstrated the presence of extranuclear DNA colocalizing with antimicrobial proteins, histones, and bacteria. Then, proteomic analysis by LTQ-Orbitrap Velos mass spectrometry provided detailed information on protein abundance changes occurring in milk upon infection. As a result, 1095 unique proteins were identified, of which 287 being significantly more abundant in mastitic milk. Upon protein ontology classification, the most represented localization classes for upregulated proteins were the cytoplasmic granule, the nucleus, and the mitochondrion, while function classes were mostly related to immune defence and inflammation pathways. All known NET markers were massively increased, including histones, granule proteases, and antimicrobial proteins. Of note was the detection of protein arginine deiminases (PAD3 and PAD4). These enzymes are responsible for citrullination, the post-translational modification that is known to trigger NET formation by inducing chromatin decondensation and extracellular release of NETs. As a further observation, citrullinated residues were detected by tandem mass spectrometry in histones of samples from mastitic animals. In conclusion, this work provides novel microscopic and proteomic information on NETs formed in vivo in the mammary gland, and reports the most complete database of proteins increased in milk upon bacterial mastitis. PMID:26088507

  7. The Effects of Extracellular Matrix on Tissue Engineering Construction of Cartilage in Vitro

    YU Li; LI Fa-tao; TANG Ming-qiao; YAN Wei-qun

    2006-01-01

    The effects of various cartilage extracellular matrix on the construction of rabbit growth plate cartilage tissue in vitro were studied. The results show that collagen, proteoglycan and hyaluronic acid can promote the growth of cultured chondrocytes but the effects of various cartilage extracellular matrix(ECM)on chondrocyte differentiation are different. Collagen can promote the hypertrophy of chondrocytes while proteoglycan and hyaluronic acid inhibit the transition of mature chondrocytes into hypertrophied chondrocytes.

  8. Extracellular enzymes of Legionella pneumophila.

    Thorpe, T C; Miller, R. D.

    1981-01-01

    All strains of Legionella pneumophila tested produced detectable levels of extracellular protease, phosphatase, lipase, deoxyribonuclease, ribonuclease, and beta-lactamase activity. Weak starch hydrolysis was also demonstrated for all strains. Elastase, collagenase, phospholipase C, hyaluronidase, chondroitinase, neuraminidase, or coagulase were not detected in any of these laboratory-maintained strains.

  9. Enzymatic Production of Extracellular Reactive Oxygen Species by Marine Microorganisms

    Diaz, J. M.; Andeer, P. F.; Hansel, C. M.

    2014-12-01

    Reactive oxygen species (ROS) serve as intermediates in a myriad of biogeochemically important processes, including cell signaling pathways, cellular oxidative stress responses, and the transformation of both nutrient and toxic metals such as iron and mercury. Abiotic reactions involving the photo-oxidation of organic matter were once considered the only important sources of ROS in the environment. However, the recent discovery of substantial biological ROS production in marine systems has fundamentally shifted this paradigm. Within the last few decades, marine phytoplankton, including diatoms of the genus Thalassiosira, were discovered to produce ample extracellular quantities of the ROS superoxide. Even more recently, we discovered widespread production of extracellular superoxide by phylogenetically and ecologically diverse heterotrophic bacteria at environmentally significant levels (up to 20 amol cell-1 hr-1), which has introduced the revolutionary potential for substantial "dark" cycling of ROS. Despite the profound biogeochemical importance of extracellular biogenic ROS, the cellular mechanisms underlying the production of this ROS have remained elusive. Through the development of a gel-based assay to identify extracellular ROS-producing proteins, we have recently found that enzymes typically involved in antioxidant activity also produce superoxide when molecular oxygen is the only available electron acceptor. For example, large (~3600 amino acids) heme peroxidases are involved in extracellular superoxide production by a bacterium within the widespread Roseobacter clade. In Thalassiosira spp., extracellular superoxide is produced by flavoproteins such as glutathione reductase and ferredoxin NADP+ reductase. Thus, extracellular ROS production may occur via secreted and/or cell surface enzymes that modulate between producing and degrading ROS depending on prevailing geochemical and/or ecological conditions.

