WorldWideScience

Sample records for cellulase

  1. Engineering Cellulases for Biorefinery

    Energy Technology Data Exchange (ETDEWEB)

    Manoj Kumar, PhD

    2010-06-27

    Lignocellulosic biomass is the most abundant, least expensive renewable natural biological resource for the production of biobased products and bioenergy is important for the sustainable development of human civilization in 21st century. For making the fermentable sugars from lignocellulosic biomass, a reduction in cellulase production cost, an improvement in cellulase performance, and an increase in sugar yields are all vital to reduce the processing costs of biorefineries. Improvements in specific cellulase activities for non-complexed cellulase mixtures can be implemented through cellulase engineering based on rational design or directed evolution for each cellulase component enzyme, as well as on the reconstitution of cellulase components. In this paper, we will provide DSM's efforts in cellulase research and developments and focus on limitations. Cellulase improvement strategies based on directed evolution using screening on relevant substrates, screening for higher thermal tolerance based on activity screening approaches such as continuous culture using insoluble cellulosic substrates as a powerful selection tool for enriching beneficial cellulase mutants from the large library. We will illustrate why and how thermostable cellulases are vital for economic delivery of bioproducts from cellulosic biomass using biochemical conversion approach.

  2. Thermostable Cellulases: Why & How?

    Energy Technology Data Exchange (ETDEWEB)

    Manoj Kumar, PhD

    2010-04-19

    Lignocellulosic biomass is the most abundant, least expensive renewable natural biological resource for the production of biobased products and bioenergy is important for the sustainable development of human civilization in 21st century. For making the fermentable sugars from lignocellulosic biomass, a reduction in cellulase production cost, an improvement in cellulase performance, and an increase in sugar yields are all vital to reduce the processing costs of biorefineries. Improvements in specific cellulase activities for non-complexed cellulase mixtures can be implemented through cellulase engineering based on rational design or directed evolution for each cellulase component enzyme, as well as on the reconstitution of cellulase components. In this paper, we will provide DSM's efforts in cellulase research and developments and focus on limitations. Cellulase improvement strategies based on directed evolution using screening on relevant substrates, screening for higher thermal tolerance based on activity screening approaches such as continuous culture using insoluble cellulosic substrates as a powerful selection tool for enriching beneficial cellulase mutants from the large library. We will illustrate why and how thermostable cellulases are vital for economic delivery of bioproducts from cellulosic biomass using biochemical conversion approach.

  3. Thermostable Cellulases: Why & How?

    Energy Technology Data Exchange (ETDEWEB)

    Manoj Kumar, PhD

    2010-03-24

    Lignocellulosic biomass is the most abundant, least expensive renewable natural biological resource for the production of biobased products and bioenergy is important for the sustainable development of human civilization in 21st century. For making the fermentable sugars from lignocellulosic biomass, a reduction in cellulase production cost, an improvement in cellulase performance, and an increase in sugar yields are all vital to reduce the processing costs of biorefineries. Improvements in specific cellulase activities for non-complexed cellulase mixtures can be implemented through cellulase engineering based on rational design or directed evolution for each cellulase component enzyme, as well as on the reconstitution of cellulase components. In this paper, we will provide DSM's efforts in cellulase research and developments and focus on limitations. Cellulase improvement strategies based on directed evolution using screening on relevant substrates, screening for higher thermal tolerance based on activity screening approaches such as continuous culture using insoluble cellulosic substrates as a powerful selection tool for enriching beneficial cellulase mutants from the large library. We will illustrate why and how thermostable cellulases are vital for economic delivery of bioproducts from cellulosic biomass using biochemical conversion approach.

  4. Measurement of saccharifying cellulase

    Directory of Open Access Journals (Sweden)

    Andreotti Raymond

    2009-09-01

    Full Text Available Abstract This article sets forth a simple cellulase assay procedure. Cellulose is variable in nature, insoluble and resistant to enzymatic attack. As a result there have been a bevy of bewildering cellulase assays published that yielded irrational results. Certain protocols focused on the rapidity of the assay while ignoring that only the most readily susceptible cellulose regions were being hydrolyzed. Other assays simplified the system by using modified soluble substrates and yielded results that bore no relationship to the real world hydrolysis of insoluble cellulose. In this study Mandels, Andreotti and Roche utilized a common substrate, Whatman filter paper. Hydrolysis of a 50 mg sample of the paper was followed to roughly 4% degradation, which circumvented the problems of attack of only the most susceptible zones. This common hydrolysis target range also resulted in some balance with regard to the interaction of the several cellulase components. The method was subsequently widely adopted. Douglas E Eveleigh

  5. Chimeric enzymes with improved cellulase activities

    Science.gov (United States)

    Xu, Qi; Baker, John O; Himmel, Michael E

    2015-03-31

    Nucleic acid molecules encoding chimeric cellulase polypeptides that exhibit improved cellulase activities are disclosed herein. The chimeric cellulase polypeptides encoded by these nucleic acids and methods to produce the cellulases are also described, along with methods of using chimeric cellulases for the conversion of cellulose to sugars such as glucose.

  6. Cellulase hydrolysis of unsorted MSW

    DEFF Research Database (Denmark)

    Jensen, Jacob Wagner; Felby, Claus; Jørgensen, Henning

    2011-01-01

    of calcium, potassium, sodium, chloride and others that may affect cellulolytic enzymes. Cellulase performance showed no effect of adding the metal ion-chelating agent EDTA to the solution. The cellulases were stable, tolerated and functioned in the presence of several contaminants....

  7. Bioprospecting thermophiles for cellulase production: a review

    Directory of Open Access Journals (Sweden)

    Somen Acharya

    2012-09-01

    Full Text Available Most of the potential bioprospecting is currently related to the study of the extremophiles and their potential use in industrial processes. Recently microbial cellulases find applications in various industries and constitute a major group of industrial enzymes. Considerable amount of work has been done on microbial cellulases, especially with resurgence of interest in biomass ethanol production employing cellulases and use of cellulases in textile and paper industry. Most efficient method of lignocellulosic biomass hydrolysis is through enzymatic saccharification using cellulases. Significant information has also been gained about the physiology of thermophilic cellulases producers and process development for enzyme production and biomass saccharification. The review discusses the current knowledge on cellulase producing thermophilic microorganisms, their physiological adaptations and control of cellulase gene expression. It discusses the industrial applications of thermophilic cellulases, their cost of production and challenges in cellulase research especially in the area of improving process economics of enzyme production.

  8. Biotechnological applications of bacterial cellulases

    Directory of Open Access Journals (Sweden)

    Esther Menendez

    2015-08-01

    Full Text Available Cellulases have numerous applications in several industries, including biofuel production, food and feed industry, brewing, pulp and paper, textile, laundry, and agriculture.Cellulose-degrading bacteria are widely spread in nature, being isolated from quite different environments. Cellulose degradation is the result of a synergic process between an endoglucanase, an exoglucanase and a,β-glucosidase. Bacterial endoglucanases degrade ß-1,4-glucan linkages of cellulose amorphous zones, meanwhile exoglucanases cleave the remaining oligosaccharide chains, originating cellobiose, which is hydrolyzed by ß-glucanases. Bacterial cellulases (EC 3.2.1.4 are comprised in fourteen Glycosil Hydrolase families. Several advantages, such as higher growth rates and genetic versatility, emphasize the suitability and advantages of bacterial cellulases over other sources for this group of enzymes. This review summarizes the main known cellulolytic bacteria and the best strategies to optimize their cellulase production, focusing on endoglucanases, as well as it reviews the main biotechnological applications of bacterial cellulases in several industries, medicine and agriculture.

  9. Exo-endo cellulase fusion protein

    Science.gov (United States)

    Bower, Benjamin S.; Larenas, Edmund A.; Mitchinson, Colin

    2012-01-17

    The present invention relates to a heterologous exo-endo cellulase fusion construct, which encodes a fusion protein having cellulolytic activity comprising a catalytic domain derived from a fungal exo-cellobiohydrolase and a catalytic domain derived from an endoglucanase. The invention also relates to vectors and fungal host cells comprising the heterologous exo-endo cellulase fusion construct as well as methods for producing a cellulase fusion protein and enzymatic cellulase compositions.

  10. Highly Efficient Thermostable DSM Cellulases: Why & How?

    Energy Technology Data Exchange (ETDEWEB)

    Manoj Kumar, PhD

    2011-04-26

    Lignocellulosic biomass is the most abundant, least expensive renewable natural biological resource for the production of biobased products and bioenergy is important for the sustainable development of human civilization in 21st century. For making the fermentable sugars from lignocellulosic biomass, a reduction in cellulase production cost, an improvement in cellulase performance, and an increase in sugar yields are all vital to reduce the processing costs of biorefineries. Improvements in specific cellulase activities for non-complexed cellulase mixtures can be implemented through cellulase engineering based on rational design or directed evolution for each cellulase component enzyme, as well as on the reconstitution of cellulase components. In this paper, we will provide DSM's efforts in cellulase research and developments and focus on limitations. Cellulase improvement strategies based on directed evolution using screening on relevant substrates, screening for higher thermal tolerance based on activity screening approaches such as continuous culture using insoluble cellulosic substrates as a powerful selection tool for enriching beneficial cellulase mutants from the large library. We will illustrate why and how thermostable cellulases are vital for economic delivery of bioproducts from cellulosic biomass using biochemical conversion approach.

  11. Engineering Cellulase Enzymes for Bioenergy

    Science.gov (United States)

    Atreya, Meera Elizabeth

    Sustainable energy sources, such as biofuels, offer increasingly important alternatives to fossil fuels that contribute less to global climate change. The energy contained within cellulosic biofuels derives from sunlight energy stored in the form of carbon-carbon bonds comprising sugars such as glucose. Second-generation biofuels are produced from lignocellulosic biomass feedstocks, including agricultural waste products and non-food crops like Miscanthus, that contain lignin and the polysaccharides hemicellulose and cellulose. Cellulose is the most abundant biological material on Earth; it is a polymer of glucose and a structural component of plant cell walls. Accessing the sugar is challenging, as the crystalline structure of cellulose resists degradation; biochemical and thermochemical means can be used to depolymerize cellulose. Cellulase enzymes catalyze the biochemical depolymerization of cellulose into glucose. Glucose can be used as a carbon source for growth of a biofuel-producing microorganism. When it converts glucose to a hydrocarbon fuel, this microbe completes the biofuels process of transforming sunlight energy into accessible, chemical energy capable of replacing non-renewable transportation fuels. Due to strong intermolecular interactions between polymer chains, cellulose is significantly more challenging to depolymerize than starch, a more accessible polymer of glucose utilized in first-generation biofuels processes (often derived from corn). While most mammals cannot digest cellulose (dietary fiber), certain fungi and bacteria produce cellulase enzymes capable of hydrolyzing it. These organisms secrete a wide variety of glycoside hydrolase and other classes of enzymes that work in concert. Because cellulase enzymes are slow-acting and expensive to produce, my aim has been to improve the properties of these enzymes as a means to make a cellulosic biofuels process possible that is more efficient and, consequently, more economical than current

  12. Reaction mechanism of dicofol removal by cellulase.

    Science.gov (United States)

    Wang, Ziyuan; Yang, Ting; Zhai, Zihan; Zhang, Boya; Zhang, Jianbo

    2015-10-01

    It remains unclear whether dicofol should be defined as a persistent organic pollutant. Its environmental persistence has gained attention. This study focused on its degradation by cellulase. Cellulase was separated using a gel chromatogram, and its degradation activity towards dicofol involved its endoglucanase activity. By analyzing the kinetic parameters of cellulase reacting with mixed substrates, it was shown that cellulase reacted on dicofol and carboxyl methyl cellulose through two different active centers. Thus, the degradation of dicofol was shown to be an oxidative process by cellulase. Next, by comparing the impacts of tert-butyl alcohol (a typical OH free-radical inhibitor) on the removal efficiencies of dicofol under both cellulase and Fenton reagent systems, it was shown that the removal of dicofol was initiated by OH free radicals produced by cellulase. Finally, 4,4'-dichloro-dibenzophenone and chloride were detected using gas chromatography mass spectrometry and ion chromatography analysis, which supported our hypothesis. The reaction mechanism was analyzed and involved an attack by OH free radicals at the orthocarbon of dicofol, resulting in the degradation product 4,4'-dichloro-dibenzophenone. PMID:26456602

  13. The realm of cellulases in biorefinery development.

    Science.gov (United States)

    Chandel, Anuj K; Chandrasekhar, G; Silva, Messias Borges; Silvério da Silva, Silvio

    2012-09-01

    Geopolitical concerns (unstable supply of gasoline, environmental pollution, and regular price hikes), economic, and employment concerns have been prompting researchers, entrepreneurs, and policy makers to focus on harnessing the potential of lignocellulosic feedstock for fuel ethanol production and its commercialization. The carbohydrate skeleton of plant cell walls needs to be depolymerised into simpler sugars for their application in fermentation reactions as a chief carbon source of suitable ethnologic strains for ethanol production. The role of cellulolytic enzymes in the degradation of structural carbohydrates of the plant cell wall into ready-to-fermentable sugar stream is inevitable. Cellulase synergistically acts upon plant cell wall polysaccharides to release glucose into the liquid media. Cellulase predominantly dominates all the plant cell wall degrading enzymes due to their vast and diverse range of applications. Apart from the major applications of cellulases such as in detergent formulations, textile desizing, and development of monogastric feed for ruminants, their role in biorefinery is truly remarkable. This is a major area where new research tools based upon fermentation based formulations, biochemistry, and system biology to expedite the structure-function relationships of cellulases including cellulosomes and new designer enzymatic cocktails are required. In the last two decades, a considerable amount of research work has been performed on cellulases and their application in biomass saccharification. However, there are still technical and economic impediments to the development of an inexpensive commercial cellulase production process. Advancements in biotechnology such as screening of microorganisms, manipulation of novel cellulase encoding traits, site-specific mutagenesis, and modifications to the fermentation process could enhance the production of cellulases. Commercially, cheaper sources of carbohydrates and modified fermentation

  14. Hidden cellulases in termites: revision of an old hypothesis

    OpenAIRE

    Tokuda, Gaku; Watanabe, Hirofumi

    2007-01-01

    The intestinal flagellates of termites produce cellulases that contribute to cellulose digestion of their host termites. However, 75% of all termite species do not harbour the cellulolytic flagellates; the endogenous cellulase secreted from the midgut tissue has been considered a sole source of cellulases in these termites. Using the xylophagous flagellate-free termites Nasutitermes takasagoensis and Nasutitermes walkeri, we successfully solubilized cellulases present in the hindgut pellets. ...

  15. Cellulase Inhibition by High Concentrations of Monosaccharides

    DEFF Research Database (Denmark)

    Hsieh, Chia-Wen; Cannella, David; Jørgensen, Henning;

    2014-01-01

    that low free water availability contributes to cellulase inhibition. Of the hydrolytic enzymes involved, those acting on the cellulose substrate, that is, exo- and endoglucanases, were the most inhibited. The β -glucosidases were shown to be less sensitive to high monosaccharide concentrations except......Biological degradation of biomass on an industrial scale culminates in high concentrations of end products. It is known that the accumulation of glucose and cellobiose, end products of hydrolysis, inhibit cellulases and decrease glucose yields. Aside from these end products, however, other...

  16. A steady state theory for processive cellulases

    DEFF Research Database (Denmark)

    Cruys-Bagger, Nicolaj; Olsen, Jens Elmerdahl; Præstgaard, Eigil;

    2013-01-01

    remains to be fully developed. In this paper, we suggest a deterministic kinetic model that relies on a processive set of enzyme reactions and a quasi steady-state assumption. It is shown that this approach is practicable in the sense that it leads to mathematically simple expressions for the steady....... This has significant kinetic implications, for example the maximal specific rate (Vmax/E0) for processive cellulases is much lower than the catalytic rate constant (kcat). We discuss how relationships based on this theory may be used in both comparative and mechanistic analyses of cellulases....

  17. CELLULASES FROM THE BASIDIO - MYCETES CULTURAL LIQUID

    Directory of Open Access Journals (Sweden)

    К. G. Dreval

    2013-04-01

    Full Text Available Adsorption of cellulases on substrate taking place during the cultivation process was determined. Adsorbed enzymes can be eluted by buffer solution with high ionic strength, but for determine their activity they should be transferred into the aqueous solution. On the basis of the results a method for obtaining of cellulases preparations from cultural liquids of basidiomycetes was developed. This method is the elution of cellulases from the cultivation substrate of basidiomycetes. It was found that using of the last allows to obtain enzymatic preparations with a high degree of purification in 3 stages (salting out of proteins — dialysis — gelchromatography. Cellulase preparations received original products of basidiomycetes strains К-1, А-Дон-02, Д-1 Irpex lacteus and AnSc-1 Daedaleopsis confragosa f. confragosa were obtained. They contained different proteins, enzymes with specific peaks out of column and their activity. However, common to them was a distinct maximum of outing from the column by endoglucanases or cellobiases, which may indicate that the studied cellulolytic complexes of basidiomycetes do not contain multiple forms of cellulases with different molecular mass. This method allowed to obtain preparations with different degree of purification in comparing with the original culture filtrate 7,3 for endoglucanase and 33,3 for cellobiase of strain А-Дон-02 I. lacteus; 13,1 for endoglucanase and 25,5 for cellobiase of strain Д-1 I. lacteus; 29,9 for endoglucanase and 90,1 for cellobiase of strain К-1 I. lacteus; 2,1 for endoglucanase and 30,6 for cellobiase of strain AnSc-1 D. confragosa f. confragosa.

  18. Product inhibition of five Hypocrea jecorina cellulases

    DEFF Research Database (Denmark)

    Murphy, Leigh; Westh, Peter; Bohlin, Christina;

    2013-01-01

    Product inhibition of cellulolytic enzymes has been deemed a critical factor in the industrial saccharification of cellulosic biomass. Several investigations have addressed this problem using crude enzyme preparations or commercial (mixed) cellulase products, but quantitative information...... cellulose may be monitored by calorimetry. The key advantage of this approach is that it directly measures the rate of hydrolysis while being essentially blind to the background of added product. We investigated the five major cellulases from Hypocrea jecorina (anamorph: Tricoderma reesei), Cel7A (formerly...... by cellobiose, but showed the highest sensitivity to glucose among all investigated enzymes. The endoglucanases Cel12A and Cel7B were moderately inhibited by cellobiose (IC50 = 60–80 mM), and weakly inhibited by glucose (IC50 = 350–380 mM). The highest resistance to both products was found for Cel5A, which...

  19. Continous monitoring of cellulase action on microcrystalline cellulose

    Energy Technology Data Exchange (ETDEWEB)

    Kremer, S.M.; Wood, P.M. (Bristol Univ. (United Kingdom). Dept. of Biochemistry)

    1992-09-01

    Cellobiose oxidase from Phanerochaete chrysosporium was used for continuous monitoring of cellulase action on microcrystalline cellulose (Avicel). Two protocols are described, the parameter monitored being either the decline in electrode potential as ferricyanide is reduced or consumption of dioxygen. Most experiments used a commercial cellulase preparation from Trichoderma reesei and ferricyanide as acceptor. Within 1 min of an addition of cellulase, ferricyanide reduction reached a steady rate. This was converted into a rate of production of substrate for cellobiose oxidase, in [mu]mol.min[sup -1]. Experiments were conducted either with a constant concentration of cellulase and increasing Avicel, or with constant Avicel and increasing cellulase. Kinetic analysis of the experiments with constant cellulase indicated a K[sub m] of 4.8 [+-] 1.0 (g cellulose).1[sup -1], which was close to the value predicted from binding studies. The specific activity of the cellulase was measured as 375 [+-] 25 [mu]mol.(g cellulase)[sup -1].min[sup -1] in experiments with a high cellulose concentration, but was less than half this value when the cellulose was saturated with cellulase. The maximal rate of cellulose degradation was 9.6 [+-] 1.3 [mu]mol.(g cellulose)[sup -1].min[sup -1]. (orig.).

  20. HYDROLYSIS OF PAPER-DISHWARE WASTES BY CELLULASE

    Institute of Scientific and Technical Information of China (English)

    JieLu; ShulanShi; RunanYang; FuzhengLiang

    2004-01-01

    The optimum conditions of hydrolysis of cellulosic wastes by cellulase were studied. The results show that the optimum conditions of sulfuric acid pretreatment were sulfuric acid consistency 0.3M, pretreatment temperature 100℃, pretreatment time 4hours. After sulfuric acid pretreatment, the optimum conditions of hydrolysis by cellulase were enzymatic temperature 50℃, enzymatic time 48hours,pH4.8,the charge of cellulase 100IU/g and the substraste consistency 60g/l. Meanwhile this paper studies that the structural change of cellulose during sulfuric acid pretreatment and cellulase hydrolysis by analyzing the infrared spectra.

  1. HYDROLYSIS OF PAPER-DISHWARE WASTES BY CELLULASE

    Institute of Scientific and Technical Information of China (English)

    Jie Lu; Shulan Shi; Runan Yang; Fuzheng Liang

    2004-01-01

    The optimum conditions of hydrolysis of cellulosic wastes by cellulase were studied. The results show that the optimum conditions of sulfuric acid pretreatment were sulfuric acid consistency 0.3M,pretreatment temperature 100℃, pretreatment time 4hours. After sulfuric acid pretreatment, the optimum conditions of hydrolysis by cellulase were enzymatic temperature 50℃ ,enzymatic time 48hours,pH4.8,the charge of cellulase 100IU/g and the substraste consistency 60g/l. Meanwhile this paper studies that the structural change of cellulose during sulfuric acid pretreatment and cellulase hydrolysis by analyzing the infrared spectra.

  2. Comparison of the thermostability of cellulases from various thermophilic fungi

    Energy Technology Data Exchange (ETDEWEB)

    Wojtczak, G.; Breuil, C.; Yamada, J.; Saddler, J.N.

    1987-10-01

    The cellulase activities of six thermophilic fungi were compared. Although the thermophilic fungi grew at relatively high temperatures (> 45/sup 0/C) the optimum temperatures for assaying the various cellulase activities were only slightly higher than the optimum temperatures for the mesophilic fungi, Trichoderma harzianum. Over prolonged incubation (> 24 h) the thermophilic strains demonstrated a higher hydrolytic potential as a result of the greater thermostability of the cellulase components. Although the extracellular cellulase activities had similar pH and temperature optima, in some cases the thermostability of the extracellular components were considerably lower.

  3. Xyloglucan oligosaccharides promote growth and activate cellulase: Evidence for a role of cellulase in cell expansion

    International Nuclear Information System (INIS)

    Oligosaccharides produced by the action of fungal cellulase on xyloglucans promoted the elongation of etiolated pea (Pisum sativum L.) stem segments in a straight-growth bioassay designed for the determination of auxins. The oligosaccharides were most active at about 1 micromolar. We tested the relative growth-promoting activities of four HPLC-purified oligosaccharides which shared a common glucose4·xylose3 (XG7) core. The substituted oligosaccharides XG8 (glucose4·xylose3·galactose) and XG9n (glucose4·xylose3·galactose2) were more effective than XG7 itself and XG9 (glucose4·xylose3·galactose·fucose). The same oligosaccharides also promoted the degradation, assayed viscometrically, of xyloglucan by an acidic cellulase from bean (Phaseolus vulgaris L.) leaves. The oligosaccharides were highly active at 10-4 molar, causing up to a fourfold increase in activity, but the effect was still detectable at 1 micromolar. Those oligosaccharides (XG8 and XG9n) which best promoted growth, stimulated cellulase activity to the greatest extent. The oligosaccharides did not stimulate the action of the cellulase in an assay based on the conversion of [3H]xyloglucan to ethanol-soluble fragments. This suggests that the oligosaccharides enhanced the midchain hydrolysis of xyloglucan molecules (which would rapidly reduce the viscosity of the solution), at the expense of cleavage near the termini (which would yield ethanol-soluble products)

  4. Purification and characterization of a carboxymethyl cellulase from Artemia salina.

    Science.gov (United States)

    Zin, Hyun Woo; Park, Kwang-Hyun; Choi, Tae Jin

    2014-01-01

    Brine shrimp (Artemia salina) belong to a group of crustaceans that feed on microalgae and require a cellulase enzyme that can be used in ethanol production from marine algae. Protein with potential cellulase activity was purified and the activity analyzed under different conditions. After initial identification of cellulase activity by CMC cellulase, surface sterilization and PCR using 16s rRNA primers was conducted to confirm that the cellulase activity was not produced from contaminating bacteria. The enzyme was purified by ammonium sulfate fractionation, gel filtration, and ion exchange chromatography. After the final purification, a 70-fold increase in specific enzyme activity was observed. SDS-PAGE results revealed that the cellulase enzyme had a molecular mass of 96 kDa. Temperature, pH, and salinity values were found to be optimal at 55 °C, pH 8.0, and 600 mM NaCl, respectively. Specifically, the enzyme showed a fivefold increase in enzyme activity in seawater compared to 600 mM NaCl in phosphate buffer. Further analysis of the purified enzyme by molecular spectrometry showed no match to known cellulases, indicating this enzyme could be a novel halophilic cellulase that can be used for the production of bioethanol from marine macroalgae. PMID:24291747

  5. Immobilization of cellulase on functionalized cobalt ferrite nanoparticles

    Energy Technology Data Exchange (ETDEWEB)

    Bohara, Raghvendra Ashok; Thorat, Nanasaheb Devappa; Pawar, Shivaji Hariba [Center for Interdisciplinary Research, D. Y. Patil University, Kolhapur (India)

    2016-01-15

    Amine functionalized cobalt ferrite (AF-CoFe{sub 2}O{sub 4}) magnetic nanoparticles (MNPs) were used for immobilization of cellulase enzyme via 1-ethyl-3-[3-dimethylaminopropyl] carbodiimide hydrochloride (EDS) and N-hydroxysuccinimide (NHS) coupling reaction. The structural, morphological and magnetic properties of AF-CoFe{sub 2}O{sub 4} were determined. TEM micrograph revealed a mean diameter of -8 nm and showed that the AF-CoFe{sub 2}O{sub 4} remain distinct with no significant change in size after binding with cellulase. Fourier transform infrared (FT-IR) spectroscopy confirmed the binding of cellulase to AF-CoFe{sub 2}O{sub 4}. The properties of immobilized cellulase were investigated by optimizing binding efficiency, pH, temperature and reusability. The results showed that the immobilized cellulase has higher thermal stability than free cellulase, which might be due to covalent interaction between cellulase and AF-CoFe{sub 2}O{sub 4} surface. The immobilized cellulase also showed good reusability after recovery. Therefore, AF-CoFe{sub 2}O{sub 4} MNPs can be considered as promising candidate for enzyme immobilization.

  6. Production Of Cellulase In Plastids Of Transgenic Plants

    Energy Technology Data Exchange (ETDEWEB)

    Lamppa, Gayle (Chicago, IL)

    2002-08-06

    A genetic construct encoding a fusion protein including endogluconase E1 and a transit peptide is used to transform plants. The plants produce cellulase by expressing the genetic construct. The cellulase is targeted to plastids and can be collected and purified.

  7. Synergy between cellulases and pectinases in the hydrolysis of hemp.

    Science.gov (United States)

    Zhang, Junhua; Pakarinen, Annukka; Viikari, Liisa

    2013-02-01

    The impact of pectinases in the hydrolysis of fresh, steam-exploded and ensiled hemp was investigated and the synergy between cellulases, pectinases and xylanase in the hydrolysis was evaluated. About half; 59.3% and 46.1% of pectin in the steam-exploded and ensiled hemp, respectively, could be removed by a low dosage of pectinases used. Pectinases were more efficient than xylanase in the hydrolysis of fresh and ensiled hemp whereas xylanase showed higher hydrolytic efficiency than the pectinase preparation used in the hydrolysis of steam-exploded hemp. Clear synergistic action between cellulases and xylanase could be observed in the hydrolysis of steam-exploded hemp. Supplementation of pectinase resulted in clear synergism with cellulases in the hydrolysis of all hemp substrates. Highest hydrolysis yield of steam-exploded hemp was obtained in the hydrolysis with cellulases and xylanase. In the hydrolysis of ensiled hemp, the synergistic action between cellulases and pectinases was more obvious for efficient hydrolysis.

  8. Enhanced cellulose degradation using cellulase-nanosphere complexes.

    Directory of Open Access Journals (Sweden)

    Craig Blanchette

    Full Text Available Enzyme catalyzed conversion of plant biomass to sugars is an inherently inefficient process, and one of the major factors limiting economical biofuel production. This is due to the physical barrier presented by polymers in plant cell walls, including semi-crystalline cellulose, to soluble enzyme accessibility. In contrast to the enzymes currently used in industry, bacterial cellulosomes organize cellulases and other proteins in a scaffold structure, and are highly efficient in degrading cellulose. To mimic this clustered assembly of enzymes, we conjugated cellulase obtained from Trichoderma viride to polystyrene nanospheres (cellulase:NS and tested the hydrolytic activity of this complex on cellulose substrates from purified and natural sources. Cellulase:NS and free cellulase were equally active on soluble carboxymethyl cellulose (CMC; however, the complexed enzyme displayed a higher affinity in its action on microcrystalline cellulose. Similarly, we found that the cellulase:NS complex was more efficient in degrading natural cellulose structures in the thickened walls of cultured wood cells. These results suggest that nanoparticle-bound enzymes can improve catalytic efficiency on physically intractable substrates. We discuss the potential for further enhancement of cellulose degradation by physically clustering combinations of different glycosyl hydrolase enzymes, and applications for using cellulase:NS complexes in biofuel production.

  9. Statistical analysis of cellulase production in Bacillus amyloliquefaciens UNPDV-22

    Directory of Open Access Journals (Sweden)

    Vasudeo Zambare

    2011-06-01

    Full Text Available The production of cellulase in Bacillus amyloliquefaciens UNPDV-22 was optimized usingresponse surface methodology (RSM. Central composite design (CCD was used to study the interactiveeffect of fermentation medium components (wheat bran, soybean meal, and malt dextrin on cellulaseactivity. Results suggested that wheat bran, soybean meal, and malt dextrin all have significant impacton cellulase production. The use of RSM resulted in a 70% increase in the cellulase activity over thecontrol of non-optimized basal medium. Optimum cellulase production of 11.23 U/mL was obtained in afermentation medium containing wheat bran (1.03%, w/v, soybean meal (2.43%, w/v, and maltdextrin (2.95%, w/v.

  10. Microbial cellulases and their applications in textile processing

    OpenAIRE

    Mojsov, Kiro

    2012-01-01

    Basic and applied research on microbial cellulases has not only generated significant scientific knowledge but has also revealed their enormous potential in biotechnology.At present, cellulases and related enzymes are used in food, brewery and wine, animal feed, textile and laundry, pulp and paper industries, as well as in agriculture and for research purposes. Textile processing is a growing industry that traditionally has used a lot of water, energy and harsh chemicals. Due to the ever-grow...

  11. Regulation of Biosynthesis of Individual Cellulases in Thermomonospora fusca

    OpenAIRE

    Spiridonov, Nikolay A.; Wilson, David B.

    1998-01-01

    Regulation of the biosynthesis of the six cellulases comprising the cellulolytic system of the thermophilic soil bacterium Thermomonospora fusca ER1 was studied. The levels of the individual enzymes produced on different noninducing and inducing carbon sources were determined. The lowest level of cellulase synthesis (3 nM) was observed with xylose as a carbon source, and the highest level (247 to 1,670 nM for different enzymes) was found in cultures grown on microcrystalline cellulose. Endoce...

  12. Investigation of newly developed solid state fermenter on carboxymethyl cellulase production

    OpenAIRE

    Lee, C K; Ibrahim, D.; Che Omar, I.

    2013-01-01

    Aims: Enzyme (cellulase) contributes 10% to overall cost in bioethanol production from lignocellulosic biomass. This means that the cost for bioethanol production will be reduced if cellulase can be produced using cheaper method. Compared with submerged fermentation, it is recognized that the cost for cellulase production using solid state fermentation (SSF) process is much cheaper. The present study aimed to optimize cellulase production via SSF process using agro-industrial residual as subs...

  13. Exploring the Mechanism Responsible for Cellulase Thermostability by Structure-Guided Recombination

    OpenAIRE

    Chia-Jung Chang; Cheng-Chung Lee; Yueh-Te Chan; Trudeau, Devin L.; Mei-Huey Wu; Chih-Hsuan Tsai; Su-May Yu; Tuan-Hua David Ho; Andrew H-J Wang; Chwan-Deng Hsiao; Arnold, Frances H.; Yu-Chan Chao

    2016-01-01

    Cellulases from Bacillus and Geobacillus bacteria are potentially useful in the biofuel and animal feed industries. One of the unique characteristics of these enzymes is that they are usually quite thermostable. We previously identified a cellulase, GsCelA, from thermophilic Geobacillus sp. 70PC53, which is much more thermostable than its Bacillus homolog, BsCel5A. Thus, these two cellulases provide a pair of structures ideal for investigating the mechanism regarding how these cellulases can ...

  14. Thermostable cellulases, and mutants thereof, capable of hydrolyzing cellulose in ionic liquid

    Energy Technology Data Exchange (ETDEWEB)

    Sapra, Rajat; Datta, Supratim; Chen, Zhiwei; Holmes, Bradley M.; Simmons, Blake A.; Blanch, Harvey W.

    2016-04-26

    The present invention provides for a composition comprising an ionic liquid and a thermostable cellulose, and a method of hydrolyzing a cellulose, comprising: (a) providing a composition comprising a solution comprising an ionic liquid and a cellulose, and (b) introducing a thermostable cellulase to the solution, such that the cellulose is hydrolyzed by the cellulase. The present invention also provides for a Thermatoga maritima thermostable cellulase mutant with increased cellulase activity.

  15. Ethanol from wood. Cellulase enzyme production

    Energy Technology Data Exchange (ETDEWEB)

    Szengyel, Zsolt

    2000-03-01

    Conversion of biomass to liquid fuels, such as ethanol, has been investigated during the past decades. First due to the oil crisis of the 1970s and lately because of concerns about greenhouse effect, ethanol has been found to be a suitable substitute for gasoline in transportation. Although ethanol is produced in large quantities from corn starch, the conversion of lignocellulosic biomass to ethanol is rather problematic. However, cellulosic raw materials are important as they are available in large quantities from agriculture and forestry. One of the most extensively investigated processes is the enzymatic process, in which fungal cellulolytic enzymes are used to convert the cellulose content of the biomass to glucose, which is then fermented to ethanol. In order to make the raw material accessible to biological attack, it has to be pretreated first. The most successful method, which has been evaluated for various lignocellulosic materials, is the steam pretreatment. In this thesis the utilization of steam pretreated willow (hardwood) and spruce (softwood) was examined for enzyme production using a filamentous fungus T. reesei RUT C30. Various carbon sources originating from the steam pretreated materials have been investigated. The replacement of the solid carbon source with a liquid carbon source, as well as the effect of pH, was studied. The effect of toxic compounds generated during pretreatment was also examined. Comparative study of softwood and hardwood showed that steam pretreated hardwood is a better carbon source than softwood. The hydrolytic potential of enzyme solutions produced on wood derived carbon sources was better compared to commercial cellulases. Also enzyme solutions produced on steam pretreated spruce showed less sensitivity towards toxic compounds formed during steam pretreatment.

  16. 21 CFR 173.120 - Carbohydrase and cellulase derived from Aspergillus niger.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Carbohydrase and cellulase derived from Aspergillus... cellulase derived from Aspergillus niger. Carbohydrase and cellulase enzyme preparation derived from Aspergillus niger may be safely used in food in accordance with the following prescribed conditions:...

  17. Production of Citric Acid from Apple Pomace Enzymolyzed by Cellulase

    Institute of Scientific and Technical Information of China (English)

    宋纪蓉; 黄洁; 徐抗震; 赵巧云

    2003-01-01

    Cellulase can evidently increase the content of glucose and has a significant effect on the production of citric acid from apple pomace by Aspergillus niger. Based on experiments, a cellulolytic enzyme named cellulase A6 was found able to produce about 170 g glucose from 1 kg dried apple pomace after 12 h reaction, with cellulase concentration of 20 U/g in the medium at 50℃, natural pH without pretreatment of alkali. Using the treated apple pomace as a liquid state substrate, Aspergillus niger-C selected out was able to produce about 256 g citric acid from 1 kg dried apple pomace at 35℃ in 3 d or 30℃ in 5 d with flask rotation speed of 210 r/min, and the conversion of citric acid could reach 80% based on the amount of sugar consumed.

  18. The catalytic inactivation of cellulase enzyme components by palladium complexes

    Energy Technology Data Exchange (ETDEWEB)

    Woodward, J.; Gooch, M.G.; Shultz, M.D.

    1993-10-01

    It has been discovered that sodium hexachloropalladate is a strong inhibitor of cellobiohydrolase I (CBH I) from Trichoderma reesei having an I{sub 50} of <50 {mu}M with p-nitrophenylcellobioside (PNPC) as the substrate. Similar complexes of the metals platinum, osmium, iridium, and rhodium have little effect on CBH I. Other cellulase activities (Avicelase, {beta}-glucanase) are also inhibited by the palladium complex, suggesting that inhibition of two major types of catalytic activity in cellulase are affected. Preliminary data on the kinetics of inhibition of CBH I by sodium hexachloropalladate indicate that the inhibition is reversible and, possibly, uncompetitive. It is anticipated that sodium hexachloropalladate and other palladium complexes will be useful for determining the effect of the binding of catalytically inactivated CBH I and other cellulase components on the structure of cellulose fibers.

  19. Optimization of solid fermentation of cellulase from Trichoderma koningii

    Institute of Scientific and Technical Information of China (English)

    LI Pei-jun; JING De-bing; ZHOU Qi-xing; ZHANG Chun-gui

    2004-01-01

    To exploit peashrub resources in Ordos as fodders, it is very crucial to realize industrial production of cheap cellulase of high activity by optimizing culture technology, especially culture substrate. In this study, a new prescription experiment based on uniform design ideal was invented and successfully applied in the solid fermentation of Trichoderma koningii F244, which being performed with two different temperature degrees. The activities of FPA, cotton lyase, CMCase and β-glucosidase were assayed and then mathematical models of enzymatic activities, which were figured out by Unconstraint Mathematical Programming, were developed by Multivariate Regression Program of SPSS10.0. Enzymatic activities of optimized substrate prescriptions corresponding to mathematical models were forecasted to determine an ideal substrate prescription. It is revealed that in solid fermentation, Tween80 has negative effect on cellulase production. Furthermore, the ideal prescription for cellulase complex production by Trichoderma koningii F244 was straw powder 16.9%,wheat bran 26.5%, (NH4)2SO4 9.5% and water 47.1%, whose corresponding cellulase activity was expected to be at the same high level with that of Trichoderma reesei Q9414 on its own recommended substrate. Especially, goats mainly fed on peashrub tissues mixed with cellulase complex of this prescription and culture technology, got an incremental ratio of 0.3 kg/d, which brought a very promising feeding prospect for local peashrub resource. By populization of this cellulase complex, it can integrate living standard, economic construction of local residents into vegetational restoration tightly and thus this paper will be very meaningful to be use for reference for western China like Ordos to realize its sustainable development of economy, society and environment.

  20. Regulation of Laccase and Cellulase Genes Transcription in Agaricus bisporus

    OpenAIRE

    Ohga, Shoji; Wood, David A.

    1998-01-01

    A time course for laccase and cellulase genes transcription of Agaricus bisporus compost culture are examined. The results of assays for laccase gene leel show that the expression of this gene increased in the compost until pinning stage of development. In the fruiting cultures the amount of leel declined rapidly over a 4-5 d period immediately. Cellulase gene celS expression contrasted sharply appeared with leel expression by remaining at a low level until after the pins were seen. The cel3...

  1. Cellulase production by halophytic fungi. Pt. 2. The role of some cultural conditions on cellulase production by Ulocladium chartarum

    Energy Technology Data Exchange (ETDEWEB)

    Sallam, L.A.R.; EL-Refai, A.M.H.; El-Sayed, S.M.

    1988-01-01

    Cellulase activities of Ulocladium chartarum were found to be influenced by the mode of incubation as well as by the age of the fungal culture. Maximal cellulase activities were, however, maintained after 3 weeks using the surface culture mode of incubation. U. chartarum was grown at different temperatures, where maximal CMCase and FPase activities were achieved at 30/sup 0/C. The cellulase activities were also tested using initially adjusted pH as well as buffered media. The highest CMCase and FPase activities (0.289 and 0.043 unit/ml respectively) were obtained with pH 6.5 and the phosphate buffer gave better results as compared with the acetate buffer.

  2. Enzymic hydrolysis of cellulosic substances by the crude cellulase from Aspergillus aculeatus

    Energy Technology Data Exchange (ETDEWEB)

    Sakamoto, R.; Hayashi, H.; Moriyama, K.; Arai, M.; Murao, S.

    1982-01-01

    The activity of crude cellulase powder from A. aculeatus F-50 was investigated on rice straw, citrus peel, and kraft pulp. After treatment of straw with 1% NaOH for 3 h, cellulase at 37 degrees for 3 days produced 85% solution, with a yield of 8% reducing sugar, which consisted of glucose and xylose. When cellulase from Trichoderma reesei was combined with A. aculeatus cellulase, a yield of 8% reducing sugars was reached in 1 day. Solubilization of orange peel was greater than 70% in 12 hours and of bleached kraft pulp, 73% in 3 days by A. aculeatus cellulase. The only reducing sugar produced was glucose.

  3. Exploring the Synergy between Cellobiose Dehydrogenase from Phanerochaete chrysosporium and Cellulase from Trichoderma reesei.

    Science.gov (United States)

    Wang, Min; Lu, Xuefeng

    2016-01-01

    Recent demands for the production of lignocellulose biofuels boosted research on cellulase. Hydrolysis efficiency and production cost of cellulase are two bottlenecks in "biomass to biofuels" process. The Trichoderma cellulase mixture is one of the most commonly used enzymes for cellulosic hydrolysis. During hydrolytic process cellobiose accumulation causes feedback inhibition against most cellobiohydrolases and endoglucanases. In this study, we demonstrated the synergism effects between cellobiose dehydrogenase (CDH) and cellulase both in vitro and in vivo. The CDH from Phanerochaete chrysosporium was heterologously expressed in Pichia pastoris. Supplementation of the purified CDH in Trichoderma cellulase increased the cellulase activities. Especially β-glucosidase activity was increased by 30-100% varying at different time points. On the other hand, the cdh gene was heterologously expressed in Trichoderma reesei to explore the synergism between CDH and cellulases in vivo. The analyses of gene expression and enzymatic profiles of filter paper activity, carboxymethylcellulase (CMCase) and β-glucosidase show the increased cellulase activity and the enhanced cellulase production in the cdh-expressing strains. The results elucidate a possible mechanism for diminishing the cellobiose inhibition of cellulase by CDH. These findings provide a novel perspective to make more economic enzyme cocktails for commercial application or explore alternative strategies for generating cellulase-producing strains with higher efficiency. PMID:27199949

  4. Recycling cellulase towards industrial application of enzyme treatment on hardwood kraft-based dissolving pulp.

    Science.gov (United States)

    Wang, Qiang; Liu, Shanshan; Yang, Guihua; Chen, Jiachuan; Ji, Xingxiang; Ni, Yonghao

    2016-07-01

    Cost-effectiveness is vital for enzymatic treatment of dissolving pulp towards industrial application. The strategy of cellulase recycling with fresh cellulase addition was demonstrated in this work to activate the dissolving pulp, i.e. decreasing viscosity and increasing Fock reactivity. Results showed that 48.8-35.1% of cellulase activity can be recovered from the filtered liquor in five recycle rounds, which can be reused for enzymatic treatment of dissolving pulp. As a result, the recycling cellulase with addition fresh cellulase of 1mg/g led to the pulp of viscosity 470mL/g and Fock reactivity 80%, which is comparable with cellulase charge of 2mg/g. Other pulp properties such as alpha-cellulose, alkaline solubility and molecular weight distribution were also determined. Additionally, a zero-release of recycling cellulase treatment was proposed to integrate into the dissolving pulp production process. PMID:27099940

  5. CBH1 homologs and varian CBH1 cellulase

    Energy Technology Data Exchange (ETDEWEB)

    Goedegebuur, Frits; Gualfetti, Peter; Mitchinson, Colin; Neefe, Paulien

    2014-07-01

    Disclosed are a number of homologs and variants of Hypocrea jecorina Cel7A (formerly Trichoderma reesei cellobiohydrolase I or CBH1), nucleic acids encoding the same and methods for producing the same. The homologs and variant cellulases have the amino acid sequence of a glycosyl hydrolase of family 7A wherein one or more amino acid residues are substituted and/or deleted.