  10. Vitamin A Deficiency and Alterations in the Extracellular Matrix

    Teresa Barber

    2014-11-01

    Full Text Available Vitamin A or retinol which is the natural precursor of several biologically active metabolites can be considered the most multifunctional vitamin in mammals. Its deficiency is currently, along with protein malnutrition, the most serious and common nutritional disorder worldwide. It is necessary for normal embryonic development and postnatal tissue homeostasis, and exerts important effects on cell proliferation, differentiation and apoptosis. These actions are produced mainly by regulating the expression of a variety of proteins through transcriptional and non-transcriptional mechanisms. Extracellular matrix proteins are among those whose synthesis is known to be modulated by vitamin A. Retinoic acid, the main biologically active form of vitamin A, influences the expression of collagens, laminins, entactin, fibronectin, elastin and proteoglycans, which are the major components of the extracellular matrix. Consequently, the structure and macromolecular composition of this extracellular compartment is profoundly altered as a result of vitamin A deficiency. As cell behavior, differentiation and apoptosis, and tissue mechanics are influenced by the extracellular matrix, its modifications potentially compromise organ function and may lead to disease. This review focuses on the effects of lack of vitamin A in the extracellular matrix of several organs and discusses possible molecular mechanisms and pathologic implications.

  11. Separation of xylo-oligosaccharides from enzymatic hydrolytes using membrane reactor

    杨富国; 方正; 徐勇; 姚春才; 余世袁; 朱琼霞

    2003-01-01

    The time course of xylo-oligosaccharides concentration and xylo-oligosaccharides yield in the separation of xylo-oligosaccharides from enzymatic hydrolytes was studied using a membrane reactor with constant permeate flux of 4 L @ m-2 @ h-1. The results show that xylanases retain 90% of its activity in the reactor. The concentration of xylo-oligosaccharides achieves the maximum, about 5.48 g/L in 30 min. The difference of xylo-oligosaccharides in the retentate and permeate stream is low, <0.62 g/L, therefore it can permeate through membrane. Under the operating conditions that xylan concentration is 30.0 g/L, pH 5.0, operating pressure 16 kPa, temperature 48 ℃,feed velocity 400 mL/min, reaction volume 400 mL, enzyme dosage 10%(volume fraction), dilution rate 1 h -1, and enzymatic hydrolysis time 195 min, the yield of xylo-oligosaccharides reaches 31.69 %.

  12. Hydrolytic Activation Kinetics of the Herbicide Benzobicyclon in Simulated Aquatic Systems.

    Williams, Katryn L; Tjeerdema, Ronald S

    2016-06-22

    Herbicide resistance is a growing concern for weeds in California rice fields. Benzobicyclon (BZB; 3-(2-chloro-4-(methylsulfonyl)benzoyl)-2-phenylthiobicyclo[3.2.1]oct-2-en-4-one) has proven successful against resistant rice field weeds in Asia. A pro-herbicide, BZB forms the active agent, benzobicyclon hydrolysate (BH), in water; however, the transformation kinetics are not understood for aquatic systems, particularly flooded California rice fields. A quantitative experiment was performed to assess the primary mechanism and kinetics of BZB hydrolysis to BH. Complete conversion to BH was observed for all treatments. Basic conditions (pH 9) enhanced the reaction, with half-lives ranging from 5 to 28 h. Dissolved organic carbon (DOC) hindered transformation, which is consistent with other base-catalyzed hydrolysis reactions. BH was relatively hydrolytically stable, with 18% maximum loss after 5 days. Results indicate BZB is an efficient pro-herbicide under aqueous conditions such as those of a California rice field, although application may be best suited for fields with recirculating tailwater systems. PMID:27248841

  13. A novel hydrolytic product from flesh of Mactra veneriformis and its bioactivities in calcium supplement

    Wang, Lingchong; Chen, Shiyong; Liu, Rui; Wu, Hao

    2012-09-01

    To prepare calcium-binding peptides, the flesh residue of Mactra Veneriformis was subjected to enzymatic hydrolysis. By comparing the capability of combining calcium of the hydrolyzates, pepsin was confirmed to be the most suitable enzyme for hydrolyzing the flesh residue to release calcium-binding peptides among the seven tested proteases. The pepsin hydrolyzate (PHM) was divided into three fractions according to the molecule weight of its composition, which ranged from 0.5 to 15 kDa. The low-molecule-weight fraction named PHM-3 had the highest capability in combining calcium. The peptides existing in the PHM-3 fraction consisted of higher contents of Glu, Ala and Leu, and could produce one type of calcium-peptide complex by powerfully chelating calcium ions. PHM-3 products could effectively increase calcium absorption and retention while they decreased the calcium excretion in animal tests. Additionally, symptoms caused by low calcium bioavailability in ovariectomized rats, such as bone mineral density reduction and mechanical strength loss could be significantly ameliorated by the hydrolytic products addition in diet.