  6. A multipurpose immobilized biocatalyst with pectinase, xylanase and cellulase activities

    Directory of Open Access Journals (Sweden)

    Gupta Munishwar

    2007-06-01

    Full Text Available Abstract Background The use of immobilized enzymes for catalyzing various biotransformations is now a widely used approach. In recent years, cross-linked enzyme aggregates (CLEAs have emerged as a novel and versatile biocatalyst design. The present work deals with the preparation of a CLEA from a commercial preparation, Pectinex™ Ultra SP-L, which contains pectinase, xylanase and cellulase activities. The CLEA obtained could be used for any of the enzyme activities. The CLEA was characterized in terms of kinetic parameters, thermal stability and reusability in the context of all the three enzyme activities. Results Complete precipitation of the three enzyme activities was obtained with n-propanol. When resulting precipitates were subjected to cross-linking with 5 mM glutaraldehyde, the three activities initially present (pectinase, xylanase and cellulase were completely retained after cross-linking. The Vmax/Km values were increased from 11, 75 and 16 to 14, 80 and 19 in case of pectinase, xylanase and cellulase activities respectively. The thermal stability was studied at 50°C, 60°C and 70°C for pectinase, xylanase and cellulase respectively. Half-lives were improved from 17, 22 and 32 minutes to 180, 82 and 91 minutes for pectinase, xylanase and cellulase respectively. All three of the enzymes in CLEA could be reused three times without any loss of activity. Conclusion A single multipurpose biocatalyst has been designed which can be used for carrying out three different and independent reactions; 1 hydrolysis of pectin, 2 hydrolysis of xylan and 3 hydrolysis of cellulose. The preparation is more stable at higher temperatures as compared to the free enzymes.

  7. Recycling cellulases for cellulosic ethanol production at industrial relevant conditions

    DEFF Research Database (Denmark)

    Lindedam, Jane; Haven, Mai Østergaard; Chylenski, Piotr;

    2013-01-01

    Different versions of two commercial cellulases were tested for their recyclability of enzymatic activity at high dry matter processes (12% or 25% DM). Recyclability was assessed by measuring remaining enzyme activity in fermentation broth and the ability of enzymes to hydrolyse fresh, pretreated...... wheat straw. Industrial conditions were used to study the impact of hydrolysis temperature (40 or 50. °C) and residence time on recyclability. Enzyme recycling at 12% DM indicated that hydrolysis at 50. °C, though ideal for ethanol yield, should be kept short or carried out at lower temperature...... to preserve enzymatic activity. Best results for enzyme recycling at 25% DM was 59% and 41% of original enzyme load for a Celluclast:Novozyme188 mixture and a modern cellulase preparation, respectively. However, issues with stability of enzymes and their strong adsorption to residual solids still pose...

  8. Optimization of cultivation conditions for the cellulase producer aspergillus terreus

    Energy Technology Data Exchange (ETDEWEB)

    Svistova, I.D.; Zherebtsov, N.A.

    1984-01-01

    Aspergillus terreus was able to synthesize C1-cellulase, Cx-cellulase, endoglucanase and cellobiase in a synthetic minimal medium containing agricultural wastes (sawdust, straw, sugar beet pulp, barley husks, etc) as a sole source of C. The highest enzyme activity was observed with sugar beet pulp whereas the lowest activity occurrred with sawdust. Of various N sources tested, NaNO3 and peptone were most effective. Addition of corn meal, malt sprout, or protein-vitamin concentration (PVC) significantly stimulated enzyme biosynthesis. The optimum culture medium, determined by the mathematical method of experimental factor design was composed of sugar beet pulp 30, KH2PO4 1-4, NaNO3 7.5, and PVC 15 g/L.

  9. Inhibitory effect of vanillin on cellulase activity in hydrolysis of cellulosic biomass.

    Science.gov (United States)

    Li, Yun; Qi, Benkun; Wan, Yinhua

    2014-09-01

    Pretreatment of lignocellulosic material produces a wide variety of inhibitory compounds, which strongly inhibit the following enzymatic hydrolysis of cellulosic biomass. Vanillin is a kind of phenolics derived from degradation of lignin. The effect of vanillin on cellulase activity for the hydrolysis of cellulose was investigated in detail. The results clearly showed that vanillin can reversibly and non-competitively inhibit the cellulase activity at appropriate concentrations and the value of IC50 was estimated to be 30 g/L. The inhibition kinetics of cellulase by vanillin was studied using HCH-1 model and inhibition constants were determined. Moreover, investigation of three compounds with similar structure of vanillin on cellulase activity demonstrated that aldehyde group and phenolic hydroxyl groups of vanillin had inhibitory effect on cellulase. These results provide valuable and detailed information for understanding the inhibition of lignin derived phenolics on cellulase.

  10. A multipurpose immobilized biocatalyst with pectinase, xylanase and cellulase activities

    OpenAIRE

    Gupta Munishwar; Sharma Aparna; Dalal Sohel

    2007-01-01

    Abstract Background The use of immobilized enzymes for catalyzing various biotransformations is now a widely used approach. In recent years, cross-linked enzyme aggregates (CLEAs) have emerged as a novel and versatile biocatalyst design. The present work deals with the preparation of a CLEA from a commercial preparation, Pectinex™ Ultra SP-L, which contains pectinase, xylanase and cellulase activities. The CLEA obtained could be used for any of the enzyme activities. The CLEA was characterize...

  11. Inhibition of lignin-derived phenolic compounds to cellulase

    OpenAIRE

    Qin, Lei; Li, Wen-Chao; Liu, Li; Zhu, Jia-Qing; Li, Xia; Li, Bing-zhi; Yuan, Ying-Jin

    2016-01-01

    Background Lignin-derived phenolic compounds are universal in the hydrolysate of pretreated lignocellulosic biomass. The phenolics reduce the efficiency of enzymatic hydrolysis and increase the cost of ethanol production. We investigated inhibition of phenolics on cellulase during enzymatic hydrolysis using vanillin as one of the typical lignin-derived phenolics and Avicel as cellulose substrate. Results As vanillin concentration increased from 0 to 10 mg/mL, cellulose conversion after 72-h e...

  12. Effects of lignin and surfactant on adsorption and hydrolysis of cellulases on cellulose

    OpenAIRE

    Yanfei LI; Sun, Zongping; Ge, Xiaoyan; Zhang, Junhua

    2016-01-01

    Background Considerable works have been reported concerning the obstruction of enzymatic hydrolysis efficiency by lignin. However, there is a lack of information about the influence of lignin on the adsorption of cellulases on cellulose, along with the hydrolytic activity of the cellulases adsorbed on lignin. In addition, limited discovery has been reported about the influence of additives on cellulase desorption from lignin and lignocellulosic materials. In this work, the effects of lignin o...

  13. Aspergillus nidulans protein kinase A plays an important role in cellulase production

    OpenAIRE

    de Assis, Leandro José; Ries, Laure Nicolas Annick; Savoldi, Marcela; dos Reis, Thaila Fernanda; Brown, Neil Andrew; Goldman, Gustavo Henrique

    2015-01-01

    Background The production of bioethanol from lignocellulosic feedstocks is dependent on lignocellulosic biomass degradation by hydrolytic enzymes. The main component of lignocellulose is cellulose and different types of organisms are able to secrete cellulases. The filamentous fungus Aspergillus nidulans serves as a model organism to study cellulase production and the available tools allow exploring more in depth the mechanisms governing cellulase production and carbon catabolite repression. ...

  14. Production of Resveratrol by Piceid Deglycosylation Using Cellulase

    Directory of Open Access Journals (Sweden)

    Chia-Hung Kuo

    2016-02-01

    Full Text Available Resveratrol is a dietary polyphenolic compound widely used in medicine, food, and cosmetic products. The glycoside form of resveratrol, piceid, is also present in several plant materials but is less bioavailable. In this study, enzymatic transformation of piceid into resveratrol using inexpensive cellulase was investigated. Response surface methodology was used to evaluate the effect of reaction parameters, including reaction temperature, reaction time, enzyme amount and pH. The optimal conditions for biotransformation of piceid to resveratrol are: a reaction temperature of 50 °C, reaction time of 4.75 h, enzyme amount of 2.5 fungal β-glucanase (FBG units and pH of 4.3. In addition, the extracts from Polygonum cuspidatum root contained high amounts of piceid were treated with cellulase in order to deglycosylation that increased resveratrol yield. After treatment, the resveratrol yield significantly increased from 2.72 to 9.49 mg/g, while the piceid contents decreased from 8.60 to 0 mg/g. The result provides an efficient method to convert piceid in the extracts of P. cuspidatum root into resveratrol by cellulase.

  15. Cellulase-assisted extraction of oligosaccharides from defatted rice bran.

    Science.gov (United States)

    Patindol, J; Wang, L; Wang, Y-J

    2007-11-01

    Defatted rice bran was subjected to cellulase treatment in order to increase its extractable oligosaccharides. Various combinations of enzyme concentration (0%, 0.5%, 1.0%, and 2.0%), temperature (room, 30, 40, and 50 degrees C), and time (1, 3, 5, and 16 h) were tested to identify the optimum extraction conditions. The saccharide content and composition of the extracts were analyzed by high-performance anion-exchange chromatography with pulsed amperometric detection (HPAEC-PAD). Digestibility was assayed in vitro with human salivary and porcine pancreatic alpha-amylases. Extraction yield ranged from 13.4% (without cellulase) to 39.9% (with 2% cellulase). Total carbohydrates, reducing sugars, and crude protein of the dried extracts ranged from 69.2% to 87.2%, 18.7% to 62.3%, and 7.1% to 22.3%, respectively. Mono- and disaccharides constituted more than 50% of the total carbohydrates in the extracts. Inherent oligosaccharides and those produced by cellulolysis made up less than 25%. The in vitro digestibility of the extracts by alpha-amylases was lower compared with that of the original rice bran sample and potato dextrin, which could be attributed to the increased concentrations of oligosaccharides and reducing sugars. PMID:18034713

  16. Xyloglucan oligosaccharides promote growth and activate cellulase: Evidence for a role of cellulase in cell expansion. [Pisum sativum L

    Energy Technology Data Exchange (ETDEWEB)

    McDougall, G.J.; Fry, S.C. (Univ. of Edinburgh (England))

    1990-07-01

    Oligosaccharides produced by the action of fungal cellulase on xyloglucans promoted the elongation of etiolated pea (Pisum sativum L.) stem segments in a straight-growth bioassay designed for the determination of auxins. The oligosaccharides were most active at about 1 micromolar. We tested the relative growth-promoting activities of four HPLC-purified oligosaccharides which shared a common glucose{sub 4} {center dot} xylose{sub 3} (XG7) core. The substituted oligosaccharides XG8 (glucose{sub 4} {center dot} xylose{sub 3} {center dot} galactose) and XG9n (glucose{sub 4} {center dot} xylose{sub 3} {center dot} galactose{sub 2}) were more effective than XG7 itself and XG9 (glucose{sub 4} {center dot} xylose{sub 3} {center dot} galactose {center dot} fucose). The same oligosaccharides also promoted the degradation, assayed viscometrically, of xyloglucan by an acidic cellulase from bean (Phaseolus vulgaris L.) leaves. The oligosaccharides were highly active at 10{sup {minus}4} molar, causing up to a fourfold increase in activity, but the effect was still detectable at 1 micromolar. Those oligosaccharides (XG8 and XG9n) which best promoted growth, stimulated cellulase activity to the greatest extent. The oligosaccharides did not stimulate the action of the cellulase in an assay based on the conversion of ({sup 3}H)xyloglucan to ethanol-soluble fragments. This suggests that the oligosaccharides enhanced the midchain hydrolysis of xyloglucan molecules (which would rapidly reduce the viscosity of the solution), at the expense of cleavage near the termini (which would yield ethanol-soluble products).

  17. Strain Improvement of Bacillus coagulans and Geobacillus stearothermophilus for Enhanced Thermostable Cellulase Production and the Effect of Different Metal Ions on Cellulase Activity

    Directory of Open Access Journals (Sweden)

    Vikas Sharma

    2012-11-01

    Full Text Available The current study was focused on the strain improvement of Bacillus coagulans and Geobacillus stearothermophilus for thermostable cellulase production with higher enzyme activity. For strain improvement UV radiations, NTG and Sodium azide were used as mutagenic agents.NTG was found to be best mutagenic agent among all in term of highest cellulase activity. Mutant strain C11 exhibited the highest cellulase specific activity at 45 U/mg followed by C15 (39 U/mg in case of B.coagulans while Mutant strain S18 exhibited thehighest cellulase specific activity at 69 U/mg followed by S12 (62 U/mg in case of G. stearothermophilus. Specific activity of cellulase was 92 U/mg in case of B.coagulans C11 and 118 U/mg in case of G. stearothermophilus S18. Ag+, Mg+, Se2+,Ca2+,Co2+,Mn2+,K+, Zn2+ ,Fe3+, Hg2+ and Cu2+ showed positive change in specific activity while Na+, Ni2+ negative change in specific activity of cellulase with respect to specific activity of cellulase in absence of any additive in case of B.coagulans C11 and Ag+, Mg+, Se2+,Co2+,Mn2+ andHg2+ showed positive change in specific activity, Na+, K+ showed no change in specific activity while Ca2+, Zn2+, Ni2+, Fe3+ and Cu2+ showed negative change in specific activity of cellulase with respect to specific activity of cellulase in absence of any additive in case of G. stearothermophilus S18.

  18. Characterization of cellulase enzyme produced by Chaetomium sp. isolated from books and archives

    Directory of Open Access Journals (Sweden)

    Moza Mohammed AL-Kharousi

    2015-12-01

    Full Text Available Background: Cellulase is an important industrial enzyme used to degrade cellulosic biomass. The demand for cellulase enzyme is continuously increasing because of its applications in various industries. Hence, screening of cellulase producing microorganisms from different sources has gained significant importance. Material and Methods: In this study, fungi isolated from books and archives were screened for their cellulase producing abilities. Four different fungi were isolated from books and archives using potato dextrose agar. Screening of these isolates for cellulase production was carried out using carboxymethyl cellulose broth. The most efficient fungus was subjected to cellulase fermentation and enzymes produced were purified and partially characterized. Results: Four different fungi, Chaetomium sp., Aspergillus niger, Aspergillus nidulans and Penicillium sp., were isolated from books and archives. All the isolates were tested for their ability to producecellulase enzyme. During the primary screening Chaetomium sp. showed good growth and highercellulase activity (155.3±25.6 U/mL in carboxymethyl cellulose medium than the other fungi. The cellulase fermentation study was conducted with Chaetomium sp. using carboxymethyl cellulose asa substrate. During the stationary phase (144 h of the growth, the cellulase activity of Chaetomium sp. was significantly high. The maximum mycelial weight of this fungi was obtained at 168 h. Viscosity of the Chaetomium sp. inoculated fermentation medium continuously decreased until 144 h because of the degradation of carboxymethyl cellulose. During cellulase fermentation, pHincreased from the initial neutral pH to 8.5. Purified cellulase showed a specific activity of 7.3 U/mg. It exhibited maximum activity at 20°C and was stable between pH 5 and 9. Conclusions: Books and archives could be a good source for the isolation of cellulase producing fungi.

  19. Nucleosome transactions on the Hypocrea jecorina (Trichoderma reesei) cellulase promoter cbh2 associated with cellulase induction.

    Science.gov (United States)

    Zeilinger, S; Schmoll, M; Pail, M; Mach, R L; Kubicek, C P

    2003-10-01

    The 5' regulatory region of the cbh2 gene of Hypocrea jecorina contains the cbh2 activating element (CAE) which is essential for induction of cbh2 gene expression by sophorose and cellulose. The CAE consists of two motifs, a CCAAT box on the template strand and a GTAATA box on the coding strand, which cooperate during induction. Northern analyses of cbh2 gene expression has revealed an absolute dependence on induction, but no direct effect of Cre1-mediated carbon catabolite repression. Investigation of the chromatin structure in the wild-type strain showed that, under repressing conditions, there is a nucleosome free region (nfr) around the CAE, which is flanked by strictly positioned nucleosomes. Induction results in a loss of positioning of nucleosomes -1 and -2 downstream of the CAE, thus making the TATA box accessible. Simultaneous mutation of both motifs of the CAE, or of the CCAAT-box alone, also leads to shifting of nucleosome -1, which normally covers the TATA-box under repressing conditions, whereas mutation of the GTAATA element results in a narrowing of the nfr, indicating that the proteins that bind to both motifs in the CAE interact with chromatin, although in different ways. A cellulase-negative mutant strain, which has previously been shown to be altered in protein binding to the CAE, still displayed the induction-specific changes in nucleosome structure, indicating that none of the proteins that directly interact with CAE are affected, and that nucleosome rearrangement and induction of cbh2 expression are uncoupled. Interestingly, the carbon catabolite repressor Cre1 is essential for strict nucleosome positioning in the 5' regulatory sequences of cbh2 under all of the conditions tested, and induction can occur in a promoter that lacks positioned nucleosomes. These data suggest that Cre1, the Hap2/3/5 complex and the GTAATA-binding protein are all involved in nucleosome assembly on the cbh2 promoter, and that the latter two respond to inducing

  20. Adsorption and mechanism of cellulase enzymes onto lignin isolated from corn stover pretreated with liquid hot water

    OpenAIRE

    Lu, Xianqin; Zheng, Xiaoju; Li, Xuezhi; Zhao, Jian

    2016-01-01

    Background In the bioconversion of lignocellulosic substrates, the adsorption behavior of cellulase onto lignin has a negative effect on enzymatic hydrolysis of cellulose, decreasing glucose production during enzymatic hydrolysis, thus decreasing the yield of fermentation and the production of useful products. Understanding the interaction between lignin and cellulase is necessary to optimize the components of cellulase mixture, genetically engineer high-efficiency cellulase, and reduce cost ...

  1. Location, formation and biosynthetic regulation of cellulases in the gliding bacteria Cytophaga hutchinsonii

    Directory of Open Access Journals (Sweden)

    Elijah Johnson

    2006-01-01

    Full Text Available An analysis of the recently published genome sequence of Cytophagahutchinsonii revealed an unusual collection of genes for an organism that can attackcrystalline cellulose. Consequently, questions were being raised by cellulase scientists, as towhat mechanism this organism uses to degrade its insoluble substrates. Cellulose, being ahighly polymeric compound and insoluble in water, cannot enter the cell walls ofmicroorganisms. Cellulose-degrading enzymes have therefore to be located on the surface ofthe cell wall or released extracellularly. The location of most cellulase enzymes has beenstudied. However, basic information on C. hutchinsonii cellulases is almost non-existent. Inthe present study, the location, formation and biosynthetic regulation of cellulases in C.hutchinsonii were demonstrated on different substrates. Various fractions isolated from C.hutchinsonii after cell rupture were assayed for carboxymethyl-cellulase activity (CMC.The cellulases were found to be predominantly cell-free during active growth on solka-flok,although 30% of activity was recorded on cell-bound enzymes. Relatively little CM-cellulase was formed when cells were grown on glucose and cellobiose. Apparently glucoseor labile substrates such as cellobiose seem to repress the formation of CM-cellulase. Thesefindings should provide some insight into possible hydrolysis mechanisms by C.hutchinsonii.

  2. 21 CFR 184.1250 - Cellulase enzyme preparation derived from Trichoderma longibrachiatum.

    Science.gov (United States)

    2010-04-01

    ... incorporated by reference in accordance with 5 U.S.C. 552(a) and 1 CFR part 51. Copies are available from the... Trichoderma longibrachiatum. 184.1250 Section 184.1250 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT....1250 Cellulase enzyme preparation derived from Trichoderma longibrachiatum. (a) Cellulase...

  3. Charge engineering of cellulases improves ionic liquid tolerance and reduces lignin inhibition.

    Science.gov (United States)

    Nordwald, Erik M; Brunecky, Roman; Himmel, Michael E; Beckham, Gregg T; Kaar, Joel L

    2014-08-01

    We report a novel approach to concurrently improve the tolerance to ionic liquids (ILs) as well as reduce lignin inhibition of Trichoderma reesei cellulase via engineering enzyme charge. Succinylation of the cellulase enzymes led to a nearly twofold enhancement in cellulose conversion in 15% (v/v) 1-butyl-3-methylimidazolium chloride ([BMIM][Cl]). The improvement in activity upon succinylation correlated with the apparent preferential exclusion of the [Cl] anion in fluorescence quenching assays. Additionally, modeling analysis of progress curves of Avicel hydrolysis in buffer indicated that succinylation had a negligible impact on the apparent KM of cellulase. As evidence of reducing lignin inhibition of T. reesei cellulase, succinylation resulted in a greater than twofold increase in Avicel conversion after 170 h in buffer with 1 wt% lignin. The impact of succinylation on lignin inhibition of cellulase further led to the reduction in apparent KM of the enzyme cocktail for Avicel by 2.7-fold. These results provide evidence that naturally evolved cellulases with highly negative surface charge densities may similarly repel lignin, resulting in improved cellulase activity. Ultimately, these results underscore the potential of rational charge engineering as a means of enhancing cellulase function and thus conversion of whole biomass in ILs. PMID:24522957

  4. Novel Penicillium cellulases for total hydrolysis of lignocellulosics.

    Science.gov (United States)

    Marjamaa, Kaisa; Toth, Karolina; Bromann, Paul Andrew; Szakacs, George; Kruus, Kristiina

    2013-05-10

    The (hemi)cellulolytic systems of two novel lignocellulolytic Penicillium strains (Penicillium pulvillorum TUB F-2220 and P. cf. simplicissimum TUB F-2378) have been studied. The cultures of the Penicillium strains were characterized by high cellulase and β-glucosidase as well moderate xylanase activities compared to the Trichoderma reesei reference strains QM 6a and RUTC30 (volumetric or per secreted protein, respectively). Comparison of the novel Penicillium and T. reesei secreted enzyme mixtures in the hydrolysis of (ligno)cellulose substrates showed that the F-2220 enzyme mixture gave higher yields in the hydrolysis of crystalline cellulose (Avicel) and similar yields in hydrolysis of pre-treated spruce and wheat straw than enzyme mixture secreted by the T. reesei reference strain. The sensitivity of the Penicillium cellulase complexes to softwood (spruce) and grass (wheat straw) lignins was lignin and temperature dependent: inhibition of cellulose hydrolysis in the presence of wheat straw lignin was minor at 35°C while at 45°C by spruce lignin a clear inhibition was observed. The two main proteins in the F-2220 (hemi)cellulase complex were partially purified and identified by peptide sequence similarity as glycosyl hydrolases (cellobiohydrolases) of families 7 and 6. Adsorption of the GH7 enzyme PpCBH1 on cellulose and lignins was studied showing that the lignin adsorption of the enzyme is temperature and pH dependent. The ppcbh1 coding sequence was obtained using PCR cloning and the translated amino acid sequence of PpCBH1 showed up to 82% amino acid sequence identity to known Penicillium cellobiohydrolases.

  5. Production of extremophilic bacterial cellulase enzymes in aspergillus niger.

    Energy Technology Data Exchange (ETDEWEB)

    Gladden, John Michael

    2013-09-01

    Enzymes can be used to catalyze a myriad of chemical reactions and are a cornerstone in the biotechnology industry. Enzymes have a wide range of uses, ranging from medicine with the production of pharmaceuticals to energy were they are applied to biofuel production. However, it is difficult to produce large quantities of enzymes, especially if they are non-native to the production host. Fortunately, filamentous fungi, such as Aspergillus niger, are broadly used in industry and show great potential for use a heterologous enzyme production hosts. Here, we present work outlining an effort to engineer A. niger to produce thermophilic bacterial cellulases relevant to lignocellulosic biofuel production.

  6. Useful halophilic, thermostable and ionic liquids tolerant cellulases

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Tao; Datta, Supratim; Simmons, Blake A.; Rubin, Edward M.

    2016-06-28

    The present invention provides for an isolated or recombinant polypeptide comprising an amino acid sequence having at least 70% identity with the amino acid sequence of a Halorhabdus utahensis cellulase, such as Hu-CBH1, wherein said amino acid sequence has a halophilic thermostable and/or thermophilic cellobiohydrolase (CBH) activity. In some embodiments, the polypeptide has a CBH activity that is resistant to up to about 20% of ionic liquids. The present invention also provides for compositions comprising and methods using the isolated or recombinant polypeptide.

  7. Separation and quantification of cellulases and hemicellulases by capillary electrophoresis

    DEFF Research Database (Denmark)

    Jørgensen, Henning; Kutter, Jörg Peter; Olsson, Lisbeth

    2003-01-01

    . Current methods are limited in their ability to quantify all of these enzymes when all are present simultaneously in a mixture. Five different cellulases (two cellobiohydrolases and three endoglucanases) and one hemicellulase (endoxylanase) were separated using capillary electrophoresis (CE) in a fused...... silica capillary at pH values close to neutral. The improvement of the separation of these six proteins by the addition of alpha, omega-diaminoalkanes with chain lengths from three to seven carbon units was investigated. Dynamically coating the capillary with 1,3-diaminopropane resulted in separation of...

  8. Optimization of cellulase production by Penicillium oxalicum using banana agrowaste as a substrate.

    Science.gov (United States)

    Shah, Shilpa P; Kalia, Kiran S; Patel, Jagdish S

    2015-01-01

    The purpose of this study was to produce a higher amount of cellulase by using an alternative carbon source, such as banana agrowaste, and to optimize the fermentation parameters for a high yield. In the present study, cellulase-producing Penicillium was isolated from a decaying wood sample. Different nutritional and environmental factors were investigated to assess their effect on cellulase production. The highest crude enzyme production was observed at a pH 6.0 and a temperature of 28°C in a medium that was supplemented with banana agrowaste as the carbon source. Pretreatment with 2N NaOH, at 7% substrate (banana agrowaste) concentration yielded the highest cellulase activity. Further to this, the effect of other parameters such as inoculum age, inoculum size, static and agitated conditions were also studied. It is concluded that Penicillium oxalicum is a powerful cellulase-producer strain under our tested experimental conditions using banana agrowaste as the carbon source.

  9. Nutrient control for stationary phase cellulase production in Trichoderma reesei Rut C-30.

    Science.gov (United States)

    Callow, Nicholas V; Ray, Christopher S; Kelbly, Matthew A; Ju, Lu-Kwang

    2016-01-01

    This work describes the use of nutrient limitations with Trichoderma reesei Rut C-30 to obtain a prolonged stationary phase cellulase production. This period of non-growth may allow for dependable cellulase production, extended fermentation periods, and the possibility to use pellet morphology for easy product separation. Phosphorus limitation was successful in halting growth and had a corresponding specific cellulase production of 5±2 FPU/g-h. Combined with the addition of Triton X-100 for fungal pellet formation and low shear conditions, a stationary phase cellulase production period in excess of 300 h was achieved, with a constant enzyme production rate of 7±1 FPU/g-h. While nitrogen limitation was also effective as a growth limiter, it, however, also prevented cellulase production.

  10. Cellulase-lignin interactions in the enzymatic hydrolysis of lignocellulose

    Energy Technology Data Exchange (ETDEWEB)

    Rahikainen, J.

    2013-11-01

    Today, the production of transportation fuels and chemicals is heavily dependent on fossil carbon sources, such as oil and natural gas. Their limited availability and the environmental concerns arising from their use have driven the search for renewable alternatives. Lignocellulosic plant biomass is the most abundant, but currently underutilised, renewable carbon-rich resource for fuel and chemical production. Enzymatic degradation of structural polysaccharides in lignocellulose produces soluble carbohydrates that serve as ideal precursors for the production of a vast amount of different chemical compounds. The difficulty in full exploitation of lignocellulose for fuel and chemical production lies in the complex and recalcitrant structure of the raw material. Lignocellulose is mainly composed of structural polysaccharides, cellulose and hemicellulose, but also of lignin, which is an aromatic polymer. Enzymatic degradation of cellulose and hemicellulose is restricted by several substrate- and enzyme-related factors, among which lignin is considered as one of the most problematic issues. Lignin restricts the action of hydrolytic enzymes and enzyme binding onto lignin has been identified as a major inhibitory mechanism preventing efficient hydrolysis of lignocellulosic feedstocks. In this thesis, the interactions between cellulase enzymes and lignin-rich compounds were studied in detail and the findings reported in this work have the potential to help in controlling the harmful cellulase-lignin interactions, and thus improve the biochemical processing route from lignocellulose to fuels and chemicals.

  11. Optimizing cellulase mixtures for maximum rate and extent of hydrolysis. Final report

    Energy Technology Data Exchange (ETDEWEB)

    Walker, L.P.; Wilson, D.B. [Cornell Univ., Ithaca, NY (United States)

    1997-03-01

    Pure Thomomonospora fusca and Trichoderma reesei cellulases and their mixtures were studied to determine the optimal set of cellulases for biomass hydrolysis. The objective was to reduce the cost of cellulase in order to help lower the overall processing cost of the enzymatic conversion of biomass cellulose to sugars, which can then be fermented into fuels and other energy-intensive chemicals. No cellulase mixture was obtained that was much better than the best commercially available preparations. However, the study has greatly increased knowledge of T. fusca cellulases, synergism, and cellulose binding, and provide evidence that future work will produce cellulases with higher activity in degrading crystalline cellulose. T. fusca cellulases may have good industrial potential because: (1) they are compatible with industrial processes that operate at elevated temperatures; (2) they retain 90% of their activity under neutral or basic conditions, which provides a great deal of flexibility in reactor design and operation; and (3) tools are now available to change specific amino acid residues in their catalytic domains and to assess how these changes influence catalysis. 74 refs.

  12. Investigation and Isolation of Cellulase-Producing microorganisms in the Red Sea

    KAUST Repository

    Fatani, Siham

    2016-05-01

    Cellulolytic microorganisms are considered to be key players in biorefinery, especially for the utilization of plant biomass. These organisms have been isolated from various environments. The Red Sea is one of the seas with high biodiversity and a unique environment, characterized by high water temperature and high salinity . However, there is little information regarding cellulases in Red Sea environments. The aim of the present study is to evaluate the Red Sea as a gene resource for microbial cellulase. I first surveyed microbial cellulases in the Red Sea using a method called metagenomes, and then investigated their abundance and diversity. My survey revealed that the Red Sea biome has a substantial abundance and a wide range of cellulase enzymes with substantial abundance, when compared with those in other environments. Next, I tried to isolate cellulase-active microorganisms from the Red Sea and I successfully obtained seven strains of four different taxonomic groups. These strains showed a similarity of 99% identity to Aspergillus ustus, 99% to Staphylococcus pasteuri, 99% to Bacillus aerius and 99% to Bacillus subtilis. The enzyme assay I conducted, revealed that these strains actually secreted active cellulases. These results suggest that the Red Sea environment can be, indeed, an excellent gene resource of microbial cellulases.

  13. Cellulase immobilization on superparamagnetic nanoparticles for reuse in cellulosic biomass conversion

    Directory of Open Access Journals (Sweden)

    Fernando Segato

    2016-07-01

    Full Text Available Current cellulosic biomass hydrolysis is based on the one-time use of cellulases. Cellulases immobilized on magnetic nanocarriers offer the advantages of magnetic separation and repeated use for continuous hydrolysis. Most immobilization methods focus on only one type of cellulase. Here, we report co-immobilization of two types of cellulases, β-glucosidase A (BglA and cellobiohydrolase D (CelD, on sub-20 nm superparamagnetic nanoparticles. The nanoparticles demonstrated 100% immobilization efficiency for both BglA and CelD. The total enzyme activities of immobilized BglA and CelD were up to 67.1% and 41.5% of that of the free cellulases, respectively. The immobilized BglA and CelD each retained about 85% and 43% of the initial immobilized enzyme activities after being recycled 3 and 10 times, respectively. The effects of pH and temperature on the immobilized cellulases were also investigated. Co-immobilization of BglA and CelD on MNPs is a promising strategy to promote synergistic action of cellulases while lowering enzyme consumption.

  14. Selection of Trichoderma mutants with enhanced cellulase production and resistant to catabolite repression

    Institute of Scientific and Technical Information of China (English)

    Szakacs G; Megyeri L; Kovacs K; Zacchi G

    2004-01-01

    @@ Due to high cost and relatively low efficiency of cellulase enzymes used for the saccharification of pretreated lignocelluloses, the improvement of cellulase secreting microorganisms is of vital importance. Trichoderma reesei QM 6a, an excellent source of cellulase was selected in the late 1960's at Natick Laboratories by its performance on pure cellulose (Solka Floc, Avicel) . QM 6a is the wild parent strain of best existing hypercellulolytic mutants such as Rut C30, VTT-D-80133,L27, CL-847 and others. Utilization of cheaper carbon sources (e. g. , pretreated wood or straw) both in enzyme production and in hydrolysis necessitates to investigate fungal species other than T. reesei.

  15. Comparative performance of precommercial cellulases hydrolyzing pretreated corn stover

    Directory of Open Access Journals (Sweden)

    Mohagheghi Ali

    2011-09-01

    Full Text Available Abstract Background Cellulases and related hydrolytic enzymes represent a key cost factor for biochemical conversion of cellulosic biomass feedstocks to sugars for biofuels and chemicals production. The US Department of Energy (DOE is cost sharing projects to decrease the cost of enzymes for biomass saccharification. The performance of benchmark cellulase preparations produced by Danisco, DSM, Novozymes and Verenium to convert pretreated corn stover (PCS cellulose to glucose was evaluated under common experimental conditions and is reported here in a non-attributed manner. Results Two hydrolysis modes were examined, enzymatic hydrolysis (EH of PCS whole slurry or washed PCS solids at pH 5 and 50°C, and simultaneous saccharification and fermentation (SSF of washed PCS solids at pH 5 and 38°C. Enzymes were dosed on a total protein mass basis, with protein quantified using both the bicinchoninic acid (BCA assay and the Bradford assay. Substantial differences were observed in absolute cellulose to glucose conversion performance levels under the conditions tested. Higher cellulose conversion yields were obtained using washed solids compared to whole slurry, and estimated enzyme protein dosages required to achieve a particular cellulose conversion to glucose yield were extremely dependent on the protein assay used. All four enzyme systems achieved glucose yields of 90% of theoretical or higher in SSF mode. Glucose yields were reduced in EH mode, with all enzymes achieving glucose yields of at least 85% of theoretical on washed PCS solids and 75% in PCS whole slurry. One of the enzyme systems ('enzyme B' exhibited the best overall performance. However in attaining high conversion yields at lower total enzyme protein loadings, the relative and rank ordered performance of the enzyme systems varied significantly depending upon which hydrolysis mode and protein assay were used as the basis for comparison. Conclusions This study provides extensive

  16. Possibilities for recycling cellulases after use in cotton processing: part I: Effects of end-product inhibition, thermal and mechanical deactivation, and cellulase depletion by adsorption.

    Science.gov (United States)

    Azevedo, Helena; Bishop, David; Cavaco-Paul, Artur

    2002-04-01

    Preliminary recycling experiments with cellulase enzymes after cotton treatments at 50 degrees C showed that activity remaining in the treatment liquors was reduced by about 80% after five recycling steps. The potential problems of end-product inhibition, thermal and mechanical deactivation, and the loss of some components of the cellulase complex by preferential and or irreversible adsorption to cotton substrates were studied. End-product inhibition studies showed that the build-up of cellobiose and glucose would be expected to cause no more than 40% activity loss after five textile treatment cycles. Thermal and mechanical treatments of cellulases suggested that the enzymes start to be deactivated at 60 degrees C and agitation levels similar to those used in textile processing did not cause significant enzyme deactivation. Analysis of cellulase solutions, by fast protein liquid chromatography, before and after adsorption on cotton fabrics, suggested that the cellobiohydrolase II (Cel6A) content of the cellulase complex was reduced, relative to the other components, by preferential adsorption. This would lead to a marked reduction in activity after several treatment cycles and top-up with pure cellobiohydrolase II would be necessary unless this component is easily recoverable from the treated fabric.

  17. 2009 Cellulosomes, Cellulases & Other Carbohydrate Modifying Enzymes GRC

    Energy Technology Data Exchange (ETDEWEB)

    Harry Gilbert

    2009-07-26

    The 2009 Gordon Conference on Cellulosomes, Cellulases & Other Carbohydrate Modifying Enzymes will present cutting-edge research on the enzymatic degradation of cellulose and other plant cell wall polysaccharides. The Conference will feature a wide range of topics that includes the enzymology of plant structural degradation, regulation of the degradative apparatus, the mechanism of protein complex assembly, the genomics of cell wall degrading organisms, the structure of the substrate and the industrial application of the process particularly within the biofuel arena. Indeed the deployment of plant cell wall degrading enzymes in biofuel processes will be an important feature of the meeting. It should be emphasized that the 2009 Conference will be expanded to include, in addition to cellulase research, recent advances in other plant cell wall degrading enzymes, and contributions from people working on hemicellulases and pectinases will be particularly welcome. Invited speakers represent a variety of scientific disciplines, including biochemistry, structural biology, genetics and cell biology. The interplay between fundamental research and its industrial exploitation is a particularly important aspect of the meeting, reflecting the appointment of the chair and vice-chair from academia and industry, respectively. The meeting will provide opportunities for junior scientists and graduate students to present their work in poster format and exchange ideas with more established figures in the field. Indeed, some poster presenters will be selected for short talks. The collegial atmosphere of this Conference, with programmed discussion sessions as well as opportunities for informal gatherings in the afternoons and evenings, provides an avenue for scientists from different disciplines to brainstorm and promotes cross-disciplinary collaborations in the various research areas represented. The Conference is likely to be heavily subscribed so we would recommend that you submit

  18. Enzymatic lignocellulose hydrolysis: Improved cellulase productivity by insoluble solids recycling

    Directory of Open Access Journals (Sweden)

    Weiss Noah

    2013-01-01

    Full Text Available Abstract Background It is necessary to develop efficient methods to produce renewable fuels from lignocellulosic biomass. One of the main challenges to the industrialization of lignocellulose conversion processes is the large amount of cellulase enzymes used for the hydrolysis of cellulose. One method for decreasing the amount of enzyme used is to recycle the enzymes. In this study, the recycle of enzymes associated with the insoluble solid fraction after the enzymatic hydrolysis of cellulose was investigated for pretreated corn stover under a variety of recycling conditions. Results It was found that a significant amount of cellulase activity could be recovered by recycling the insoluble biomass fraction, and the enzyme dosage could be decreased by 30% to achieve the same glucose yields under the most favorable conditions. Enzyme productivity (g glucose produced/g enzyme applied increased between 30 and 50% by the recycling, depending on the reaction conditions. While increasing the amount of solids recycled increased process performance, the methods applicability was limited by its positive correlation with increasing total solids concentrations, reaction volumes, and lignin content of the insoluble residue. However, increasing amounts of lignin rich residue during the recycle did not negatively impact glucose yields. Conclusions To take advantage of this effect, the amount of solids recycled should be maximized, based on a given processes ability to deal with higher solids concentrations and volumes. Recycling of enzymes by recycling the insoluble solids fraction was thus shown to be an effective method to decrease enzyme usage, and research should be continued for its industrial application.

  19. Enhanced cellulase production from Trichoderma reesei QM 9414 on physically treated wheat straw

    Energy Technology Data Exchange (ETDEWEB)

    Acebal, C.; Castillon, M.P.; Estrada, P.; Mata, I.; Costa, E.; Aguado, J.; Romero, D.; Jimenez, F.

    1986-06-01

    Trichoderma reesei QM 9414 was grown on wheat straw as the sole carbon source. The straw was pretreated by physical and chemical methods. The particle size of straw was less than 0.177 mm. Growth of T. reesei QM 9414 was maximal with alkali-pretreated straw whereas cellulase production was optimal when physically pretreated straw was used as substrate. Cellulase yields expressed as IU enzyme activity/g cellulose present in the cultures were considerably higher when alkali pretreatment of wheat straw was omitted. Cellulase yields of 666 IU/g cellulose for filter paper activity (FPA) are the highest described for cultures of T. reesei QM 9414 carried out in analogous conditions. Crystallinity index of the cellulose contained in wheat straw increased slightly after alkali pretreatment. This increase did not decrease cellulose accessibility to the fungus. Delignification of wheat straw was not necessary to achieve the best cellulase production.

  20. Comparison between the cellulase systems of Trichoderma harzianum E58 and Trichoderma reesei C30

    Energy Technology Data Exchange (ETDEWEB)

    Saddler, J.N.; Hogan, C.M.; Louis-Seize, G.

    1985-06-01

    Nearly all of the filter paper, endoglucanase and ..beta..-glucosidase activities of T. harzianum E58 were located extracellularly, with low amounts of these activities detected in the cell extracts and relatively little associated with the cell wall. Most of the filter paper and endoglucanase activities of T. reesei C30 were detected extracellularly. The half lives of the different cellulase activities were assayed at various temperatures over a period of time. When the pH of the filtrate was adjusted to 4.8, the cellulase activities were considerably enhanced, with the average half-life at 50/sup 0/C extended to 25 hrs. When various lignocellulosic substrates were hydrolyzed by T. harzianum E58 cellulases approximately 90% of the reducing sugars were present as glucose while 50 - 60% of the reducing sugars were detected as glucose when T. reesei C30 cellulases were used.

  1. [Cellulase and xylanase activities of Fusarium Lk:Fr. genus fungi of different trophic groups].

    Science.gov (United States)

    Kurchenko, I M; Sokolova, O V; Zhdanova, N M; Iarynchyn, A M; Iovenko, O M

    2008-01-01

    A comparative analysis of cellulase and xylanase activities of 26 fungal strains of phytopathogenic, saprophytic and endophytic Fusarium species has been realized using the qualitative reactions. The rare of their linear growth on the media with carboxymethyl cellulose or xylane has been studied. It was shown that the fungi of genus Fusarium belonging to different trophic groups possessed low activities of investigated enzymes as a whole, but in endophytic strains their levels were lower than in phytopathogenic ones. At the same time the distinct strain dependence of cellulase and xylanase activities was fixed in the fungi of different trophic groups. As far as the cellulase and xylanase activities in phytopathogenic isolates varied from complete absence to high levels, and since the activity maximum for each of the investigated strains was observed in different growth terms the conclusion was made that the cellulase and xylanase activities could not be considered as possible markers of the fungal isolate pathogenicity on the strain level.

  2. [Cellulase and xylanase activity of phytopathogenic and endophytic fungal strains of Alternaria alternata (Fr.) Keissler].

    Science.gov (United States)

    Kurchenko, I M; Sokolova, O V; Zhdanova, N M; Iarynchyn, A M; Iovenko, O M

    2008-01-01

    A comparative analysis of cellulase and xylanase activity of 25 fungal strains of phytopathogenic and endophytic Alternaria alternata had been realized for the first time using the qualitative reactions. The rate of their linear growth on the media with carboxymethylcellulose or xylane had been studied. The cellulase and xylanase activities clearly depended on the distinct strain. The absence of distinct dependence of cellulase and xylanase activities on the species and organs of host plants was demonstrated. The majority of investigated strains of A. alternata did not possess a cellulase activity or the latter was low, but as a whole the phytopathogenic strains were more active than endophytic ones. Xylanase activity was considerable for the fungal strains of all trophyc groups. It was shown that the level of xylanase activity cannot become a biochemical marker of the A. alternata isolate pathogenicity.

  3. Immobilization of cellulases on magnetic particles to enable enzyme recycling during hydrolysis of lignocellulose

    DEFF Research Database (Denmark)

    Alftrén, Johan

    for biochemicals and biofuels production because of the great abundance of the feedstock and the fact that it is a waste material and does not directly compete with food production. Lignocellulose consists of cellulose (the most prevalent component), hemicellulose and lignin and the polymeric sugars (in cellulose...... approach to decrease the costs of the cellulases could be to immobilize the enzymes on particles and thereby enable enzyme re-use. However, recycling of immobilized cellulases using common separation unit operations such as centrifugation or filtration may be difficult when dealing with lignocellulosic...... feedstocks containing insolubles. This could potentially be overcome by immobilizing the cellulases on magnetically susceptible particles. Consequently, the immobilized cellulases could be magnetically recovered and recycled for a new cycle of enzymatic hydrolysis of cellulose. The main objective...

  4. Prediction of Color Properties of Cellulase-Treated 100% Cotton Denim Fabric

    Directory of Open Access Journals (Sweden)

    C. W. Kan

    2013-01-01

    Full Text Available Artificial neural network (ANN model was used for predicting colour properties of 100% cotton denim fabrics, including colour yield (in terms of K/S value and CIE L*, a*, b*, C*, and h° values, under the influence of cellulase treatment with various combinations of cellulase processing parameters. Variables examined in the ANN model included treatment temperature, treatment time, pH, mechanical agitation, and fabric yarn twist level. The ANN model was compared with a linear regression model where the ANN model produced superior results in the prediction of colour properties of cellulase-treated 100% cotton denim fabrics. The relative importance of the examined factors influencing colour properties was also investigated. The analysis revealed that cellulase treatment processing parameters played an important role in affecting the colour properties of the treated 100% denim cotton fabrics.

  5. Incorporation of fungal cellulases in bacterial minicellulosomes yields viable, synergistically acting celluloytic complexes

    NARCIS (Netherlands)

    Mingardon, F.; Chanal, A.; Lopez Contreras, A.M.; Dray, C.; Bayer, E.A.; Fierobe, H.P.

    2007-01-01

    Artificial designer minicellulosomes comprise a chimeric scaffoldin that displays an optional cellulose-binding module (CBM) and bacterial cohesins from divergent species which bind strongly to enzymes engineered to bear complementary dockerins. Incorporation of cellulosomal cellulases from Clostrid

  6. Character of Cellulase Activity in the Guts of Flagellate-Free Termites with Different Feeding Habits

    OpenAIRE

    Li, Zhi-qiang; Liu, Bing-Rong; Zeng, Wen-Hui; Xiao, Wei-Liang; Li, Qiu-Jian; Zhong, Jun-Hong

    2013-01-01

    Cellulose digestion in termites (Isoptera) is highly important for ecological reasons and applications in biofuel conversion. The speciose Termitidae family has lost flagellates in the hindgut and developed diverse feeding habits. To address the response of cellulase activity to the differentiation of feeding habits, a comparative study of the activity and distribution of composite cellulases, endo-β-1, 4-glucanase, and β-glucosidase was performed in seven common flagellate-free termites with...

  7. Cellulase Activity in Solid State Fermentation of Palm Kernel Cake with Trichoderma sp.

    Directory of Open Access Journals (Sweden)

    Massaud, M. B. N.

    2012-01-01

    Full Text Available Aims: The effect of different types of fungal inocula to the cellulase activity measured on palm kernel cake (PKC was studied. Methodology and Results: Isolate Pro-A1 which was identified as Trichoderma sp. was selected as a potential producer of cellulase via solid state fermentation technique (SSF. Two types of PKCs were used; raw PKC (containing residual oil and defatted PKC. The PKCs were inoculated with different concentrations of conidia and varying amounts (g of solid mycelia plugs (SMP for SSF. The effect of ultrafiltered crude fungal filtrate (CFF as inocula was also being tested. The highest cellulase activity of 2.454 FPU/mL was detected with 60% (wt/wt SMP applied to the raw PKC. Conversely, 2.059 FPU/mL of cellulase activity was measured when 80% (wt/wt of SMP was applied to the defatted PKC which is 62.3% higher than the untreated defatted PKC; and more than 100% increase in enzymatic activity compared to raw PKC. The cellulase activity in the SSF inoculated with 8 x 106 conidia /mL and 12 x 106 conidia /mL were 1.704 FPU/mL for raw PKC and 1.856 FPU/mL for defatted PKC, an enhancement of about 46% from uninoculated batch. Inoculation with CFF bears corresponding maximum improvement of the cellulase activity on both PKCs of 13.58% (raw and 2.86% (defatted. Conclusion, significance and impact of study: The current study proves that Trichoderma sp. in the form of SMP can enhance the cellulase activity on PKCs effectively with more than 100% increment. Fungal conidia are also a better choice in enhancing cellulase activity of Trichoderma sp. permitted that the PKC used is devoid of oil. From this study, Trichoderma sp. holds the potential of converting lignocellulosic materials into products of commercial and industrial values such as glucose and other biofuels.

  8. Production of Alkaline Cellulase by Fungi Isolated from an Undisturbed Rain Forest of Peru

    OpenAIRE

    Karin Vega; Gretty K. Villena; Sarmiento, Victor H.; Yvette Ludeña; Nadia Vera; Marcel Gutiérrez-Correa

    2012-01-01

    Alkaline cellulase producing fungi were isolated from soils of an undisturbed rain forest of Peru. The soil dilution plate method was used for the enumeration and isolation of fast growing cellulolytic fungi on an enriched selective medium. Eleven out of 50 different morphological colonies were finally selected by using the plate clearing assay with CMC as substrate at different pH values. All 11 strains produced cellulases in liquid culture with activities at alkaline pH values without an ap...

  9. Efficient leaching of cellulases produced by Trichoderma harzianum in solid state fermentation

    OpenAIRE

    Roussos, Sevastianos; Raimbault, Maurice; Saucedo-Castaneda, G.; Lonsane, B.K.

    1992-01-01

    Recovery of cellulases from solid state culture of #Trichoderma harzianum$ was efficiently achieved by hydraulic pressing. Pressing of fermented solids yielded carboxymethyl-cellulase (CMCase) extraction efficiency of 71% and a ratio of leachate to fermented solids of 0.58 (v/w). Addition of water to pressed solids and second pressing improved the efficiency (95%) with simultaneous increase in the ratio to 1.16 (v/w). The overall extraction of filter paper activity was lower (85%) than that o...

  10. Synergistic and Dose-Controlled Regulation of Cellulase Gene Expression in Penicillium oxalicum

    OpenAIRE

    Zhonghai Li; Guangshan Yao; Ruimei Wu; Liwei Gao; Qinbiao Kan; Meng Liu; Piao Yang; Guodong Liu; Yuqi Qin; Xin Song; Yaohua Zhong; Xu Fang; Yinbo Qu

    2015-01-01

    Filamentous fungus Penicillium oxalicum produces diverse lignocellulolytic enzymes, which are regulated by the combinations of many transcription factors. Here, a single-gene disruptant library for 470 transcription factors was constructed and systematically screened for cellulase production. Twenty transcription factors (including ClrB, CreA, XlnR, Ace1, AmyR, and 15 unknown proteins) were identified to play putative roles in the activation or repression of cellulase synthesis. Most of these...

  11. Cellulase recycling after high-solids simultaneous saccharification and fermentation of combined pretreated corncob

    Directory of Open Access Journals (Sweden)

    Ruoyu eDu

    2014-06-01

    Full Text Available Despite the advantageous prospect of second-generation bioethanol, its final commercialization must overcome the primary cost impediment due to enzyme assumption. To solve this problem, this work achieves high-concentration ethanol fermentation and multi-round cellulase recycling through process integration. The optimal time and temperature of the re-adsorption process were determined by monitoring the adsorption kinetics of cellulases. Both glucose and cellobiose inhibited cellulase adsorption. After 96 h of ethanol fermentation, 40% of the initial cellulase remained in the broth, from which 62.5% of the cellulase can be recycled and reused in fresh substrate re-adsorption for 90 min. Under optimum conditions, i.e., pH 5.0, dry matter loading of 15 wt%, cellulase loading of 45 FPU/g glucan, two cycles of fermentation and re-adsorption can yield two-fold increased ethanol outputs and reduce enzyme costs by over 50%. The ethanol concentration in each cycle can be achieved at levels greater than 40 g/L.

  12. Production of Alkaline Cellulase by Fungi Isolated from an Undisturbed Rain Forest of Peru

    Directory of Open Access Journals (Sweden)

    Karin Vega

    2012-01-01

    Full Text Available Alkaline cellulase producing fungi were isolated from soils of an undisturbed rain forest of Peru. The soil dilution plate method was used for the enumeration and isolation of fast growing cellulolytic fungi on an enriched selective medium. Eleven out of 50 different morphological colonies were finally selected by using the plate clearing assay with CMC as substrate at different pH values. All 11 strains produced cellulases in liquid culture with activities at alkaline pH values without an apparent decrease of them indicating that they are true alkaline cellulase producers. Aspergillus sp. LM-HP32, Penicillium sp. LM-HP33, and Penicillium sp. LM-HP37 were the best producers of FP cellulase (>3 U mL−1 with higher specific productivities (>30 U g−1 h−1. Three strains have been found suitable for developing processes for alkaline cellulase production. Soils from Amazonian rain forests are good sources of industrial fungi with particular characteristics. The results of the present study are of commercial and biological interest. Alkaline cellulases may be used in the polishing and washing of denim processing of the textile industry.

  13. The Effects of Bioprocess Parameters on Cellulase Production with Trichoderma viride CMIT35

    Directory of Open Access Journals (Sweden)

    Teodor Vintila

    2010-05-01

    Full Text Available Fungal cellulases are well-studied, and have various applications in industry, health or agriculture. Species of Trichoderma can produce substantial amounts of endoglucanase, exoglucanase (saccharifying cellulases, and some strains are able to produce important quantities of β-glucosidase. A number of fungi were isolated abroad and screened for cellulolytic potential. In this study, the kinetics of cellulase production from an indigenous strain of T. viride CMIT35 is reported. Product formation parameters of different types of cellulases indicate that the studied strain of T. viride is capable of producing important levels of cellulases when grown on Mandels medium with wheat bran as carbon source. Furthermore, it was observed that production of endoglucanase reaches its maximum during exponential phase of growth, while exoglucanase during the stationary phase. Enzyme production by solid-state fermentation was also investigated and found to be more efficient than liquid state fermentation. High production of cellulase was noted at the following parameters for liquid cultures: 4% wheat bran, 5% inoculum, 180 r.p.m. agitation, pH 5; and 60% humidity in the case of solid state fermentation.

  14. Synergistic effect of cellulase and xylanase during hydrolysis of natural lignocellulosic substrates.

    Science.gov (United States)

    Song, Hui-Ting; Gao, Yuan; Yang, Yi-Min; Xiao, Wen-Jing; Liu, Shi-Hui; Xia, Wu-Cheng; Liu, Zi-Lu; Yi, Li; Jiang, Zheng-Bing

    2016-11-01

    Synergistic combination of cellulase and xylanase has been performed on pre-treated substrates in many previous studies, while few on natural substrates. In this study, three unpretreated lignocellulosic substrates were studied, including corncob, corn stover, and rice straw. The results indicated that when the mixed cellulase and xylanase were applied, reducing sugar concentrations were calculated as 19.53, 15.56, and 17.35mg/ml, respectively, based on the 3,5 dinitrosalicylic acid (DNS) method. Compared to the treatment with only cellulose, the hydrolysis yields caused by mixed cellulase and xylanase were improved by 133%, 164%, and 545%, respectively. In addition, the conversion yield of corncob, corn stover, and rice straw by cellulase-xylanase co-treatment reached 43.9%, 48.5%, and 40.2%, respectively, based on HPLC analysis, which confirmed the synergistic effect of cellulase-xylanase that was much higher than either of the single enzyme treatment. The substrate morphology was also evaluated to explore the synergistic mechanism of cellulase-xylanase. PMID:27560367

  15. Catalysis of Rice Straw Hydrolysis by the Combination of Immobilized Cellulase from Aspergillus niger on β-Cyclodextrin-Fe3O4 Nanoparticles and Ionic Liquid

    Directory of Open Access Journals (Sweden)

    Po-Jung Huang

    2015-01-01

    Full Text Available Cellulase from Aspergillus niger was immobilized onto β-cyclodextrin-conjugated magnetic particles by silanization and reductive amidation. The immobilized cellulase gained supermagnetism due to the magnetic nanoparticles. Ninety percent of cellulase was immobilized, but the activity of immobilized cellulase decreased by 10%. In this study, ionic liquid (1-butyl-3-methylimidazolium chloride was introduced into the hydrolytic process because the original reaction was a solid-solid reaction. The activity of immobilized cellulase was improved from 54.87 to 59.11 U g immobilized cellulase−1 at an ionic liquid concentration of 200 mM. Using immobilized cellulase and ionic liquid in the hydrolysis of rice straw, the initial reaction rate was increased from 1.629 to 2.739 g h−1 L−1. One of the advantages of immobilized cellulase is high reusability—it was usable for a total of 16 times in this study. Compared with free cellulase, magnetized cellulase can be recycled by magnetic field and the activity of immobilized cellulase was shown to remain at 85% of free cellulase without denaturation under a high concentration of glucose (15 g L−1. Therefore, immobilized cellulase can hydrolyze rice straw continuously compared with free cellulase. The amount of harvested glucose can be up to twentyfold higher than that from the hydrolysis by free cellulase.

  16. SCREENING AND OPTIMIZATION OF CULTURE CONDITIONS FOR CELLULASE PRODUCTION BY ASPERGILLUS NIGER NSPR012 IN SUBMERGED FERMENTATION

    Directory of Open Access Journals (Sweden)

    Juliet Bamidele Akinyele

    2014-12-01

    Full Text Available This study aimed at screening of selected fungal strains and optimization of process parameters for cellulases production in submerged fermentation. Aspergillus niger NSPR012 was selected for further studies as the most potent in producing cellulase of high activity. Utilization of various agro-wastes as substitute tocarboxy methyl cellulose (CMC for cellulase production was also investigated. Among tested carbon sources, banana peels at a concentration of 5% was found to be the most effective carbon source. The cellulase production by Aspergillus niger NSPR012 in mineral salt medium attained maximum after 96 h of incubation. Maximum cellulase activity (0.466µmol/min/mL was obtained with locust beans as the best organic nitrogen source. The optimum incubation temperature and initial pH were 37°C and 5.5, respectively. With this information, banana peels could have good biotechnological potential for bio-products formation in which cellulase is one.

  17. Effect of Different Cellulase and Pectinase Enzyme Treatments on Protoplast Isolation and Viability in Lilium ledebeourii Bioss.

    Directory of Open Access Journals (Sweden)

    Esmaeil CHAMANI

    2012-11-01

    Full Text Available For overcoming interspecific incompatibility, protoplast combination method is a proper procedure for making a new plant withdesired traits. For this purpose, protoplast preparation is a first and important step. Hence, experiments were conducted to evaluatevarious combinations of cellulose, pectinase and their treatment times on protoplast production and protoplast viability in Liliumledebeourii Bioss. The results of experiment revealed that the protoplast yield was significantly affected by different treatment levels.Cellulase at 4% gave the highest numbers of protoplasts at 3.71×105 protoplast/g FW. Pectinase at 1% gave the highest numbers ofprotoplast. For treatment times, the highest yield of protoplast was with leaf explants treated for 24 h. Analysis of variance indicated thatconcentration, time and three-way interaction of cellulase, pectinase and time were significant at p<0.01. Cellulase at 4% and pectinase at0.2% for 24 h gave the highest viability. Interactions of cellulase × pectinase, cellulase × time, pectinase × time and cellulase × pectinase× treatment time were significant at P≤0.05 for protoplast number. The highest and lowest protoplast numbers were produced in mediacontaining 4% cellulase and 1% pectinase for 24 h (6.65×105 protoplast/g FW and 1% cellulase and 0.2% pectinase for 12 h, respectively.It’s concluded that, the best treatment for isolation of Lilium protoplast was 4% cellulase and 1% pectinase for 24 h.

  18. Glycosylation Helps Cellulase Enzymes Bind to Plant Cell Walls (Fact Sheet)

    Energy Technology Data Exchange (ETDEWEB)

    2012-06-01

    Computer simulations suggest a new strategy to design enhanced enzymes for biofuels production. Large-scale computer simulations predict that the addition of glycosylation on carbohydrate-binding modules can dramatically improve the binding affinity of these protein domains over amino acid mutations alone. These simulations suggest that glycosylation can be used as a protein engineering tool to enhance the activity of cellulase enzymes, which are a key component in the conversion of cellulose to soluble sugars in the production of biofuels. Glycosylation is the covalent attachment of carbohydrate molecules to protein side chains, and is present in many proteins across all kingdoms of life. Moreover, glycosylation is known to serve a wide variety of functions in biological recognition, cell signaling, and metabolism. Cellulase enzymes, which are responsible for deconstructing cellulose found in plant cell walls to glucose, contain glycosylation that when modified can affect enzymatic activity-often in an unpredictable manner. To gain insight into the role of glycosylation on cellulase activity, scientists at the National Renewable Energy Laboratory (NREL) used computer simulation to predict that adding glycosylation on the carbohydrate-binding module of a cellulase enzyme dramatically boosts the binding affinity to cellulose-more than standard protein engineering approaches in which amino acids are mutated. Because it is known that higher binding affinity in cellulases leads to higher activity, this work suggests a new route to designing enhanced enzymes for biofuels production. More generally, this work suggests that tuning glycosylation in cellulase enzymes is a key factor to consider when engineering biochemical conversion processes, and that more work is needed to understand how glycosylation affects cellulase activity at the molecular level.

  19. The prospects of cellulase-producing bacteria for the bioconversion of lignocellulosic biomass

    Directory of Open Access Journals (Sweden)

    Miranda Maki, Kam Tin Leung, Wensheng Qin

    2009-01-01

    Full Text Available Lignocellulosic biomass is a renewable and abundant resource with great potential for bioconversion to value-added bioproducts. However, the biorefining process remains economically unfeasible due to a lack of biocatalysts that can overcome costly hurdles such as cooling from high temperature, pumping of oxygen/stirring, and, neutralization from acidic or basic pH. The extreme environmental resistance of bacteria permits screening and isolation of novel cellulases to help overcome these challenges. Rapid, efficient cellulase screening techniques, using cellulase assays and metagenomic libraries, are a must. Rare cellulases with activities on soluble and crystalline cellulose have been isolated from strains of Paenibacillus and Bacillus and shown to have high thermostability and/or activity over a wide pH spectrum. While novel cellulases from strains like Cellulomonas flavigena and Terendinibacter turnerae, produce multifunctional cellulases with broader substrate utilization. These enzymes offer a framework for enhancement of cellulases including: specific activity, thermalstability, or end-product inhibition. In addition, anaerobic bacteria like the clostridia offer potential due to species capable of producing compound multienzyme complexes called cellulosomes. Cellulosomes provide synergy and close proximity of enzymes to substrate, increasing activity towards crystalline cellulose. This has lead to the construction of designer cellulosomes enhanced for specific substrate activity. Furthermore, cellulosome-producing Clostridium thermocellum and its ability to ferment sugars to ethanol; its amenability to co-culture and, recent advances in genetic engineering, offer a promising future in biofuels. The exploitation of bacteria in the search for improved enzymes or strategies provides a means to upgrade feasibility for lignocellulosic biomass conversion, ultimately providing means to a 'greener' technology.

  20. Synergistic and Dose-Controlled Regulation of Cellulase Gene Expression in Penicillium oxalicum.

    Science.gov (United States)

    Li, Zhonghai; Yao, Guangshan; Wu, Ruimei; Gao, Liwei; Kan, Qinbiao; Liu, Meng; Yang, Piao; Liu, Guodong; Qin, Yuqi; Song, Xin; Zhong, Yaohua; Fang, Xu; Qu, Yinbo

    2015-09-01

    Filamentous fungus Penicillium oxalicum produces diverse lignocellulolytic enzymes, which are regulated by the combinations of many transcription factors. Here, a single-gene disruptant library for 470 transcription factors was constructed and systematically screened for cellulase production. Twenty transcription factors (including ClrB, CreA, XlnR, Ace1, AmyR, and 15 unknown proteins) were identified to play putative roles in the activation or repression of cellulase synthesis. Most of these regulators have not been characterized in any fungi before. We identified the ClrB, CreA, XlnR, and AmyR transcription factors as critical dose-dependent regulators of cellulase expression, the core regulons of which were identified by analyzing several transcriptomes and/or secretomes. Synergistic and additive modes of combinatorial control of each cellulase gene by these regulatory factors were achieved, and cellulase expression was fine-tuned in a proper and controlled manner. With one of these targets, the expression of the major intracellular β-glucosidase Bgl2 was found to be dependent on ClrB. The Bgl2-deficient background resulted in a substantial gene activation by ClrB and proved to be closely correlated with the relief of repression mediated by CreA and AmyR during cellulase induction. Our results also signify that probing the synergistic and dose-controlled regulation mechanisms of cellulolytic regulators and using it for reconstruction of expression regulation network (RERN) may be a promising strategy for cellulolytic fungi to develop enzyme hyper-producers. Based on our data, ClrB was identified as focal point for the synergistic activation regulation of cellulase expression by integrating cellulolytic regulators and their target genes, which refined our understanding of transcriptional-regulatory network as a "seesaw model" in which the coordinated regulation of cellulolytic genes is established by counteracting activators and repressors. PMID:26360497

  1. Proteomics based compositional analysis of complex cellulase-hemicellulase mixtures

    Energy Technology Data Exchange (ETDEWEB)

    Chundawat, Shishir P.; Lipton, Mary S.; Purvine, Samuel O.; Uppugundla, Nirmal; Gao, Dahai; Balan, Venkatesh; Dale, Bruce E.

    2011-10-07

    Efficient deconstruction of cellulosic biomass to fermentable sugars for fuel and chemical production is accomplished by a complex mixture of cellulases, hemicellulases and accessory enzymes (e.g., >50 extracellular proteins). Cellulolytic enzyme mixtures, produced industrially mostly using fungi like Trichoderma reesei, are poorly characterized in terms of their protein composition and its correlation to hydrolytic activity on cellulosic biomass. The secretomes of commercial glycosyl hydrolase producing microbes was explored using a proteomics approach with high-throughput quantification using liquid chromatography-tandem mass spectrometry (LC-MS/MS). Here, we show that proteomics based spectral counting approach is a reasonably accurate and rapid analytical technique that can be used to determine protein composition of complex glycosyl hydrolase mixtures that also correlates with the specific activity of individual enzymes present within the mixture. For example, a strong linear correlation was seen between Avicelase activity and total cellobiohydrolase content. Reliable, quantitative and cheaper analytical methods that provide insight into the cellulosic biomass degrading fungal and bacterial secretomes would lead to further improvements towards commercialization of plant biomass derived fuels and chemicals.

  2. Surface activation of dyed fabric for cellulase treatment.

    Science.gov (United States)

    Schimper, Christian B; Ibanescu, Constanta; Bechtold, Thomas

    2011-10-01

    Surface activation of fabric made from cellulose fibres, such as viscose, lyocell, modal fibres and cotton, can be achieved by printing of a concentrated NaOH-containing paste. From the concentration of reducing sugars formed in solution, an increase in intensity of the cellulase hydrolysis by a factor of six to eight was observed, which was mainly concentrated at the activated parts of the fabric surface. This method of local activation is of particular interest for modification of materials that have been dyed with special processes to attain an uneven distribution of dyestuff within the yarn cross-section, e.g., indigo ring-dyed denim yarn for jeans production. Fabrics made from regenerated cellulose fibres were used as model substrate to express the effects of surface activation on indigo-dyed material. Wash-down experiments on indigo-dyed denim demonstrated significant colour removal from the activated surface at low overall weight loss of 4-5%. The method is of relevance for a more eco-friendly processing of jeans in the garment industry.

  3. Ruminococcus flavefaciens 007C cellulosomes and cellulase consortium

    Directory of Open Access Journals (Sweden)

    Maša VODOVNIK

    2015-11-01

    Full Text Available Ruminococcus flavefaciens is among the most important cellulolytic bacterial species in rumen and gastrointestinal tract of monogastric herbivorous animals. Its efficiency in degradation of (hemicellulosic substrates is associated with the production of remarkably intricate extracellular multienzyme complexes, named cellulosomes. In the present work we investigated the cellulolytic system of 007C. The bioinformatic analysis of the draft genome sequence revealed identical organization of sca gene cluster as has previously been found in four other strains of R. flavefaciens. The cluster consists of five genes in the following order: scaC-scaA-scaB-cttA-scaE. The cellulases of R. flavefaciens 007C belong to four families of glycoside hydrolases, namely GH48, GH44, GH9 in GH5. Majority of these enzymes are putative endoglucanases, belonging to families GH5 and GH9, whereas only one gene encoding GH44 and GH48 was found. Apart from catalytic domains, most of these proteins also contain dockerins – signature sequences, which indicate their attachement to cellulosomes. On the other hand, carbohydrate-binding modules were only found coupled to GH9 catalytic domains. Zymogram analysis showed that larger endoglucanases were mostly constitutively expressed, wheras smaller enzymes were only detected in later phases of Avicel-grown cultures.

  4. Genome shuffling of Aspergillus glaucus HGZ-2 for enhanced cellulase production.

    Science.gov (United States)

    Zhao, Yuping; Jiang, Changxing; Yu, Hupeng; Fang, Fang; Yang, Jingzhu

    2014-10-01

    The production of cellulase from Aspergillus glaucus HGZ-2 was improved by using genome shuffling. The starting populations, obtained by UV irradiation, were subjected to recursive protoplast fusion. The optimal conditions for protoplast formation and regeneration were 7 mg/ml snailase and 5 mg/ml cellulase at 34 °C for 3.0 h using 0.7 M NaCl as an osmotic stabilizer. The protoplasts were inactivated under UV for 30 min or heated at 50 °C for 50 min, and a fusant probability of about 100 % was observed. The positive colonies were created by fusing the inactivated protoplasts. The optimal conditions for protoplast fusion were PEG6000 concentration of 35 %, CaCl2 concentration of 0.02 M, and incubation time of 12 min. After two rounds of genome shuffling, one strain (Y) was obtained. Its filter paper cellulase (FPase) and carboxymethyl cellulase (CMCase) activity reached 71 and 70 U/ml, respectively, which were increased by 1.95-fold and 1.72-fold in comparison with that of its ancestor strain. The results indicated that genome shuffling was an efficient means for the improved production of cellulases by A. glaucus HGZ-2. PMID:25099375

  5. Structure of the catalytic domain of the Clostridium thermocellum cellulase CelT.

    Science.gov (United States)

    Kesavulu, Muppuru M; Tsai, Jia Yin; Lee, Hsiao Lin; Liang, Po Huang; Hsiao, Chwan Deng

    2012-03-01

    Cellulases hydrolyze cellulose, a major component of plant cell walls, to oligosaccharides and monosaccharides. Several Clostridium species secrete multi-enzyme complexes (cellulosomes) containing cellulases. C. thermocellum CelT, a family 9 cellulase, lacks the accessory module(s) necessary for activity, unlike most other family 9 cellulases. Therefore, characterization of the CelT structure is essential in order to understand its catalytic mechanism. Here, the crystal structure of free CelTΔdoc, the catalytic domain of CelT, is reported at 2.1 Å resolution. Its structure differs in several aspects from those of other family 9 cellulases. CelTΔdoc contains an additional α-helix, α-helices of increased length and two additional surface-exposed β-strands. It also contains three calcium ions instead of one as found in C. cellulolyticum Cel9M. CelTΔdoc also has two flexible loops at the open end of its active-site cleft. Movement of these loops probably allows the substrate to access the active site. CelT is stable over a wide range of pH and temperature conditions, suggesting that CelT could be used to convert cellulose biomass into biofuel.

  6. Application of Statistical Design for the Production of Cellulase by Trichoderma reesei Using Mango Peel.

    Science.gov (United States)

    Saravanan, P; Muthuvelayudham, R; Viruthagiri, T

    2012-01-01

    Optimization of the culture medium for cellulase production using Trichoderma reesei was carried out. The optimization of cellulase production using mango peel as substrate was performed with statistical methodology based on experimental designs. The screening of nine nutrients for their influence on cellulase production is achieved using Plackett-Burman design. Avicel, soybean cake flour, KH(2)PO(4), and CoCl(2)·6H(2)O were selected based on their positive influence on cellulase production. The composition of the selected components was optimized using Response Surface Methodology (RSM). The optimum conditions are as follows: Avicel: 25.30 g/L, Soybean cake flour: 23.53 g/L, KH(2)PO(4): 4.90 g/L, and CoCl(2)·6H(2)O: 0.95 g/L. These conditions are validated experimentally which revealed an enhanced Cellulase activity of 7.8 IU/mL.

  7. Application of Statistical Design for the Production of Cellulase by Trichoderma reesei Using Mango Peel

    Directory of Open Access Journals (Sweden)

    P. Saravanan

    2012-01-01

    Full Text Available Optimization of the culture medium for cellulase production using Trichoderma reesei was carried out. The optimization of cellulase production using mango peel as substrate was performed with statistical methodology based on experimental designs. The screening of nine nutrients for their influence on cellulase production is achieved using Plackett-Burman design. Avicel, soybean cake flour, KH2PO4, and CoCl2·6H2O were selected based on their positive influence on cellulase production. The composition of the selected components was optimized using Response Surface Methodology (RSM. The optimum conditions are as follows: Avicel: 25.30 g/L, Soybean cake flour: 23.53 g/L, KH2PO4: 4.90 g/L, and CoCl2·6H2O: 0.95 g/L. These conditions are validated experimentally which revealed an enhanced Cellulase activity of 7.8 IU/mL.

  8. Antimicrobial and Antioxidant Activity of Chitosan/Hydroxypropyl Methylcellulose Film-Forming Hydrosols Hydrolyzed by Cellulase

    Directory of Open Access Journals (Sweden)

    Anna Zimoch-Korzycka

    2016-09-01

    Full Text Available The aim of this study was to evaluate the impact of cellulase (C on the biological activity of chitosan/hydroxypropyl methylcellulose (CH/HPMC film-forming hydrosols. The hydrolytic activity of cellulase in two concentrations (0.05% and 0.1% was verified by determination of the progress of polysaccharide hydrolysis, based on viscosity measurement and reducing sugar-ends assay. The 2,2-diphenyl-1-picrylhydrazyl (DPPH free radical scavenging effect, the ferric reducing antioxidant power (FRAP, and microbial reduction of Pseudomonas fluorescens, Yersinia enterocolitica, Bacillus cereus, and Staphylococcus aureus were studied. During the first 3 h of reaction, relative reducing sugar concentration increased progressively, and viscosity decreased rapidly. With increasing amount of enzyme from 0.05% to 0.1%, the reducing sugar concentration increased, and the viscosity decreased significantly. The scavenging effect of film-forming solutions was improved from 7.6% at time 0 and without enzyme to 52.1% for 0.1% cellulase after 20 h of reaction. A significant effect of cellulase addition and reaction time on antioxidant power of the tested film-forming solutions was also reported. Film-forming hydrosols with cellulase exhibited a bacteriostatic effect on all tested bacteria, causing a total reduction.

  9. Strategies to increase cellulase production with submerged fermentation using fungi isolated from the Brazilian biome

    Directory of Open Access Journals (Sweden)

    Genilton da Silva Faheina Junior

    2015-03-01

    Full Text Available Studies on new microbial sources of cellulase and accurate assessment of the steps that increase cellulase production are essential strategies to reduce costs of various processes using such enzymes. This study aimed at the selection of cellulase-producing filamentous fungi, and at the research of parameters involving cellulase production by submerged fermentation. The first test consisted of selecting the best cellulase-producing microorganisms (FPase in Erlenmeyer flasks containing 200 mL of specific growth medium. The next test was designed to further investigate the enzyme production in fermentation with four types of soluble sugars: glucose, lactose, sucrose and xylose. In bioreactor tests, three different inoculation strategies were analyzed. The best FPase activity was presented by the strain Trichoderma sp. CMIAT 041 (49.9 FPU L-1 and CMCase by the fungus Lasiodiplodia theobromae CMIAT 096 (350.0 U L-1. Sucrose proved to be the best option among the soluble sugars tested, with higher rates of FPase activity (49.9 FPU L-1 and CMCase (119.7 U L-1. The best inoculation strategy for the bioreactor was a spore suspension obtained from a semi-solid state fermentation of wheat bran for 72h.

  10. Optimization Studies on Cellulase Production from Bacillus Anthracis and Ochrobactrum Anthropic (YZ1 Isolated from Soil

    Directory of Open Access Journals (Sweden)

    Mohammad Badrud Duza

    2015-06-01

    Full Text Available The present study was carried out to demonstrate the optimization of growth conditions of bacteria with high cellulase activity. Cellulose degrading bacteria were isolated from soil samples collected from different areas of Guntur district, A.P. The bacteria were isolated using serial dilution and pour plate methods. The isolated bacteria were identified by morphological, biochemical and molecular procedures. The isolated bacterial species were screened for cellulase production in sub-merged fermentation process. The two tested bacterial species showed maximum yield for cellulase production. These two bacteria were identified as Bacillus anthracis and Ochrobactrum anthropi (YZ1. Supplementation of glucose, peptone, tyrosine and EDTA to the fermentation medium is favoured enzyme secretion. The optimum pH and temperature for the activity of crude enzyme was 8 and 45°C, respectively for Ochrobactrum anthropi (YZ1 while for Bacillus anthracis, it was 8 and 4°C, respectively.14% of inoculum level and 96 h of incubation period showed the maximum yield by both the species bacteria for cellulase production. The results of present study indicated that favorable fermentation conditions and the selection of a suitable growth medium played a key role in the production of cellulase from newly isolated Bacillus anthracis and Ochrobactrum anthropi (YZ1.

  11. Production, Optimization, and Characterization of Organic Solvent Tolerant Cellulases from a Lignocellulosic Waste-Degrading Actinobacterium, Promicromonospora sp. VP111.

    Science.gov (United States)

    Thomas, Lebin; Ram, Hari; Kumar, Alok; Singh, Ved Pal

    2016-07-01

    High costs of natural cellulose utilization and cellulase production are an industrial challenge. In view of this, an isolated soil actinobacterium identified as Promicromonospora sp. VP111 showed potential for production of major cellulases (CMCase, FPase, and β-glucosidase) utilizing untreated agricultural lignocellulosic wastes. Extensive disintegration of microcrystalline cellulose and adherence on it during fermentation divulged true cellulolytic efficiency of the strain. Conventional optimization resulted in increased cellulase yield in a cost-effective medium, and the central composite design (CCD) analysis revealed cellulase production to be limited by cellulose and ammonium sulfate. Cellulase activities were enhanced by Co(+2) (1 mM) and retained up to 60 °C and pH 9.0, indicating thermo-alkaline tolerance. Cellulases showed stability in organic solvents (25 % v/v) with log P ow  ≥ 1.24. Untreated wheat straw during submerged fermentation was particularly degraded and yielded about twofold higher levels of cellulases than with commercial cellulose (Na-CMC and avicel) which is especially economical. Thus, this is the first detailed report on cellulases from an efficient strain of Promicromonospora that was non-hemolytic, alkali-halotolerant, antibiotic (erythromycin, kanamycin, rifampicin, cefaclor, ceftazidime) resistant, multiple heavy metal (Mo(+6) = W(+6) > Pb(+2) > Mn(+2) > Cr(+3) > Sn(+2)), and organic solvent (n-hexane, isooctane) tolerant, which is industrially and environmentally valuable. PMID:26956574

  12. POTENTIAL OF THERMOSTABLE CELLULASES IN BIOPROCESSING OF SWITCHGRASS TO ETHANOL

    Directory of Open Access Journals (Sweden)

    Kasivishavanathan Muthukumarappan

    2011-04-01

    Full Text Available Switchgrass (Panicum virgatum, a perennial grass native to North America, is a promising energy crop for bioethanol production. The aim of this study was to optimize the enzymatic saccharification of thermo-mechanically pretreated switchgrass using a thermostable cellulase from Geobacillus sp. in a three-level, four-variable central composite design of response surface methodology. Different combinations of solids loadings (5 to 20%, enzyme loadings (5 to 20 FPU g-1 DM, temperature (50 to 70 oC, and time (36 to 96 h were investigated in a total of 30 experiments to model glucose release from switchgrass. All four factors had a significant impact on the cellulose conversion yields with a high coefficient of determination of 0.96. The use of higher solids loadings (20% and temperatures (70 oC during enzymatic hydrolysis proved beneficial for the significant reduction of hydrolysis times (2.67-times and enzyme loadings (4-times, with important implications for reduced capital and operating costs of ethanol production. At 20% solids, the increase of temperature of enzymatic hydrolysis from 50 oC to 70 oC increased glucose concentrations by 34%. The attained maximum glucose concentration of 23.52 g L-1 translates into a glucose recovery efficiency of 46% from the theoretical yield. Following red yeast fermentation, a maximum ethanol concentration of 11 g L-1 was obtained, accounting for a high glucose to ethanol fermentation efficiency of 92%. The overall conversion efficiency of switchgrass to ethanol was 42%.

  13. Interrelationships between cellulase activity and cellulose particle morphology

    Energy Technology Data Exchange (ETDEWEB)

    Olsen, Johan P.; Donohoe, Bryon S.; Borch, Kim; Westh, Peter; Resch, Michael G.

    2016-06-11

    It is well documented that the enzymatic hydrolysis of cellulose follows a reaction pattern where an initial phase of relatively high activity is followed by a gradual slow-down over the entire course of the reaction. This phenomenon is not readily explained by conventional factors like substrate depletion, product inhibition or enzyme instability. It has been suggested that the underlying reason for the loss of enzyme activity is connected to the heterogeneous structure of cellulose, but so far attempts to establish quantitative measures of such a correlation remain speculative. Here, we have carried out an extensive microscopy study of Avicel particles during extended hydrolysis with Hypocrea jecorina cellobiohydrolase 1 (CBH1) and endoglucanase 1 and 3 (EG1 and EG3) alone and in mixtures. We have used differential interference contrast microscopy and transmission electron microscopy to observe and quantify structural features at um and nm resolution, respectively. We implemented a semi-automatic image analysis protocol, which allowed us to analyze almost 3000 individual micrographs comprising a total of more than 300,000 particles. From this analysis we estimated the temporal development of the accessible surface area throughout the reaction. We found that the number of particles and their size as well as the surface roughness contributed to surface area, and that within the investigated degree of conversion (<30 %) this measure correlated linearly with the rate of reaction. Based on this observation we argue that cellulose structure, specifically surface area and roughness, plays a major role in the ubiquitous rate loss observed for cellulases.

  14. Recyclable thermoresponsive polymer-cellulase bioconjugates for biomass depolymerization.

    Science.gov (United States)

    Mackenzie, Katherine J; Francis, Matthew B

    2013-01-01

    -derived cellulases or to the separation of multiple species using polymers with different recovery temperatures. PMID:23270527

  15. Heterologous expression of cellulase genes in natural Saccharomyces cerevisiae strains.

    Science.gov (United States)

    Davison, Steffi A; den Haan, Riaan; van Zyl, Willem Heber

    2016-09-01

    Enzyme cost is a major impediment to second-generation (2G) cellulosic ethanol production. One strategy to reduce enzyme cost is to engineer enzyme production capacity in a fermentative microorganism to enable consolidated bio-processing (CBP). Ideally, a strain with a high secretory phenotype, high fermentative capacity as well as an innate robustness to bioethanol-specific stressors, including tolerance to products formed during pre-treatment and fermentation of lignocellulosic substrates should be used. Saccharomyces cerevisiae is a robust fermentative yeast but has limitations as a potential CBP host, such as low heterologous protein secretion titers. In this study, we evaluated natural S. cerevisiae isolate strains for superior secretion activity and other industrially relevant characteristics needed during the process of lignocellulosic ethanol production. Individual cellulases namely Saccharomycopsis fibuligera Cel3A (β-glucosidase), Talaromyces emersonii Cel7A (cellobiohydrolase), and Trichoderma reesei Cel5A (endoglucanase) were utilized as reporter proteins. Natural strain YI13 was identified to have a high secretory phenotype, demonstrating a 3.7- and 3.5-fold higher Cel7A and Cel5A activity, respectively, compared to the reference strain S288c. YI13 also demonstrated other industrially relevant characteristics such as growth vigor, high ethanol titer, multi-tolerance to high temperatures (37 and 40 °C), ethanol (10 % w/v), and towards various concentrations of a cocktail of inhibitory compounds commonly found in lignocellulose hydrolysates. This study accentuates the value of natural S. cerevisiae isolate strains to serve as potential robust and highly productive chassis organisms for CBP strain development. PMID:27470141

  16. Induction of cellulases and hemicellulases by tamarind (Tamarindus indica) kernel polysaccharide

    Energy Technology Data Exchange (ETDEWEB)

    Ghosh, B.S.; Kundu, A.B.

    1980-01-01

    Tamarind kernel polysaccharide (TKP) which is available in India in abundance could be used as an excellent substrate for production of cellulases, hemicellulases, ..beta..-glucosidase and ..beta..-xylosidase. A growth medium of a known cellulytic strain, Aspergillus terreus containing TKP has been optimized to get enhanced yields of cellulase and hemicellulase for practical application in jute manufacturing units around Calcutta. Of the large number of sugars and treated and untreated cellulosics tested, TKP was found to produce the highest amounts of cellulases, hemicellulases, ..beta..-glucosidase, ..beta..-xylosidase and extra-cellular protein. The effects of the use of different inorganic nitrogenous substances, nutrients and surfactants in the optimized medium have been tested. Both initial pH of the medium and final pH of the culture filtrate have been found to have a marked effect on enzyme production, especially ..beta..-glucosidase production.

  17. A pyranose dehydrogenase-based biosensor for kinetic analysis of enzymatic hydrolysis of cellulose by cellulases

    DEFF Research Database (Denmark)

    Cruys-Bagger, Nicolaj; Badino, Silke Flindt; Tokin, Radina Naytchova;

    2014-01-01

    A novel electrochemical enzyme biosensor was developed for real-time detection of cellulase activity when acting on their natural insoluble substrate, cellulose. The enzyme biosensor was constructed with pyranose dehydrongease (PDH) from Agaricus meleagris that was immobilized on the surface......-biosensor was shown to be anomer unspecific and it can therefore be used in kinetic studies over broad time-scales of both retaining- and inverting cellulases (in addition to enzyme cocktails). The biosensor was used for real-time measurements of the activity of the inverting cellobiohydrolase Cel6A from Hypocrea...... equation for processive cellulases, and it was found that the turnover for HjCel6A at saturating substrate concentration (i.e. maximal apparent specific activity) was similar (0.39–0.40 s−1) for the two substrates. Conversely, the substrate load at half-saturation was much lower for BMCC compared to Avicel...

  18. Is an organic nitrogen source needed for cellulase production by Trichoderma reesei Rut-C30?

    DEFF Research Database (Denmark)

    Rodríguez Gómez, Divanery; Hobley, Timothy John

    2013-01-01

    The effect of organic and inorganic nitrogen sources on Trichoderma reesei Rut-C30 cellulase production was investigated in submerged cultivations. Stirred tank bioreactors and shake flasks, with and without pH control, respectively, were employed. The experimental design involved the addition...... of individual organic nitrogen sources (soy peptone, glutamate, glycine and alanine) within a basal medium containing Avicel (i.e. micro crystalline cellulose) and ammonium sulphate. It was found that in the shake flask experiments, the highest cellulase activities (~0.1 ± 0.02 FPU ml−1) were obtained...... with media containing soy peptone (3–6 g l−1) and glutamate (3.6 g l−1). However, these improvements in the cellulase titers in the presence of the organic nitrogen sources appeared to be related to smaller changes in the pH of the medium. This was confirmed using stirred tank bioreactors with pH control...

  19. Cellulose hydrolysis by fungi. 2. Cellulase production by Trichoderma harzianum in liquid medium fermentation

    Energy Technology Data Exchange (ETDEWEB)

    Roussos, S.; Raimbault, M. (Laboratoire de Microbiologie ORSTOM, Centre de Recherche IRCHA, 91 - Vert-le-Petit (France))

    Microcrystalline cellulose (cellulose Avicel, Merck) supported growth of Trichoderma harzianum and induced production of cellulases in liquid cultures. After 50 h growth, the total cellulasic activities present in both the supernatant and the mycelium were 3,000 IU/l of carboxymethyl cellulose, 400 IU/l of filter paper activity, and 4 IU/l of cotton activity corresponding to 1.7 g/l of proteins. Cellulase production could be increased by a preliminary treatment of cellulose, and pH regulation during growth. The influence of inoculum concentration was studied and an optimum of 3 X 10/sup 7/ conidia/g dry weight of substrate was demonstrated. Using a synthetic culture medium, a soluble factor of germination was demonstrated which could be leached out by 3 successive washings of conidia.

  20. Deciphering the molecular mechanisms behind cellulase production in Trichoderma reesei, the hyper-cellulolytic filamentous fungus.