  14. A Novel Hydrolytic Product from Flesh of Mactra veneriformis and Its Bioactivities in Calcium Supplement

    WANG Lingchong; CHEN Shiyong; LIU Rui; WU Hao

    2012-01-01

    To prepare calcium-binding peptides,the flesh residue of Mactra Veneriformis was subjected to enzymatic hydrolysis.By comparing the capability of combining calcium of the hydrolyzates,pepsin was confirmed to be the most suitable enzyme for hydrolyzing the flesh residue to release calcium-binding peptides among the seven tested proteases.The pepsin hydrolyzate(PHM)was divided into three fractions according to the molecule weight of its composition,which ranged from 0.5 to 15 kDa.The low-molecule-weight fraction named PHM-3 had the highest capability in combining calcium.The peptides existing in the PHM-3fraction consisted of higher contents of Glu,Ala and Leu,and could produce one type of calcium-peptide complex by powerfully chelating calcium ions.PHM-3 products could effectively increase calcium absorption and retention while they decreased the calcium excretion in animal tests.Additionally,symptoms caused by low calcium bioavailability in ovariectomized rats,such as bone mineral density reduction and mechanical strength loss could be significantly ameliorated by the hydrolytic products addition in diet.

  15. An in vitro evaluation of hydrolytic enzymes as dental plaque control agents.

    Ledder, Ruth G; Madhwani, Tejal; Sreenivasan, Prem K; De Vizio, William; McBain, Andrew J

    2009-04-01

    The plaque-control potential of commercially available amylase, lipase and protease was evaluated by observing their effects on coaggregation and on bacterial viability within various plaque microcosms. A quantitative coaggregation assay indicated that protease significantly inhibited the extent of coaggregation of Actinomyces naeslundii and Streptococcus oralis (P naeslundii versus Fusobacterium nucleatum and A. naeslundii versus P. gingivalis. Concomitant challenge of constant-depth film fermenter-grown plaques with the enzymes did not result in detectable ecological perturbations (assessed by differential culture and denaturing gradient gel electrophoresis). Similar dosing and analysis of multiple Sorbarod devices did not reveal increases in bacterial dispersion which could result from disaggregation of extant plaques. A short-term hydroxyapatite colonization model was therefore used to investigate possible enzyme effects on early-stage plaque development. Whilst culture did not indicate significant reductions in adhesion or plaque accumulation, a vital visual assay revealed significantly increased aggregation frequency following enzyme exposure. In summary, although hydrolytic enzymes negatively influenced binary coaggregation, they did not cause statistically significant changes in bacterial viability within plaque microcosms. In contrast, enzyme exposure increased aggregation within extant plaques. PMID:19273645

  16. Lignocellulosic Fermentation of Wild Grass Employing Recombinant Hydrolytic Enzymes and Fermentative Microbes with Effective Bioethanol Recovery

    Saprativ P. Das

    2013-01-01

    Full Text Available Simultaneous saccharification and fermentation (SSF studies of steam exploded and alkali pretreated different leafy biomass were accomplished by recombinant Clostridium thermocellum hydrolytic enzymes and fermentative microbes for bioethanol production. The recombinant C. thermocellum GH5 cellulase and GH43 hemicellulase genes expressed in Escherichia coli cells were grown in repetitive batch mode, with the aim of enhancing the cell biomass production and enzyme activity. In batch mode, the cell biomass (A600 nm of E. coli cells and enzyme activities of GH5 cellulase and GH43 hemicellulase were 1.4 and 1.6 with 2.8 and 2.2 U·mg−1, which were augmented to 2.8 and 2.9 with 5.6 and 3.8 U·mg−1 in repetitive batch mode, respectively. Steam exploded wild grass (Achnatherum hymenoides provided the best ethanol titres as compared to other biomasses. Mixed enzyme (GH5 cellulase, GH43 hemicellulase mixed culture (Saccharomyces cerevisiae, Candida shehatae system gave 2-fold higher ethanol titre than single enzyme (GH5 cellulase single culture (Saccharomyces cerevisiae system employing 1% (w/v pretreated substrate. 5% (w/v substrate gave 11.2 g·L−1 of ethanol at shake flask level which on scaling up to 2 L bioreactor resulted in 23 g·L−1 ethanol. 91.6% (v/v ethanol was recovered by rotary evaporator with 21.2% purification efficiency.