    Science.gov (United States)

    Shida, Yosuke; Furukawa, Takanori; Ogasawara, Wataru

    2016-09-01

    The filamentous fungus Trichoderma reesei is a potent cellulase producer and the best-studied cellulolytic fungus. A lot of investigations not only on glycoside hydrolases produced by T. reesei, but also on the machinery controlling gene expression of these enzyme have made this fungus a model organism for cellulolytic fungi. We have investigated the T. reesei strain including mutants developed in Japan in detail to understand the molecular mechanisms that control the cellulase gene expression, the biochemical and morphological aspects that could favor this phenotype, and have attempted to generate novel strains that may be appropriate for industrial use. Subsequently, we developed recombinant strains by combination of these insights and the heterologous-efficient saccharifing enzymes. Resulting enzyme preparations were highly effective for saccharification of various biomass. In this review, we present some of the salient findings from the recent biochemical, morphological, and molecular analyses of this remarkable cellulase hyper-producing fungus. PMID:27075508

  1. Synthesizing a Cellulase like Chimeric Protein by Recombinant Molecular Biology Techniques

    Science.gov (United States)

    Banerjee, Hirendra Nath; Krauss, Christopher; Smith, Valerie; Mahaffey, Kelly; Boston, Ava

    2016-01-01

    In order to meet the Renewable Fuels Standard demands for 30 billion gallons of biofuels by the end of 2020, new technologies for generation of cellulosic ethanol must be exploited. Breaking down cellulose by cellulase enzyme is very important for this purpose but this is not thermostable and degrades at higher temperatures in bioreactors. Towards creation of a more ecologically friendly method of rendering bioethanol from cellulosic waste, we attempted to produce recombinant higher temperature resistant cellulases for use in bioreactors. The project involved molecular cloning of genes for cellulose-degrading enzymes based on bacterial source, expressing the recombinant proteins in E. coli and optimizing enzymatic activity. We were able to generate in vitro bacterial expression systems to produce recombinant His-tag purified protein which showed cellulase like activity. PMID:27468362

  2. IMMOBILIZATION OF GLUCOSE OXIDASE AND CELLULASE BY CHITOSAN-POLYACRYLIC ACID COMPLEX

    Institute of Scientific and Technical Information of China (English)

    WANG Lingzhi; JIANG Yingyan; ZHANG Changde; HUANG Dexiu

    1990-01-01

    This study is concerned with chitosan-polyacrylic acid complex as a carrier to immobilize glucose oxidase (GOD) and cellulase. The optimum temperature of the immobilized GOD (IG) was determined to be 60 ℃ which is higher than that of the native GOD about 40 ℃ . The optimum temperature of the immobilized cellulase (IC) was determined to be about 30 ℃ higher than that of native cellulase. Both of the optimum pH of IG and IC shifted one pH unit to acid. Immobilized enzyme may be used in more wide pH range. Their storage life are much longer compared with their native states. Both of them can be reused at least 12 times.

  3. Purification and characterization of five cellulases and one xylanase from Penicillium brasilianum IBT 20888

    DEFF Research Database (Denmark)

    Jørgensen, Henning; Eriksson, T.; Borjesson, J.;

    2003-01-01

    The filamentous fungus Penicillium brasilianum IBT 20888 was cultivated on a mixture of 30 g l(-1) cellulose and 10 g l(-1) xylan for 111 h and the resulting culture filtrate was used for protein purification. From the cultivation broth, five cellulases and one xylanase were purified. Hydrolysis...... the cellulose-binding domain or an essential part of it. The basic xylanase (pI > 9) was only active towards xylan. Two of the purified cellulases with endoglucanase activity were partly sequenced and based on sequence homology with known enzymes they were classified as belonging to families 5 and 12...

  4. Cellulases, nucleic acids encoding them and methods for making and using them

    Science.gov (United States)

    Blum, David; Gemsch Cuenca, Joslin; Dycaico, Mark

    2013-04-23

    This invention relates to molecular and cellular biology and biochemistry. In one aspect, the invention provides polypeptides having cellulase activity, e.g., endoglucanase, cellobiohydrolase, mannanase and/or .beta.-glucosidase activity, polynucleotides encoding these polypeptides, and methods of making and using these polynucleotides and polypeptides. In one aspect, the invention is directed to polypeptides cellulase activity, e.g., endoglucanase, cellobiohydrolase, mannanase and/or .beta.-glucosidase activity, including thermostable and thermotolerant activity, and polynucleotides encoding these enzymes, and making and using these polynucleotides and polypeptides. The polypeptides of the invention can be used in a variety of pharmaceutical, agricultural, food and feed processing and industrial contexts.

  5. Cellulase occurs in multiple active forms in ripe avocado fruit mesocarp.

    Science.gov (United States)

    Kanellis, A K; Kalaitzis, P

    1992-02-01

    The existence of multiple forms of avocado (Persea americana Mill. cv Hass) cellulase in crude protein extracts of ripe avocado fruit is reported. Cellulase was separated into at least 11 multiple forms by native isoelectric focusing in the pH range between 4 and 7 and visualized by both activity staining using Congo red and immunostaining. The enzyme components were acidic proteins with isoelectric points in the range of pH 5.10 to 6.80, the predominant forms having isoelectric points of 5.60, 5.80, 5.95, and 6.20. All 11 forms were immunologically related with molecular masses of 54 kilodaltons.

  6. Effect of variations in growth parameters on cellulase activity of Trichoderma viride NSPR006 cultured on different wood-dusts

    Directory of Open Access Journals (Sweden)

    Olaniyi, O. O.

    2013-01-01

    Full Text Available Aims: The biotechnology research into agro wastes has been driven by the need to screen organisms for hyper-production of novel extracellular enzymes in which cellulase plays a significant role. Therefore, the aim of the study was to pre-screen selected fungal strains and optimize cultural conditions for cellulase production by Trichoderma viride NSPR006 cultured on pretreated sawdust as lignocellulosic substrate. Methodology and results: The selected fungal isolates namely Trichoderma viride NSPR006, Botrydiplodia NSPR007 and Acremonium butyri NSPR06B obtained from the culture collection of the Nigerian Stored Products Research Institute Ilorin, Kwara State, Nigeria were screened for the production of cellulase in mineral salt medium in which carboxymethylcellulose (CMC had been incorporated as the sole carbon source. All the tested fungal isolates produced cellulase with differences in the amount of enzyme production. Of all the selected fungal isolates screened, Trichoderma viride NSPR006 was found to yield highest cellulase activity compared to the other isolates. Among tested carbon sources, Pachyslasma tessmani wood dust at 3% level proved to the best for cellulase production. Of the entire tested organic nitrogen sources, locust beans were observed to yield maximum cellulase activity (0.194 µmol/min/mL. The optimum temperature, incubation time and pH for maximum cellulase production were 28 °C, 72 h and 6.5, respectively. Conclusion, significance and impact of study: Outcome of this study shows the effectiveness of pre-treatment of wood dust as low cost system for hyper-production of cellulase for industrial application. Also, the work revealed the use of pretreated wood dust as substitute to commercial substrate known to be expensive in cellulase production.

  7. Standard Assays Do Not Predict the Efficiency of Commercial Cellulase Preparations Towards Plant Materials

    NARCIS (Netherlands)

    Kabel, Mirjam A.; Maarel, Marc J.E.C. van der; Klip, Gert; Voragen, Alphons G.J.; Schols, Henk A.

    2006-01-01

    Commercial cellulase preparations are potentially effective for processing biomass feedstocks in order to obtain bioethanol. In plant cell walls, cellulose fibrils occur in close association with xylans (monocotyls) or xyloglucans (dicotyls). The enzymatic conversion of cellulose/xylans is a complex

  8. Standard assays do not predict the efficiency of commercial cellulase preparations towards plant materials

    NARCIS (Netherlands)

    Kabel, M.A.; Maarel, van der M.J.E.C.; Klip, G.; Voragen, A.G.J.; Schols, H.A.

    2006-01-01

    Commercial cellulase preparations are potentially effective for processing biomass feedstocks in order to obtain bioethanol. In plant cell walls, cellulose fibrils occur in close association with xylans (monocotyls) or xyloglucans (dicotyls). The enzymatic conversion of cellulose/xylans is a complex

  9. Cellulases and hemicellulases from endophytic Acremonium species and its application on sugarcane bagasse hydrolysis

    Science.gov (United States)

    The aim of this work was to have cellulase activity and hemicellulase activity screenings of endophyte Acremonium species (Acremonium zeae EA0802 and Acremonium sp. EA0810). Both fungi were cultivated in submerged culture (SC) containing L-arabinose, D-xylose, oat spelt xylan, sugarcane bagasse, or...

  10. Regulation of cellulase expression, sporulation, and morphogenesis by velvet family proteins in Trichoderma reesei.

    Science.gov (United States)

    Liu, Kuimei; Dong, Yanmei; Wang, Fangzhong; Jiang, Baojie; Wang, Mingyu; Fang, Xu

    2016-01-01

    Homologs of the velvet protein family are encoded by the ve1, vel2, and vel3 genes in Trichoderma reesei. To test their regulatory functions, the velvet protein-coding genes were disrupted, generating Δve1, Δvel2, and Δvel3 strains. The phenotypic features of these strains were examined to identify their functions in morphogenesis, sporulation, and cellulase expression. The three velvet-deficient strains produced more hyphal branches, indicating that velvet family proteins participate in the morphogenesis in T. reesei. Deletion of ve1 and vel3 did not affect biomass accumulation, while deletion of vel2 led to a significantly hampered growth when cellulose was used as the sole carbon source in the medium. The deletion of either ve1 or vel2 led to the sharp decrease of sporulation as well as a global downregulation of cellulase-coding genes. In contrast, although the expression of cellulase-coding genes of the ∆vel3 strain was downregulated in the dark, their expression in light condition was unaffected. Sporulation was hampered in the ∆vel3 strain. These results suggest that Ve1 and Vel2 play major roles, whereas Vel3 plays a minor role in sporulation, morphogenesis, and cellulase expression.

  11. Evaluation of Minimal Trichoderma reesei Cellulase Mixtures on Differently Pretreated Barley Straw Substrate

    DEFF Research Database (Denmark)

    Rosgaard, Lisa; Pedersen, Sven; Langston, J;

    2007-01-01

    The commercial cellulase product Celluclast 1.5, derived from Trichoderma reesei (Novozymes A/S, Bagsv ae rd, Denmark), is widely employed for hydrolysis of lignocellulosic biomass feedstocks. This enzyme preparation contains a broad spectrum of cellulolytic enzyme activities, most notably...

  12. Evaluation of minimal Trichoderma reesei cellulase mixtures on differently pretreated barley straw substrates

    DEFF Research Database (Denmark)

    Rosgaard, Lisa; Pedersen, Sven; Langston, Jim;

    2007-01-01

    The commercial cellulase product Celluclast 1.5, derived from Trichoderma reesei (Novozymes A/S, Bagsv ae rd, Denmark), is widely employed for hydrolysis of lignocellulosic biomass feedstocks. This enzyme preparation contains a broad spectrum of cellulolytic enzyme activities, most notably...

  13. Combinatorial Screening for Transgenic Yeasts with High Cellulase Activities in Combination with a Tunable Expression System.

    Directory of Open Access Journals (Sweden)

    Yoichiro Ito

    Full Text Available Combinatorial screening used together with a broad library of gene expression cassettes is expected to produce a powerful tool for the optimization of the simultaneous expression of multiple enzymes. Recently, we proposed a highly tunable protein expression system that utilized multiple genome-integrated target genes to fine-tune enzyme expression in yeast cells. This tunable system included a library of expression cassettes each composed of three gene-expression control elements that in different combinations produced a wide range of protein expression levels. In this study, four gene expression cassettes with graded protein expression levels were applied to the expression of three cellulases: cellobiohydrolase 1, cellobiohydrolase 2, and endoglucanase 2. After combinatorial screening for transgenic yeasts simultaneously secreting these three cellulases, we obtained strains with higher cellulase expressions than a strain harboring three cellulase-expression constructs within one high-performance gene expression cassette. These results show that our method will be of broad use throughout the field of metabolic engineering.

  14. The production of cellulase in a spouted bed fermentor using cells immobilized in biomass support particles.

    Science.gov (United States)

    Webb, C; Fukuda, H; Atkinson, B

    1986-01-01

    Continuous cellulase production by Trichoderma viride QM 9123, immobilized in 6 mm diameter, spherical, stainless steel biomass support particles, has been achieved using a medium containing glucose as the main carbon source. Experiments were carried out in a 10-L spouted bed fermentor. In this type of reactor-recycled broth is used to create a jet at the base of a bed of particles, causing the particles to spout and circulate. During the circulation, particles pass through a region of high shear near the jet inlet. This effectively prevents a buildup of excess biomass and thus enables steady-state conditions to be achieved during continuous operation. Continuous production of cellulase was achieved at significantly higher yield and productivity than in conventional systems. At a dilution rate of 0.15 h(-1) (nominal washout rate for freely suspended cells is 0.012 h(-1)), the yield of cellulase on glucose was 31% higher than that measured during batch operation, while the volumetric productivity (31.5 FPA U/L. h) was 53% greater than in the batch system. The specific cellulase productivity of the immobilized cells was more than 3 times that of freely suspended cells, showing that diffusional limitations can be beneficial. This offers significant opportunity for the further development of biomass support particles and associated bioreactors. PMID:18553840

  15. Plant carbohydrate binding module enhances activity of hybrid microbial cellulase enzyme

    Directory of Open Access Journals (Sweden)

    Caitlin Siobhan Byrt

    2012-11-01

    Full Text Available A synthetic, highly active cellulase enzyme suitable for in planta production may be a valuable tool for biotechnological approaches to develop transgenic biofuel crops with improved digestibility. Here, we demonstrate that the addition of a plant derived carbohydrate binding module (CBM to a synthetic glycosyl hydrolase (GH improved the activity of the hydrolase in releasing sugar from plant biomass. A CEL-HYB1-CBM enzyme was generated by fusing a hybrid microbial cellulase, CEL-HYB1, with the carbohydrate-binding module (CBM of the tomato (Solanum lycopersicum SlCel9C1 cellulase. CEL-HYB1 and CEL-HYB1-CBM enzymes were produced in vitro using Pichia pastoris and the activity of these enzymes was tested using CMC, MUC and native crystalline cellulose assays. The presence of the CBM substantially improved the endo-glucanase activity of CEL-HYB1, especially against the native crystalline cellulose encountered in Sorghum plant cell walls. These results indicate that addition of an endogenous plant derived CBM to cellulase enzymes may enhance hydrolytic activity.

  16. Cellulase production from treated oil palm empty fruit bunch degradation by locally isolated Thermobifida fusca

    Directory of Open Access Journals (Sweden)

    M. Nazli Naim

    2013-02-01

    Full Text Available The aim of this research was to evaluate the production of cellulases from locally isolated bacteria, Thermobifida fusca, using thermal and chemical treated oil palm empty fruit bunch (OPEFB as substrate in liquid-state fermentation (LSF. T. fusca was successfully isolated and was a dominant cellulase producer in OPEFB composting at the thermophilic stage. Analysis of the surface morphology of OPEFB samples using Scanning Electron Microscopy (SEM showed that the most significant changes after the combination of thermal and chemical pretreatment was the removal of silica bodies, and this observation was supported by X-ray Diffraction analysis (XRD, Fourier Transform Infrared (FTIR, and Thermogravimetric analysis (TG showing changes on the hemicelluloses, cellulose, and lignin structures throughout the pretreatment process. As a result of the pretreatment, higher cellulase production by T. fusca was obtained. The highest activity for CMCase, FPase, and β-glucosidase using optimally treated OPEFB were 0.24 U/mL, 0.34 U/mL, and 0.04 U/mL, respectively. Therefore, it can be suggested that the combination of chemical and thermal pretreatments enhances the degradation of OPEFB for subsequent use as fermentation substrate, contributing to a higher cellulases yield by T. fusca.

  17. Assessment of methods to determine minimal cellulase concentrations for efficient hydrolysis of cellulose

    Energy Technology Data Exchange (ETDEWEB)

    Hogan, C.M.; Mes-Hartree, M.; Saddler, J.N. (Forintek Canada Corp., Ottawa, ON (Canada). Biotechnology and Chemistry Dept.); Kushner, D.J. (Toronto Univ., Ontario (Canada). Dept. of Microbiology)

    1990-02-01

    The enzyme loading needed to achieve substrate saturation appeared to be the most economical enzyme concentration to use for hydrolysis, based on percentage hydrolysis. Saturation was reached at 25 filter paper units per gram substrate on Solka Floc BW300, as determined by studying (a) initial adsorption of the cellulase preparation onto the substrate, (b) an actual hydrolysis or (c) a combined hydrolysis and fermentation (CHF) process. Initial adsorption of the cellulases onto the substrate can be used to determine the minimal cellulase requirements for efficient hydrolysis since enzymes initially adsorbed to the substrate have a strong role in governing the overall reaction. Trichoderma harzianum E58 produces high levels of {beta}-glucosidase and is able to cause high conversion of Solka Floc BW300 to glucose without the need for exogenous {beta}-glucosidase. End-product inhibition of the cellulase and {beta}-glucosidase can be more effectively reduced by employing a CHF process than by supplemental {beta}-glucosidase. (orig.).

  18. Enhanced cellulase production by Trichoderma harzianum by cultivation on glycerol followed by induction on cellulosic substrates.

    Science.gov (United States)

    Delabona, Priscila da Silva; Lima, Deise Juliana; Robl, Diogo; Rabelo, Sarita Cândida; Farinas, Cristiane Sanchez; Pradella, José Geraldo da Cruz

    2016-05-01

    The use of glycerol obtained as an intermediate of the biodiesel manufacturing process as carbon source for microbial growth is a potential alternative strategy for the production of enzymes and other high-value bioproducts. This work evaluates the production of cellulase enzymes using glycerol for high cell density growth of Trichoderma harzianum followed by induction with a cellulosic material. Firstly, the influence of the carbon source used in the pre-culture step was investigated in terms of total protein secretion and fungal morphology. Enzymatic productivity was then determined for cultivation strategies using different types and concentrations of carbon source, as well as different feeding procedures (batch and fed-batch). The best strategy for cellulase production was then further studied on a larger scale using a stirred tank bioreactor. The proposed strategy for cellulase production, using glycerol to achieve high cell density growth followed by induction with pretreated sugarcane bagasse, achieved enzymatic activities up to 2.27 ± 0.37 FPU/mL, 106.40 ± 8.87 IU/mL, and 9.04 ± 0.39 IU/mL of cellulase, xylanase, and β-glucosidase, respectively. These values were 2 times higher when compared to the control experiments using glucose instead of glycerol. This novel strategy proved to be a promising approach for improving cellulolytic enzymes production, and could potentially contribute to adding value to biomass within the biofuels sector. PMID:26883662

  19. Hydrodynamic and kinetic study of cellulase production by Trichoderma reesei with pellet morphology.

    Science.gov (United States)

    Yu, Liang; Chao, Yapeng; Wensel, Pierre; Chen, Shulin

    2012-07-01

    Numerical simulations and experimental validation were performed to understand the effects of hydrodynamics on pellet formation and cellulase production by filamentous T. reesei. The constructed model combined a steady-state multiple reference frame (MRF) approach describing mechanical mixing, oxygen mass transfer, and non-Newtonian flow field with a transient sliding mesh approach and kinetics of oxygen consumption, pellet formation, and enzyme production. The model was experimentally validated at various agitation speeds in a two-impeller Rushton turbine fermentor. Results from simulation and experimentation showed that higher agitation speeds led to increases in the pellet diameter and the proportion of pelletized (vs. filamentous) forms of the biomass. It also led to increase in dissolved oxygen mass transfer rate in shear-thinning fluid and cellulase productivity. The extent of these increases varied considerably among agitation speeds. Pellet formation and morphology were presumably affected within a viscosity-dependent shear-rate range. Cellulase activity and cell viability were shown to be sensitive to impeller shear. A maximum cellulase activity of 3.5 IU/mL was obtained at 400 rpm, representing a twofold increase over that at 100 rpm. PMID:22252572

  20. Modification Effect of Cellulase on the Physicochemical Characteristic of Polysaccharides Edible Films

    Directory of Open Access Journals (Sweden)

    Anna Zimoch-Korzycka

    2015-01-01

    Full Text Available This study was conducted to assess hydrolytic influence of cellulase (C on the physicochemical stability of chitosan (CH/hydroxypropyl methylcellulose (HPMC films in time of storage (T. Initially, nine films were physically characterized by contact angle, water vapour permeability (WVP, water activity (aw, tensile test, dynamic mechanical thermal analysis (DMTA, and thermogravimetric analysis (TGA and chemically by Fourier Transform Infrared Spectrometry (FTIR. The contact angle results varied from 53.67° to 78.33°. The presence of the enzyme and passing time reduced the WVP from 8.46E-09 to 7.41E-09 g/s·m·Pa. The enzyme treatment improved elasticity but decreased tensile strength of films. After adding cellulase Tg was shifted to a higher temperature. Thermal stability of the films decreased with addition of cellulase and after prolonging storage time. FTIR analysis proved that chemical changes in polysaccharides structure were caused by cellulase incorporation in films composition, which may be observed in appearance of 1656 cm−1 band. The aw values did not change.

  1. ENZYMATIC KINETICS OF CELLULASES ISOLATED FROM SOIL BACTE RIA OF DOON VALLEY , UTTARAKHAND

    OpenAIRE

    Vinit; Ashutosh; Amit; Sonia

    2015-01-01

    Cellulases refers to a suite of enzymes produced chiefly by fungi , bacteria , and protozoans that catalyze cellulolysis which is the hydrolysis of cellulose . Cellulose is the most abundant natural polymer on earth . It is the structural component of the plant cell walls which helps in the hydrolysis of 1, 4 - beta - D - glycosidic linkages in cellulose, lichenin and cereal beta - D - glu...

  2. Immobilization of cellulase mixtures on magnetic particles for hydrolysis of lignocellulose and ease of recycling

    DEFF Research Database (Denmark)

    Alftrén, Johan; Hobley, Timothy John

    2014-01-01

    In the present study whole cellulase mixtures were covalently immobilized on non-porous magnetic particles to enable enzyme reuse. It was shown that CellicCTec2 immobilized on magnetic particles activated with cyanuric chloride gave the highest bead activity measured by mass of reducing sugar...

  3. Exploring the Mechanism Responsible for Cellulase Thermostability by Structure-Guided Recombination.

    Directory of Open Access Journals (Sweden)

    Chia-Jung Chang

    Full Text Available Cellulases from Bacillus and Geobacillus bacteria are potentially useful in the biofuel and animal feed industries. One of the unique characteristics of these enzymes is that they are usually quite thermostable. We previously identified a cellulase, GsCelA, from thermophilic Geobacillus sp. 70PC53, which is much more thermostable than its Bacillus homolog, BsCel5A. Thus, these two cellulases provide a pair of structures ideal for investigating the mechanism regarding how these cellulases can retain activity at high temperature. In the present study, we applied the SCHEMA non-contiguous recombination algorithm as a novel tool, which assigns protein sequences into blocks for domain swapping in a way that lessens structural disruption, to generate a set of chimeric proteins derived from the recombination of GsCelA and BsCel5A. Analyzing the activity and thermostability of this designed library set, which requires only a limited number of chimeras by SCHEMA calculations, revealed that one of the blocks may contribute to the higher thermostability of GsCelA. When tested against swollen Avicel, the highly thermostable chimeric cellulase C10 containing this block showed significantly higher activity (22%-43% and higher thermostability compared to the parental enzymes. With further structural determinations and mutagenesis analyses, a 310 helix was identified as being responsible for the improved thermostability of this block. Furthermore, in the presence of ionic calcium and crown ether (CR, the chimeric C10 was found to retain 40% residual activity even after heat treatment at 90°C. Combining crystal structure determinations and structure-guided SCHEMA recombination, we have determined the mechanism responsible for the high thermostability of GsCelA, and generated a novel recombinant enzyme with significantly higher activity.

  4. Investigation of newly developed solid state fermenter on carboxymethyl cellulase production

    Directory of Open Access Journals (Sweden)

    Lee, C. K.

    2013-01-01

    Full Text Available Aims: Enzyme (cellulase contributes 10% to overall cost in bioethanol production from lignocellulosic biomass. This means that the cost for bioethanol production will be reduced if cellulase can be produced using cheaper method. Compared with submerged fermentation, it is recognized that the cost for cellulase production using solid state fermentation (SSF process is much cheaper. The present study aimed to optimize cellulase production via SSF process using agro-industrial residual as substrate.Methodology and result: Newly developed solid state bioreactor, FERMSOSTAT had been evaluated in cellulase production using local isolate Aspergillus niger USM AI 1 grown on sugarcane baggase and palm kernel cake as substrates at 1:1 (w/w ratio. Under optimized SSF conditions of 0.5 kg substrate; 70% (w/w moisture content; 30 °C; aeration at 4 L/h.g fermented substrate for 5 min and mixing at 0.5 rpm for 5 min, about 62.6 U/g of CMCase activity obtained. At the same time, comparative studies of the enzyme production under the same SSF conditions indicated that CMCase produced by Trichoderma reesei was about 9% lower compared with A. niger USM AI 1.Conclusion, significance and impact of study: It can be concluded that the performance of newly developed SSF fermenter is good since it can used to produce CMCase enzyme with reasonable good title (863% increased in CMCase production after optimization. Thus, this newly developed SSF bioreactor has highly potential be used as prototype for larger scale bioreactor design.

  5. Cellulase Production by Native Bacteria Using Water Hyacinth as Substrate under Solid State Fermentation

    Directory of Open Access Journals (Sweden)

    Suresh Chandra Kurup, R.

    2005-01-01

    Full Text Available Most of the freshwater systems in tropical countries are infested with one kind of aquatic weed or the other causing serious environmental problems. All efforts to control the growth and spread of these weeds have failed miserably and hence the concept of eradication through utilization is being adopted by many researchers. Solid state fermentation, the culturing of microorganisms on moist solid substrates in the absence or near absence of free water, has generatedgreat deal of interest among researchers because of its various advantages over the submerged fermentation technique. Cellulase enzyme is used extensively in various industries, especially in textile, food and in the bioconversion of lignocellulosic wastes to alcohol. The extensive use of cellulase in industries depends on the cost of the enzyme and hence considerable research is being carried out to isolate better microbial strains and also to develop new fermentationprocesses with the aim to reduce the product cost. The objective of the present study is to determine whether water hyacinth, one of the commonly found aquatic weeds, can be used as a substrate for cellulase production, by three native bacterial isolates named WHB 3, WHB 4 and SMB 3, under the process of solid state fermentation. Results indicatethat all the three isolates produced cellulase enzyme by using water hyacinth as the solid support. Under optimized conditions of moisture, pH, temperature, incubation time and inoculum concentration, the enzyme yield increased from 16.8 to 94.8 units for SMB 3, from 25.2 to 110.4 units for WHB 3 and from 18.0 to 127.2 units for WHB 4. The addition of nitrogen and carbon sources resulted in a significant increase in cellulase yield and WHB 3 produced the maximum amount of 216 units followed by SMB 3 and WHB 4.

  6. Exploring the Mechanism Responsible for Cellulase Thermostability by Structure-Guided Recombination.

    Science.gov (United States)

    Chang, Chia-Jung; Lee, Cheng-Chung; Chan, Yueh-Te; Trudeau, Devin L; Wu, Mei-Huey; Tsai, Chih-Hsuan; Yu, Su-May; Ho, Tuan-Hua David; Wang, Andrew H-J; Hsiao, Chwan-Deng; Arnold, Frances H; Chao, Yu-Chan

    2016-01-01

    Cellulases from Bacillus and Geobacillus bacteria are potentially useful in the biofuel and animal feed industries. One of the unique characteristics of these enzymes is that they are usually quite thermostable. We previously identified a cellulase, GsCelA, from thermophilic Geobacillus sp. 70PC53, which is much more thermostable than its Bacillus homolog, BsCel5A. Thus, these two cellulases provide a pair of structures ideal for investigating the mechanism regarding how these cellulases can retain activity at high temperature. In the present study, we applied the SCHEMA non-contiguous recombination algorithm as a novel tool, which assigns protein sequences into blocks for domain swapping in a way that lessens structural disruption, to generate a set of chimeric proteins derived from the recombination of GsCelA and BsCel5A. Analyzing the activity and thermostability of this designed library set, which requires only a limited number of chimeras by SCHEMA calculations, revealed that one of the blocks may contribute to the higher thermostability of GsCelA. When tested against swollen Avicel, the highly thermostable chimeric cellulase C10 containing this block showed significantly higher activity (22%-43%) and higher thermostability compared to the parental enzymes. With further structural determinations and mutagenesis analyses, a 310 helix was identified as being responsible for the improved thermostability of this block. Furthermore, in the presence of ionic calcium and crown ether (CR), the chimeric C10 was found to retain 40% residual activity even after heat treatment at 90°C. Combining crystal structure determinations and structure-guided SCHEMA recombination, we have determined the mechanism responsible for the high thermostability of GsCelA, and generated a novel recombinant enzyme with significantly higher activity. PMID:26986867

  7. Evaluating leachate recirculation with cellulase addition to enhance waste biostabilisation and landfill gas production.

    Science.gov (United States)

    Frank, R R; Davies, S; Wagland, S T; Villa, R; Trois, C; Coulon, F

    2016-09-01

    The effect of leachate recirculation with cellulase augmentation on municipal solid waste (MSW) biostabilisation and landfill gas production was investigated using batch bioreactors to determine the optimal conditions of moisture content, temperature and nutrients. Experimentation was thereafter scaled-up in 7L bioreactors. Three conditions were tested including (1) leachate recirculation only, (2) leachate recirculation with enzyme augmentation and (3) no leachate recirculation (control). Cumulative biogas production of the batch tests indicated that there was little difference between the leachate and control test conditions, producing on average 0.043m(3)biogaskg(-1) waste. However the addition of cellulase at 15×10(6)Utonne(-1) waste doubled the biogas production (0.074m(3)biogaskg(-1) waste). Similar trend was observed with the bioreactors. Cellulase addition also resulted in the highest COD reduction in both the waste and the leachate samples (47% and 42% COD reduction, respectively). In both cases, the quantity of biogas produced was closer to the lower value of theoretical and data-based biogas prediction indicators (0.05-0.4m(3)biogaskg(-1) waste). This was likely due to a high concentration of heavy metals present in the leachate, in particular Cr and Mn, which are known to be toxic to methanogens. The cost-benefit analysis (CBA) based on the settings of the study (cellulase concentration of 15×10(6)Utonne(-1) waste) showed that leachate bioaugmentation using cellulase is economically viable, with a net benefit of approximately €12.1million on a 5Mt mixed waste landfill. PMID:27397800

  8. Identification of a haloalkaliphilic and thermostable cellulase with improved ionic liquid tolerance

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Tao; Datta, Supratim; Eichler, Jerry; Ivanova, Natalia; Axen, Seth D.; Kerfeld, Cheryl A.; Chen, Feng; Kyrpides, Nikos; Hugenholtz, Philip; Cheng, Jan-Fang; Sale, Kenneth L.; Simmons, Blake; Rubin, Eddy

    2011-02-17

    Some ionic liquids (ILs) have been shown to be very effective solvents for biomass pretreatment. It is known that some ILs can have a strong inhibitory effect on fungal cellulases, making the digestion of cellulose inefficient in the presence of ILs. The identification of IL-tolerant enzymes that could be produced as a cellulase cocktail would reduce the costs and water use requirements of the IL pretreatment process. Due to their adaptation to high salinity environments, halophilic enzymes are hypothesized to be good candidates for screening and identifying IL-resistant cellulases. Using a genome-based approach, we have identified and characterized a halophilic cellulase (Hu-CBH1) from the halophilic archaeon, Halorhabdus utahensis. Hu-CBH1 is present in a gene cluster containing multiple putative cellulolytic enzymes. Sequence and theoretical structure analysis indicate that Hu-CBH1 is highly enriched with negatively charged acidic amino acids on the surface, which may form a solvation shell that may stabilize the enzyme, through interaction with salt ions and/or water molecules. Hu-CBH1 is a heat tolerant haloalkaliphilic cellulase and is active in salt concentrations up to 5 M NaCl. In high salt buffer, Hu-CBH1 can tolerate alkali (pH 11.5) conditions and, more importantly, is tolerant to high levels (20percent w/w) of ILs, including 1-allyl-3-methylimidazolium chloride ([Amim]Cl). Interestingly, the tolerances to heat, alkali and ILs are found to be salt-dependent, suggesting that the enzyme is stabilized by the presence of salt. Our results indicate that halophilic enzymes are good candidates for the screening of IL-tolerant cellulolytic enzymes.

  9. Cellulase production by Trichoderma harzianum in static and mixed solid-state fermentation reactors under nonaseptic conditions

    Energy Technology Data Exchange (ETDEWEB)

    Deschamps, F.; Giuliano, C.; Asther, M.; Huet, M.C.; Roussos, S.

    1985-09-01

    Cellulase production from lignocellulosic materials was studied in solid-state cultivation by both static and mixed techniques under nonaseptic conditions. The effects of fermentation conditions, such as moisture content, pH, temperature, and aeration, on cellulase production by Trichoderma harzianum using a mixture of wheat straw (80%) and bran (20%) were investigated. With a moisture content of 74% and a pH of 5.8, 18 IU filter paper activity and 198 IU endoglucanase activity/g initial substrate content were obtained in 66 hours. The extension from static column cultivation to stirred tank reactor of 65 l capacity gave similar yields of cellulase.

  10. Overproduction of cellulase by Trichoderma reesei RUT C30 through batch-feeding of synthesized low-cost sugar mixture.

    Science.gov (United States)

    Li, Yonghao; Liu, Chenguang; Bai, Fengwu; Zhao, Xinqing

    2016-09-01

    Cellulase is a prerequisite for the bioconversion of lignocellulosic biomass, but its high cost presents the biggest challenge. In this article, low-cost mixture was produced from glucose through the transglycosylation reaction catalyzed by β-glucosidase for cellulase overproduction by Trichodema reesei RUT C30. As a result, cellulase titer of 90.3FPU/mL, which was more than 10 folds of that achieved with lactose as inducer, was achieved at 144h. Meanwhile, cellulase productivity was drastically increased to 627.1FPU/L/h, at least 3-5 folds higher than previously reported by the fungal species. The crude enzyme was further tested by hydrolyzing NaOH-pretreated corn stover with 15% solid loading, and 96.6g/L glucose was released with 92.6% sugar yield at 96h and 44.8g/L ethanol was obtained. PMID:27268435

  11. Enhancement of CO/sub 2/ and ethylene production and cellulase activity by glyphosate in Phaseolus vulgaris

    Energy Technology Data Exchange (ETDEWEB)

    Abu-Irmaileh, B.E.; Jordan, L.S.; Kumamoto, J.

    1979-01-01

    The effect of glyphosate (N-(phosphonomethyl)glycine) on carbon dioxide. (CO/sub 2/) levels, ethylene production, and cellulase activity was investigated. Production of ethylene increased within 12 h and CO/sub 2/ increased within 24 h when 12-day-old bean plants (Phaseolus vulgaris L. Red Kidney) were treated with 20 mM isopropylamine salt of glyphosate. The CO/sub 2/ cycled for 3 days and then increased around treated plants. Specific activity of cellulase was increased in debladed bean seedlings that had been retreated with 20 mM isopropylamine salt of glyphosate. Cellulase enhancement was detected 2 days after the pretreated plants were debladed. Glyphosate-enhanced ethylene production may have increased the cellulase activity. 24 references, 3 figures.

  12. Optimization of process parameters for cellulase production from Bacillus sp. JS14 in solid substrate fermentation using response surface methodology

    Directory of Open Access Journals (Sweden)

    Jagdish Singh

    2012-08-01

    Full Text Available The aim of this work was to isolate the potent bacterial strains for the production of cellulose enzyme. A total 30 bacterial isolates showed positive results for the cellulase production but highest enzyme activity was shown by isolate JS 14. From the morphological and biochemical reactions, the isolate was identified as Bacillus sp. Cellulase production was studied by this strain using response surface methodology (RSM. A central composite design (CCD quadratic response surface was applied to explicate the parameters that significantly affected cellulase production in solid substrate fermentation (SSF. The wheat bran concentration and incubation period were significant factors. The process parameters optimized with response surface methodology was wheat bran concentration 400 g/L; pH, 6.5; temperature, 400C and incubation period 5 days when inoculum 10 % (1x107 cells/ ml was used for cellulase production in SSF. Supplementation of lactose and CMC to the wheat bran medium favored the enzyme formation.

  13. Characterization of the newly isolated Geobacillus sp. T1, the efficient cellulase-producer on untreated barley and wheat straws.

    Science.gov (United States)

    Assareh, Reza; Shahbani Zahiri, Hossein; Akbari Noghabi, Kambiz; Aminzadeh, Saeed; Bakhshi Khaniki, Gholamreza

    2012-09-01

    A thermophile cellulase-producing bacterium was isolated and identified as closely related to Geobacillus subterraneus. The strain, named Geobacillus sp. T1, was able to grow and produce cellulase on cellobiose, microcrystalline cellulose, carboxymethylcellulose (CMC), barley straw, wheat straw and Whatman No. 1 filter paper. However, barley and wheat straws were significantly better substrates for cellulase production. When Geobacillus sp. T1 was cultivated in the presence of 0.5% barley straw, 0.1% Tween 80 and pH 6.5 at 50°C, the maximum level of free cellulase up to 143.50 U/mL was produced after 24h. This cellulase (≈ 54 kDa) was most active at pH 6.5 and 70°C. The enzyme in citrate phosphate buffer (10mM) was stable at 60°C for at least 1h. Geobacillus sp. T1 with efficient growth and cellulase production on straws seems a potential candidate for conversion of agricultural biomass to fuels.

  14. Trpac1, a pH response transcription regulator, is involved in cellulase gene expression in Trichoderma reesei.

    Science.gov (United States)

    He, Ronglin; Ma, Lijuan; Li, Chen; Jia, Wendi; Li, Demao; Zhang, Dongyuan; Chen, Shulin

    2014-12-01

    Fungi grow over a relatively wide pH range and adapt to extracellular pH through a genetic regulatory system mediated by a key component PacC, which is a pH transcription regulator. The cellulase production of the filamentous fungi Trichoderma reesei is sensitive to ambient pH. To investigate the connection between cellulase expression regulation and ambient pH, an ortholog of Aspergillus nidulans pacC, Trpac1, was identified and functionally characterized using a target gene deletion strategy. Deleting Trpac1 dramatically increased the cellulase production and the transcription levels of the major cellulase genes at neutral pH, which suggested Trpac1 is involved in the regulation of cellulase production. It was further observed that the expression levels of transcription factors xyr1 and ace2 also increased in the ΔTrpac1 mutant at neutral pH. In addition, the ΔTrpac1 mutant exhibited conidiation defects under neutral and alkaline pH. These results implied that Trpac1 in involved in growth and development process and cellulase gene expression in T. reesei.

  15. Addressing the Recalcitrance of Cellulose Degradation through Cellulase Discovery, Nano-scale Elucidation of Molecular Mechanisms, and Kinetic Modeling

    Energy Technology Data Exchange (ETDEWEB)

    Walker, Larry P., Bergstrom, Gary; Corgie, Stephane; Craighead, Harold; Gibson, Donna; Wilson, David

    2011-06-13

    This research project was designed to play a vital role in the development of low cost sugars from cellulosic biomass and contributing to the national effort to displace fossil fuel usage in the USA transportation sector. The goal was to expand the portfolio of cell wall degrading enzymes through innovative research at the nano-scale level, prospecting for novel cellulases and building a kinetic framework for the development of more effective enzymatic conversion processes. More precisely, the goal was to elucidate the molecular mechanisms for some cellulases that are very familiar to members of our research team and to investigate what we hope are novel cellulases or new enzyme combinations from the world of plant pathogenic fungi and bacteria. Hydrolytic activities of various cellulases and cellulase cocktails were monitored at the nanoscale of cellulose fibrils and the microscale of pretreated cellulose particles, and we integrated this insight into a heterogeneous reaction framework. The over-riding approach for this research program was the application of innovative and cutting edge optical and high-throughput screening and analysis techniques for observing how cellulases hydrolyze real substrates.

  16. PEA PEEL WASTE: A LIGNOCELLULOSIC WASTE AND ITS UTILITY IN CELLULASE PRODUCTION BY Trichoderma reesei UNDER SOLID STATE CULTIVATION

    Directory of Open Access Journals (Sweden)

    Nitin Verma

    2011-03-01

    Full Text Available A wide variety of waste bioresources are available on our planet for conversion into bioproducts. In the biological systems, microorganisms are used to utilize waste as an energy source for the synthesis of valuable products such as biomass proteins and enzymes. The large quantities of byproducts generated during the processing of plant food involve an economic and environmental problem due to their high volumes and elimination costs. After isolation of the main constituent, there are abundant remains which represent an inexpensive material that has been undervalued until now. Pea peel waste is one of the undervalued, unused sources of energy that can serve as a potential source for cellulase production. Batch experiments have been performed, using pea peel waste as a carbon source for cellulase production under solid state cultivation by Trichoderma reesei. It was observed that 30 oC temperature and pH 5.0 are the most favorable conditions for cellulase production by T. reesei. FPase activity significantly increases by incorporation of whey as well as wheat starch hydrolysate in the basal salt media used in the production study. The present study describes the utility of pea peel waste, whey as well as wheat starch hydrolysate in cellulase production by T. reesei. The utilization of economically cheap, pea peel waste for cellulase production could be a novel, cost effective, and valuable approach in cellulase production as well as in solid waste management.

  17. Cellulase-assisted extraction of polysaccharides from Cucurbita moschata and their antibacterial activity.

    Science.gov (United States)

    Qian, Zhi-Gang

    2014-01-30

    In this study, cellulase-assisted extraction of water soluble polysaccharides from pumpkin (Cucurbita moschata) and their antibacterial activity were investigated. The polysaccharides yield was monitored during the extraction process. The optimum extraction conditions were determined as follows: time, 40 min; temperature, 55°C; pH, 4.5; and cellulase amount, 4,000 U/g. The extracts were centrifuged, filtered, proteins removed by Sevag method, concentrated to approximately 15% (w/v), precipitated with 5 volumes of absolute ethanol, freeze-dried, and pulverized to yield a water soluble powder of pumpkin polysaccharides (PP). The sugar content of the product was 68.3%, and the yield was 17.34% (w/w), respectively. The PP had high antibacterial activity against Bacillus subtilis, Staphylococcus aureus, and Escherichia coli at the concentration of 100 mg/mL.

  18. Study on Preparation of the Low-Molecular-Weight Chitosan Using Cellulase

    Institute of Scientific and Technical Information of China (English)

    LI He-sheng; SUN Yu-xi; HUANG Xiao-chun; WANG Hong-fei; QIU Di-hong

    2006-01-01

    The degradation of chitosan ( DD of 72.05%) with aid of cellulase was carried out under the conditions of 45℃, pH 5.0 and a ratio of 1:15( chitosan/enzyme). The results showed cellulase could degrade chitosan efficiently. Viscosity of chitosan was decreased very quickly and reducing sugar released was increased with time during degradation. By using the membrane, the separation of the hydrolysis mixture was studied. Rejection of protein can be reached to be 99.74%.65.9% of low-molecular-weight chitosans was less than 2 kDa. Solubility of low-molecular-weight chitosan was found to be better than chitosan and transmittance could reach to be more than 95 % in entire range of pH 1 ~ 13.

  19. Production of Cellulase from Oil Palm Biomass as Substrate by Solid State Bioconversion

    Directory of Open Access Journals (Sweden)

    Md. Z. Alam

    2005-01-01

    Full Text Available Solid state bioconversion (SSB of lignocellulosic material oil palm biomass (OPB generated from palm oil industries as waste was conducted by evaluating the enzyme production through filamentous fungus in lab-scale experiment. OPB in the form of empty fruit bunches (EFB was used as the solid substrate and treated with the fungus Trichoderma harzianum to produce cellulase. The results presented in this study revealed that the higher cellulase activity of 0.0413 unit was achieved at the day 3 of fermentation. While the optimum study indicated the enzyme production of 0.0433 unit with moisture content of 50%, 0.0413 unit with 5% v/w of inoculum size and 0.0413 unit with co-substrate concentration of 2% (w/w at days 9, 9 and 12 of fungal treatment, respectively. The parameters glucosamine and reducing sugar were observed to evaluate the growth and substrate utilization in the experiment.

  20. PURIFICATION AND SOME PROPERTIES OF CELLULASE FROM ODONTOTERMES FORMOSANUS (ISOPTERA: TERMITIDAE)

    Institute of Scientific and Technical Information of China (English)

    Tian-ciYang; Jian-chuMo; Jia-anCheng

    2004-01-01

    The purification of the cellulase from Odontotermes forrnosanus workers was achieved by using anion-exchange column of UNOsphere Q, BioLogic DuoFlow chromatography system. The purified cellulase was identified as an endoglucanase and some of its properties were investigated. The EGase activity was 807.5-fold as high as the initial enzyme activity using CMC as substrate and 14.4-fold using salicin as substrate. The enzyme preparations were homogeneous as judged by SDS-PAGE electrophoresis, molecular weight of which was 80 kDa and confirmed by 2-DE zymogram analysis. The enzyme was isoelectric at pH 6.4, which was active on CMC substrate.

  1. Production of nanotubes in delignified porous cellulosic materials after hydrolysis with cellulase.

    Science.gov (United States)

    Koutinas, Αthanasios Α; Papafotopoulou-Patrinou, Evgenia; Gialleli, Angelika-Ioanna; Petsi, Theano; Bekatorou, Argyro; Kanellaki, Maria

    2016-08-01

    In this study, tubular cellulose (TC), a porous cellulosic material produced by delignification of sawdust, was treated with a Trichoderma reesei cellulase in order to increase the proportion of nano-tubes. The effect of enzyme concentration and treatment duration on surface characteristics was studied and the samples were analyzed with BET, SEM and XRD. Also, a composite material of gelatinized starch and TC underwent enzymatic treatment in combination with amylase (320U) and cellulase (320U) enzymes. For TC, the optimum enzyme concentration (640U) led to significant increase of TC specific surface area and pore volume along with the reduction of pore diameter. It was also shown that the enzymatic treatment did not result to a significant change of cellulose crystallinity index. The produced nano-tubular cellulose shows potential for application to drug and chemical preservative delivery systems. PMID:26996258

  2. Valorizing recycled paper sludge by a bioethanol production process with cellulase recycling.

    Science.gov (United States)

    Gomes, Daniel; Domingues, Lucília; Gama, Miguel

    2016-09-01

    The feasibility of cellulase recycling in the scope of bioethanol production from recycled paper sludge (RPS), an inexpensive byproduct with around 39% of carbohydrates, is analyzed. RPS was easily converted and fermented by enzymes and cells, respectively. Final enzyme partition between solid and liquid phases was investigated, the solid-bound enzymes being efficiently recovered by alkaline washing. RPS hydrolysis and fermentation was conducted over four rounds, recycling the cellulases present in both fractions. A great overall enzyme stability was observed: 71, 64 and 100% of the initial Cel7A, Cel7B and β-glucosidase activities, respectively, were recovered. Even with only 30% of fresh enzymes added on the subsequent rounds, solid conversions of 92, 83 and 71% were achieved for the round 2, 3 and 4, respectively. This strategy enabled an enzyme saving around 53-60%, while can equally contribute to a 40% reduction in RPS disposal costs. PMID:27289054

  3. Effects of non-ionic surfactants on the interactions between cellulases and tannic acid

    DEFF Research Database (Denmark)

    Olsen, Søren Nymand; Bohlin, Christina Helena; Murphy, Leigh;

    2011-01-01

    Addition of non-ionic surfactants (NIS) is known to accelerate enzymatic lignocellulose hydrolysis. The mechanism behind this accelerating effect is still not elucidated but has been hypothesized to originate from favorable NIS–lignin interactions which alleviate non-productive adsorption...... of cellulases to lignin. In the current work we address this hypothesis using tannic acid (TAN) as a general poly-phenolic model compound (for lignin and soluble phenolics) and measure the mutual interactions of cellulases (CBHI, CBHII, EGI, EGII and BG), TAN and NIS (Triton X-100) using isothermal titration...... calorimetry (ITC). The experimental results suggest rather strong enzyme-specific interactions with TAN in reasonable agreement with enzyme specific lignin inhibition found in the literature. Enzyme–TAN interactions were disrupted by the presence of NIS by a mechanism of strong TAN–NIS interaction...

  4. A pyranose dehydrogenase-based biosensor for kinetic analysis of enzymatic hydrolysis of cellulose by cellulases

    OpenAIRE

    Cruys-Bagger, Nicolaj; Badino, Silke Flindt; Tokin, Radina Naytchova; Gontsarik, Mark; Fathalinejad, Samin; Jensen, Kenneth; Toscano, Miguel Duarte; Sørensen, Trine Holst; Borch, Kim; Tatsumi, Hirosuke; Väljamäe, Priit; Westh, Peter

    2014-01-01

    A novel electrochemical enzyme biosensor was developed for real-time detection of cellulase activity when acting on their natural insoluble substrate, cellulose. The enzyme biosensor was constructed with pyranose dehydrongease (PDH) from Agaricus meleagris that was immobilized on the surface of a carbon paste electrode, which contained the mediator 2,6-dichlorophenolindophenol (DCIP). An oxidation current of the reduced form of DCIP, DCIPH2, produced by the PDH-catalyzed reaction with either ...

  5. Statistical Optimization of Fermentation Conditions for Cellulase Production from Palm Oil Mill Effluent

    Directory of Open Access Journals (Sweden)

    Jamal I. Daoud

    2010-01-01

    Full Text Available Problem statement: Palm oil mill effluent discharged by the oil palm industries is considered as the mixed of high polluted effluent which is abundant (about 20 million tonnes year-1 and its effect contributes to the serious environmental problems through the pollution of water bodies. Approach: The aim of this study was to identify the potential of low cost substrate such as Palm Oil Mill Effluent (POME for the production of cellulase enzyme by liquid state bioconversion. The filamentous fungus Trichoderma harzianum was used for liquid state bioconversion of POME for cellulase production. Statistical optimization was carried out to evaluate the physico-chemical parameters (factors for maximum cellulase production by 2-level fractional factorial design with six central points. The polynomial regression model was developed using the experimental data including the effects of linear, quadratic and interaction of the factors. The factors involved were substrate (POME and co-substrate (wheat flour concentrations, temperature, pH, inoculum and agitation. Results: Statistical analysis showed that the optimum conditions were: Temperature of 30°C, substrate concentration of 2%, wheat flour concentration of 3%, pH of 4, inoculum of 3% and agitation of 200 rpm. Under these conditions, the model predicted the enzyme production to be about 14 FPU mL-1. Analysis Of Variance (ANOVA of the design showed a high coefficient of determination (R2 value of 0.999, thus ensuring a high satisfactory adjustment of the quadratic model with the experimental data. Conclusion/Recommendations: This study indicates a better solution for waste management through the utilization of POME for cellulase production that could be used in the industrial applications such as bioethanol production.

  6. Computer Simulations Reveal Multiple Functions for Aromatic Residues in Cellulase Enzymes (Fact Sheet)

    Energy Technology Data Exchange (ETDEWEB)

    2012-07-01

    NREL researchers use high-performance computing to demonstrate fundamental roles of aromatic residues in cellulase enzyme tunnels. National Renewable Energy Laboratory (NREL) computer simulations of a key industrial enzyme, the Trichoderma reesei Family 6 cellulase (Cel6A), predict that aromatic residues near the enzyme's active site and at the entrance and exit tunnel perform different functions in substrate binding and catalysis, depending on their location in the enzyme. These results suggest that nature employs aromatic-carbohydrate interactions with a wide variety of binding affinities for diverse functions. Outcomes also suggest that protein engineering strategies in which mutations are made around the binding sites may require tailoring specific to the enzyme family. Cellulase enzymes ubiquitously exhibit tunnels or clefts lined with aromatic residues for processing carbohydrate polymers to monomers, but the molecular-level role of these aromatic residues remains unknown. In silico mutation of the aromatic residues near the catalytic site of Cel6A has little impact on the binding affinity, but simulation suggests that these residues play a major role in the glucopyranose ring distortion necessary for cleaving glycosidic bonds to produce fermentable sugars. Removal of aromatic residues at the entrance and exit of the cellulase tunnel, however, dramatically impacts the binding affinity. This suggests that these residues play a role in acquiring cellulose chains from the cellulose crystal and stabilizing the reaction product, respectively. These results illustrate that the role of aromatic-carbohydrate interactions varies dramatically depending on the position in the enzyme tunnel. As aromatic-carbohydrate interactions are present in all carbohydrate-active enzymes, the results have implications for understanding protein structure-function relationships in carbohydrate metabolism and recognition, carbon turnover in nature, and protein engineering

  7. Cellulase production by Trichoderma longi, Aspergillus niger and Saccharomyces cerevisae cultured on waste materials from orange.

    Science.gov (United States)

    Omojasola, P F; Jilani, O P

    2008-10-15

    The wastes materials from the sweet orange (Citrus sinensis) were used as substrate for the production of cellulase. The rind, the pericarp or albedo and the pulp were hydrolyzed by cellulolytic enzymes of Trichoderma longibrachiatum, Aspergillus niger and Saccharomyces cerevisiae after they were treated with alkali and steam. The amount of glucose released from the substrates following the secretion of cellulase by the three microorganisms was measured. The orange wastes released amounts of glucose ranging from 0.76-0.96 mg mL(-1) by Trichoderma longibrachiatum, 0.90-1.08 mg mL(-1) by A. niger and 0.60-0.76 mg mL(-1) by S. cerevisiae after five days of fermentation. The conditions of the fermentation were then varied to determine their effect on cellulase production. Fermentation parameters varied were time, pH, substrate concentration, temperature and inoculum size. After this, conditions that produced highest amounts of glucose were combined in an optimization experiment. Glucose production under optimized conditions were 0.94 mg mL(-1) by T. longibrachiatum, 0.83 mg mL(-1) by A. niger and 0.67 mg mL(-1) by S. cerevisae. The activity of the test organisms' cellulase against CMC on the orange wastes was also determined with T. longibrachiatum producing 3.86 mg mL(-1), A. niger 2.94 mg mL(-1) and S. cerevisiae 2.30 mg mL(-1) glucose amounts all from orange pulp. PMID:19137846

  8. Potential of Biosynthesized Silver Nanoparticles as Nanocatalyst for Enhanced Degradation of Cellulose by Cellulase

    OpenAIRE

    Salunke, Bipinchandra K.; Sawant, Shailesh S.; Tae Koo Kang; Deok Yun Seo; Youngjong Cha; Sun A. Moon; Bassam Alkotaini; Ezhaveni Sathiyamoorthi; Beom Soo Kim

    2015-01-01

    Silver nanoparticles (AgNPs) as a result of their excellent optical and electronic properties are promising catalytic materials for various applications. In this study, we demonstrate a novel approach for enhanced degradation of cellulose using biosynthesized AgNPs in an enzyme catalyzed reaction of cellulose hydrolysis by cellulase. AgNPs were synthesized through reduction of silver nitrate by extracts of five medicinal plants (Mentha arvensis var. piperascens, Buddleja officinalis Maximowic...

  9. Performance and Nutrient Utilization of Layers Fed Diet Supplemented with Microbial Phytase and Cellulase

    Institute of Scientific and Technical Information of China (English)

    2003-01-01

    A 31-week feeding trial was conducted to investigated the effects of dietary supplementation of microbial phytase and cellulase on performance,nutrients utilization and tibia quality of laying hens fed maize and soybean meal diets.192 18-week-old Hisex layers were used in the trial A 2×2×2 factorial design was used in the experiment with three factors of two levels each:0.38% and 0.16% of dietary non-phytate P(nP).0 and 300 U*kg-1 of phytase (Ph),and 0 and 0.10% of cellulase (Ce).The results showed that supplementation of 300 U*kg-1 phytase significantly improved utilization of dietary crude ash,CP,Ca,total P and copper (P<0.05),and improved tibia breaking strength (P<0.05).No effect of phytase on performance was observed.Addition of 0.10% cellulase decreased feed intake (P<0.05),increased utilization of CF (P<0.05) and Ca(P<0.01),and decreased total tibia ash weight (P<0.05).300 U*kg-1 phytase and 0.10% cellulase exhibited obvious positive interactions to enhance utilization of dietary phytic P and copper (P<0.05).0.16% nP did not reduce performance of the layers,but improved egg shell quality at 20 wks,increased utilization of dietary total P,phytic P and Copper (P<0.01),decreased utilization of dietary CP,increased tibia breaking strength and Ca,Mn contents of tibia(P<0.01)

  10. Evaluating leachate recirculation with cellulase addition to enhance waste biostabilisation and landfill gas production

    OpenAIRE

    Frank, R. R.; S. Davies; Wagland, Stuart T.; Villa, Raffaella; Trois, C.; Coulon, Frederic

    2016-01-01

    The effect of leachate recirculation with cellulase augmentation on municipal solid waste (MSW) biostabilisation and landfill gas production was investigated using batch bioreactors to determine the optimal conditions of moisture content, temperature and nutrients. Experimentation was thereafter scaled-up in 7 L bioreactors. Three conditions were tested including (1) leachate recirculation only, (2) leachate recirculation with enzyme augmentation and (3) no leachate recirculation (control). C...

  11. Ultrasounds pretreatment of olive pomace to improve xylanase and cellulase production by solid-state fermentation

    OpenAIRE

    Leite, A; Salgado, José Manuel; Venâncio, Armando; Domínguez, José Manuel; Belo, Isabel

    2016-01-01

    Abstract Olive mills generate a large amount of waste that can be revaluated. This work aim to improve the production lignocellulolytic enzymes by solid-state fermentation using ultrasounds pretreated olive mill wastes. The composition of olive mill wastes (crude and exhausted olive pomace) was compared and several physicochemical characteristics were significantly different. The use of both wastes in SSF was evaluated and a screening of fungi for xylanase and cellulase production was carried...

  12. ENZYMATIC KINETICS OF CELLULASES ISOLATED FROM SOIL BACTE RIA OF DOON VALLEY , UTTARAKHAND

    Directory of Open Access Journals (Sweden)

    Vinit

    2015-05-01

    Full Text Available Cellulases refers to a suite of enzymes produced chiefly by fungi , bacteria , and protozoans that catalyze cellulolysis which is the hydrolysis of cellulose . Cellulose is the most abundant natural polymer on earth . It is the structural component of the plant cell walls which helps in the hydrolysis of 1, 4 - beta - D - glycosidic linkages in cellulose, lichenin and cereal beta - D - glucans . Cellulases are used for clarif ication of fruit juice, vegetable juice, roots, treatment of wine, extraction of oils and improving the quality of the bakery products . Eight soil samples were collected for cellulose preliminary screening from Gullarghati, Doon valley at different pH and temperatures, because maximum diversity was possible there as there was no interference by the humans . 110 colonies were isolated by the activity zone plate method containing CMC as a substrate using Congo red dye . Best twelve colonies were selected and ch ecked using DNS method at 540 A 0 . Four strains BR - 1, BR - 2, BR - 3 and BR - 4 were used on the basis of spectrophotometerically and characterized with the study of substrate . Maximum velocity (Vmax was observed for BR - 2 i . e . 170 units per mg protein with Km of 49 . 50mg/ml . Strain BR - 1 gave to pH optima at 4 . 5 and 6 . 5, strain BR - 2 gave maximum activity at 4 . 5 and 7 . 0 pH, BR - 3 strain gave maximum activity at pH 5 . 0 and 6 . 5 with the highest yield of cellulases w ere obtained at pH 4 . 5, 5 . 5 and 7 . 0 in bacterial s train BR - 4 . The results also shows the effect of temperature bacterial strain BR - 1, BR - 2 and BR - 4 with maximum cellulases activity at 45 0 C and bacterial strain BR - 3 maximum activity at 25 0 C .

  13. PRODUCTION OF FERMENTABLE SUGARS FROM OIL PALM EMPTY FRUIT BUNCH USING CRUDE CELLULASE COCKTAILS WITH TRICHODERMA ASPERELLUM UPM1 AND ASPERGILLUS FUMIGATUS UPM2 FOR BIOETHANOL PRODUCTION

    OpenAIRE

    Nurul Kartini Abu Bakar,; Zuraidah Zanirun; Suraini Abd-Aziz; Farinazleen Mohd Ghazali; Mohd Ali Hassan

    2012-01-01

    Utilization of oil palm empty fruit bunch (OPEFB) for bioethanol production with crude cellulase cocktails from locally isolated fungi was studied. Enzymatic saccharification of alkaline pretreated OPEFB was done using different cellulase enzyme preparations. Crude cellulase cocktails from Trichoderma asperellum UPM1 and Aspergillus fumigatus UPM2 produced 8.37 g/L reducing sugars with 0.17 g/g yield. Production of bioethanol from OPEFB hydrolysate using Baker’s yeast produced approximately 0...

  14. PERFORMANCE OF LAYER HEN FED FERMENTED Jatropha Curcas L. MEAL SUPPLEMENTED WITH CELLULASE AND PHYTASE ENZYME

    Directory of Open Access Journals (Sweden)

    S. Sumiati

    2014-10-01

    Full Text Available The objective of the experiment was to study the effect of feeding fermented Jatropha curcas L.meal (JCM supplemented with cellulase and phytase on the performances of ISA-Brown laying henaged 25-30 weeks. The Jatropha curcas meal was fermented using Rizhopus oligosporus. In this study200 laying hens were used and distributed to 5 treatments and 4 replications in Completely RandomizedDesign. The diet treatments were: R0 = control diet (without JCM, R1; diet contained fermented JCM7.5%, R2; diet contained fermented JCM 7.5% + celullase 200 g/ton, R3; diet contained fermented JCM7.5% + phytase 200 g/ton and R4; diet contained fermented JCM 7.5% + cellulase 200 g/ton + phytase200 g/ton. The parameters observed were feed consumption, hen day egg production, egg massproduction, egg weight and feed conversion ratio. The results showed that feeding fermented JCM 7.5%,both enzyme supplemented as well as unsupplemented significantly decreased (P<0.05 the feedconsumption, hen day egg and egg mass production. However, the treatments did not influence the eggweight. Supplementation of cellulase (R2 or phytase (R3 improved the feed conversion ratio with thevalue as same as the R0 diet.

  15. Analysis of Casein Biopolymers Adsorption to Lignocellulosic Biomass as a Potential Cellulase Stabilizer

    Directory of Open Access Journals (Sweden)

    Anahita Dehkhoda Eckard

    2012-01-01

    Full Text Available Although lignocellulosic materials have a good potential to substitute current feedstocks used for ethanol production, conversion of these materials to fermentable sugars is still not economical through enzymatic hydrolysis. High cost of cellulase has prompted research to explore techniques that can prevent from enzyme deactivation. Colloidal proteins of casein can form monolayers on hydrophobic surfaces that alleviate the de-activation of protein of interest. Scanning electron microscope (SEM, fourier transform infrared spectroscopy (FT-IR, capillary electrophoresis (CE, and Kjeldahl and BSA protein assays were used to investigate the unknown mechanism of action of induced cellulase activity during hydrolysis of casein-treated biomass. Adsorption of casein to biomass was observed with all of the analytical techniques used and varied depending on the pretreatment techniques of biomass. FT-IR analysis of amides I and II suggested that the substructure of protein from casein or skim milk were deformed at the time of contact with biomass. With no additive, the majority of one of the cellulase mono-component, 97.1 ± 1.1, was adsorbed to CS within 24 h, this adsorption was irreversible and increased by 2% after 72 h. However, biomass treatment with skim-milk and casein reduced the adsorption to 32.9% ± 6.0 and 82.8% ± 6.0, respectively.

  16. Studies on influence of natural biowastes on cellulase production by Aspergillus niger.

    Science.gov (United States)

    Kiranmayi, M Usha; Poda, Sudhakar; Vijayalakshmi, M; Krishna, P V

    2011-11-01

    The objective of this study was to determine the influence of natural biowaste substrates such as banana peel powder and coir powder at varying environmental parameters of pH (4-9) and temperature (20-50 degrees C) on the cellulase enzyme production by Aspergillus niger. The cellulase enzyme production was analyzed by measuring the amount of glucose liberated in IU ml(-1) by using the dinitrosalicylic acid assay method. The substrates were pretreated with 1% NaOH (alkaline treatment) and autoclaved. The maximum activity of the enzyme was assayed at varying pH with temperatures being constant and varying temperatures with pH being constant. The highest activity of the enzyme at varying pH was recorded at pH 6 for banana peel powder (0.068 +/- 0.002 IU ml) and coir powder (0.049 +/- 0.002 IU ml(-1)) and the maximum activity of the enzyme at varying temperature was recorded at 35 degrees C for both banana peel powder (0.072 +/- 0.001 IU ml(-1)) and coir powder (0.046 +/- 0.003 IU ml(-1)). At varying temperatures and pH the high level of enzyme production was obtained at 35 degrees C and pH 6 by using both the substrates, respectively. However among the two substrates used for the production of cellulases by Aspergillus niger banana peel powder showed maximum enzymatic activity than coir powder as substrate.

  17. Production of cellulase enzymes during the solid-state fermentation of empty palm fruit bunch fiber.

    Science.gov (United States)

    Kim, Seonghun; Kim, Chul Ho

    2012-01-01

    Penicillium verruculosum COKE4E is a fungal strain isolated from bituminous coal. The microorganism cultivated in a minimal medium supplemented with Avicel, carboxymethylcellulose, and oat spelt xylan produced cellulase enzymes as exhibiting carboxymethylcellulase (CMCase), Avicelase, xylanase, and cellobiosidase activities. In this study, the productivity of the extracellular enzymes in the strain was evaluated by using empty palm fruit bunch fiber (EPFBF), a lignocellulosic biomass, as a substrate for solid-state bioconversion. The highest cellulase activities were observed after 6 days of fermentation at pH 6.0 and 30 °C. The enzymes were secreted as cellulosomes for the degradation of EPFBF as a sole carbon source. Focused ion beam analysis showed that P. verruculosum COKE4E produced cellulolytic enzymes that were able to effectively biodegrade EPFBF during solid-state fermentation. In this process, 6.5 U of CMCase, 6.8 U of Avicelase, and 8.8 U of xylanase per gram of dry solid EPFBF were produced. These results demonstrate that EPFBF may be a potential raw material in solid-state fermentation for the production of cellulase enzymes to be used for biofuel production. PMID:22052232

  18. Analysis of Casein Biopolymers Adsorption to Lignocellulosic Biomass as a Potential Cellulase Stabilizer

    Science.gov (United States)

    Eckard, Anahita Dehkhoda; Muthukumarappan, Kasiviswanathan; Gibbons, William

    2012-01-01

    Although lignocellulosic materials have a good potential to substitute current feedstocks used for ethanol production, conversion of these materials to fermentable sugars is still not economical through enzymatic hydrolysis. High cost of cellulase has prompted research to explore techniques that can prevent from enzyme deactivation. Colloidal proteins of casein can form monolayers on hydrophobic surfaces that alleviate the de-activation of protein of interest. Scanning electron microscope (SEM), fourier transform infrared spectroscopy (FT-IR), capillary electrophoresis (CE), and Kjeldahl and BSA protein assays were used to investigate the unknown mechanism of action of induced cellulase activity during hydrolysis of casein-treated biomass. Adsorption of casein to biomass was observed with all of the analytical techniques used and varied depending on the pretreatment techniques of biomass. FT-IR analysis of amides I and II suggested that the substructure of protein from casein or skim milk were deformed at the time of contact with biomass. With no additive, the majority of one of the cellulase mono-component, 97.1 ± 1.1, was adsorbed to CS within 24 h, this adsorption was irreversible and increased by 2% after 72 h. However, biomass treatment with skim-milk and casein reduced the adsorption to 32.9% ± 6.0 and 82.8% ± 6.0, respectively. PMID:23118515

  19. In vitro flow cytometry-based screening platform for cellulase engineering.

    Science.gov (United States)

    Körfer, Georgette; Pitzler, Christian; Vojcic, Ljubica; Martinez, Ronny; Schwaneberg, Ulrich

    2016-01-01

    Ultrahigh throughput screening (uHTS) plays an essential role in directed evolution for tailoring biocatalysts for industrial applications. Flow cytometry-based uHTS provides an efficient coverage of the generated protein sequence space by analysis of up to 10(7) events per hour. Cell-free enzyme production overcomes the challenge of diversity loss during the transformation of mutant libraries into expression hosts, enables directed evolution of toxic enzymes, and holds the promise to efficiently design enzymes of human or animal origin. The developed uHTS cell-free compartmentalization platform (InVitroFlow) is the first report in which a flow cytometry-based screened system has been combined with compartmentalized cell-free expression for directed cellulase enzyme evolution. InVitroFlow was validated by screening of a random cellulase mutant library employing a novel screening system (based on the substrate fluorescein-di-β-D-cellobioside), and yielded significantly improved cellulase variants (e.g. CelA2-H288F-M1 (N273D/H288F/N468S) with 13.3-fold increased specific activity (220.60 U/mg) compared to CelA2 wildtype: 16.57 U/mg). PMID:27184298

  20. Ultrasounds pretreatment of olive pomace to improve xylanase and cellulase production by solid-state fermentation.

    Science.gov (United States)

    Leite, Paulina; Salgado, José Manuel; Venâncio, Armando; Domínguez, José Manuel; Belo, Isabel

    2016-08-01

    Olive mills generate a large amount of waste that can be revaluated. This work aim to improve the production lignocellulolytic enzymes by solid-state fermentation using ultrasounds pretreated olive mill wastes. The composition of olive mill wastes (crude and exhausted olive pomace) was compared and several physicochemical characteristics were significantly different. The use of both wastes in SSF was evaluated and a screening of fungi for xylanase and cellulase production was carried out. After screening, the use of exhausted olive pomace and Aspergillus niger led to the highest enzyme activities, so that they were used in the study of ultrasounds pre-treatment. The results showed that the sonication led to a 3-fold increase of xylanase activity and a decrease of cellulase activity. Moreover, the liquid fraction obtained from ultrasounds treatment was used to adjust the moisture of solid and a positive effect on xylanase (3.6-fold increase) and cellulase (1.2-fold increase) production was obtained. PMID:27209456

  1. Screening and Characterization of the High-Cellulase-Producing Strain Aspergillus glaucus XC9

    Institute of Scientific and Technical Information of China (English)

    Xu Chang; Long Minnan; Wu Xiaobing; Xu Huijuan; Chen Zhongan; Zhang Fengzhang; Xu Liangshu

    2006-01-01

    Cellulose is a kind of renewable resource that is abundant in nature.It can be degraded by microorganisms such as mildew.A mildew strain with high cellulase activity was isolated from mildewy maize cob and classified as Aspergillus glaucus XC9 by morphological and 18S rRNA gene sequence analyses.We studied the effects of nitrogen source,initial pH,temperature,incubation time,medium composition,and surfactants on cellulase production.Maximal activities of carboxymethylcellulase (6,812 U/g dry koji) and filter paperase (172 U/g dry koji) were obtained in conditions as follows:initial pH,5.5-6.0;temperature,30℃;cultivation period,3-4 days;inoculum ratio,6% (vol/vol);sugarcane bagasse/wheat bran ratio,4:6.When bagasse was used as substrate and mixed with wet koji at a 1:1 (wt/wt) ratio,the yield of reducing sugars was 36.4%.The corresponding conversion rate of cellulose to reducing sugars went as high as 81.9%.The results suggest that A.glaucus XC9 is a preferred candidate for cellulase production.

  2. In vitro flow cytometry-based screening platform for cellulase engineering

    Science.gov (United States)

    Körfer, Georgette; Pitzler, Christian; Vojcic, Ljubica; Martinez, Ronny; Schwaneberg, Ulrich

    2016-01-01

    Ultrahigh throughput screening (uHTS) plays an essential role in directed evolution for tailoring biocatalysts for industrial applications. Flow cytometry-based uHTS provides an efficient coverage of the generated protein sequence space by analysis of up to 107 events per hour. Cell-free enzyme production overcomes the challenge of diversity loss during the transformation of mutant libraries into expression hosts, enables directed evolution of toxic enzymes, and holds the promise to efficiently design enzymes of human or animal origin. The developed uHTS cell-free compartmentalization platform (InVitroFlow) is the first report in which a flow cytometry-based screened system has been combined with compartmentalized cell-free expression for directed cellulase enzyme evolution. InVitroFlow was validated by screening of a random cellulase mutant library employing a novel screening system (based on the substrate fluorescein-di-β-D-cellobioside), and yielded significantly improved cellulase variants (e.g. CelA2-H288F-M1 (N273D/H288F/N468S) with 13.3-fold increased specific activity (220.60 U/mg) compared to CelA2 wildtype: 16.57 U/mg). PMID:27184298

  3. Effet de la pectolyase Y-23 et de la cellulase Onozuka RS sur le rendement en protoplastes viables de Prunus cerasus L.

    Directory of Open Access Journals (Sweden)

    Mehri-Kamoun R.

    2001-01-01

    Full Text Available Effect of pectolyase Y-23 and cellulase Onozuka RS on the yield of viable protoplasts of Prunus cerasus L. ""Montmorency"". To isolate leaf mesophyll, leaf and root callus protoplasts of Prunus cerasus L. ""Montmorency"", we have determined the optimum enzymatic mixtures to be used, and characterized the specific activity of these enzymes. The analysis of the specific activities of enzymes allows to compare the different cellulases and pectinases used to obtain protoplasts in relation with the tissue sources. This analysis concerned the FPase (degradation of filter paper and CMCase activities for cellulases Onozuka RS and R-10, and the PME (pectinmethylesterase, PL (pectate lyase and PG (polygalacturonase activities for the pectinases Macerozyme R-10 and Pectolyase Y-23. The results show that the digestion of leaf mesophyll tissues need cellulase Onozuka RS and Pectolyase Y-23 while callus protoplasts of the same material, can be isolated with cellulase Onozuka R-10 and Macerozyme R-10. The enzymes cellulase Onozuka RS and Pectolyase Y-23 (as pectinase improved significantly the yield and the viability of leaf mesophyll protoplasts compared to cellulase Onozuka R-10 and Macerozyme R-10. These results were correlated to the specific activities of the enzymes. Significant differences between the 2 pectinases are observed for PME, PL and PG activities and between the 2 cellulases for CMCase activity. From callus, the maximum amount of viable protoplasts was obtained with cellulase Onozuka R-10 (low CMCase activity and Macerozyme R-10 (low PG activity.

  4. Disruption of Trichoderma reesei cre2, encoding an ubiquitin C-terminal hydrolase, results in increased cellulase activity

    Directory of Open Access Journals (Sweden)

    Denton Jai A

    2011-11-01

    Full Text Available Abstract Background The filamentous fungus Trichoderma reesei (Hypocrea jecorina is an important source of cellulases for use in the textile and alternative fuel industries. To fully understand the regulation of cellulase production in T. reesei, the role of a gene known to be involved in carbon regulation in Aspergillus nidulans, but unstudied in T. reesei, was investigated. Results The T. reesei orthologue of the A. nidulans creB gene, designated cre2, was identified and shown to be functional through heterologous complementation of a creB mutation in A. nidulans. A T. reesei strain was constructed using gene disruption techniques that contained a disrupted cre2 gene. This strain, JKTR2-6, exhibited phenotypes similar to the A. nidulans creB mutant strain both in carbon catabolite repressing, and in carbon catabolite derepressing conditions. Importantly, the disruption also led to elevated cellulase levels. Conclusions These results demonstrate that cre2 is involved in cellulase expression. Since the disruption of cre2 increases the amount of cellulase activity, without severe morphological affects, targeting creB orthologues for disruption in other industrially useful filamentous fungi, such as Aspergillus oryzae, Trichoderma harzianum or Aspergillus niger may also lead to elevated hydrolytic enzyme activity in these species.

  5. The use of T-DNA insertional mutagenesis to improve cellulase production by the thermophilic fungus Humicola insolens Y1.

    Science.gov (United States)

    Xu, Xinxin; Li, Jinyang; Shi, Pengjun; Ji, Wangli; Liu, Bo; Zhang, Yuhong; Yao, Bin; Fan, Yunliu; Zhang, Wei

    2016-01-01

    Humicola insolens is an excellent producer of pH-neutral active, thermostable cellulases that find many industrial applications. In the present study, we developed an efficient Agrobacterium tumefaciens-mediated transformation system for H. insolens. We transformed plasmids carrying the promoter of the glyceraldehyde-3-phosphate dehydrogenase gene of H. insolens driving the transcription of genes encoding neomycin phosphotransferase, hygromycin B phosphotransferase, and enhanced green fluorescent protein. We optimized transformation efficiency to obtain over 300 transformants/10(6) conidia. T-DNA insertional mutagenesis was employed to generate an H. insolens mutant library, and we isolated a transformant termed T4 with enhanced cellulase and hemicellulase activities. The FPase, endoglucanase, cellobiohydrolase, β-glucosidase, and xylanase activities of T4, measured at the end of fermentation, were 60%, 440%, 320%, 41%, and 81% higher than those of the wild-type strain, respectively. We isolated the sequences flanking the T-DNA insertions and thus identified new genes potentially involved in cellulase and hemicellulase production. Our results show that it is feasible to use T-DNA insertional mutagenesis to identify novel candidate genes involved in cellulase production. This will be valuable when genetic improvement programs seeking to enhance cellulase production are planned, and will also allow us to gain a better understanding of the genetics of the thermophilic fungus H. insolens. PMID:27506519

  6. The use of T-DNA insertional mutagenesis to improve cellulase production by the thermophilic fungus Humicola insolens Y1

    Science.gov (United States)

    Xu, Xinxin; Li, Jinyang; Shi, Pengjun; Ji, Wangli; Liu, Bo; Zhang, Yuhong; Yao, Bin; Fan, Yunliu; Zhang, Wei

    2016-01-01

    Humicola insolens is an excellent producer of pH-neutral active, thermostable cellulases that find many industrial applications. In the present study, we developed an efficient Agrobacterium tumefaciens-mediated transformation system for H. insolens. We transformed plasmids carrying the promoter of the glyceraldehyde-3-phosphate dehydrogenase gene of H. insolens driving the transcription of genes encoding neomycin phosphotransferase, hygromycin B phosphotransferase, and enhanced green fluorescent protein. We optimized transformation efficiency to obtain over 300 transformants/106 conidia. T-DNA insertional mutagenesis was employed to generate an H. insolens mutant library, and we isolated a transformant termed T4 with enhanced cellulase and hemicellulase activities. The FPase, endoglucanase, cellobiohydrolase, β-glucosidase, and xylanase activities of T4, measured at the end of fermentation, were 60%, 440%, 320%, 41%, and 81% higher than those of the wild-type strain, respectively. We isolated the sequences flanking the T-DNA insertions and thus identified new genes potentially involved in cellulase and hemicellulase production. Our results show that it is feasible to use T-DNA insertional mutagenesis to identify novel candidate genes involved in cellulase production. This will be valuable when genetic improvement programs seeking to enhance cellulase production are planned, and will also allow us to gain a better understanding of the genetics of the thermophilic fungus H. insolens. PMID:27506519

  7. Differential cellulolytic activity of native-form and C-terminal tagged-form cellulase derived from coptotermes formosanus and expressed in E. coli

    Science.gov (United States)

    The endogenous cellulase gene (CfEG3a) of Coptotermes formosanus, an economically important pest termite, was cloned and overexpressed in both native form (nCfEG) and C-terminal His-tagged form (tCfEG) in E.coli. Both forms of recombinant cellulases showed hydrolytic activity on cellulosic substrate...

  8. The effects of adding lactic acid bacteria and cellulase in oil palm (Elais guineensis Jacq.) frond silages on fermentation quality, chemical composition and in vitro digestibility

    NARCIS (Netherlands)

    Ebrahimi, Mahdi; Rajion, Mohamed Ali; Goh, Yong Meng; Farjam, Abdoreza Soleimani; Sazili, Awis Qurni; Schonewille, Jan Thomas

    2014-01-01

    The main objective of the current study was to evaluate whether oil palm frond (OPF) can be successfully ensiled without or with the additives cellulase or lactic acid bacteria (LAB). Thus, fresh OPF was ensiled either without additives or with cellulase or LAB or their combination. Ensiling was car

  9. Fungal cellulase/xylanase production and corresponding hydrolysis using pretreated corn stover as substrates.

    Science.gov (United States)

    Zhang, Liang; Wang, Xiaoqing; Ruan, Zhenhua; Liu, Ying; Niu, Xiaorui; Yue, Zhengbo; Li, Zhimin; Liao, Wei; Liu, Yan

    2014-01-01

    Three pretreated corn stover (ammonia fiber expansion, dilute acid, and dilute alkali) were used as carbon source to culture Trichoderma reesei Rut C-30 for cellulase and xylanase production. The results indicated that the cultures on ammonia fiber expansion and alkali pretreated corn stover had better enzyme production than the acid pretreated ones. The consequent enzymatic hydrolysis was performed applying fungal enzymes on pretreated corn stover samples. Tukey's statistical comparisons exhibited that there were significant differences on enzymatic hydrolysis among different combination of fungal enzymes and pretreated corn stover. The higher sugar yields were achieved by the enzymatic hydrolysis of dilute alkali pretreated corn stover.

  10. In Situ Stability of Substrate-Associated Cellulases Studied by DSC

    DEFF Research Database (Denmark)

    Borch, Kim; Cruys-Bagger, Nicolaj; Badino, Silke Flindt;

    2014-01-01

    This work shows that differential scanning calorimetry (DSC) can be used to monitor the stability of substrate-adsorbed cellulases during long-term hydrolysis of insoluble cellulose. Thermal transitions of adsorbed enzyme were measured regularly in subsets of a progressing hydrolysis, and the size....... There was no correlation between the intrinsic stability, specified by the transition temperature in the DSC, and the long-term stability derived from the peak area. The results are discussed with respect to the role of enzyme denaturation for the ubiquitous slowdown observed in the enzymatic hydrolysis of cellulose....

  11. A pyranose dehydrogenase-based biosensor for kinetic analysis of enzymatic hydrolysis of cellulose by cellulases

    DEFF Research Database (Denmark)

    Cruys-Bagger, Nicolaj; Badino, Silke Flindt; Tokin, Radina Naytchova;

    2014-01-01

    A novel electrochemical enzyme biosensor was developed for real-time detection of cellulase activity when acting on their natural insoluble substrate, cellulose. The enzyme biosensor was constructed with pyranose dehydrongease (PDH) from Agaricus meleagris that was immobilized on the surface...... jecorina (HjCel6A) on cellulosic substrates with different morphology (bacterial microcrystalline cellulose (BMCC) and Avicel). The steady-state rate of hydrolysis increased towards a saturation plateau with increasing loads of substrate. The experimental results were rationalized using a steady-state rate....... Biosensors covered with a polycarbonate membrane showed high operational stability of several weeks with daily use....

  12. Cellulase production by two mutant strain of Trichoderma longibrachiatum Qm9414 and Rut C30

    International Nuclear Information System (INIS)

    Native or pretreated biomass from Onopordum nervosum boiss, has been examined as candidate feedstock for cellulase production by two mutant strain of trichoderma longibrachiatum QM9414 and Rut C30. Batch cultivation methods were evaluated and compared with previous experiments using ball-milled, crystalline cellulose (Solka floc). Batch cultivation of T. longibrachiatum Rut C30 on 55% (W/V) acid pretreated O. nervosum biomass yielded enzyme productivities and activities comparable to those obtained on Solka floc. However, the overall enzyme production performance was lower than on Solka floc at comparable cellulose concentrations. This fact may be due to the accumulation of pretreated by products and lignin in the fermentor.(author)

  13. Cloning and identification of novel hydrolase genes from a dairy cow rumen metagenomic library and characterization of a cellulase gene

    Directory of Open Access Journals (Sweden)

    Gong Xia

    2012-10-01

    Full Text Available Abstract Background Interest in cellulose degrading enzymes has increased in recent years due to the expansion of the cellulosic biofuel industry. The rumen is a highly adapted environment for the degradation of cellulose and a promising source of enzymes for industrial use. To identify cellulase enzymes that may be of such use we have undertaken a functional metagenomic screen to identify cellulase enzymes from the bacterial community in the rumen of a grass-hay fed dairy cow. Results Twenty five clones specifying cellulose activity were identified. Subcloning and sequence analysis of a subset of these hydrolase-positive clones identified 10 endoglucanase genes. Preliminary characterization of the encoded cellulases was carried out using crude extracts of each of the subclones. Zymogram analysis using carboxymethylcellulose as a substrate showed a single positive band for each subclone, confirming that only one functional cellulase gene was present in each. One cellulase gene, designated Cel14b22, was expressed at a high level in Escherichia coli and purified for further characterization. The purified recombinant enzyme showed optimal activity at pH 6.0 and 50°C. It was stable over a broad pH range, from pH 4.0 to 10.0. The activity was significantly enhanced by Mn2+ and dramatically reduced by Fe3+ or Cu2+. The enzyme hydrolyzed a wide range of beta-1,3-, and beta-1,4-linked polysaccharides, with varying activities. Activities toward microcrystalline cellulose and filter paper were relatively high, while the highest activity was toward Oat Gum. Conclusion The present study shows that a functional metagenomic approach can be used to isolate previously uncharacterized cellulases from the rumen environment.

  14. Interkalation der Enzyme Papain und Cellulase sowie von Graphenoxid in Hydrotalcit- und Hydrocalumit-artige Schichtstrukturen (LDHs)

    OpenAIRE

    Zou, Nan

    2014-01-01

    In dieser Arbeit wurden der Einbau, die kristallchemischen und thermischen Eigenschaften von Interkalationsverbindungen der Enzyme Papain und Cellulase in Hydrotalcit-artige sowie von Graphenoxid (GO) in Hydrocalumit-artige Layered Double Hydroxides (LDHs) untersucht. Bei der Interkalationsverbindung Mg2Al-Papain-LDH wurde ein Schichtabstand von 5,3 nm gefunden, und für Mg2Al-Cellulase-LDH ein Wert von 5,0 nm. Mittels Anionenaustausch mit Sulfat können die Enzyme aus der Interkalationsverbind...

  15. Controlled production of cellulases in plants for biomass conversion. Progress report, June 15, 1996--March 10, 1997

    Energy Technology Data Exchange (ETDEWEB)

    Danna, K.J.

    1997-06-01

    The goal of this project is to facilitate conversion of plant biomass to usable energy by developing transgenic plants that express genes for microbial cellulases, which can be activated after harvest of the plants. In particular, we want to determine the feasibility of targeting an endoglucanase and a cellobiohydrolase to the plant apoplast (cell wall milieu). The apoplast not only contains cellulose, the substrate for the enzymes, but also can tolerate large amounts of foreign protein. To avoid detrimental effects of cellulase expression in plants, we have chosen enzymes with high temperature optima; the genes for these enzymes are from thermophilic organisms that can use cellulose as a sole energy source.

  16. Rather than by direct acquisition via lateral gene transfer, GHF5 cellulases were passed on from early Pratylenchidae to root-knot and cyst nematodes

    Directory of Open Access Journals (Sweden)

    Rybarczyk-Mydłowska Katarzyna

    2012-11-01

    Full Text Available Abstract Background Plant parasitic nematodes are unusual Metazoans as they are equipped with genes that allow for symbiont-independent degradation of plant cell walls. Among the cell wall-degrading enzymes, glycoside hydrolase family 5 (GHF5 cellulases are relatively well characterized, especially for high impact parasites such as root-knot and cyst nematodes. Interestingly, ancestors of extant nematodes most likely acquired these GHF5 cellulases from a prokaryote donor by one or multiple lateral gene transfer events. To obtain insight into the origin of GHF5 cellulases among evolutionary advanced members of the order Tylenchida, cellulase biodiversity data from less distal family members were collected and analyzed. Results Single nematodes were used to obtain (partial genomic sequences of cellulases from representatives of the genera Meloidogyne, Pratylenchus, Hirschmanniella and Globodera. Combined Bayesian analysis of ≈ 100 cellulase sequences revealed three types of catalytic domains (A, B, and C. Represented by 84 sequences, type B is numerically dominant, and the overall topology of the catalytic domain type shows remarkable resemblance with trees based on neutral (= pathogenicity-unrelated small subunit ribosomal DNA sequences. Bayesian analysis further suggested a sister relationship between the lesion nematode Pratylenchus thornei and all type B cellulases from root-knot nematodes. Yet, the relationship between the three catalytic domain types remained unclear. Superposition of intron data onto the cellulase tree suggests that types B and C are related, and together distinct from type A that is characterized by two unique introns. Conclusions All Tylenchida members investigated here harbored one or multiple GHF5 cellulases. Three types of catalytic domains are distinguished, and the presence of at least two types is relatively common among plant parasitic Tylenchida. Analysis of coding sequences of cellulases suggests that root

  17. Cellulase activity and dissolved organic carbon release from lignocellulose macrophyte-derived in four trophic conditions

    Directory of Open Access Journals (Sweden)

    Flávia Bottino

    2016-06-01

    Full Text Available Abstract Considering the importance of lignocellulose macrophyte-derived for the energy flux in aquatic ecosystems and the nutrient concentrations as a function of force which influences the decomposition process, this study aims to relate the enzymatic activity and lignocellulose hydrolysis in different trophic statuses. Water samples and two macrophyte species were collected from the littoral zone of a subtropical Brazilian Reservoir. A lignocellulosic matrix was obtained using aqueous extraction of dried plant material (≈40 °C. Incubations for decomposition of the lignocellulosic matrix were prepared using lignocelluloses, inoculums and filtered water simulating different trophic statuses with the same N:P ratio. The particulate organic carbon and dissolved organic carbon (POC and DOC, respectively were quantified, the cellulase enzymatic activity was measured by releasing reducing sugars and immobilized carbon was analyzed by filtration. During the cellulose degradation indicated by the cellulase activity, the dissolved organic carbon daily rate and enzyme activity increased. It was related to a fast hydrolysable fraction of cellulose that contributed to short-term carbon immobilization (ca. 10 days. After approximately 20 days, the dissolved organic carbon and enzyme activity were inversely correlated suggesting that the respiration of microorganisms was responsible for carbon mineralization. Cellulose was an important resource in low nutrient conditions (oligotrophic. However, the detritus quality played a major role in the lignocelluloses degradation (i.e., enzyme activity and carbon release.