  17. Comparison of biogas sludge and raw crop material as source of hydrolytic cultures for anaerobic digestion.

    Weiß, Stefan; Somitsch, Walter; Klymiuk, Ingeborg; Trajanoski, Slave; Guebitz, Georg M

    2016-05-01

    Mixed fermentative/hydrolytic bacteria were enriched on lignocellulose substrates in minimal medium under semi-anaerobic mesophilic conditions in the presence or absence of natural zeolite as growth supporter to ultimately bioaugment non-adapted sludge and thereby enhance the overall anaerobic digestion (AD) of recalcitrant plant material. Desired enzyme activities, i.e. xylanases and cellulase were monitored during subsequent cultivation cycles. Furthermore, enriched microbial communities were characterized by 16S rRNA-based 454-Pyrosequencing, revealing Firmicutes, Bacteriodetes, Proteobacteria and Spirochaetes to be the predominant bacterial groups in cultures derived from anaerobic sludge and raw crop material, i.e. maple green cut and wheat straw as well. Enriched populations relevant for biopolymer hydrolysis were then compared in biological methane potential tests to demonstrate positive effects on the biogasification of renewable plant substrate material. A significant impact on methane productivity was observed with adapted mixed cultures when used in combination with clinoptilolite to augment and supplement non-adapted bioreactor sludge. PMID:26894564

  18. Inhibitors of the Hydrolytic Enzyme Dimethylarginine Dimethylaminohydrolase (DDAH: Discovery, Synthesis and Development

    Rhys B. Murphy

    2016-05-01

    Full Text Available Dimethylarginine dimethylaminohydrolase (DDAH is a highly conserved hydrolytic enzyme found in numerous species, including bacteria, rodents, and humans. In humans, the DDAH-1 isoform is known to metabolize endogenous asymmetric dimethylarginine (ADMA and monomethyl arginine (l-NMMA, with ADMA proposed to be a putative marker of cardiovascular disease. Current literature reports identify the DDAH family of enzymes as a potential therapeutic target in the regulation of nitric oxide (NO production, mediated via its biochemical interaction with the nitric oxide synthase (NOS family of enzymes. Increased DDAH expression and NO production have been linked to multiple pathological conditions, specifically, cancer, neurodegenerative disorders, and septic shock. As such, the discovery, chemical synthesis, and development of DDAH inhibitors as potential drug candidates represent a growing field of interest. This review article summarizes the current knowledge on DDAH inhibition and the derived pharmacokinetic parameters of the main DDAH inhibitors reported in the literature. Furthermore, current methods of development and chemical synthetic pathways are discussed.

  19. (R,S)-2-chlorophenoxyl pyrazolides as novel substrates for improving lipase-catalyzed hydrolytic resolution.

    Kao, Min-fang; Lu, Pei-yu; Kao, Jou-yan; Wang, Pei-yun; Wu, An-chi; Tsai, Shau-Wei

    2012-01-01

    The best reaction condition of Candida antartica lipase B as biocatalyst, 3-(2-pyridyl)pyrazole as leaving azole, and water-saturated methyl t-butyl ether as reaction medium at 45°C were first selected for performing the hydrolytic resolution of (R,S)-2-(4-chlorophenoxyl) azolides (1-4). In comparison with the kinetic resolution of (R,S)-2-phenylpropionyl 3-(2-pyridyl)pyrazolide or (R,S)-α-methoxyphenylacetyl 3-(2-pyridyl)pyrazolide at the same reaction condition, excellent enantioselectivity with more than two order-of-magnitudes higher activity for each enantiomer was obtained. The resolution was then extended to other (R,S)-3-(2-pyridyl)pyrazolides (5-7) containing 2-chloro, 3-chloro, or 2,4-dichloro substituent, giving good (E > 48) to excellent (E > 100) enantioselectivity. The thermodynamic analysis for 1, 2, and 4-7 demonstrates profound effects of the acyl or leaving moiety on varying enthalpic and entropic contributions to the difference of Gibbs free energies. A thorough kinetic analysis further indicates that on the basis of 6, the excellent enantiomeric ratio for 4 and 7 is due to the higher reactivity of (S)-4 and lower reactivity of (R)-7, respectively. PMID:22012845

  20. Evolution and Distribution of Hydrolytic Enzyme Activities during Preharvest Sprouting of Wheat (Triticum aestivum) in the Field.