  18. Hydrolysis of Ammonia-pretreated Sugar Cane Bagasse with Cellulase, β-Glucosidase, and Hemicellulase Preparations

    Science.gov (United States)

    Prior, Bernard A.; Day, Donal F.

    Sugar cane bagasse consists of hemicellulose (24%) and cellulose (38%), and bioconversion of both fractions to ethanol should be considered for a viable process. We have evaluated the hydrolysis of pretreated bagasse with combinations of cellulase, β-glucosidase, and hemicellulase. Ground bagasse was pretreated either by the AFEX process (2NH3: 1 biomass, 100 °C, 30 min) or with NH4OH (0.5 g NH4OH of a 28% [v/v] per gram dry biomass; 160 °C, 60 min), and composition analysis showed that the glucan and xylan fractions remained largely intact. The enzyme activities of four commercial xylanase preparations and supernatants of four laboratory-grown fungi were determined and evaluated for their ability to boost xylan hydrolysis when added to cellulase and β-glucosidase (10 filter paper units [FPU]: 20 cellobiase units [CBU]/g glucan). At 1% glucan loading, the commercial enzyme preparations (added at 10% or 50% levels of total protein in the enzyme preparations) boosted xylan and glucan hydrolysis in both pretreated bagasse samples. Xylanase addition at 10% protein level also improved hydrolysis of xylan and glucan fractions up to 10% glucan loading (28% solids loading). Significant xylanase activity in enzyme cocktails appears to be required for improving hydrolysis of both glucan and xylan fractions of ammonia pretreated sugar cane bagasse.

  19. Solid state fermentation for production of microbial cellulases: Recent advances and improvement strategies.

    Science.gov (United States)

    Behera, Sudhanshu S; Ray, Ramesh C

    2016-05-01

    Lignocellulose is the most plentiful non-food biomass and one of the most inexhaustible renewable resources on the planet, which is an alternative sustainable energy source for the production of second generation biofuels. Lignocelluloses are composed of cellulose, hemicellulose and lignin, in which the sugar polymers account for a large portion of the biomass. Cellulases belong to the glycoside hydrolase family and catalyze the hydrolysis of glyosidic linkages depolymerizing cellulose to fermentable sugars. They are multi-enzymatic complex proteins and require the synergistic action of three key enzymes: endoglucanase (E.C. 3.2.1.4), exoglucanase (E.C. 3.2.1.176) (E.C. 3.2.1.91) and β-glucosidase (E.C. 3.2.1.21) for the depolymerization of cellulose to glucose. Solid state fermentation, which holds growth of microorganisms on moist solid substrates in the absence of free flowing water, has gained considerable attention of late due its several advantages over submerged fermentation. The review summarizes the critical analysis of recent literature covering production of cellulase in solid state fermentation using advance technologies such as consolidated bioprocessing, metabolic engineering and strain improvement, and circumscribes the strategies to improve the enzyme yield. PMID:26601764

  20. The Relation Between Promoter Chromatin Status, Xyr1 and Cellulase Ex-pression in Trichoderma reesei.

    Science.gov (United States)

    Mello-de-Sousa, Thiago M; Rassinger, Alice; Derntl, Christian; Poças-Fonseca, Marcio J; Mach, Robert L; Mach-Aigner, Astrid R

    2016-04-01

    The ascomycete Trichoderma reesei is used for the production of plant cell wall-degrading enzymes in industrial scale. The interplay of the transactivator Xyr1 and the repressor Cre1 mainly regulates the expression of these enzymes. During induc-ing conditions, such as in the presence of sophorose, the transcription of the two major cellulase-encoding genes, cbh1 and cbh2, is activated as well as the expression of xyr1. In the presence of D-glucose carbon catabolite repression mediated by Cre1 takes place and the expression of Xyr1 and the plant cell wall-degrading enzymes is down-regulated. In this study we compare the chromatin status of xyr1, cbh1, and cbh2 promoters in the wild-type strain and the Cre1-deficient strain Rut-C30. Chromatin rearrangement occurs in the xyr1 promoter during induction on sophorose. Chromatin opening and protein-DNA interactions in the xyr1 promoter were detected especially in a region located 0.9 kb upstream the translation start co-don, which bears several putative Cre1-binding sites and a CCAAT-box. Moreover, the xyr1 promoter is overall more acces-sible in a cre1-truncated background, no matter which carbon source is present. This makes the xyr1 regulatory sequence a good target for promoter engineering aiming at the enhancement of cellulase production. PMID:27226770

  1. The Effect of Cellulose Crystal Structure and Solid-State Morphology on the Activity of Cellulases

    Energy Technology Data Exchange (ETDEWEB)

    Stipanovic, Arthur J [SUNY College of Environmental Science and Forestry

    2014-11-17

    Consistent with the US-DOE and USDA “Roadmap” objective of producing ethanol and chemicals from cellulosic feedstocks more efficiently, a three year research project entitled “The Effect of Cellulose Crystal Structure and Solid-State Morphology on the Activity of Cellulases” was initiated in early 2003 under DOE sponsorship (Project Number DE-FG02-02ER15356). A three year continuation was awarded in June 2005 for the period September 15, 2005 through September 14, 2008. The original goal of this project was to determine the effect of cellulose crystal structure, including allomorphic crystalline form (Cellulose I, II, III, IV and sub-allomorphs), relative degree of crystallinity and crystallite size, on the activity of different types of genetically engineered cellulase enzymes to provide insight into the mechanism and kinetics of cellulose digestion by “pure” enzymes rather than complex mixtures. We expected that such information would ultimately help enhance the accessibility of cellulose to enzymatic conversion processes thereby creating a more cost-effective commercial process yielding sugars for fermentation into ethanol and other chemical products. Perhaps the most significant finding of the initial project phase was that conversion of native bacterial cellulose (Cellulose I; BC-I) to the Cellulose II (BC-II) crystal form by aqueous NaOH “pretreatment” provided an increase in cellulase conversion rate approaching 2-4 fold depending on enzyme concentration and temperature, even when initial % crystallinity values were similar for both allomorphs.

  2. The influence of sorbitol on the production of cellulases and xylanases in an airlift bioreactor.

    Science.gov (United States)

    Ritter, Carla Eliana Todero; Fontana, Roselei Claudete; Camassola, Marli; da Silveira, Maurício Moura; Dillon, Aldo José Pinheiro

    2013-11-01

    The production of cellulases and xylanases by Penicillium echinulatum in an airlift bioreactor was evaluated. In batch production, we tested media with isolated or associated cellulose and sorbitol. In fed-batch production, we tested cellulose addition at two different times, 30 h and 48 h. Higher liquid circulation velocities in the downcomer were observed in sorbitol 10 g L(-1) medium. In batch production, higher FPA (filter paper activity) and endoglucanase activities were obtained with cellulose (7.5 g L(-1)) and sorbitol (2.5 g L(-1)), 1.0 U mL(-1) (120 h) and 6.4 U m L(-1) (100 h), respectively. For xylanases, the best production condition was cellulose 10 g L(-1), which achieved 5.5 U mL(-1) in 64 h. The fed-batch process was favorable for obtaining xylanases, but not for FPA and endoglucanases, suggesting that in the case of cellulases, the inducer must be added early in the process.

  3. Cellulase and Xylanase Production by Penicillium echinulatum in Submerged Media Containing Cellulose Amended with Sorbitol

    Directory of Open Access Journals (Sweden)

    Carla Eliana Todero Ritter

    2013-01-01

    Full Text Available The present work investigated the use of sorbitol as a soluble carbon source, in association with cellulose, to produce cellulases and xylanases in submerged cultures of Penicillium echinulatum 9A02S1. Because cellulose is an insoluble carbon source, in cellulase production, there are some problems with rheology and oxygen transfer. The submerged fermentations containing media composed of 0, 0.25, 0.5, 0.75, and 1% (w/v sorbitol and cellulose that were added at different times during the cultivation; 0.2% (w/v soy bran; 0.1% (w/v wheat bran; and a solution of salts. The highest filter paper activity (FPA ( IU·mL−1 was obtained on the seventh day in the medium containing 0.5% (w/v sorbitol and 0.5% (w/v cellulose added 24 h after the start of cultivation. However, the CMCases showed an activity peak on the sixth day ( IU·mL−1 in the medium containing 0.75% (w/v sorbitol and 0.75% (w/v cellulose added after 12 h of cultivation. The xylanases showed the highest activity in the medium with 0.75% (w/v sorbitol and 0.25% (w/v cellulose added 36 h after the start of cultivation. This strategy enables the reduction of the cellulose concentration, which in high concentrations can cause rheological and oxygen transfer problems.

  4. Understanding the Role of Physical Properties of Cellulose on Its Hydrolyzability by Cellulases

    Science.gov (United States)

    O'Dell, Patrick Jonathan

    Cellulose has long been explored as a potential feedstock for biofuel, however the recalcitrance of cellulose makes its conversion into biofuel much more challenging and economically unfavorable compared to well-established processes for converting starch or sugar feedstocks into biofuel. Enzymes capable of hydrolyzing cellulose into soluble sugars, glucose and cellobiose, have been found to work processively along cellulose microfibrils starting from reducing end groups. For this study, cellulose was produced and purified in-house from Gluconacetobacter xylinum cultures, and characterized by quantifying functional groups (aldehyde, ketone, and carboxyl groups) to determine the extent of oxidation of cellulose due to the processing steps. The main goal of this study was to look at the impacts of ultrasonication on cellulose's structure and the enzymatic hydrolyzability of cellulose. A completely randomized experimental design was used to test the effect of ultrasonication time and amplitude (intensity) on changes in cellulose fibril length, degree of polymerization, and rates and extents of hydrolysis. Results indicated that sonication time does significantly impact both the fibril length and average degree of polymerization of cellulose. The impact of ultrasonication on the hydrolyzability of cellulose by commercial cellulase and beta-glucosidase preparations could not be effectively resolved due to high variability in the experimental results. These studies serve as a basis for future studies understanding the role of cellulose microstructure in the mechanism of cellulase hydrolysis of cellulose.

  5. Entrapment of cross-linked cellulase colloids in alginate beads for hydrolysis of cellulose.

    Science.gov (United States)

    Nguyen, Le Truc; Lau, Yun Song; Yang, Kun-Lin

    2016-09-01

    Entrapment of enzymes in calcium alginate beads is a popular enzyme immobilization method. However, leaching of immobilized enzymes from the alginate beads is a common problem because enzyme molecules are much smaller than the pore size of alginate beads (∼200nm). To address this issue, we employ a millifluidic reactor to prepare cross-linked cellulase aggregate (XCA) colloids with a uniform size (∼300nm). Subsequently, these colloids are immobilized in calcium alginate beads as biocatalysts to hydrolyze cellulose substrates. By using fluorescent microscopy, we conclude that the immobilized XCA colloids distribute uniformly inside the beads and do not leach out from the beads after long-term incubation. Meanwhile, the pore size of the alginate beads is big enough for the cellulose substrates and fibers to diffuse into the beads for hydrolysis. For example, palm oil fiber and microcrystalline cellulose can be hydrolyzed within 48h and release reducing sugar concentrations up to 2.48±0.08g/l and 4.99±0.09g/l, respectively. Moreover, after 10 cycles of hydrolysis, 96.4% of the XCA colloids remain inside the alginate beads and retain 67% of the original activity. In contrast, free cellulase immobilized in the alginate beads loses its activity completely after 10 cycles. The strategy can also be used to prepare other types of cross-linked enzyme aggregates with high uniformity. PMID:27318817

  6. Endo-xylanase and endo-cellulase-assisted extraction of pectin from apple pomace.

    Science.gov (United States)

    Wikiera, Agnieszka; Mika, Magdalena; Starzyńska-Janiszewska, Anna; Stodolak, Bożena

    2016-05-20

    Pectins were extracted from apple pomace with monoactive preparation of endo-xylanase and endo-cellulase. The process was conducted for 10 h in conditions of pH 5.0 at 40 °C, with constant shaking. Endo-xylanase application resulted in the highest extraction efficiency of pectins (19.8%). The obtained polymer was characterised by a very high molecular mass, high level of neutral sugars - mainly arabinose, galactose and glucose, and very high DM (73.4). It also contained the highest level of protein and phenols. Pectin extracted with endo-cellulase had 1.5 fold lower molecular mass but contained significantly more GalA (70.5%) of a high degree of methylation (66.3%). The simultaneous application of both enzymatic preparations resulted in their cooperation, leading to a decrease of both the extraction efficiency and the molecular mass of pectin. However, this pectin was distinguished by the highest GalA (74.7%) and rhamnose contents.

  7. A minimal set of bacterial cellulases for consolidated bioprocessing of lignocellulose.

    Science.gov (United States)

    Liao, Hehuan; Zhang, Xiao-Zhou; Rollin, Joseph A; Zhang, Yi-Heng Percival

    2011-11-01

    Cost-effective release of fermentable sugars from non-food biomass through biomass pretreatment/enzymatic hydrolysis is still the largest obstacle to second-generation biorefineries. Therefore, the hydrolysis performance of 21 bacterial cellulase mixtures containing the glycoside hydrolase family 5 Bacillus subtilis endoglucanase (BsCel5), family 9 Clostridium phytofermentans processive endoglucanase (CpCel9), and family 48 C. phytofermentans cellobiohydrolase (CpCel48) was studied on partially ordered low-accessibility microcrystalline cellulose (Avicel) and disordered high-accessibility regenerated amorphous cellulose (RAC). Faster hydrolysis rates and higher digestibilities were obtained on RAC than on Avicel. The optimal ratios for maximum cellulose digestibility were dynamic for Avicel but nearly fixed for RAC. Processive endoglucanase CpCel9 was the most important for high cellulose digestibility regardless of substrate type. This study provides important information for the construction of a minimal set of bacterial cellulases for the consolidated bioprocessing bacteria, such as Bacillus subtilis, for converting lignocellulose to biocommodities in a single step.

  8. Cellulase and xylanase activity during the decomposition of three aquatic macrophytes in a tropical oxbow lagoon.

    Science.gov (United States)

    Sciessere, L; Cunha-Santino, M B; Bianchini, I

    2011-07-01

    Due to the connection between enzymatic activity and degradation of different fractions of organic matter, enzyme assays can be used to estimate degradation rates of particulate and dissolved organic carbon in freshwater systems. The aim of this study was to quantify and model the enzymatic degradation involving the decomposition of macrophytes, describing temporal activity of cellulases (EC 3.2.1.4 and EC 3.2.1.91) and xylanase (EC 3.2.1.8) during in situ decomposition of three aquatic macrophytes (Salvinia sp., Eichhornia azurea and Cyperus giganteus) on the surface and water-sediment interface (w-s interface) of an oxbow lagoon (Óleo lagoon) within a natural Brazilian Savanna Reserve. Overall, the enzymatic degradation of aquatic macrophytes in Óleo lagoon occurred during the whole year and was initiated together with leaching. Xylanase production was ca. 5 times higher than cellulase values due to easy access to this compound by cellulolytic microorganisms. Enzymatic production and detritus mass decay were similar on the surface and w-s interface. Salvinia sp. was the most recalcitrant detritus, with low mass decay and enzymatic activity. E. azurea and C. giganteus decomposition rates and enzymatic production were high and similar. Due to the physicochemical homogeneity observed in the Óleo lagoon, the differences between the decay rates of each species are mostly related with detritus chemical quality. PMID:24031706

  9. Neurospora crassa tox-1 Gene Encodes a pH- and Temperature-Tolerant Mini-Cellulase.

    Science.gov (United States)

    Xiao, Yue; Zhang, Qiongsi; Luo, Yiquan; Zhang, Ying; Luo, Xi; Wang, Yuchuan; Cao, Weiguo; Pinto, Vito De; Liu, Qiuyun; Li, Gang

    2016-06-15

    Cellulases that endure extreme conditions are essential in various industrial sectors. This study reports a mini-cellulase gene tox-1 from Neurospora crassa. The gene tox-1 was cloned in Escherichia coli after chimerization with the YebF gene and substitutions of certain isoleucine and valine with leucine residues. The yeast transformants could grow on rice straw-agar medium. The 44-amino acid peptide and its two mutant variants displayed potent cellulase activities in Congo Red assay and enzymatic assays. Conservative replacements with leucine have substantially increased the stabilities and half-lives of the peptides at alkaline pH and low and high temperatures and also the tolerance to organic solvents and surfactants, on the basis of activities toward cellose. The small size of the mini-cellulase would allow for commercially viable automatic chemical peptide synthesis. This work suggests that conservative leucine replacements may serve as a general strategy in the engineering of more robust enzymes with special features with little loss of activities. PMID:27229865

  10. Effect of Pre-harvest Treatments on the Cellulase Activity and Quality of Ber Fruit Under Cold Storage Conditions

    Directory of Open Access Journals (Sweden)

    Sukhjit Kaur JAWANDHA

    2009-12-01

    Full Text Available Studies were carried out to find out the effect of various pre-harvest treatments such as CaCl2 (@ 0.5%, 1.0% and 2.0%, Ca(NO32 (@0.5%, 1.0% and 2.0%, GA3 (@ 20, 40 and 60 ppm and Bavistin (@ 0.1% on the cellulase activity and quality of 'Umran' ber fruits during cold storage. Marked trees were sprayed at colour break stage with the test chemicals. Fruits were packed in CFB boxes and placed in cold storage (3-5 oC and 85 � 90 % RH for 30 days. The fruits were evaluated after 10, 20 and 30 days interval for various parameters such as cellulase activity, phenolics content, palatability rating and rotting percentage. Cellulase activity registered a gradual increase upto 20 days of storage thereafter a decline was noted in all the treatments. The palatability rating increased up to 10 days of storage in all the treatments, except control but subsequently it decreased with an advancement in storage period. Among the various pre-harvest treatments CaCl2 (2% recorded minimum cellulase activity and rotting percentage and have also registered high palatability rating and phenolics content during cold storage conditions. Studies showed that pre-harvest application of CaCl2 (2% maintained very good fruit quality and prolonged shelf-life for 20 days under cold storage conditions.

  11. Cost evaluation of cellulase enzyme for industrial-scale cellulosic ethanol production based on rigorous Aspen Plus modeling.

    Science.gov (United States)

    Liu, Gang; Zhang, Jian; Bao, Jie

    2016-01-01

    Cost reduction on cellulase enzyme usage has been the central effort in the commercialization of fuel ethanol production from lignocellulose biomass. Therefore, establishing an accurate evaluation method on cellulase enzyme cost is crucially important to support the health development of the future biorefinery industry. Currently, the cellulase cost evaluation methods were complicated and various controversial or even conflict results were presented. To give a reliable evaluation on this important topic, a rigorous analysis based on the Aspen Plus flowsheet simulation in the commercial scale ethanol plant was proposed in this study. The minimum ethanol selling price (MESP) was used as the indicator to show the impacts of varying enzyme supply modes, enzyme prices, process parameters, as well as enzyme loading on the enzyme cost. The results reveal that the enzyme cost drives the cellulosic ethanol price below the minimum profit point when the enzyme is purchased from the current industrial enzyme market. An innovative production of cellulase enzyme such as on-site enzyme production should be explored and tested in the industrial scale to yield an economically sound enzyme supply for the future cellulosic ethanol production.

  12. Production and Partial Characterization of Cellulases from Trichoderma sp. IS-05 Isolated from Sandy Coastal Plains of Northeast Brazil

    Directory of Open Access Journals (Sweden)

    Jackeline Pereira Andrade

    2011-01-01

    Full Text Available This study evaluated the production of cellulolytic enzymes by Trichoderma sp. IS-05 strain, isolated from sand dunes, according to its ability to grow on cellulose as carbon source. Wheat bran was tested as the carbon source and peptone tested as the nitrogen source. Different concentrations of carbon and nitrogen were tested using a factorial design to identify optimal cellulase activity production. The results showed that media containing wheat bran 4.0% (w/v and peptone 0.25% (w/v lead to the highest production, 564.0 U L−1 of cellulase, obtained after 2 days of fermentation. The pH and temperature profile showed optimal activity at pH 3.0 and 60∘C. As for thermostability, the cellulase was most tolerant at 60∘C, retaining more than 59.6% of maximal activity even after 4 hours of incubation. The combination of acid pH, high temperature tolerance, and production of cellulase from agro-industrial residues by Trichoderma sp. IS-05 offers possibilities condition for the biomass hydrolysis process to produce bioethanol.

  13. Utilization of spent coffee grounds for isolation and stabilization of Paenibacillus chitinolyticus CKS1 cellulase by immobilization.

    Science.gov (United States)

    Buntić, Aneta V; Pavlović, Marija D; Antonović, Dušan G; Šiler-Marinković, Slavica S; Dimitrijević-Branković, Suzana I

    2016-08-01

    This study has explored the feasibility of using spent coffee grounds as a good supporting material for the Paenibacillus chitinolyticus CKS1 cellulase immobilization. An optimal operational conditions in a batch-adsorption system were found to be: carrier mass of 12 g/L, under the temperature of 45 °C and no pH adjustments. The immobilization yield reached about 71%. An equilibrium establishment between the cellulase and the carrier surface occurred within 45 min, whereas the process kinetics may be predicted by the pseudo-second-order model. An immobilized cellulase preparation expressed very good avicelase activity, this reached up to 2.67 U/g, and revealed an improved storage stability property, compared to free enzyme sample counterpart. The addition of metal ions, such as K(+) and Mg(2+) did not affect positively immobilization yield results, but on the contrary, contributed to an improved bio-activities of the immobilized cellulase, thus may be employed before each enzyme application. The method developed in this study offers a cheap and effective alternative for immediate enzyme isolation from the production medium and its stabilization, compared to other carriers used for the immobilization. PMID:27626091

  14. Suppression of cellulase and polygalacturonase and induction of alcohol dehydrogenase isoenzymes in avocado fruit mesocarp subjected to low oxygen stress.

    Science.gov (United States)

    Kanellis, A K; Solomos, T; Roubelakis-Angelakis, K A

    1991-05-01

    Expression of polygalacturonase and cellulase, two hydrolytic enzymes of avocado (Persea americana, cv Hass) fruit which are synthesized de novo during ripening, and alcohol dehydrogenase, a known anaerobic protein, were studied under different O(2) regimes. Low O(2) concentrations (2.5-5.5%) diminished the accumulation of polygalacturonase and cellulase proteins and the expression of their isoenzymes. This pattern of change in cellulase protein was also reflected in the steady-state amount of its mRNA. In contrast, 7.5 and 10% O(2) did not alter the changes observed in fruits ripened in air. On the other hand, alcohol dehydrogenase was induced in 2.5, 3.5, and 5.5% O(2) but not in 7.5 or 10% O(2). The recovery from the hypoxic stress upon returning the fruits back to air for 24 hours, was also a function of O(2) tensions under which the fruits were kept. Thus, the synthesis of polygalacturonase and cellulase was directly related to O(2) levels, while the activity of the isoenzymes of alcohol dehydrogenase was inversely related to O(2) levels. The results indicate that hypoxia exerts both negative and positive effects on the expression of certain genes and that these effects are initiated at the same levels of O(2).

  15. Screening and characterization of a novel cellulase gene from the gut microflora of Hermetia illucens using metagenomic library.

    Science.gov (United States)

    Lee, Chang-Muk; Lee, Young-Seok; Seo, So-Hyeon; Yoon, Sang-Hong; Kim, Soo-Jin; Hahn, Bum-Soo; Sim, Joon-Soo; Koo, Bon-Sung

    2014-09-01

    A metagenomic fosmid library was constructed using genomic DNA isolated from the gut microflora of Hermetia illucens, a black soldier fly. A cellulase-positive clone, with the CS10 gene, was identified by extensive Congo-red overlay screenings for cellulase activity from the fosmid library of 92,000 clones. The CS10 gene was composed of a 996 bp DNA sequence encoding the mature protein of 331 amino acids. The deduced amino acids of CS10 showed 72% sequence identity with the glycosyl hydrolase family 5 gene of Dysgonomonas mossii, displaying no significant sequence homology to already known cellulases. The purified CS10 protein presented a single band of cellulase activity with a molecular mass of approximately 40 kDa on the SDS-PAGE gel and zymogram. The purified CS10 protein exhibited optimal activity at 50°C and pH 7.0, and the thermostability and pH stability of CS10 were preserved at the ranges of 20~50°C and pH 4.0~10.0. CS10 exhibited little loss of cellulase activity against various chemical reagents such as 10% polar organic solvents, 1% non-ionic detergents, and 0.5 M denaturing agents. Moreover, the substrate specificity and the product patterns by thinlayer chromatography suggested that CS10 is an endo-β-1,4-glucanase. From these biochemical properties of CS10, it is expected that the enzyme has the potential for application in industrial processes. PMID:25022521

  16. Construction of a cellulase hyper-expression system in Trichoderma reesei by promoter and enzyme engineering

    Directory of Open Access Journals (Sweden)

    Zou Gen

    2012-02-01

    Full Text Available Abstract Background Trichoderma reesei is the preferred organism for producing industrial cellulases. However, a more efficient heterologous expression system for enzymes from different organism is needed to further improve its cellulase mixture. The strong cbh1 promoter of T. reesei is frequently used in heterologous expression, however, the carbon catabolite repressor CREI may reduce its strength by binding to the cbh1 promoter at several binding sites. Another crucial point to enhance the production of heterologous enzymes is the stability of recombinant mRNA and the prevention of protein degradation within the endoplasmic reticulum, especially for the bacteria originated enzymes. In this study, the CREI binding sites within the cbh1 promoter were replaced with the binding sites of transcription activator ACEII and the HAP2/3/5 complex to improve the promoter efficiency. To further improve heterologous expression efficiency of bacterial genes within T. reesei, a flexible polyglycine linker and a rigid α-helix linker were tested in the construction of fusion genes between cbh1 from T. reesei and e1, encoding an endoglucanase from Acidothermus cellulolyticus. Results The modified promoter resulted in an increased expression level of the green fluorescent protein reporter by 5.5-fold in inducing culture medium and 7.4-fold in repressing culture medium. The fusion genes of cbh1 and e1 were successfully expressed in T. reesei under the control of promoter pcbh1m2. The higher enzyme activities and thermostability of the fusion protein with rigid linker indicated that the rigid linker might be more suitable for the heterologous expression system in T. reesei. Compared to the parent strain RC30-8, the FPase and CMCase activities of the secreted enzyme mixture from the corresponding transformant R1 with the rigid linker increased by 39% and 30% at 60°C, respectively, and the reduced sugar concentration in the hydrolysate of pretreated corn stover

  17. Cellulase variants

    Energy Technology Data Exchange (ETDEWEB)

    Blazej, Robert; Toriello, Nicholas; Emrich, Charles; Cohen, Richard N.; Koppel, Nitzan

    2015-07-14

    This invention provides novel variant cellulolytic enzymes having improved activity and/or stability. In certain embodiments the variant cellulotyic enzymes comprise a glycoside hydrolase with or comprising a substitution at one or more positions corresponding to one or more of residues F64, A226, and/or E246 in Thermobifida fusca Cel9A enzyme. In certain embodiments the glycoside hydrolase is a variant of a family 9 glycoside hydrolase. In certain embodiments the glycoside hydrolase is a variant of a theme B family 9 glycoside hydrolase.

  18. Produção de celulases por Aspergillus niger e cinética da desativação celulásica=Cellulases production by Aspergillus niger and cellulase deactivation kinetic

    Directory of Open Access Journals (Sweden)

    Caroline Mariana de Aguiar

    2011-10-01

    Full Text Available O presente trabalho teve como objetivo a avaliação da cinética de produção de enzimas celulases pelo fungo Aspergillus niger e da cinética de desativação das celulases. Foi utilizado bagaço de cana-de-açúcar pré-tratado como fonte de carbono na fermentação para a produção do complexo celulásico e também como substrato da hidrólise enzimática. A. niger foi cultivado em três bateladas, cada uma contendo 10, 50 e 100 g L-1 de bagaço pré-tratado com NaOH 4% (m v-1. A cinética da produção das celulases foi obtida determinando-se a atividade enzimática das amostras coletadas ao longo do tempo. As variações do pH também foram determinadas. A deativação enzimática foi avaliada determinando-se periodicamente a atividade das amostras armazenadas nas condições de resfriamento (4°C e de congelamento (-18ºC. Conclui-se que o A. niger produz celulases quando cultivado em meio de cultivo contendo bagaço de cana-de-açúcar pré-tratado e que o tempo ideal para coleta do caldo enzimático foi de aproximadamente sete dias, com produtividade máxima de 0,0013 U mL-1∙h para a batelada com 10 g L-1 e 0,0018 U mL-1∙h para as bateladas com 50 e 100 g L-1. O complexo celulásico não sofre desativação se armazenado à temperatura de -18°C por 43 dias, mas perde cerca de 40% da sua atividade após 48h se armazenado a 4°C.This work aimed to evaluate the kinetic for the cellulase production by Aspergillus niger and the deactivation kinetic of the cellulase enzymes. Cellulase were produced in three different batches using NaOH 4% (w v-1 pre-treated sugarcane bagasse as the carbon source in the fermentation broth. The amount of the bagasse in each batch was 10, 50 and 100 g L-1. The kinetic of the cellulase production was accomplished by periodically determining the cellulasic activity of the fermentation broth using pre-treated bagasse as the hydrolysis substrate. Changes in the pH also were determined. The cellulase

  19. Cellulases and beyond: the first 70 years of the enzyme producer Trichoderma reesei.

    Science.gov (United States)

    Bischof, Robert H; Ramoni, Jonas; Seiboth, Bernhard

    2016-01-01

    More than 70 years ago, the filamentous ascomycete Trichoderma reesei was isolated on the Solomon Islands due to its ability to degrade and thrive on cellulose containing fabrics. This trait that relies on its secreted cellulases is nowadays exploited by several industries. Most prominently in biorefineries which use T. reesei enzymes to saccharify lignocellulose from renewable plant biomass in order to produce biobased fuels and chemicals. In this review we summarize important milestones of the development of T. reesei as the leading production host for biorefinery enzymes, and discuss emerging trends in strain engineering. Trichoderma reesei has very recently also been proposed as a consolidated bioprocessing organism capable of direct conversion of biopolymeric substrates to desired products. We therefore cover this topic by reviewing novel approaches in metabolic engineering of T. reesei. PMID:27287427

  20. Production of cellulases and hemicellulases by an anaerobic mixed culture from lignocellulosic biomass.

    Science.gov (United States)

    Tabassum, R; Rajoka, M I; Malik, K A

    1990-03-01

    A comparison of different habitats, biogas plant, rumen fluid and sewage sludge, for cellulolytic organisms indicated sewage studge was the best source. Enrichment cultura gave a mixed culture which exhibited CMCase activity as well as extracellular Avicelase, xylanase, β-glucosidase, β-xylosidase activities and cell-bound β-glucosidase, and β-xylosidase production in a synthetic medium with eleven different cellulosic and lignocellulosic substrates. The activity of extracellular β-glucosidase and β-xylosidase production was significantly higher than endogenous activities. Hemicellulases were induced better than cellulases. The anzyme system was stable under aerobic conditions. Of the different lignocellulosic substrates, kallar grass was the best inducer of extracellular enzymes.

  1. Efficiency of new fungal cellulase systems in boosting enzymatic degradation of barley straw lignocellulose

    DEFF Research Database (Denmark)

    Rosgaard, L.; Pedersen, S.; Meyer, Anne Boye Strunge

    2006-01-01

    This study examined the cellulytic effects on steam-pretreated barley straw of cellulose-degrading enzyme systems from the five thermophilic fungi Chaetomium thermophilum, Thielavia terrestris, Thermoascus aurantiacus, Corynascus thermophilus, and Myceliophthora thermophila and from the mesophile...... Penicillum funiculosum. The catalytic glucose release was compared after treatments with each of the crude enzyme systems when added to a benchmark blend of a commercial cellulase product, Celluclast, derived from Trichoderma reesei and a P-glucosidase, Novozym 188, from Aspergillus niger. The enzymatic...... treatments were evaluated in an experimental design template comprising a span of pH (3.5-6.5) and temperature (35-65 degrees C) reaction combinations. The addition to Celluclast + Novozym 188 of low dosages of the crude enzyme systems, corresponding to 10 wt % of the total enzyme protein load, increased...

  2. Effect of repeated applications of buprofezin and acephate on soil cellulases, amylase, and invertase.

    Science.gov (United States)

    Raju, M Naga; Venkateswarlu, K

    2014-10-01

    The impact of repeated applications of buprofezin and acephate, at concentrations ranging from 0.25 to 1.0 kg ha(-1), on activities of cellulases, amylase, and invertase in unamended and nitrogen, phosphorous, and potassium (NPK) fertilizer-amended soil planted with cotton was studied. The nontarget effect of selected insecticides, when applied once, twice, or thrice on soil enzyme activities, was dose-dependent; the activities decreased with increasing concentrations of insecticides. However, there was a rapid decline in activities of enzymes after three repeated applications of insecticides in unamended or NPK-amended soil. Our data clearly suggest that insecticides must be applied judiciously in pest management in order to protect the enzymes largely implicated in soil fertility. PMID:24869954

  3. The Activity of Cellulase from Thermophilic Fungi Isolated from CaneBagasses

    International Nuclear Information System (INIS)

    The activity of cellulase from thermophilic fungi isolated from canebagasses has been measured. This wild strain, named fungal strain PJ-2,secreted a large amount of cellulolytic enzyme components consisting of 0.46units of avicelase, 0.8 units of carboxymethyl cellulose hydrolizing enzyme(CMCase) and 0.5 units of β-glucosidase per ml of culture broth oncultivation in Mandels Reese medium for 7 days at 500 oC. These cellulasesproduction was lower than that of Trichoderma reesei NRRL 3653 producing 0.5units/ml avicelase, 1.6 units/ml CMCase and 0.4 units/ml ofβ-glucosidase cultivated in the same medium at 30 oC. However,thermophilic fungi may be potential to be exploited in lignocellulosedegradation at the tropical areas as the process usually needs temperature ofabove 50 oC. (author)

  4. Cellulase-producing bacteria from Thai higher termites, Microcerotermes sp.: enzymatic activities and ionic liquid tolerance.

    Science.gov (United States)

    Taechapoempol, Kitipong; Sreethawong, Thammanoon; Rangsunvigit, Pramoch; Namprohm, Weerachart; Thamprajamchit, Bandhit; Rengpipat, Sirirat; Chavadej, Sumaeth

    2011-05-01

    The three highest hydrolysis-capacity-value isolates of Bacillus subtilis (A 002, M 015, and F 018) obtained from Thai higher termites, Microcerotermes sp., under different isolation conditions (aerobic, anaerobic, and anaerobic/aerobic) were tested for cellulase activities--FPase, endoglucanase, and β-glucosidase--at 37 °C and pH 7.2 for 24 h. Their tolerance to an ionic liquid, 1-butyl-3-methylimidazolium chloride ([BMIM]Cl), was also investigated. The results showed that the isolate M 015 provided the highest endoglucanase activity whereas the highest FPase and β-glucosidase activities were observed for the isolate F 018. The isolate F 018 also showed the highest tolerance to [BMIM]Cl in the range of 0.1-1.0 vol.%. In contrast, the isolate A 002 exhibited growth retardation in the presence of 0.5-1.0 vol.% [BMIM]Cl.

  5. Cellulase production by halophytic fungi. Pt. 1. Screening experiments and salt relations

    Energy Technology Data Exchange (ETDEWEB)

    Sallam, L.A.R.; EL-Refai, A.M.H.; El-Sayed, S.M.

    1988-01-01

    The potentiality of 3 local halophytic fungi, isolated from the salt marshes at Burg El-Arab area in the Western Mediterranean Coast of Egypt, to produce cellulose-hydrolysing enzymes has been studied under static culture conditions using 4 different media charged with avicel as a sole carbon source. Ulocladium chartarum was selected as the most potent for cellulase production. This fungus has not been proviously reported to possess cellulolytic activities. Dispensing the basal medium 2 mm depth and incubation of U. chartarum for 3 weeks were the most favourable conditions for best cellulolytic activities. The production of cellulolytic enzymes was markedly affected by the salt supplied and its concentration. Best CMCase and FPase activities (0.289 and 0.043 unit/ml respectively) were noticed on using 60 g Na/sub 2/SO/sub 4//l.

  6. Enhanced cellulase and β-glucosidase production by a mutant of Alternaria alternata

    International Nuclear Information System (INIS)

    The cellulolytic activity of the wild type and a mutant strain of A. alternata was investigated. Mutants were induced by gamma radiation. A suspension of about 105 condidia/mL in 0.05M phosphate buffer pH 5 were irradiated in a gamma-cell-type (Cammacell 220, Atomic Energy of Canada Limited, Ottawa, Canada) 60Co source with a dose rate of 2.5 krad/min. The amount of radiation given was 70 krad which resulted in about 10% survival level. The stock culture was maintained on a sterile growth medium supplemented with 1% cellulose 123 and 0.3% agar. Following the incubation period, the fungal biomass was harvested by centrifugation (5000g for 10 min) and the clarified supernatant was used as the source of cellulase and β-glucosidase

  7. Potential of Biosynthesized Silver Nanoparticles as Nanocatalyst for Enhanced Degradation of Cellulose by Cellulase

    Directory of Open Access Journals (Sweden)

    Bipinchandra K. Salunke

    2015-01-01

    Full Text Available Silver nanoparticles (AgNPs as a result of their excellent optical and electronic properties are promising catalytic materials for various applications. In this study, we demonstrate a novel approach for enhanced degradation of cellulose using biosynthesized AgNPs in an enzyme catalyzed reaction of cellulose hydrolysis by cellulase. AgNPs were synthesized through reduction of silver nitrate by extracts of five medicinal plants (Mentha arvensis var. piperascens, Buddleja officinalis Maximowicz, Epimedium koreanum Nakai, Artemisia messer-schmidtiana Besser, and Magnolia kobus. An increase of around twofold in reducing sugar formation confirmed the catalytic activity of AgNPs as nanocatalyst. The present study suggests that immobilization of the enzyme onto the surface of the AgNPs can be useful strategy for enhanced degradation of cellulose, which can be utilized for diverse industrial applications.

  8. Cellulose chain binding free energy drives the processive move of cellulases on the cellulose surface.

    Science.gov (United States)

    Wang, Yefei; Zhang, Shujun; Song, Xiangfei; Yao, Lishan

    2016-09-01

    Processivity is essential for cellulases in their catalysis of cellulose hydrolysis. But what drives the processive move is not well understood. In this work, we use Trichoderma reesei Cel7B as a model system and show that its processivity is directly correlated to the binding free energy difference of a cellulose chain occupying the binding sites -7 to +2 and that occupying sites -7 to -1. Several mutants that have stronger interactions with glycosyl units in sites +1 and +2 than the wild type enzyme show higher processivity. The results suggest that after the release of the product cellobiose located in sites +1 and +2, the enzyme pulls the cellulose chain to fill the vacant sites, which propels its processive move on the cellulose surface. Biotechnol. Bioeng. 2016;113: 1873-1880. © 2016 Wiley Periodicals, Inc. PMID:26928155

  9. Cellulase production by two mutant strain of Trichoderma longibranchiatum QM 9414 and Rut C30

    International Nuclear Information System (INIS)

    Native or pretreated biomass from Onopordum nervosum Boiss, has been examined as candidate feedstock for cellulase production by two mutant strain of Trichoderma Ionqibrachiatum QM9414 and Rut C30. Batch cultivation methods were evaluated and compared with previous experiments using ball-milled, crystalline cellulose (Solka floc). Batch cultivation of T. Ionqibrachiatum Rut C30 on 5% (w/v) acid pretreated O. nervosum biomass yielded enzyme productivities and activities comparable to those obtained on Solka floc. However, the overall enzyme production performance was lower than on Solka floc at comparable cellulose concentrations. This fact may be due to the accumulation of pretreated by products and lignin in the ferment. (Author) 40 refs

  10. Molecular Dynamics and Metadynamics Simulations of the Cellulase Cel48F

    Directory of Open Access Journals (Sweden)

    Osmair Vital de Oliveira

    2014-01-01

    Full Text Available Molecular dynamics (MD and metadynamics techniques were used to study the cellulase Cel48F-sugar. Cellulase is enzyme that breaks cellulose fibers into small sugar units and is potentially useful in second generation alcohol production. In MD simulations, the overall structure of equilibrated Cel48F did not significantly change along the trajectory, retaining root mean square deviation below 0.15 nm. A set of 15 residues interacting with the sugar chains via hydrogen bonding throughout the simulation was observed. The free energy of dissociation (ΔGdiss. of the chains in the catalytic tunnel of Cel48F was determined by metadynamics. The ΔGdiss. values of the chains entering and leaving the wild-type Cel48F cavity were 13.9 and 62.1 kcal/mol, respectively. We also mutated the E542 and Q543 to alanine residue and obtained ΔGdiss. of 41.8 and 45.9 kcal/mol, respectively. These mutations were found to facilitate smooth dissociation of the sugar chain across the Cel48F tunnel. At the entry of the Cel48F tunnel, three residues were mutated to alanine: T110, T213, and L274. Contrary to the T110A-Cel48F, the mutants T213-Cel48F and L274-Cel48F prevented the sugar chain from passing across the leaving site. The present results can be a guideline in mutagenesis studies to improve processing by Cel48F.

  11. Monocentric and polycentric anaerobic fungi produce structrally related cellulases and xylanases

    Energy Technology Data Exchange (ETDEWEB)

    Li, Xin-Liang; Chen, Huizhong; Ljungdahl, L.G. [Univ. of Georgia, Athens, GA (United States)

    1997-02-01

    Cellulase and xylanase cDNAs were isolated from a cDNA library of the polycentric anaerobic fungus Orpinomyces sp. strain PC-2 constructed in Escherichia coli. The cellulase cDNA (celB) was 1.8 kb long with an open reading frame (ORF) coding for a polypeptide of 471 amino acids, and the xylanase cDNA (xynA) was 1.2 kb long with an ORF encoding a polypeptide of 362 amino acids. Single transcripts of 1.9 kb for celB and 1.5 kb for xynA were detected in total RNA of Orpinomyces grown on Avicel. Genomic DNA regions coding for CelA and XynA were devoid of introns. The enzymes were highly homologous (80 to 85% identity) to the corresponding enzymes of the monocentric anaerobic fungus Neocallimastix patriciarum and, like those, contained in addition to a catalytic domain, a noncatalytic repeated peptide domain (NCRPD). The Orpinomyces xylanase contained one catalytic domain and thus differed from the Neocallimastix xylanase, which had two similar catalytic domains. Two peptides corresponding to the catalytic domain and the NCRPD of XynA were synthesized, and antibodies against them were raised and affinity column purified. The antibodies against the catalytic domain peptide reacted specifically with the xylanases of Orpinomyces and Neocallimastix, while the antibodies against the NCRPD reacted with many (at least eight) extracellular proteins of Orpinomyces and Neocallimastix, suggesting that the NCRPD is present in a number of polypeptides. 36 refs., 8 figs., 2 tabs.

  12. Cellulose crystallinity index: measurement techniques and their impact on interpreting cellulase performance

    Directory of Open Access Journals (Sweden)

    Parilla Philip A

    2010-05-01

    Full Text Available Abstract Although measurements of crystallinity index (CI have a long history, it has been found that CI varies significantly depending on the choice of measurement method. In this study, four different techniques incorporating X-ray diffraction and solid-state 13C nuclear magnetic resonance (NMR were compared using eight different cellulose preparations. We found that the simplest method, which is also the most widely used, and which involves measurement of just two heights in the X-ray diffractogram, produced significantly higher crystallinity values than did the other methods. Data in the literature for the cellulose preparation used (Avicel PH-101 support this observation. We believe that the alternative X-ray diffraction (XRD and NMR methods presented here, which consider the contributions from amorphous and crystalline cellulose to the entire XRD and NMR spectra, provide a more accurate measure of the crystallinity of cellulose. Although celluloses having a high amorphous content are usually more easily digested by enzymes, it is unclear, based on studies published in the literature, whether CI actually provides a clear indication of the digestibility of a cellulose sample. Cellulose accessibility should be affected by crystallinity, but is also likely to be affected by several other parameters, such as lignin/hemicellulose contents and distribution, porosity, and particle size. Given the methodological dependency of cellulose CI values and the complex nature of cellulase interactions with amorphous and crystalline celluloses, we caution against trying to correlate relatively small changes in CI with changes in cellulose digestibility. In addition, the prediction of cellulase performance based on low levels of cellulose conversion may not include sufficient digestion of the crystalline component to be meaningful.