    Olaerts, Heleen; Roye, Chiara; Derde, Liesbeth J; Sinnaeve, Georges; Meza, Walter R; Bodson, Bernard; Courtin, Christophe M

    2016-07-20

    To date, research on preharvest sprouted (PHS) wheat has mostly been conducted on kernels germinated under laboratory conditions, which differ widely from conditions in the field. To obtain detailed knowledge of the evolution of hydrolytic enzyme activities in PHS wheat (Triticum aestivum), a broad collection of samples from three varieties was obtained by harvesting before, at, and after maturity. Delaying harvest time coupled with periods of heavy rainfall caused sprouting in the kernels, observed as a drop in Falling Number and an increase in α-amylase activity. The appearance of α- and β-amylase, peptidase, and endoxylanase activity during field sprouting was independent from each other. Consequently, Falling Number could not be used to predict activity of other hydrolytic enzymes. When differentiating endogenous from kernel-associated microbial enzymes, results showed that α- and β-amylase and peptidase activity of PHS kernels were predominantly of endogenous origin, whereas endoxylanase activity was largely from microbial origin. PMID:27341479

  1. Biochar amendment to lead-contaminated soil: Effects on fluorescein diacetate hydrolytic activity and phytotoxicity to rice.

    Tan, Xiaofei; Liu, Yunguo; Gu, Yanling; Zeng, Guangming; Hu, Xinjiang; Wang, Xin; Hu, Xi; Guo, Yiming; Zeng, Xiaoxia; Sun, Zhichao

    2015-09-01

    The amendment effects of biochar on total microbial activity was measured by fluorescein diacetate (FDA) hydrolytic activity, and phytotoxicity in Pb(II)-contaminated soils was examined by the application of 4 different biochars to soil, with rice as a test plant. The FDA hydrolytic activities of biochar-amended soils were much higher than that of the control. The survival rate of rice in lead-contaminated biochar-amended soils showed significant improvement over the control, especially for bamboo biochar-amended soil (93.3%). In addition, rice grown in lead-contaminated control sediment displayed lower biomass production than that in biochar-amended soil. The immobilization of Pb(II) and the positive effects of biochar amendment on soil microorganisms may account for these effects. The results suggest that biochar may have an excellent ability to mitigate the toxic effects of Pb(II) on soil microorganisms and rice. PMID:25900615

  2. Preventing hypoxia-induced cell death in beta cells and islets via hydrolytically activated, oxygen-generating biomaterials

    Pedraza, Eileen; Coronel, Maria M.; Fraker, Christopher A.; Ricordi, Camillo; Stabler, Cherie L

    2012-01-01

    A major hindrance in engineering tissues containing highly metabolically active cells is the insufficient oxygenation of these implants, which results in dying or dysfunctional cells in portions of the graft. The development of methods to increase oxygen availability within tissue-engineered implants, particularly during the early engraftment period, would serve to allay hypoxia-induced cell death. Herein, we designed and developed a hydrolytically activated oxygen-generating biomaterial in t...

  3. PREPARATION AND PROPERTIES OF EXTRACELLULAR BIOPOLYMER FLOCCULANT

    2007-01-01

    The biopolymer flocculant (named PS-2) producing by Pseudomonas fluorescens was investigated. The PS-2 had high efficiency with small dosage, when dealing with kaolin suspension, formed larger floc, with big sedimentation rate, over a wide range of temperatures. Distributing of flocculating activity test showed that the biopolymer flocculant was an extracellular product. The composition analysis of purified biopolymer flocculant showed that it composed mainly of polysaccharide and nucleic acid. The content of polysaccharide was 86.7%, which determined by using phenol-vitriol method, and the content of nucleic acid was 7.8%, which determined by UV absorption method. The biopolymer flocculant as a powder form showed much better stability than that as a supernatant. The character of biopolymer flocculant was stable even it was heated to 100℃ when it in acidic condition. The optimal conditions to flocculate kaolin suspension were as follows: pH 8~12, flocculant dosage 1mL/L, and Ca2+ as the optimal cation.