  13. Cellulase and Xylanase Production by Penicillium echinulatum in Submerged Media Containing Cellulose Amended with Sorbitol.

    Science.gov (United States)

    Todero Ritter, Carla Eliana; Camassola, Marli; Zampieri, Denise; Silveira, Mauricio Moura; Dillon, Aldo José Pinheiro

    2013-01-01

    The present work investigated the use of sorbitol as a soluble carbon source, in association with cellulose, to produce cellulases and xylanases in submerged cultures of Penicillium echinulatum 9A02S1. Because cellulose is an insoluble carbon source, in cellulase production, there are some problems with rheology and oxygen transfer. The submerged fermentations containing media composed of 0, 0.25, 0.5, 0.75, and 1% (w/v) sorbitol and cellulose that were added at different times during the cultivation; 0.2% (w/v) soy bran; 0.1% (w/v) wheat bran; and a solution of salts. The highest filter paper activity (FPA) (1.95  ±  0.04 IU·mL(-1)) was obtained on the seventh day in the medium containing 0.5% (w/v) sorbitol and 0.5% (w/v) cellulose added 24 h after the start of cultivation. However, the CMCases showed an activity peak on the sixth day (9.99 ± 0.75 IU·mL(-1)) in the medium containing 0.75% (w/v) sorbitol and 0.75% (w/v) cellulose added after 12 h of cultivation. The xylanases showed the highest activity in the medium with 0.75% (w/v) sorbitol and 0.25% (w/v) cellulose added 36 h after the start of cultivation. This strategy enables the reduction of the cellulose concentration, which in high concentrations can cause rheological and oxygen transfer problems.

  14. Genome sequencing of the Trichoderma reesei QM9136 mutant identifies a truncation of the transcriptional regulator XYR1 as the cause for its cellulase-negative phenotype

    Energy Technology Data Exchange (ETDEWEB)

    Lichius, Alexander; Bidard, Frederique; Buchholz, Franziska; Le Crom, Stphane; Martin, Joel X.; Schackwitz, Wendy; Austerlitz, Tina; Grigoriev, Igor V.; Baker, Scott E.; Margeot, Antoine; Seiboth, Bernhard; Kubicek, Christian P.

    2015-12-01

    Background: Trichoderma reesei is the main industrial source of cellulases and hemicellulases required for the hydrolysis of biomass to simple sugars, which can then be used in the production of biofuels and biorefineries. The highly productive strains in use today were generated by classical mutagenesis. As byproducts of this procedure, mutants were generated that turned out to be unable to produce cellulases. In order to identify the mutations responsible for this inability, we sequenced the genome of one of these strains, QM9136, and compared it to that of its progenitor T. reesei QM6a. Results: In QM9136, we detected a surprisingly low number of mutagenic events in the promoter and coding regions of genes, i.e. only eight indels and six single nucleotide variants. One of these indels led to a frame-shift in the Zn2Cys6 transcription factor XYR1, the general regulator of cellulase and xylanase expression, and resulted in its C-terminal truncation by 140 amino acids. Retransformation of strain QM9136 with the wild-type xyr1 allele fully recovered the ability to produce cellulases, and is thus the reason for the cellulase-negative phenotype. Introduction of an engineered xyr1 allele containing the truncating point mutation into the moderate producer T. reesei QM9414 rendered this strain also cellulase-negative. The correspondingly truncated XYR1 protein was still able to enter the nucleus, but failed to be expressed over the basal constitutive level. Conclusion: The missing 140 C-terminal amino acids of XYR1 are therefore responsible for its previously observed auto-regulation which is essential for cellulases to be expressed. Our data present a working example of the use of genome sequencing leading to a functional explanation of the QM9136 cellulase-negative phenotype.

  15. IMPROVED CELLULASE PRODUCTION BY Aspergillus terreus USING OIL PALM EMPTY FRUIT BUNCH FIBRE AS SUBSTRATE IN A STIRRED TANK BIOREACTOR THROUGH OPTIMIZATION OF THE FERMENTATION CONDITIONS

    Directory of Open Access Journals (Sweden)

    Mahdi Shahriarinour

    2011-05-01

    Full Text Available Response surface methodology (RSM was performed to evaluate the effects of dissolved oxygen tension (DOT and initial pH on the production of carboxymethyl cellulase (CMCase, filter-paper hydrolase (FPase, and β-glucosidase by Aspergillus terreus in a 2 L stirred tank bioreactor. Delignified oil palm empty fruit bunch (OPEFB fibre was used as the main substrate under submerged fermentation. Growth of A. terreus and the production of three main components of cellulase were optimized by central composite design (CCD design. Statistical analysis of results showed that the individual terms of these two variables (DOT and pH had significant effects on growth and the production of all components of cellulase. Maximum growth (13.07 g/L and cellulase activity (CMCase = 50.33 U/mL, FPase = 2.29 U/mL and β-glucosidase = 15.98 U/ml were obtained when the DOT and initial culture pH were set at 55% and 5.5, respectively. A high proportion of β-glucosidase to FPase (8:1 in cellulase of A. terreus could be beneficial for efficient hydrolysis of cellulosic materials. The use of OPEFB as a main substrate would reduce the cost of fermentation for the production of cellulase.

  16. Effects of dietary addition of cellulase and a Saccharomyces cerevisiae fermentation product on nutrient digestibility, rumen fermentation and enteric methane emissions in growing goats.

    Science.gov (United States)

    Lu, Qi; Wu, Jian; Wang, Min; Zhou, Chuanshe; Han, Xuefeng; Odongo, Edwin Nicholas; Tan, Zhiliang; Tang, Shaoxun

    2016-01-01

    This study was designed to assess the effectiveness of dietary cellulase (243 U/g, derived from Neocallimastix patriciarum) and a Saccharomyces cerevisiae fermentation product (yeast product) on ruminal fermentation characteristics, enteric methane (CH4) emissions and methanogenic community in growing goats. The experiment was conducted in a 5 × 5 Latin square design using five Xiangdong black wether goats. The treatments included a Control and two levels of cellulase (0.8 g and 1.6 g/kg dry matter intake (DMI), i.e. 194 U/kg and 389 U/kg DMI, respectively) crossed over with two levels (6 g or 12 g/kg DMI) of the yeast product. There were no significant differences regarding feed intake, apparent digestibility of organic matter, neutral detergent fibre and acid detergent fibre among all the treatments. In comparison with the Control, the ruminal ammonia N concentration was decreased (p = 0.001) by cellulase and yeast product addition. The activities of carboxymethylcellulase and xylanase were decreased after cellulase addition. Moreover, dietary cellulase and yeast product addition led to a significant reduction (p diversity and copy numbers of methanogens among treatments were not dissimilar. The present results indicate that the combination of cellulase and yeast fermentation product can reduce the production of CH4 energy and mitigate the enteric CH4 emissions to a certain degree. PMID:27032031

  17. Recycling cellulases for cellulosic ethanol production at industrial relevant conditions: potential and temperature dependency at high solid processes.

    Science.gov (United States)

    Lindedam, Jane; Haven, Mai Østergaard; Chylenski, Piotr; Jørgensen, Henning; Felby, Claus

    2013-11-01

    Different versions of two commercial cellulases were tested for their recyclability of enzymatic activity at high dry matter processes (12% or 25% DM). Recyclability was assessed by measuring remaining enzyme activity in fermentation broth and the ability of enzymes to hydrolyse fresh, pretreated wheat straw. Industrial conditions were used to study the impact of hydrolysis temperature (40 or 50°C) and residence time on recyclability. Enzyme recycling at 12% DM indicated that hydrolysis at 50°C, though ideal for ethanol yield, should be kept short or carried out at lower temperature to preserve enzymatic activity. Best results for enzyme recycling at 25% DM was 59% and 41% of original enzyme load for a Celluclast:Novozyme188 mixture and a modern cellulase preparation, respectively. However, issues with stability of enzymes and their strong adsorption to residual solids still pose a challenge for applicable methods in enzyme recycling.

  18. "Newton's cradle" proton relay with amide-imidic acid tautomerization in inverting cellulase visualized by neutron crystallography.

    Science.gov (United States)

    Nakamura, Akihiko; Ishida, Takuya; Kusaka, Katsuhiro; Yamada, Taro; Fushinobu, Shinya; Tanaka, Ichiro; Kaneko, Satoshi; Ohta, Kazunori; Tanaka, Hiroaki; Inaka, Koji; Higuchi, Yoshiki; Niimura, Nobuo; Samejima, Masahiro; Igarashi, Kiyohiko

    2015-08-01

    Hydrolysis of carbohydrates is a major bioreaction in nature, catalyzed by glycoside hydrolases (GHs). We used neutron diffraction and high-resolution x-ray diffraction analyses to investigate the hydrogen bond network in inverting cellulase PcCel45A, which is an endoglucanase belonging to subfamily C of GH family 45, isolated from the basidiomycete Phanerochaete chrysosporium. Examination of the enzyme and enzyme-ligand structures indicates a key role of multiple tautomerizations of asparagine residues and peptide bonds, which are finally connected to the other catalytic residue via typical side-chain hydrogen bonds, in forming the "Newton's cradle"-like proton relay pathway of the catalytic cycle. Amide-imidic acid tautomerization of asparagine has not been taken into account in recent molecular dynamics simulations of not only cellulases but also general enzyme catalysis, and it may be necessary to reconsider our interpretation of many enzymatic reactions. PMID:26601228

  19. ROLE OF ALKALINE-TOLERANT FUNGAL CELLULASES IN RELEASE OF TOTAL ANTIOXIDANTS FROM AGRO-WASTES UNDER SOLID STATE FERMENTATION

    Directory of Open Access Journals (Sweden)

    Chinnarajan Ravindran

    2011-07-01

    Full Text Available The alkaline-tolerant marine-derived fungus Chaetomium globosum was tested for the production of enhanced levels of cellulases and free phenolics under highly alkaline conditions using agro wastes (cotton seed, sugar cane bagasse as substrates under solid state fermentation (SSF processes. In both the agro wastes used, an increase in cellulases (β-endoglucanase, β-Glucosidase, and β-exoglucanase production was observed with increase in pH. This enhanced carbohydrate-hydrolyzing enzymes (β-endoglucanase, β-Glucosidase and β-exoglucanase and thereby enriched the total phenolic release from agro-wastes under SSF conditions of higher pH. A linear correlation was observed between released total phenolic contents of agro-wastes and total antioxidant property. The increased antioxidant activity on free radical scavenging was also observed with the increase in pH. Thus, the present study makes it possible to produce nutraceutical ingredients cost-effectively from agricultural wastes.

  20. Regulation of amylase, cellulase and chitinase secretion in the digestive tract of the two-spotted field cricket, Gryllus bimaculatus.

    Science.gov (United States)

    Weidlich, Sandy; Müller, Sonja; Hoffmann, Klaus H; Woodring, Joseph

    2013-06-01

    The secretion of amylase and cellulase in Gryllus bimaculatus is determined by increased food intake, whereby shortly after molting food consumption increases. About half of the standing amylase concentration (activity) in the endothelial cells can be secreted within 30 min. The peak of amylase and cellulase secretion that occurs in the photophase is related to the feeding peak in the previous scotophase. The secretion of chitinase on the other hand is primarily controlled by the molting cycle. Only amylase secretion was affected by calcium in the incubation medium, suggesting an apocrine release mechanism. Refeeding experiments (after 5 days without food) suggest that the release of amylase in response to a nutrient in the lumen (glucose) is not due to simple stimulation of exocytosis, but rather a stimulation of synthesis. PMID:23585293

  1. Artificial Intelligence Techniques to Optimize the EDC/NHS-Mediated Immobilization of Cellulase on Eudragit L-100

    OpenAIRE

    Min-Chao He; Yun-Yun Liu,; Wei Qi; Zhen-Hong Yuan; Jing-Liang Xu,; Yu Zhang

    2012-01-01

    Two artificial intelligence techniques, namely artificial neural network (ANN) and genetic algorithm (GA) were combined to be used as a tool for optimizing the covalent immobilization of cellulase on a smart polymer, Eudragit L-100. 1-Ethyl-3-(3-dimethyllaminopropyl) carbodiimide (EDC) concentration, N-hydroxysuccinimide (NHS) concentration and coupling time were taken as independent variables, and immobilization efficiency was taken as the response. The data of the centr...

  2. Thermostability Mechanism for the Hyperthermophilicity of Extremophile Cellulase TmCel12A: Implied from Molecular Dynamics Simulation.

    Science.gov (United States)

    Lian, Peng; Yuan, Congmin; Xu, Qin; Fu, Wei

    2016-08-01

    Thermostability is of considerable importance for the application of cellulase in cellulosic ethanol production. The cellulase 12A from the hyperthermophile Thermotoga maritima (TmCel12A) is an ideal candidate to study thermostability of cellulases. Optimal temperature of the wild-type enzyme is 85 °C. Recently, it has been observed that surface loop mutation Y61G not only accelerates the hydrolysis rate but also extends the half-life of the enzyme at high temperature. However, the mechanism of how Y61G enhances thermostability of TmCel12A has not been revealed. Here, molecular dynamics simulation together with dynamic correlation network analysis was used to explore thermostability mechanism of TmCel12A. A hydrophobic cluster constructed by Y61, W176, V62, and L144 in the binding pocket was found to play a pivotal role in modulating thermostability as well as catalytic capability of TmCel12A. It stabilizes the apoenzyme at high temperature; however, it impedes the substrate binding. Y61G mutation disturbs the hydrophobic cluster as the counterpart amino acid W176 forms a cation-π interaction with R60 instead of the π-π interaction with Y61 in WT. Moreover, Y61G mutation makes the enzyme more rigid and more extended via altering the amino acid communities at the hinge part of the enzyme. An earlier hypothesis proposed from crystallographic observation that Y61G may accelerate the products releasing has been also confirmed by our simulations. These findings may provide a new direction for both theoretical and experimental scientists to improve the thermostability of other cellulases that can be potentially applied in biofuel industry. PMID:27384708

  3. Optimisation of Cellulase Production by Penicillium funiculosum in a Stirred Tank Bioreactor Using Multivariate Response Surface Analysis

    OpenAIRE

    Marcelle Lins de Albuquerque de Carvalho; Daniele Fernandes Carvalho; Edelvio de Barros Gomes; Roberto Nobuyuki Maeda; Lidia Maria Melo Santa Anna; Aline Machado de Castro; Nei Pereira

    2014-01-01

    Increasing interest in the production of second-generation ethanol necessitates the low-cost production of enzymes from the cellulolytic complex (endoglucanases, exoglucanases, and β-glucosidases), which act synergistically in cellulose breakdown. The present work aimed to optimise a bioprocess to produce these biocatalysts from the fungus Penicillium funiculosum ATCC11797. A statistical full factorial design (FFD) was employed to determine the optimal conditions for cellulase production. The...

  4. Kinetic transcriptome analysis reveals an essentially intact induction system in a cellulase hyper-producer Trichoderma reesei strain

    OpenAIRE

    Poggi-Parodi, Dante; Bidard, Frédérique; Pirayre, Aurélie; Portnoy, Thomas; Blugeon, Corinne; Seiboth, Bernhard; Kubicek, Christian P.; Le Crom, Stéphane; Margeot, Antoine

    2014-01-01

    Background The filamentous fungus Trichoderma reesei is the main industrial cellulolytic enzyme producer. Several strains have been developed in the past using random mutagenesis, and despite impressive performance enhancements, the pressure for low-cost cellulases has stimulated continuous research in the field. In this context, comparative study of the lower and higher producer strains obtained through random mutagenesis using systems biology tools (genome and transcriptome sequencing) can ...

  5. Molecular characterization of Portuguese populations of the pinewood nematode Bursaphelenchus xylophilus using cytochrome b and cellulase genes.

    Science.gov (United States)

    Valadas, V; Laranjo, M; Mota, M; Oliveira, S

    2013-12-01

    Bursaphelenchus xylophilus is the causal agent of pine wilt disease and a worldwide pest with high economic impact. Since its first diagnosis in Portugal in 1999, it has been subjected to quarantine measures with impact on forest health and ecosystem stability, significantly affecting international trade of wood products. The disease was detected in the north and centre of continental Portugal and, since 2008, the whole country has been considered an affected area. Recently, it was detected in Madeira Island. In order to avoid new outbreaks, it has become of major importance to understand the patterns of spread, introduction points and to characterize the new populations from continental Portugal and Madeira Island. Mitochondrial cytochrome b (cytb) and parasitic cellulase gene sequences were used to evaluate the genetic relationships among isolates that could indicate possible origins of the new outbreaks. Portuguese isolates were compared with isolates from USA, China, Japan and South Korea, in order to investigate possible infection pathways and disease spread patterns in Portugal. Phylogenetic trees based on both genes show that Portuguese isolates group with Asian isolates. Isolates from USA are in a separate position in both gene trees. However, the phylogenetic tree based on the cellulase gene sequences shows higher differentiation among Portuguese isolates than that of cytb. These results agree with those previously obtained using inter-simple sequence repeats (ISSR). This was the first study to use cytb and cellulase genes to characterize pinewood nematode (PWN) populations. This study suggests that cellulase is a better marker than cytb to study genetic diversity in B. xylophilus. PMID:23067571

  6. Optimization and characterization of alkaline protease and carboxymethyl-cellulase produced by Bacillus pumillus grown on Ficus nitida wastes

    OpenAIRE

    Eman Zakaria Gomaa

    2013-01-01

    The potentiality of 23 bacterial isolates to produce alkaline protease and carboxymethyl-cellulase (CMCase) on Ficus nitida wastes was investigated. Bacillus pumillus ATCC7061 was selected as the most potent bacterial strain for the production of both enzymes. It was found that the optimum production of protease and CMCase were recorded at 30 °C, 5% Ficus nitida leaves and incubation period of 72 h. The best nitrogen sources for protease and CMCase production were yeast extract and casein, re...

  7. Penicillium oxalicum PoFlbC regulates fungal asexual development and is important for cellulase gene expression.

    Science.gov (United States)

    Yao, Guangshan; Li, Zhonghai; Wu, Ruimei; Qin, Yuqi; Liu, Guodong; Qu, Yinbo

    2016-01-01

    Filamentous fungi can initiate vegetative growth on complex plant polysaccharides in nature through secreting a large amount of lignocellulose-degrading enzymes. These fungi develop a large amount of asexual spores to disperse and survive under harsh conditions, such as carbon and nitrogen depletion. Numerous studies report the presence of a cross-talk between asexual development and extracellular enzyme production, especially at the regulation level. This study identified and characterized a C2H2-type transcription factor called PoFlbC, which is an Aspergillus FlbC ortholog, in cellulolytic fungus Penicillium oxalicum. Results showed that the native level of PoFlbC was crucial for the normal growth and asexual development of P. oxalicum. Importantly, deletion of the PoflbC gene substantially reduced cellulase and hemicellulase productions. Comparative transcriptome analysis by RNA sequencing revealed a global downregulation of genes encoding cellulases, hemicellulases, and other proteins with functions in lignocellulose degradation. A similar defect was also observed in the OEPoflbC strain, suggesting that the production of cellulolytic enzymes was maintained by native expression of the PoflbC. In this study, an essential activator for both fungal asexual development and cellulase production was established in P. oxalicum.

  8. The correlation between mannanase and cellulase activities towards fibre content of palm oil sludge fermented with Aspergillus niger

    Directory of Open Access Journals (Sweden)

    T. Purwadaria

    1998-12-01

    Full Text Available Enzyme (mannanase and cellulase activities and fibre (hemicellulose, cellulose and lignin contents were determined during the fermentation course of palm oil sludge with Aspergillus niger TL (wild type and A. niger ES I (an asporogenous mutant. The analyses were carried out at the incubation time of 3 and 4 days of aerobic fennentation and at 2 days of anaerobic fermentation afterward. The correlations between mamlanase activity with hemicellulose content and cellulose activity with cellulose content were calculated by linear regression . The activities of matutanase and cellulase are increasing during the aerobic fennentation, while in the anaerobic fennentation the enzyme activities are decreasing due to instability of the enzymes. The enzyme activities of ESI are higher than the TL. The regression coefficient is highly significant for correlation between mamlanase and hemicellulose content of fermented product by ESI (r = 0.83; P0 .05 . Marutanase and cellulase activities were also detected after the fermented product dried at 60°C which indicated the enzymes are quite stable .

  9. Highly thermostable and pH-stable cellulases from Aspergillus niger NS-2: properties and application for cellulose hydrolysis.

    Science.gov (United States)

    Bansal, Namita; Janveja, Chetna; Tewari, Rupinder; Soni, Raman; Soni, Sanjeev Kumar

    2014-01-01

    Optimization of cultural conditions for enhanced cellulase production by Aspergillus niger NS-2 were studied under solid-state fermentation. Significant increase in yields (CMCase 463.9 ± 20.1 U/g, FPase 101.1 ± 3.5 U/g and β-glucosidase 99 ± 4.0 U/g) were obtained under optimized conditions. Effect of different nutritional parameters was studied to induce the maximum production of cellulase complex. Scale-up studies for enzyme production process were carried out. Characterization studies showed that enzymes produced by A. niger NS-2 were highly temperature- and pH stable. At 50 °C, the half life for CMCase, FPase, β-glucosidase were approximately 240 h. Cellulases from A. niger NS-2 were stable at 35 °C for 24 h over a broader pH range of 3.0-9.0. We examined the feasibility of using steam pretreatment to increase the saccharification yields from various lignocellulosic residues for sugar release which can potentially be used in bioethanol production. Saccharification of pretreated dry potato peels, carrot peels, composite waste mixture, orange peels, onion peels, banana peels, pineapple peels by crude enzyme extract from A. niger NS-2, resulted in very high cellulose conversion efficiencies of 92-98 %. PMID:24052336

  10. An integrative process of bioconversion of oil palm empty fruit bunch fiber to ethanol with on-site cellulase production.

    Science.gov (United States)

    Zhu, Youshuang; Xin, Fengxue; Zhao, Ying; Chang, Yunkang

    2014-11-01

    The aim of this study was to efficiently convert oil palm empty fruit bunch fiber (OPEFB), one of the most commonly generated lingo-wastes in Southeast Asia, into both cellulase and bioethanol. The unprocessed cellulase crude (37.29%) produced under solid-state fermentation using OPEFB as substrate showed a better reducing sugar yield using filter paper than the commercial enzyme blend (34.61%). Organosolv pretreatment method could efficiently reduce hemicellulose (24.3-18.6%) and lignin (35.2-22.1%) content and increase cellulose content (40.5-59.3%) from OPEFB. Enzymatic hydrolysis of pretreated OPEFB using the crude cellulase with 20% solid content, enzyme loading of 15 FPU/g OPEFB at 50 °C, and pH 5.5 resulted in a OPEFB hydrolysate containing 36.01 g/L glucose after 72 h. Fermentation of the hydrolysate medium produced 17.64 g/L ethanol with 0.49 g/g yield from glucose and 0.088 g/g yield from OPEFB at 8 h using Saccharomyces cerevisiae. PMID:24839153

  11. Correlation between Agar Plate Screening and Solid-State Fermentation for the Prediction of Cellulase Production by Trichoderma Strains

    Directory of Open Access Journals (Sweden)

    Camila Florencio

    2012-01-01

    Full Text Available The viability of converting biomass into biofuels and chemicals still requires further development towards the reduction of the enzyme production costs. Thus, there is a growing demand for the development of efficient procedures for selection of cellulase-producing microorganisms. This work correlates qualitative screening using agar plate assays with quantitative measurements of cellulase production during cultivation under solid-state fermentation (SSF. The initial screening step consisted of observation of the growth of 78 preselected strains of the genus Trichoderma on plates, using microcrystalline cellulose as carbon source. The 49 strains that were able to grow on this substrate were then subjected to a second screening step using the Congo red test. From this test it was possible to select 10 strains that presented the highest enzymatic indices (EI, with values ranging from 1.51 to 1.90. SSF cultivations using sugarcane bagasse and wheat bran as substrates were performed using selected strains. The CG 104NH strain presented the highest EGase activity (25.93 UI·g−1. The EI results obtained in the screening procedure using plates were compared with cellulase production under SSF. A correlation coefficient (R2 of 0.977 was obtained between the Congo red test and SSF, demonstrating that the two methodologies were in good agreement.

  12. Highly thermostable and pH-stable cellulases from Aspergillus niger NS-2: properties and application for cellulose hydrolysis.

    Science.gov (United States)

    Bansal, Namita; Janveja, Chetna; Tewari, Rupinder; Soni, Raman; Soni, Sanjeev Kumar

    2014-01-01

    Optimization of cultural conditions for enhanced cellulase production by Aspergillus niger NS-2 were studied under solid-state fermentation. Significant increase in yields (CMCase 463.9 ± 20.1 U/g, FPase 101.1 ± 3.5 U/g and β-glucosidase 99 ± 4.0 U/g) were obtained under optimized conditions. Effect of different nutritional parameters was studied to induce the maximum production of cellulase complex. Scale-up studies for enzyme production process were carried out. Characterization studies showed that enzymes produced by A. niger NS-2 were highly temperature- and pH stable. At 50 °C, the half life for CMCase, FPase, β-glucosidase were approximately 240 h. Cellulases from A. niger NS-2 were stable at 35 °C for 24 h over a broader pH range of 3.0-9.0. We examined the feasibility of using steam pretreatment to increase the saccharification yields from various lignocellulosic residues for sugar release which can potentially be used in bioethanol production. Saccharification of pretreated dry potato peels, carrot peels, composite waste mixture, orange peels, onion peels, banana peels, pineapple peels by crude enzyme extract from A. niger NS-2, resulted in very high cellulose conversion efficiencies of 92-98 %.

  13. Isolation of Homogeneous Polysaccharide Monooxygenases from Fungal Sources and Investigation of Their Synergism with Cellulases when Acting on Cellulose.

    Science.gov (United States)

    Bulakhov, A G; Gusakov, A V; Chekushina, A V; Satrutdinov, A D; Koshelev, A V; Matys, V Yu; Sinitsyn, A P

    2016-05-01

    Lytic polysaccharide monooxygenases (PMO) discovered several years ago are enzymes classified as oxidoreductases. In nature, they participate in microbial degradation of cellulose together with cellulases that belong to the hydrolytic type of enzymes (class of hydrolases). Three PMO from ascomycetes - Thielavia terrestris, Trichoderma reesei, and Myceliophthora thermophila - were isolated and purified to homogeneous state using various types of chromatography. The first two enzymes are recombinant proteins heterologously expressed by the Penicillium verruculosum fungus, while the third is a native PMO secreted by M. thermophila. When acting on microcrystalline cellulose, all these PMOs displayed synergism with the cellulase complex of the P. verruculosum fungus. Replacing 10% of cellulases (by protein concentration) with PMO in the presence of 6.25 mM gallic acid or 2.5 µM of cellobiose dehydrogenase from M. thermophila, used as electron donors for PMO, resulted in the 17-31% increase in the yield of reducing sugars after 24-48 h of the enzymatic reaction. PMID:27297903

  14. Two major facilitator superfamily sugar transporters from Trichoderma reesei and their roles in induction of cellulase biosynthesis.

    Science.gov (United States)

    Zhang, Weixin; Kou, Yanbo; Xu, Jintao; Cao, Yanli; Zhao, Guolei; Shao, Jing; Wang, Hai; Wang, Zhixing; Bao, Xiaoming; Chen, Guanjun; Liu, Weifeng

    2013-11-15

    Proper perception of the extracellular insoluble cellulose is key to initiating the rapid synthesis of cellulases by cellulolytic Trichoderma reesei. Uptake of soluble oligosaccharides derived from cellulose hydrolysis represents a potential point of control in the induced cascade. In this study, we identified a major facilitator superfamily sugar transporter Stp1 capable of transporting cellobiose by reconstructing a cellobiose assimilation system in Saccharomyces cerevisiae. The absence of Stp1 in T. reesei resulted in differential cellulolytic response to Avicel versus cellobiose. Transcriptional profiling revealed a different expression profile in the Δstp1 strain from that of wild-type strain in response to Avicel and demonstrated that Stp1 somehow repressed induction of the bulk of major cellulase and hemicellulose genes. Two other putative major facilitator superfamily sugar transporters were, however, up-regulated in the profiling. Deletion of one of them identified Crt1 that was required for growth and enzymatic activity on cellulose or lactose, but was not required for growth or hemicellulase activity on xylan. The essential role of Crt1 in cellulase induction did not seem to rely on its transporting activity because the overall uptake of cellobiose or sophorose by T. reesei was not compromised in the absence of Crt1. Phylogenetic analysis revealed that orthologs of Crt1 exist in the genomes of many filamentous ascomycete fungi capable of degrading cellulose. These data thus shed new light on the mechanism by which T. reesei senses and transmits the cellulose signal and offers potential strategies for strain improvement.

  15. Soluble inhibitors generated during hydrothermal pretreatment of oil palm mesocarp fiber suppressed the catalytic activity of Acremonium cellulase.

    Science.gov (United States)

    Zakaria, Mohd Rafein; Hirata, Satoshi; Fujimoto, Shinji; Ibrahim, Izzudin; Hassan, Mohd Ali

    2016-01-01

    Oil palm mesocarp fiber was subjected to hydrothermal pretreatment under isothermal and non-isothermal conditions. The pretreated slurries were separated by filtration, pretreated liquids and solids were characterized. An enzymatic digestibility study was performed for both pretreated slurries and solids to understand the effect of soluble inhibitors generated during the pretreatment process. The highest glucose yield obtained from pretreated slurries was 70.1%, and gradually decreased with higher pretreatment severities. The highest glucose yield obtained in pretreated solids was 100%, after pretreatment at 210°C for 20min. In order to study the inhibitory effects of compounds generated during pretreatment with cellulase, technical grade solutions that mimic the pretreated liquid were prepared and their effect on Acremonium cellulase activity was monitored using Avicel. Xylo-oligomers and tannic acid were identified as powerful inhibitors of Acremonium cellulase, and the lowest hydrolysis rate of Avicel of 0.18g/g-glucose released/L/h was obtained from tannic acid. PMID:26524253

  16. Composition and microstructure alteration of triticale grain surface after processing by enzymes of cellulase complex

    Directory of Open Access Journals (Sweden)

    Elena Kuznetsova

    2016-01-01

    Full Text Available It is found that the pericarp tissue of grain have considerable strength and stiffness, that has an adverse effect on quality of whole-grain bread. Thereby, there exists the need for preliminary chemical and biochemical processing of durable cell walls before industrial use. Increasingly used in the production of bread finds an artificial hybrid of the traditional grain crops of wheat and rye - triticale, grain which has high nutritional value. The purpose of this research was to evaluate the influence of cellulose complex (Penicillium canescens enzymes on composition and microstructure alteration of triticale grain surface, for grain used in baking. Triticale grain was processed by cellulolytic enzyme preparations with different composition (producer is Penicillium canescens. During experiment it is found that triticale grain processing by enzymes of cellulase complex leads to an increase in the content of water-soluble pentosans by 36.3 - 39.2%. The total amount of low molecular sugars increased by 3.8 - 10.5 %. Studies show that under the influence of enzymes the microstructure of the triticale grain surface is changing. Microphotographs characterizing grain surface structure alteration in dynamic (every 2 hours during 10 hours of substrate hydrolysis are shown. It is found that the depth and direction of destruction process for non-starch polysaccharides of grain integument are determined by the composition of the enzyme complex preparation and duration of exposure. It is found, that xylanase involved in the modification of hemicelluloses fiber having both longitudinal and radial orientation. Hydrolysis of non-starch polysaccharides from grain shells led to increase of antioxidant activity. Ferulic acid was identified in alcoholic extract of triticale grain after enzymatic hydrolysis under the influence of complex preparation containing cellulase, xylanase and β-glucanase. Grain processing by independent enzymes containing in complex

  17. The effects of adding lactic acid bacteria and cellulase in oil palm (Elais guineensis Jacq. frond silages on fermentation quality, chemical composition and in vitro digestibility

    Directory of Open Access Journals (Sweden)

    Mahdi Ebrahimi

    2014-07-01

    Full Text Available The main objective of the current study was to evaluate whether oil palm frond (OPF can be successfully ensiled without or with the additives cellulase or lactic acid bacteria (LAB. Thus, fresh OPF was ensiled either without additives or with cellulase or LAB or their combination. Ensiling was carried out by storing 2 kg samples in airtight glass jars at 25- 30°C for 12 weeks. Thereafter, the silage samples were subjected to proximate analyses, an in vitro digestibility assay and measures on selected indices of fermentation. Fermentation of OPF without additives appeared to be unsuccessful as both pH and ammonia content were too high (4.9 and 9.9%, respectively. In contrast, the use of cellulase or LAB resulted in silages with a pH<4.5 and ammonia fractions <8.4%, but the lowest values were found when both cellulase and LAB were used, i.e. pH=4.1 and ammonia fraction=6.7%. In vitro digestibility of dry matter was significantly higher in the cellulase treated silages. The process of ensiling was associated with both a significant decrease of the fat content of OPF and a significant change of the fatty acid profile. However, the proportions of major fatty acids (C16:0 and C18:2n-6 were not affected by the process of ensiling. In conclusion, the use of cellulase additive appears a practical tool to safeguard the process of fermentation. Using a cellulase enzyme or its combination with LAB improves the fermentation profile and increases the nutritional value of the OPF silage.

  18. Characterization, optimization, and scale-up of cellulases production by trichoderma reesei cbs 836.91 in solid-state fermentation using agro-industrial products.

    Science.gov (United States)

    Ortiz, Gastón E; Guitart, María E; Cavalitto, Sebastián F; Albertó, Edgardo O; Fernández-Lahore, Marcelo; Blasco, Martín

    2015-11-01

    The application of cellulases in saccharification processes is restricted by its production cost. Consequently, new fungal strains able to elaborate higher cellulases titers and with special activity profiles are required to make the process economical. The aim of this investigation was to find a promising wild-type Trichoderma strain for cellulases production. The Trichoderma reesei strain 938 (CBS 836.91) was selected among twenty strains on the basis of cellulase-agar-plate screening. Evaluation of the selected strain on six solid substrates indicated the highest activities to be obtained from wheat bran. Statistical analyses of the experimental design indicated a significant effect of pH and moisture on the generation of endoglucanase (EGA) and filter-paper (FPA) activity. Furthermore, a central-composite design-based optimization revealed that pH values between 6.4 and 6.6 and moisture from 74 to 94% were optimal for cellulases production. Under these conditions, 8-10 IU gds(-1) of FPA and 15.6-17.8 IU gds(-1) of EGA were obtained. In addition, cultivation in a rotating-drum reactor under optimal conditions gave 8.2 IU gds(-1) FPA and 13.5 IU gds(-1) EGA. Biochemical characterization of T. reesei 938 cellulases indicated a substantially higher resistance to 4 mM Fe(+2) and a slightly greater tolerance to alkaline pH in comparison to Celluclast(®). These results suggest that T. reesei 938 could be a promising candidate for improved cellulases production through direct-evolution strategies.

  19. Characterization, optimization, and scale-up of cellulases production by trichoderma reesei cbs 836.91 in solid-state fermentation using agro-industrial products.

    Science.gov (United States)

    Ortiz, Gastón E; Guitart, María E; Cavalitto, Sebastián F; Albertó, Edgardo O; Fernández-Lahore, Marcelo; Blasco, Martín

    2015-11-01

    The application of cellulases in saccharification processes is restricted by its production cost. Consequently, new fungal strains able to elaborate higher cellulases titers and with special activity profiles are required to make the process economical. The aim of this investigation was to find a promising wild-type Trichoderma strain for cellulases production. The Trichoderma reesei strain 938 (CBS 836.91) was selected among twenty strains on the basis of cellulase-agar-plate screening. Evaluation of the selected strain on six solid substrates indicated the highest activities to be obtained from wheat bran. Statistical analyses of the experimental design indicated a significant effect of pH and moisture on the generation of endoglucanase (EGA) and filter-paper (FPA) activity. Furthermore, a central-composite design-based optimization revealed that pH values between 6.4 and 6.6 and moisture from 74 to 94% were optimal for cellulases production. Under these conditions, 8-10 IU gds(-1) of FPA and 15.6-17.8 IU gds(-1) of EGA were obtained. In addition, cultivation in a rotating-drum reactor under optimal conditions gave 8.2 IU gds(-1) FPA and 13.5 IU gds(-1) EGA. Biochemical characterization of T. reesei 938 cellulases indicated a substantially higher resistance to 4 mM Fe(+2) and a slightly greater tolerance to alkaline pH in comparison to Celluclast(®). These results suggest that T. reesei 938 could be a promising candidate for improved cellulases production through direct-evolution strategies. PMID:26256022

  20. Cellulase production by Streptomyces viridobrunneus SCPE-09 using lignocellulosic biomass as inducer substrate.

    Science.gov (United States)

    Da Vinha, Fábio Nuno Marques; Gravina-Oliveira, Mônica Pires; Franco, Marcella Novaes; Macrae, Andrew; da Silva Bon, Elba Pinto; Nascimento, Rodrigo Pires; Coelho, Rosalie Reed Rodrigues

    2011-06-01

    An actinomycete strain, isolated from a soil sample under a sugar cane plantation in Brazil and identified as Streptomyces viridobrunneus SCPE-09, was selected as a promising cellulolytic strain, and tested for its ability to produce cellulases from agro-industrial residues. Sugar cane bagasse or wheat bran was tested as carbon source, and corn steep liquor tested as nitrogen source. Different concentrations of carbon and nitrogen were tested using factorial design to identify optimal cellulose production. The results showed that media containing wheat bran 2.0% (w/v) and corn steep liquid 0.19% (w/v) lead to the highest production, 2.0 U mL(-1) of CMCase, obtained on the fifth day of fermentation. The pH and temperature profile showed optimal activity at pH 4.9 and 50°C. As for thermostability, endoglucanases were most tolerant at 50°C, retaining more than 80% of maximal activity even after 2 h of incubation. Zymogram analyses using supernatant from growth under optimized conditions revealed the presence of two CMCase bands with apparent molecular masses of 37 and 119 kDa. The combination of pH tolerance and CMCase production from agro-industrial residues by S. viridobrunneus SCPE-09 offers promise for future bioethanol biotechnologies.

  1. Probing the nature of AFEX-pretreated corn stover derived decomposition products that inhibit cellulase activity.

    Science.gov (United States)

    Humpula, James F; Uppugundla, Nirmal; Vismeh, Ramin; Sousa, Leonardo; Chundawat, Shishir P S; Jones, A Daniel; Balan, Venkatesh; Dale, Bruce E; Cheh, Albert M

    2014-01-01

    Sequential fractionation of AFEX-pretreated corn stover extracts was carried out using ultra-centrifugation, ultra-filtration, and solid phase extraction to isolate various classes of pretreatment products to evaluate their inhibitory effect on cellulases. Ultra-centrifugation removed dark brown precipitates that caused no appreciable enzyme inhibition. Ultra-filtration of ultra-centrifuged AFEX-pretreated corn stover extractives using a 10 kDa molecular weight cutoff (MWCO) membrane removed additional high molecular weight components that accounted for 24-28% of the total observed enzyme inhibition while a 3 kDa MWCO membrane removed 60-65%, suggesting significant inhibition is caused by oligomeric materials. Solid phase extraction (SPE) of AFEX-pretreated corn stover extractives after ultra-centrifugation removed 34-43% of the inhibition; ultra-filtration with a 5 kDa membrane removed 44-56% of the inhibition and when this ultra-filtrate was subjected to SPE a total of 69-70% of the inhibition were removed. Mass spectrometry found several phenolic compounds among the hydrophobic inhibition removed by SPE adsorption.

  2. A thermophilic ionic liquid-tolerant cellulase cocktail for the production of cellulosic biofuels.

    Directory of Open Access Journals (Sweden)

    Joshua I Park

    Full Text Available Generation of biofuels from sugars in lignocellulosic biomass is a promising alternative to liquid fossil fuels, but efficient and inexpensive bioprocessing configurations must be developed to make this technology commercially viable. One of the major barriers to commercialization is the recalcitrance of plant cell wall polysaccharides to enzymatic hydrolysis. Biomass pretreatment with ionic liquids (ILs enables efficient saccharification of biomass, but residual ILs inhibit both saccharification and microbial fuel production, requiring extensive washing after IL pretreatment. Pretreatment itself can also produce biomass-derived inhibitory compounds that reduce microbial fuel production. Therefore, there are multiple points in the process from biomass to biofuel production that must be interrogated and optimized to maximize fuel production. Here, we report the development of an IL-tolerant cellulase cocktail by combining thermophilic bacterial glycoside hydrolases produced by a mixed consortia with recombinant glycoside hydrolases. This enzymatic cocktail saccharifies IL-pretreated biomass at higher temperatures and in the presence of much higher IL concentrations than commercial fungal cocktails. Sugars obtained from saccharification of IL-pretreated switchgrass using this cocktail can be converted into biodiesel (fatty acid ethyl-esters or FAEEs by a metabolically engineered strain of E. coli. During these studies, we found that this biodiesel-producing E. coli strain was sensitive to ILs and inhibitors released by saccharification. This cocktail will enable the development of novel biomass to biofuel bioprocessing configurations that may overcome some of the barriers to production of inexpensive cellulosic biofuels.

  3. Cold adaptation of a mesophilic cellulase, EG III from Trichoderma reesei, by directed evolution

    Institute of Scientific and Technical Information of China (English)

    XIAO; Zhizhuang(肖志壮); WANG; Pan(王攀); QU; Yinbo(曲音波); GAO; Peiji(高培基); WANG; Tianhong(汪天虹)

    2002-01-01

    Cold-active enzymes have received little research attention although they are very useful in industries. Since the structure bases of cold adaptation of enzymes are still unclear, it is also very difficult to obtain cold-adapted enzymes for industrial applications using routine protein engineering methods. In this work, we employed directed evolution method to randomly mutate a mesophilic cellulase, endoglucanase III (EG III) from Trichoderma reesei, and obtained a cold- adapted mutant, designated as w-3. DNA sequence analysis indicates that w-3 is a truncated form of native EG III with a deletion of 25 consecutive amino acids at C-terminus. Further examination of enzymatic kinetics and thermal stability shows that mutant w-3 has a higher Kcat value and becomes more thermolabile than its parent. In addition, activation energies of w-3 and wild type EG III calculated from Arrhenius equation are 13.3 kJ@mol-1 and 26.2 kJ@mol-1, respectively. Therefore, the increased specific activity of w-3 at lower temperatures could result from increased Kcat value and decreased activation energy.

  4. Accessibility of Enzymatically Delignified Bambusa bambos for Efficient Hydrolysis at Minimum Cellulase Loading: An Optimization Study

    Directory of Open Access Journals (Sweden)

    Arindam Kuila

    2011-01-01

    Full Text Available In the present investigation, Bambusa bambos was used for optimization of enzymatic pretreatment and saccharification. Maximum enzymatic delignification achieved was 84%, after 8 h of incubation time. Highest reducing sugar yield from enzyme-pretreated Bambusa bambos was 818.01 mg/g dry substrate after 8 h of incubation time at a low cellulase loading (endoglucanase, β-glucosidase, exoglucanase, and xylanase were 1.63 IU/mL, 1.28 IU/mL, 0.08 IU/mL, and 47.93 IU/mL, respectively. Enzyme-treated substrate of Bambusa bambos was characterized by analytical techniques such as Fourier transformed infrared spectroscopy (FTIR, X-ray diffraction (XRD, and scanning electron microscopy (SEM. The FTIR spectrum showed that the absorption peaks of several functional groups were decreased after enzymatic pretreatment. XRD analysis indicated that cellulose crystallinity of enzyme-treated samples was increased due to the removal of amorphous lignin and hemicelluloses. SEM image showed that surface structure of Bambusa bambos was distorted after enzymatic pretreatment.

  5. [Screening and Enzyme Production Characteristics of Thermophilic Cellulase-producing Strains].