  4. PREPARATION AND PROPERTIES OF EXTRACELLULAR BIOPOLYMER FLOCCULANT

    LI Chunxiang; LIU Binbin; XIONG Jinshui; YAN Jingchun

    2007-01-01

    The biopolymer flocculant (named PS-2) producing by Pseudomonas fluorescens was investigated. The PS-2 had high efficiency with small dosage, when dealing with kaolin suspension,formed larger floc, with big sedimentation rate, over a wide range of temperatures. Distributing of flocculating activity test showed that the biopolymer flocculant was an extracellular product. The composition analysis of purified biopolymer flocculant showed that it composed mainly of polysaccharide and nucleic acid. The content of polysaccharide was 86.7%, which determined by using phenol-vitriol method, and the content of nucleic acid was 7.8%, which determined by UV absorption method. The biopolymer flocculant as a powder form showed much better stability than that as a supernatant. The character of biopolymer flocculant was stable even it was heated to 100 ℃ when it in acidic condition. The optimal conditions to flocculate kaolin suspension were as follows:pH 8~12, flocculant dosage 1mL/L, and Ca2+ as the optimal cation.

  5. Extracellular enzyme kinetics scale with resource availability

    Microbial community metabolism relies on external digestion, mediated by extracellular enzymes that break down complex organic matter into molecules small enough for cells to assimilate. We analyzed the kinetics of 40 extracellular enzymes that mediate the degradation and assimi...

  6. Solid Heteropolyacids (HPAs) in Hydrolytic Conversion of Biomass%固体杂多酸在生物质水解转化中的应用

    张建明; 翟尚儒; 黄德智; 翟滨; 安庆大

    2012-01-01

    With global oil production flattening out, attention is being increasingly paid to a kind of renewable clean energy--biomass. Heteropolyacids are important catalysts in the so-called clean technologies. They possess strong acidity, structural flexibility and fairly high thermal stability. It would be preferable to carry out the heteropolyacids-catalyzed reaction in biomass hydrolytic conversion. The performance of heteropolyacids towards hydrolysis of biomass in pure water, organic solvents and biphasic systems exhibit different advantages and limitations. In this paper, we reviewed the latest progress in the hydrolytic conversion of biomass into valuable chemicals using heteropolyacids in different catalytic systems. Highly effective utilization of biomass has positive effects on solving energy problems and achieving sustainable development of energy and chemical industry. Heteropolyacids used as excellent green catalyst will possess extensive application prospect in biomass conversion.%随着化石资源的日益枯竭,寻求可替代清洁能源已成为全球重大课题。生物质是一种可再生的清洁能源,目前人们尝试通过利用生物质转化缓解日益增长的能源需求。杂多酸是应用在清洁工艺中的重要催化剂,结构和酸度的设计调变性及较高的热稳定性,使其广泛用于生物质的水解转化反应平台。目前固体杂多酸在水溶剂、有机溶剂及两相体系中降解生物质有着各自不同的优缺点。本文综述了杂多酸在不同反应体系中水解转化生物质制备精细化学品的研究进展,并对其在生物质水解转化利用中的应用前景进行了展望。

  7. A field pilot-scale study of biological treatment of heavy oil-produced water by biological filter with airlift aeration and hydrolytic acidification system.

    Zhang, Min; Wang, Junming; Zhang, Zhongzhi; Song, Zhaozheng; Zhang, Zhenjia; Zhang, Beiyu; Zhang, Guangqing; Wu, Wei-Min

    2016-03-01

    Heavy oil-produced water (HOPW) is a by-product during heavy oil exploitation and can cause serious environmental pollution if discharged without adequate treatment. Commercial biochemical treatment units are important parts of HOPW treatment processes, but many are not in stable operation because of the toxic and refractory substances, salt, present. Therefore, pilot-scale experiments were conducted to evaluate the performance of hydrolytic acidification-biological filter with airlift aeration (HA-BFAA), a novel HOPW treatment system. Four strains isolated from oily sludge were used for bioaugmentation to enhance the biodegradation of organic pollutants. The isolated bacteria were evaluated using 3-day biochemical oxygen demand, oil, dodecyl benzene sulfonic acid, and chemical oxygen demand (COD) removals as evaluation indices. Bioaugmentation enhanced the COD removal by 43.5 mg/L under a volume load of 0.249 kg COD/m(3) day and hydraulic retention time of 33.6 h. The effluent COD was 70.9 mg/L and the corresponding COD removal was 75.0 %. The optimum volumetric air-to-water ratio was below 10. The removal ratios of the total extractable organic pollutants, alkanes, and poly-aromatic hydrocarbons were 71.1, 94.4, and 94.0 %, respectively. Results demonstrated that HA-BFAA was an excellent HOPW treatment system. PMID:26549708

  8. Extracellular matrix in ovarian follicles.

    Rodgers, R J; Irving-Rodgers, H F; van Wezel, I L

    2000-05-25

    A lot is known about the control of the development of ovarian follicles by growth factors and hormones, but less is known about the roles of extracellular matrix in the control of follicular growth and development. In this review we focus on the specialized extracellular matrix of the basal laminas that are present in ovarian follicles. These include the follicular basal lamina itself, the Call-Exner bodies of the membrana granulosa, the subendothelial and arteriole smooth muscle basal laminas in the theca, and the basal lamina-like material of the thecal matrix. We discuss the evidence that during follicle development the follicular basal lamina changes in composition, that many of its components are produced by the granulosa cells, and that the follicular basal laminas of different follicles have different ultrastructural appearances, linked to the shape of the aligning granulosa cells. All these studies suggest that the follicular basal lamina is extremely dynamic during follicular development. PMID:10963877

  9. Extracellular metalloproteinases in Phytomonas serpens.

    Vermelho, Alane B; Almeida, Flávia V S; Bronzato, Leandro S; Branquinha, Marta H

    2003-03-01

    The detection of extracellular proteinases in Phytomonas serpens, a trypanosomatid isolated from tomato fruits, is demonstrated in this paper. Maximal production occurred at the end of the logarithmic phase of growth. These enzymes exhibited selective substrate utilization in SDS-PAGE, being more active with gelatin; hemoglobin and bovine serum albumin were not degraded. Three proteinases were detected in SDS-PAGE-gelatin, with apparent molecular masses between 94 and 70 kDa. The proteolytic activity was completely blocked by 1,10-phenanthroline and strongly inhibited by EDTA, whereas a partial inhibition was observed with trans-epoxysuccinyl-L-leucylamido-(4-guanidino) butane (E-64) and soybean trypsin inhibitor; phenylmethylsulfonyl fluoride weakly inhibited the enzymes. This inhibition profile indicated that these extracellular proteinases belong to the metalloproteinase class. PMID:12795409

  10. Immobilization of cholesterol esterase in mesoporous silica materials and its hydrolytic activity toward diethyl phthalate

    Cholesterol esterase (CE, cholesteryl ester hydrolase, EC 3.1.1.13) from porcine pancreas (molecular weight 400–500 kDa) exhibits hydrolytic activity toward various toxic organic phthalate esters. CE was confined in the nanospace (diameter 3–30 nm) of five types of mesoporous silica (MPS) that differ in structural properties such as pore diameter, pore volume, and particle morphology. These structural properties were characterized by transmission electron microscopy, small-angle X-ray diffraction, N2 adsorption–desorption experiments, solid-state 13C nuclear magnetic resonance (NMR), and solid-state 29Si NMR. Catalytic activities of immobilized and free CE were evaluated by the hydrolysis of diethyl phthalate in phosphate buffer solutions containing an organic cosolvent. Optimal activity recovery was achieved when CE was immobilized in n-decane-functionalized MPS, which had a large pore size (22.5 nm). The immobilization also protected against effects of temperature within the range 30 °C–60 °C; CE immobilized in n-decyl-functionalized MPS exhibited better thermal stability than in non-functionalized MPS or free CE. Moreover, it retained approximately 60% of its catalytic activity even after six catalytic cycles. - Highlights: ► The highest activity of immobilized CE was shown in MPS with a pore size of 22.5 nm. ► Catalytic efficiency improved when MPS was functionalized by n-decyl substitution. ► Immobilized CE exhibited good thermal stability and reusability. ► Organic co-solvent and the substrate structures affected enzyme activities.