    Science.gov (United States)

    Feng, Hong-mei; Qin, Yong-sheng; Li, Xiao-fan; Zhou, Jin-xing; Peng, Xia-wei

    2016-04-15

    A total of 6 thermophilic cellulase-producing strainswere isolated from organic garden waste mixed chicken composting at thermophilic period. These isolates were identified as Streptomyces thermoviolaceus, S. thermodiastaticus, S. thermocarboxydus, S. albidoflavus, S. thermovulgaris and Brevibacillus borstelensis through 16S rRNA gene sequence alignment and phylogenetic tree analysis. The cellulose-degrading microbial community has been investigated in few researches so far both at home and abroad. In this study, the mixed strains M-1 was made up of the 6 cellulose-decomposing microorganisms. The CMCase activity of the mixed strains M- 1 was stronger than any of the 6 single strains. Production of CMCase from mixed strains M-1 was studied by optimizing different physico-chemical parameters. The Maximum CMCase production (135.9 U · mL⁻¹) of strains M-1 was achieved at 45 °C in a liquid medium (pH 4) inoculated with 1% (volume fraction), containing a mixture of wheat bran and starch, corn flour and KNO₃. After optimization of separation conditions, CMCase production capacity was improved by 1.8 times. PMID:27548981

  6. Cloning and expression of cellulase XF-818 of Xylella fastidiosa in Escherichia Coli

    Directory of Open Access Journals (Sweden)

    Wulff Nelson Arno

    2003-01-01

    Full Text Available Xylella fastidiosa's genome was the first of a plant pathogen to be completely sequenced. Through comparative sequence analysis many genes were identified and, among them, several potentially involved in plant-pathogen interaction. However, the biological role of each gene should be assigned experimentally. On this regard, heterologous protein expression is a powerful tool to produce proteins from such genes, allowing their characterization. X. fastidiosa lives inside xylem vessels and eventually would degrade pit membranes from xylem cells to move radialy into the host. The identification of several putative plant cell wall degrading enzymes on X. fastidiosa genome prompted the assession of the function of such proteins. The open reading frame (ORF Xf-818 was cloned into expression vector pET20b and E. coli cells harboring such plasmid exhibited cellulase activity. Using IPTG at 0.4 mmol L-1 with a 12 h incubation at 32°C are the best conditions to produce higher amounts of heterologous protein. The enzyme degrades cellulose confirming the endoglucanase activity of Xf-818.

  7. Research on Extraction and Characterization of Cellulase from Commercial Enzyme Preparation

    Directory of Open Access Journals (Sweden)

    Guowei Shu

    2013-07-01

    Full Text Available The extraction of cellulase from commercial enzyme preparation by ammonium sulfate precipitation and its enzymatic characterization were studied. The results were as follows: the conditions of precipitation was 60% saturation of ammonium sulfate, the recovery rate of carboxymethylcellulase (CMCase and Filter Paper enzyme Activity (FPA were 73.8 and 71.4%, respectively. The enzyme has optimal temperature of 55°C, optimal pH of 4.8, the ions of Co2+, Mn2+ in the buffer lowered the activity of CMCase, but the Cu2+ in low concentration activated the CMCase. The ions of Co2+, Mn2+ and Al2+ in the buffer lowered the activity of FPA, but the Cu2+ and K+ in low concentration activated the FPA; Km and Vmax of the enzyme were 2.22×10-3 g/mL and 1.11×104 U/h using Lineweaver-Burk graphic method, respectively.

  8. Hydrolytic properties of two cellulases of Trichoderma reesei expressed in yeast.

    Science.gov (United States)

    Bailey, M J; Siika-aho, M; Valkeajärvi, A; Penttilä, M E

    1993-02-01

    Two cellulases of the filamentous fungus Trichoderma reesei, cellobiohydrolase II (CBHII, EC 3.2.1.91) and endoglucanase I (EGI, EC 3.2.1.4), produced in recombinant strains of the yeast Saccharomyces cerevisiae, were tested in the hydrolysis of cellulose, xylan and other polymeric substrates. Both enzymes were active against unsubstituted, insoluble cellulose. CBHII had greater activity than EGI against crystalline cellulose, whereas in the case of amorphous substrate the order was reversed. Evidence for synergism was obtained when mixtures of the two enzymes were used with a constant total protein dosage. The EGI was also active against soluble substituted cellulose derivatives, whereas the activity of CBHII against these substrates was insignificant. Both enzymes were active against barley (1-->3,1-->4)-beta-glucan, but were inactive against (1-->3,1-->6)-beta-glucan (laminarin). An apparent low mannan-degrading activity of EGI against locust-bean (Ceratonia siliqua) gum galactomannan was not confirmed when homopolymeric mannan was used as substrate in a prolonged hydrolysis test. EGI exhibited considerably greater activity against insoluble, unsubstituted hardwood xylan than against amorphous cellulose. Soluble 4-O-methyl-glucuronoxylan was also attacked by EGI, although to a somewhat lesser extent than the unsubstituted xylan. By comparison with two purified xylanases of T. reesei, EGI produced xylo-oligosaccharides with a longer mean chain length when acting on both substituted and unsubstituted xylan substrates. CBHII was inactive against xylan.

  9. Fermentation Optimization and Unstructured Kinetic Model for Cellulase Production by Rhizopus stolonifer var. reflexus TP-02 on Agriculture By-Products.

    Science.gov (United States)

    Li, Song; Tang, Bin; Xu, Zhongyuan; Chen, Tao; Liu, Long

    2015-12-01

    Agricultural by-products, rice straw, wheat bran juice, and soybean residue, were used as substrates for cellulase production using Rhizopus stolonifer var. reflexus TP-02. The culture medium was optimized though uniform design experimentation during shaking flask fermentation, and the ideal formulation obtained for filter paper enzyme (FPase) production was 10 % bran diffusion juice, 1 % rice straw, 0.17 % urea, 0.17 % soybean residue, 0.11 % KH2PO4, and 0.027 % Tween 80, and the maximal FPase activity in the culture supernatant was 13.16 U/mL at an incubation time of 3 days. A kinetic model for cellulase production in batch fermentation was subsequently developed. The unstructured kinetic model considered three responses, namely biomass, cellulase, and sugar. Models for the production of three types of cellulase components (i.e., endoglucanases, cellobiohydrolases, and β-glucosidases) were established to adequately describe the cellulase production pattern. It was found that the models fitted the experimental data well under pH 5.0 and 6.0, but only the avicelase production model predicted the experimental data under pH-uncontrolled conditions. PMID:26400494

  10. Proteomic profiling of cellulase-aid-extracted membrane proteins for functional identification of cellulose synthase complexes and their potential associated- components in cotton fibers.

    Science.gov (United States)

    Li, Ao; Wang, Ruyi; Li, Xianliang; Liu, Mingyong; Fan, Jian; Guo, Kai; Luo, Bing; Chen, Tingting; Feng, Shengqiu; Wang, Yanting; Wang, Bingrui; Peng, Liangcai; Xia, Tao

    2016-01-01

    Cotton fibers are an excellent model for understanding of cellulose biosynthesis in higher plants. In this study, we determined a high cellulose biosynthesis activity in vitro by optimizing biochemical reaction conditions in cotton fibers. By adding a commercial cellulase enzyme into fibers extraction process, we extracted markedly higher levels of GhCESA1 and GhCESA8 proteins and observed an increase in β-1,4-glucan and β-1,3-glucan products in vitro. LC-MS/MS analysis of anti-GhCESA8-immunoprecipitated proteins showed that 19 proteins could be found in three independent experiments including four CESAs (GhCESA1,2,7,8), five well-known non-CESA proteins, one callose synthase (CALS) and nine novel proteins. Notably, upon the cellulase treatment, four CESAs, one CALS and four novel proteins were measured at relatively higher levels by calculating total peptide counts and distinct peptide numbers, indicating that the cellulase-aid-extracted proteins most likely contribute to the increase in β-glucan products in vitro. These results suggest that the cellulase treatment may aid to release active cellulose synthases complexes from growing glucan chains and make them more amenable to extraction. To our knowledge, it is the first time report about the functional identification of the potential proteins that were associated with plant cellulose and callose synthases complexes by using the cellulase-aided protein extraction. PMID:27192945

  11. In situ stability of substrate-associated cellulases studied by DSC.

    Science.gov (United States)

    Alasepp, Kadri; Borch, Kim; Cruys-Bagger, Nicolaj; Badino, Silke; Jensen, Kenneth; Sørensen, Trine H; Windahl, Michael S; Westh, Peter

    2014-06-24

    This work shows that differential scanning calorimetry (DSC) can be used to monitor the stability of substrate-adsorbed cellulases during long-term hydrolysis of insoluble cellulose. Thermal transitions of adsorbed enzyme were measured regularly in subsets of a progressing hydrolysis, and the size of the transition peak was used as a gauge of the population of native enzyme. Analogous measurements were made for enzymes in pure buffer. Investigations of two cellobiohydrolases, Cel6A and Cel7A, from Trichoderma reesei, which is an anamorph of the fungus Hypocrea jerorina, showed that these enzymes were essentially stable at 25 °C. Thus, over a 53 h experiment, Cel6A lost less than 15% of the native population and Cel7A showed no detectable loss for either the free or substrate-adsorbed state. At higher temperatures we found significant losses in the native populations, and at the highest tested temperature (49 °C) about 80% Cel6A and 35% of Cel7A was lost after 53 h of hydrolysis. The data consistently showed that Cel7A was more long-term stable than Cel6A and that substrate-associated enzyme was less long-term stable than enzyme in pure buffer stored under otherwise equal conditions. There was no correlation between the intrinsic stability, specified by the transition temperature in the DSC, and the long-term stability derived from the peak area. The results are discussed with respect to the role of enzyme denaturation for the ubiquitous slowdown observed in the enzymatic hydrolysis of cellulose.

  12. Adsorption of cellulase on cereal brans: a simple functional model from response surface methodology

    Directory of Open Access Journals (Sweden)

    Rui Sergio F. da Silva

    1980-11-01

    Full Text Available A functional model based on Langmuirian adsorption as a limiting mechanism was proposed to explain the effect of cellulase during the enzymatic pretreatment of bran, conducted prior to extraction of proteins, by wet alkaline process from wheat and buckwheat bran materials. The proposed model provides a good fit (r = 0.99 for the data generated thru predictive model taken from the response surface methodology, permitting calculation of a affinity constant (b and capacity constant (k, for wheat bran (b = 0.255 g/IU and k = 17.42% and buckwheat bran (b = 0.066g/IUand k = 78.74%.Modelo funcional baseado na adsorção de Langmuir como mecanismo limitante proposto para explicar o efeito da celulase durante o pré-tratamento enzimático de farelos, visando à extração de proteínas, através do método alcalino-úmido. O referido modelo ajusta se muito bem (r = 0,99 aos dados gerados com base em modelo preditivo obtido da metodologia da superfície de resposta. Pode-se calcular a constante de afinidade (b e a constante de capacidade (k para o farelo de trigo e farelo de trigo mourisco (sarraceno, usando uma equação análoga à isoterma de adsorção de Langmuir. Os resultados indicaram que o farelo de trigo mourisco apresenta uma capacidade mais alta para adsorver celulase e, conseqüentemente,'pode-se esperar uma resposta maior ao pré-tratamento com esta enzima.

  13. 2007 GRC on Cellulases and Cellulosomes (July 29-August 3, 2007)

    Energy Technology Data Exchange (ETDEWEB)

    Mark Morrison

    2008-09-22

    industry have made many important and valuable contributions to the success of all the Conferences. This makes the Conference a truly interactive and productive venue for all sectors interested in the fundamental and applied sciences of cellulases, cellulosomes, and other carbohydrate active enzymes.

  14. PRODUCTION OF FERMENTABLE SUGARS FROM OIL PALM EMPTY FRUIT BUNCH USING CRUDE CELLULASE COCKTAILS WITH TRICHODERMA ASPERELLUM UPM1 AND ASPERGILLUS FUMIGATUS UPM2 FOR BIOETHANOL PRODUCTION

    Directory of Open Access Journals (Sweden)

    Nurul Kartini Abu Bakar,

    2012-06-01

    Full Text Available Utilization of oil palm empty fruit bunch (OPEFB for bioethanol production with crude cellulase cocktails from locally isolated fungi was studied. Enzymatic saccharification of alkaline pretreated OPEFB was done using different cellulase enzyme preparations. Crude cellulase cocktails from Trichoderma asperellum UPM1 and Aspergillus fumigatus UPM2 produced 8.37 g/L reducing sugars with 0.17 g/g yield. Production of bioethanol from OPEFB hydrolysate using Baker’s yeast produced approximately 0.59 g/L ethanol, corresponding to 13.8% of the theoretical yield. High reducing sugars concentration in the final fermentation samples resulted from accumulation of non-fermentable sugars such as xylose and cellobiose that were not consumed by the yeast. The results obtained support the possible utilization of OPEFB biomass for bioethanol production in the future.

  15. Simultaneous Cellulase Production, Saccharification and Detoxification Using Dilute Acid Hydrolysate of S. spontaneum with Trichoderma reesei NCIM 992 and Aspergillus niger.

    Science.gov (United States)

    Sateesh, Lanka; Rodhe, Adivikatla Vimala; Naseeruddin, Shaik; Yadav, Kothagauni Srilekha; Prasad, Yenumulagerard; Rao, Linga Venkateswar

    2012-06-01

    Bioethanol production from lignocellulosic materials has several limitations. One aspect is the high production cost of cellulases used for saccharification of substrate and inhibition of fermenting yeast due to inhibitors released in acid hydrolysis. In the present work we have made an attempt to achieve simultaneous cellulases production, saccharification and detoxification using dilute acid hydrolysate of Saccharum spontaneum with and without addition of nutrients, supplemented with acid hydrolyzed biomass prior to inoculation in one set and after 3 days of inoculation in another set. Organisms used were T. reesei NCIM 992, and Aspergillus niger isolated in our laboratory. Cellulase yield obtained was 0.8 IU/ml on fourth day with T. reesei. Sugars were found to increase from fourth to fifth day, when hydrolysate was supplemented with nutrients and acid hydrolyzed biomass followed by inoculation with T. reesei. Phenolics were also found to decrease by 67%. PMID:23729891

  16. Thermodynamics of adsorption/desorption of cellulases NS 50013 on /from Avicel PH 101 and Protobind 1000

    Directory of Open Access Journals (Sweden)

    Khurram Shahzad Baig

    2016-06-01

    Full Text Available Insight from thermodynamic parameters of enthalpy (ΔH, entropy (ΔS and Gibbs free energy (ΔG was used to predict conditions for desorption of cellulases from wheat straw. The analogues of the cellulose and lignin components of wheat straw used were Avicel PH 101 and Protobind 1000, respectively. The ΔHa for adsorption on Avicelat pH 5 was -16.10kJmol-1 , ΔSa was -50.10 Jmol-1K -1 which indicated an increase in order due to adsorption and ΔGa was negative only from 298K to 323K. Results of adsorption on Protobind 1000 were pretty much the inverse over the same range of temperature, proving the great affinity of cellulases for lignin. Over 298K to 333K, desorption from Avicel resulted in a ΔHdincrease from 17.50 to 26.20 kJmol-1 , and a ΔSdincrease from 46.89 to 75.65Jmol-1K -1when pH increased from 6 to 9indicating an enthalpy-driven, nonspontaneous desorption. For Protobind, the positive ΔHd (9.65 to 6.90 kJmol-1 with small positive ΔSd (21.70 to 9.03 Jmol-1K -1 indicated that disorder was less than that of Avicel. For both the substrates ΔGd decreased with rise in temperature in given temperature range. The minimum ΔGdof Avicel than that of Protobind proved that it was more difficult to desorb cellulases from Protobind.For bioethanol producing industries, using lignocellulosic material (e.g., wheat straw where cellulose is embedded in lignin, removal of lignin is recommend along with adsorption/hydrolysis to be conducted at 323 K and desorption from used material at 333 K and pH 9.

  17. Simultaneous production of cellulase and reducing sugar from alkali-pretreated sugarcane bagasse via solid state fermentation

    Directory of Open Access Journals (Sweden)

    Li Wan Yoon

    2012-11-01

    Full Text Available This study optimized alkali pretreatment of sugarcane bagasse (SCB and investigated the potential of alkali-pretreated SCB in producing cellulase and reducing sugar by a white-rot fungus, P. sanguineus, via solid state fermentation (SSF. The fermentability of the reducing sugar produced during SSF was examined by co-culturing yeast, Saccharomyces cerevisiae, with P. sanguineus. Central composite design (CCD was applied to optimize the pretreatment based on reducing sugar yield obtained from enzymatic hydrolysis of the pretreated SCB. The model developed from CCD fitted the data well, and the optimized conditions for alkali pretreatment were 128 °C, 0.62 M NaOH, and 30 min with a reducing sugar yield of 97.8%. The alkali-pretreated SCB after washing and drying was cultivated with P. sanguineus during SSF. It was found that cellulase and reducing sugar can be produced simultaneously from this SSF system. The maximum cellulase activities determined from filter paper assay (FPase, carboxylmethylcellulase (CMCase assay and β-glucosidase assay were 0.02 IU/mL, 0.11 IU/mL, and 0.13 IU/mL on day 8, day 3, and day 6 of cultivation, respectively. The maximum reducing sugar concentration of 19.9 mg/g pretreated SCB was obtained on day 4 of SSF. The reducing sugar produced was converted into ethanol upon the addition of yeast into the SSF system. Evidently, the reducing sugar acquired can be further utilized to produce other valuable products in subsequent processes.

  18. A carboxymethyl cellulase from a marine yeast ( Aureobasidium pullulans 98): Its purification, characterization, gene cloning and carboxymethyl cellulose digestion

    Science.gov (United States)

    Rong, Yanjun; Zhang, Liang; Chi, Zhenming; Wang, Xianghong

    2015-10-01

    We have reported that A. pullulans 98 produces a high yield of cellulase. In this study, a carboxymethyl cellulase (CMCase) in the supernatant of the culture of A. pullulans 98 was purified to homogeneity, and the maximum production of CMCase was 4.51 U (mg protein)-1. The SDS-PAGE analysis showed that the molecular mass of the purified CMCase was 67.0 kDa. The optimal temperature of the purified enzyme with considerable thermosensitivity was 40°C, much lower than that of the CMCases from other fungi. The optimal pH of the enzyme was 5.6, and the activity profile was stable in a range of acidity (pH 5.0-6.0). The enzyme was activated by Na+, Mg2+, Ca2+, K+, Fe2+ and Cu2+, however, it was inhibited by Fe3+, Ba2+, Zn2+, Mn2+ and Ag+. K m and V max values of the purified enzyme were 4.7 mg mL-1 and 0.57 µmol L-1 min-1 (mg protein)-1, respectively. Only oligosaccharides with different sizes were released from carboxymethylcellulose (CMC) after hydrolysis with the purified CMCase. The putative gene encoding CMCase was cloned from A. pullulans 98, which contained an open reading frame of 954 bp (EU978473). The protein deduced contained the conserved domain of cellulase superfamily (glucosyl hydrolase family 5). The N-terminal amino acid sequence of the purified CMCase was M-A-P-H-A-E-P-Q-S-Q-T-T-E-Q-T-S-S-G-Q-F, which was consistent with that deduced from the cloned gene. This suggested that the purified CMCase was indeed encoded by the cloned CMCase gene in this yeast.

  19. One-step purification and characterization of cellulase-free xylanase produced by alkalophilic Bacillus subtilis ash

    OpenAIRE

    Sanghi, Ashwani; Garg, Neelam; Gupta, V. K.; Mittal, Ashwani; R.C. Kuhad

    2010-01-01

    The present study describes the one-step purification and characterization of an extracellular cellulase-free xylanase from a newly isolated alkalophilic and moderately thermophilic strain of Bacillus subtilis ASH. Xylanase was purified to homogeneity by 10.5-fold with ~43% recovery using ion-exchange chromatography through CM-Sephadex C-50. The purified enzyme revealed a single band on SDS-PAGE gel with a molecular mass of 23 kDa. It showed an optimum pH at 7.0 and was stable over the pH ran...

  20. Effect of temperature on the production of cellulases, xylanases and lytic enzymes by selected Trichoderma reesei mutants

    OpenAIRE

    Piotr Janas; Zdzisław Targoński

    2014-01-01

    The effect of temperature in the rangę of 26-38°C on the production of cellulases, xylanases and lytic enzymes by four mutant strains of Trichoderma reesei was analysed. On the basis of these investigations three thermosensitive strains (M-7. RUT C 30 and VTT-D-78085) which showed reduced excretion of the above mentioned enzymes as well as protein and a thermoresistant mutant (VTT-D-79I24) which grew within a temperature range of 26-34°C were characterized. Higher temperature caused an increa...

  1. ENHANCED PRODUCTION OF CELLULASE-FREE XYLANASE BY ALKALOPHILIC BACILLUS SUBTILIS ASH AND ITS APPLICATION IN BIOBLEACHING OF KRAFT PULP

    OpenAIRE

    Ashwani Sanghi; Neelam Garg; Kalika Kuhar; Kuhad, Ramesh C.; Gupta, Vijay K

    2009-01-01

    This paper reports high level production of a cellulase-free xylanase using wheat bran, a cost-effective substrate, under submerged fermentation by alkalophilic Bacillus subtilis ASH. Production of xylanase was observed even at alkaline pH up to 11.0 and temperature 60 °C, although the highest enzyme titer was recorded at neutral pH and 37 °C. The enzyme production under optimized fermentation was 1.5-fold greater than under unoptimized conditions. Pre-treatment of unbleached pulp of 10% cons...

  2. Reversibility of substrate adsorption for the cellulases Cel7A, Cel6A and Cel7B from H. jecorina

    DEFF Research Database (Denmark)

    Pellegrini, Vanessa de Oliveira Arnoldi; Lei, Nina; Kysaram, Madhuri;

    2014-01-01

    Adsorption of cellulases on the cellulose surface is an integral part of the catalytic mechanism, and a detailed description of the adsorption process is therefore required for a fundamental understanding of this industrially important class of enzymes. However, the mode of adsorption has proven...... cellobiohydrolases (Cel7A and Cel6A) and one endoglucanase (Cel7B) on four types of pure cellulose substrates. Specifically, we monitored dilution-induced release of adsorbed enzyme in samples that had previously been brought to a steady state (constant concentration of free enzyme). In simple dilution experiments...

  3. Modulation of cellulase activity by charged lipid bilayers with different acyl chain properties for efficient hydrolysis of ionic liquid-pretreated cellulose.

    Science.gov (United States)

    Mihono, Kai; Ohtsu, Takeshi; Ohtani, Mai; Yoshimoto, Makoto; Kamimura, Akio

    2016-10-01

    The stability of cellulase activity in the presence of ionic liquids (ILs) is critical for the enzymatic hydrolysis of insoluble cellulose pretreated with ILs. In this work, cellulase was incorporated in the liposomes composed of negatively charged 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoglycerol (POPG) and zwitterionic phosphatidylcholines (PCs) with different length and degree of unsaturation of the acyl chains. The liposomal cellulase-catalyzed reaction was performed at 45°C in the acetate buffer solution (pH 4.8) with 2.0g/L CC31 as cellulosic substrate. The crystallinity of CC31 was reduced by treating with 1-butyl-3-methylimidazolium chloride ([Bmim]Cl) at 120°C for 30min. The liposomal cellulase continuously catalyzed hydrolysis of the pretreated CC31 for 48h producing glucose in the presence of 15wt% [Bmim]Cl. The charged lipid membranes were interactive with [Bmim](+), as elucidated by the [Bmim]Cl-induced alterations in fluorescence polarization of the membrane-embedded 1,6-diphenyl-1,3,5-hexatriene (DPH) molecules. The charged membranes offered the microenvironment where inhibitory effects of [Bmim]Cl on the cellulase activity was relieved. The maximum glucose productivity GP of 10.8 mmol-glucose/(hmol-lipid) was obtained at the reaction time of 48h with the cellulase incorporated in the liposomes ([lipid]=5.0mM) composed of 50mol% POPG and 1,2-dilauroyl-sn-glycero-3-phosohocholine (DLPC) with relatively short and saturated acyl chains. PMID:27318965

  4. Effects of Additives on the Thermostability of Cellulase%添加剂对纤维素酶耐温性的影响

    Institute of Scientific and Technical Information of China (English)

    姚秀清; 孙丽萍; 周丹丹

    2011-01-01

    In order to improve the thermostability of cellulase under higher temperature, the effects of five additives including polyethylene glycol (PEG) , glycerol, xylitol, calcium chloride and xanthan gum on cellulose activities were studied. The results showed that protective effect was found in the presence of polyethylene glycol, glycerol, xylitol and xanthan gum under the condition of 58 ℃ and preserved 24 h. The cellulase activities with the addition of additives were higher than that without additives. Among the four additives, protective effect of xylitol on the cellulase activities was most evident. Relative cellulase activity was increased from 0.59 to 0.94. While inhibiting effect of calcium chloride on the cellulase activities was existed with relative cellulase activity decreased from 0.59 to 0.49.%为提高纤维素酶在较高温度条件下的耐温性,考察5种添加剂(聚乙二醇、甘油、木糖醇、氯化钙、黄原胶)对纤维素酶酶活的影响.结果表明,在58℃、保存24h条件下,聚乙二醇、甘油、木糖醇、黄原胶对纤维素酶酶活有保护效应,较未添加时的相对酶活有所增加.木糖醇的保护效应最为显著,相对酶活由0.59提高到0.94;而氯化钙对酶活有抑制效应,相对酶活由0.59降低为0.49.

  5. The binding of cellulase variants to dislocations: a semi-quantitative analysis based on CLSM (confocal laser scanning microscopy) images.

    Science.gov (United States)

    Hidayat, Budi J; Weisskopf, Carmen; Felby, Claus; Johansen, Katja S; Thygesen, Lisbeth G

    2015-12-01

    Binding of enzymes to the substrate is the first step in enzymatic hydrolysis of lignocellulose, a key process within biorefining. During this process elongated plant cells such as fibers and tracheids have been found to break into segments at irregular cell wall regions known as dislocations or slip planes. Here we study whether cellulases bind to dislocations to a higher extent than to the surrounding cell wall. The binding of fluorescently labelled cellobiohydrolases and endoglucanases to filter paper fibers was investigated using confocal laser scanning microscopy and a ratiometric method was developed to assess and quantify the abundance of the binding of cellulases to dislocations as compared to the surrounding cell wall. Only Humicola insolens EGV was found to have stronger binding preference to dislocations than to the surrounding cell wall, while no difference in binding affinity was seen for any of the other cellulose variants included in the study (H. insolens EGV variants, Trichoderma reesei CBHI, CBHII and EGII). This result favours the hypothesis that fibers break at dislocations during the initial phase of hydrolysis mostly due to mechanical failure rather than as a result of faster degradation at these locations. PMID:26626331

  6. Systems analysis of lactose metabolism in Trichoderma reesei identifies a lactose permease that is essential for cellulase induction.

    Directory of Open Access Journals (Sweden)

    Christa Ivanova

    Full Text Available Trichoderma reesei colonizes predecayed wood in nature and metabolizes cellulose and hemicellulose from the plant biomass. The respective enzymes are industrially produced for application in the biofuel and biorefinery industry. However, these enzymes are also induced in the presence of lactose (1,4-0-ß-d-galactopyranosyl-d-glucose, a waste product from cheese manufacture or whey processing industries. In fact, lactose is the only soluble carbon source that induces these enzymes in T. reesei on an industrial level but the reason for this unique phenomenon is not understood. To answer this question, we used systems analysis of the T. reesei transcriptome during utilization of lactose. We found that the respective CAZome encoded all glycosyl hydrolases necessary for cellulose degradation and particularly for the attack of monocotyledon xyloglucan, from which ß-galactosides could be released that may act as the inducers of T. reesei's cellulases and hemicellulases. In addition, lactose also induces a high number of putative transporters of the major facilitator superfamily. Deletion of fourteen of them identified one gene that is essential for lactose utilization and lactose uptake, and for cellulase induction by lactose (but not sophorose in pregrown mycelia of T. reesei. These data shed new light on the mechanism by which T. reesei metabolizes lactose and offers strategies for its improvement. They also illuminate the key role of ß-D-galactosides in habitat specificity of this fungus.

  7. Optimisation of Cellulase Production by Penicillium funiculosum in a Stirred Tank Bioreactor Using Multivariate Response Surface Analysis

    Directory of Open Access Journals (Sweden)

    Marcelle Lins de Albuquerque de Carvalho

    2014-01-01

    Full Text Available Increasing interest in the production of second-generation ethanol necessitates the low-cost production of enzymes from the cellulolytic complex (endoglucanases, exoglucanases, and β-glucosidases, which act synergistically in cellulose breakdown. The present work aimed to optimise a bioprocess to produce these biocatalysts from the fungus Penicillium funiculosum ATCC11797. A statistical full factorial design (FFD was employed to determine the optimal conditions for cellulase production. The optimal composition of culture media using Avicel (10 g·L−1 as carbon source was determined to include urea (1.2 g·L−1, yeast extract (1.0 g·L−1, KH2PO4 (6.0 g·L−1, and MgSO4·7H2O (1.2 g·L−1. The growth process was performed in batches in a bioreactor. Using a different FFD strategy, the optimised bioreactor operational conditions of an agitation speed of 220 rpm and aeration rate of 0.6 vvm allowed the obtainment of an enzyme pool with activities of 508 U·L−1 for FPase, 9,204 U·L−1 for endoglucanase, and 2,395 U·L−1 for β-glucosidase. The sequential optimisation strategy was effective and afforded increased cellulase production in the order from 3.6 to 9.5 times higher than production using nonoptimised conditions.

  8. Simultaneous cloning and expression of two cellulase genes from Bacillus subtilis newly isolated from Golden Takin (Budorcas taxicolor Bedfordi)

    International Nuclear Information System (INIS)

    A bacterial strain with high cellulase activity was isolated of feces sample of Golden Takin (Budorcas taxicolor Bedfordi). The bacterium was classified and designated Bacillus subtilis LN by morphological and 16SrDNA gene sequence analysis. Two putative cellulase genes, CelL15 and CelL73, were simultaneously cloned from the isolated strain by PCR. The putative gene CelL15 consisted of an open reading frame (ORF) of 1470 nucleotides and encoded a protein of 490 amino acids with a molecular weight of 54 kDa. The CelL73 gene consisted of an open reading frame (ORF) of 741 nucleotides and encoded a protein of 247 amino acids with a molecular weight of 27 kDa. Both genes were purified and cloned into pET-28a for expression in Escherichia coli BL21 (DE3). The ability of E. coli to degrade cellulose was enhanced when the two recombinants were cultured together.

  9. Cellulase and xylanase productions by isolated Amazon Bacillus strains using soybean industrial residue based solid-state cultivation

    Directory of Open Access Journals (Sweden)

    Heck Júlio X.

    2002-01-01

    Full Text Available In Brazil, a large amount of a fibrous residue is generated as result of soybean (Glycine max protein production. This material, which is rich in hemicellulose and cellulose, can be used in solid state cultivations for the production of valuable metabolites and enzymes. In this work, we studied the bioconversion of this residue by bacteria strains isolated from water and soil collected in the Amazon region. Five strains among 87 isolated bacteria selected for their ability to produce either celullases or xylanases were cultivated on the aforementioned residue. From strain BL62, identified as Bacillus subtilis, it was obtained a preparation showing the highest specific cellulase activity, 1.08 UI/mg protein within 24 hours of growth. Concerning xylanase, the isolate BL53, also identified as Bacillus subtilis, showed the highest specific activity for this enzyme, 5.19 UI/mg protein within 72 hours of cultivation. It has also been observed the production of proteases that were associated with the loss of cellulase and xylanase activities. These results indicated that the selected microorganisms, and the cultivation process, have great biotechnological potential.

  10. CYANOBACTERIAL BIOMASS AS N-SUPPLEMENT TO OIL PALM EMPTY FRUIT BUNCH (OPEFB FIBRE FOR IMPROVEMENT OF CELLULASE PRODUCTION BY ASPERGILLUS TERREUS IN SUBMERGED FERMENTATION

    Directory of Open Access Journals (Sweden)

    Mahdi Shahriarinour

    2011-03-01

    Full Text Available The possibility of using dry biomass of a cyanobacterium, Anacystis nidulans, as nitrogen source supplement for improvement of cellulase production by Aspergillus terreus was studied in submerged fermentation using oil palm empty fruit bunch (OPEFB fibre as a carbon source. For comparison, four other nitrogen sources (ammonium sulphate, urea, peptone, and yeast extract were also tested. Growth and cellulase production were greatly enhanced in fermentation using biomass of cyanobacterium as the nitrogen source. The use of cyanobacterial biomass as a nitrogen source also reduced the inhibitory effect of high concentrations of CaCl2 to growth of A. terreus and cellulase production. The addition of 0.3 g L-1 CaCl2 to the medium containing OPEFB fibre and cyanobacterial biomass further enhanced the cellulase production, though growth remained unchanged. The final FPase, CMCase, and β-glucosidase obtained in fermentation using 10 g L-1 OPEFB fibre and 6 g/L cynaobacterial biomass with the addition of 3 mM CaCl2 was 0.97 U mL-1, 14.1 U mL-1, and 10.4 U mL-1, respectively.

  11. Isolation and characteristics of one marine psychrotrophic cellulase-generating bacterium Ar/w/b/75°/10/5 from Chuckchi Sea, Arctic

    Institute of Scientific and Technical Information of China (English)

    曾胤新; 陈波

    2002-01-01

    Microorganisms living in polar zones play an important part as the potential source of organic activity materials with low temperature characteristics in the biotechnological applications. A psychrotrophic bacterium (strain Ar/w/b/75°/10/5), producing cellulase at low temperatures during late-exponential and early-stationary phases of cell growth, was isolated from sea ice-covered surface water in Chuckchi Sea, Arctic. This bacterium, with rod cells, was Gram-negative, slightly halophilic. Colony growing on agar plate was in black. Optimum growth temperature was 15℃. No cell growth was observed at 35℃ or above. Optimum salt concentration for cell growth was between 2 and 3 % of sodium chloride in media. Maximal cellulase activity was detected at a temperature of 35℃ and pH8. Cellulase was irreversibly inactivated when incubated at 55℃ within 30 min. Enzyme can be kept stable at the temperature no higher than 25℃. Of special interest was that this bacterium produced various extracellular enzymes including cellulase, amylase, agar hydrolase and protease, at low or moderate temperature conditions, which is certainly of it potential value for applications.

  12. Rather than by direct acquisition via lateral gene transfer, GHF5 cellulases were passed on from early Pratylenchidae to root-knot and cyst nematodes

    NARCIS (Netherlands)

    Rybarczyk-Mydlowska, K.D.; Maboreke, H.R.; Megen, van H.H.B.; Elsen, van den S.J.J.; Mooijman, P.J.W.; Smant, G.; Bakker, J.; Helder, J.

    2012-01-01

    Background: Plant parasitic nematodes are unusual Metazoans as they are equipped with genes that allow for symbiont-independent degradation of plant cell walls. Among the cell wall-degrading enzymes, glycoside hydrolase family 5 (GHF5) cellulases are relatively well characterized, especially for hig

  13. Microbial β-glucosidases from cow rumen metagenome enhance the saccharification of lignocellulose in combination with commercial cellulase cocktail

    Directory of Open Access Journals (Sweden)

    Del Pozo Mercedes V

    2012-09-01

    Full Text Available Abstract Background A complete saccharification of plant polymers is the critical step in the efficient production of bio-alcohols. Beta-glucosidases acting in the degradation of intermediate gluco-oligosaccharides produced by cellulases limit the yield of the final product. Results In the present work, we have identified and then successfully cloned, expressed, purified and characterised 4 highly active beta-glucosidases from fibre-adherent microbial community from the cow rumen. The enzymes were most active at temperatures 45–55°C and pH 4.0-7.0 and exhibited high affinity and activity towards synthetic substrates such as p-nitrophenyl-beta-D-glucopyranoside (pNPbetaG and pNP-beta-cellobiose, as well as to natural cello-oligosaccharides ranging from cellobiose to cellopentaose. The apparent capability of the most active beta-glucosidase, herein named LAB25g2, was tested for its ability to improve, at low dosage (31.25 units g-1 dry biomass, using pNPbetaG as substrate, the hydrolysis of pre-treated corn stover (dry matter content of 20%; 350 g glucan kg-1 dry biomass in combination with a beta-glucosidase-deficient commercial Trichoderma reseei cellulase cocktail (5 units g-1 dry biomass in the basis of pNPbetaG. LAB25g2 increased the final hydrolysis yield by a factor of 20% (44.5 ± 1.7% vs. 34.5 ± 1.5% in control conditions after 96–120 h as compared to control reactions in its absence or in the presence of other commercial beta-glucosidase preparations. The high stability (half-life higher than 5 days at 50°C and pH 5.2 and 2–38000 fold higher (as compared with reported beta-glucosidases activity towards cello-oligosaccharides may account for its performance in supplementation assays. Conclusions The results suggest that beta-glucosidases from yet uncultured bacteria from animal digestomes may be of a potential interest for biotechnological processes related to the effective bio-ethanol production in combination with low dosage of

  14. Artificial intelligence techniques to optimize the EDC/NHS-mediated immobilization of cellulase on Eudragit L-100.

    Science.gov (United States)

    Zhang, Yu; Xu, Jing-Liang; Yuan, Zhen-Hong; Qi, Wei; Liu, Yun-Yun; He, Min-Chao

    2012-01-01

    Two artificial intelligence techniques, namely artificial neural network (ANN) and genetic algorithm (GA) were combined to be used as a tool for optimizing the covalent immobilization of cellulase on a smart polymer, Eudragit L-100. 1-Ethyl-3-(3-dimethyllaminopropyl) carbodiimide (EDC) concentration, N-hydroxysuccinimide (NHS) concentration and coupling time were taken as independent variables, and immobilization efficiency was taken as the response. The data of the central composite design were used to train ANN by back-propagation algorithm, and the result showed that the trained ANN fitted the data accurately (correlation coefficient R(2) = 0.99). Then a maximum immobilization efficiency of 88.76% was searched by genetic algorithm at a EDC concentration of 0.44%, NHS concentration of 0.37% and a coupling time of 2.22 h, where the experimental value was 87.97 ± 6.45%. The application of ANN based optimization by GA is quite successful. PMID:22942683

  15. Effect of Pre-harvest Treatments on the Cellulase Activity and Quality of Ber Fruit Under Cold Storage Conditions

    OpenAIRE

    Sukhjit Kaur JAWANDHA; Mahajan, B. V. C.; Parmpal Singh GILL

    2009-01-01

    Studies were carried out to find out the effect of various pre-harvest treatments such as CaCl2 (@ 0.5%, 1.0% and 2.0%), Ca(NO3)2 (@0.5%, 1.0% and 2.0%), GA3 (@ 20, 40 and 60 ppm) and Bavistin (@ 0.1%) on the cellulase activity and quality of 'Umran' ber fruits during cold storage. Marked trees were sprayed at colour break stage with the test chemicals. Fruits were packed in CFB boxes and placed in cold storage (3-5 oC and 85 � 90 % RH) for 30 days. The fruits were evaluated after 10, 20 and...

  16. Partial purification and properties of cellulase-free alkaline xylanase produced by Rhizopus stolonifer in solid-state fermentation

    OpenAIRE

    Antonio José Goulart; Eleonora Cano Carmona; Rubens Monti

    2005-01-01

    Rhizopus stolonifer was cultivated in wheat bran to produce a cellulase-free alkaline xylanase. The purified enzyme obtained after molecular exclusion chromatography in Sephacryl S-200 HR showed optimum temperature as 45º C and hydrolysis pHs optima as pH 6.0 and 9.0. Xylanase presented higher Vmax at pH 9.0 (0.87 µmol/mg protein) than at pH 6.0 and minor Km at pH 6.0 (7.42 mg/mL) than at pH 9.0.Rhizopus stolonifer foi cultivado em meio de farelo de trigo para produzir uma xilanase alcalina c...

  17. Artificial Intelligence Techniques to Optimize the EDC/NHS-Mediated Immobilization of Cellulase on Eudragit L-100

    Science.gov (United States)

    Zhang, Yu; Xu, Jing-Liang; Yuan, Zhen-Hong; Qi, Wei; Liu, Yun-Yun; He, Min-Chao

    2012-01-01

    Two artificial intelligence techniques, namely artificial neural network (ANN) and genetic algorithm (GA) were combined to be used as a tool for optimizing the covalent immobilization of cellulase on a smart polymer, Eudragit L-100. 1-Ethyl-3-(3-dimethyllaminopropyl) carbodiimide (EDC) concentration, N-hydroxysuccinimide (NHS) concentration and coupling time were taken as independent variables, and immobilization efficiency was taken as the response. The data of the central composite design were used to train ANN by back-propagation algorithm, and the result showed that the trained ANN fitted the data accurately (correlation coefficient R2 = 0.99). Then a maximum immobilization efficiency of 88.76% was searched by genetic algorithm at a EDC concentration of 0.44%, NHS concentration of 0.37% and a coupling time of 2.22 h, where the experimental value was 87.97 ± 6.45%. The application of ANN based optimization by GA is quite successful. PMID:22942683

  18. Proteomic and Functional Analysis of the Cellulase System Expressed by Postia placenta during Brown Rot of Solid Wood

    Energy Technology Data Exchange (ETDEWEB)

    Ryu, Jae San; Shary, Semarjit; Houtman, Carl J.; Panisko, Ellen A.; Korripally, Premsagar; St John, Franz J.; Crooks, Casey; Siika-aho, Matti; Magnuson, Jon K.; Hammel, Ken

    2011-11-01

    Abstract Brown rot basidiomycetes have an important ecological role in lignocellulose recycling and are notable for their rapid degradation of wood polymers via oxidative and hydrolytic mechanisms. However, most of these fungi apparently lack processive (exo-acting) cellulases, such as cellobiohydrolases, which are generally required for efficient cellulolysis. The recent sequencing of the Postia placenta genome now permits a proteomic approach to this longstanding conundrum. We grew P. placenta on solid aspen wood, extracted proteins from the biodegrading substrate, and analyzed tryptic digests by shotgun liquid chromatography-tandem mass spectrometry. Comparison of the data with the predicted P. placenta proteome revealed the presence of 34 likely glycoside hydrolases, but only four of these-two in glycoside hydrolase family 5, one in family 10, and one in family 12-have sequences that suggested possible activity on cellulose. We expressed these enzymes heterologously and determined that they all exhibited endoglucanase activity on phosphoric acid-swollen cellulose. They also slowly hydrolyzed filter paper, a more crystalline substrate, but the soluble/insoluble reducing sugar ratios they produced classify them as nonprocessive. Computer simulations indicated that these enzymes produced soluble/insoluble ratios on reduced phosphoric acid-swollen cellulose that were higher than expected for random hydrolysis, which suggests that they could possess limited exo activity, but they are at best 10-fold less processive than cellobiohydrolases. It appears likely that P. placenta employs a combination of oxidative mechanisms and endo-acting cellulases to degrade cellulose efficiently in the absence of a significant processive component.

  19. One-step purification and characterization of cellulase-free xylanase produced by alkalophilic Bacillus subtilis ash

    Directory of Open Access Journals (Sweden)

    Ashwani Sanghi

    2010-06-01

    Full Text Available The present study describes the one-step purification and characterization of an extracellular cellulase-free xylanase from a newly isolated alkalophilic and moderately thermophilic strain of Bacillus subtilis ASH. Xylanase was purified to homogeneity by 10.5-fold with ~43% recovery using ion-exchange chromatography through CM-Sephadex C-50. The purified enzyme revealed a single band on SDS-PAGE gel with a molecular mass of 23 kDa. It showed an optimum pH at 7.0 and was stable over the pH range 6.0-9.0. The optimum temperature for enzyme activity was 55 ºC. The purified xylanase did not lose any activity up to 45 ºC, however, it retained 80% and 51% of its activity after pre-incubation at 55 ºC and 60 ºC, respectively. The enzyme obeyed Michaelis-Menton kinetics towards birch wood xylan with apparent Km 3.33 mg/ml and Vmax 100 IU/ml. The enzyme was strongly inhibited by Hg2+ and Cu2+ while enhanced by Co2+ and Mn2+. The purified enzyme could be stored at 4 ºC for six weeks without any loss of catalytic activity. The faster and economical purification of the cellulase-free xylanase from B. subtilis ASH by one-step procedure together with its appreciable stability at high temperature and alkaline pH makes it potentially effective for industrial applications.

  20. Improved activity of a thermophilic cellulase, Cel5A, from Thermotoga maritima on ionic liquid pretreated switchgrass.

    Directory of Open Access Journals (Sweden)

    Zhiwei Chen

    Full Text Available Ionic liquid pretreatment of biomass has been shown to greatly reduce the recalcitrance of lignocellulosic biomass, resulting in improved sugar yields after enzymatic saccharification. However, even under these improved saccharification conditions the cost of enzymes still represents a significant proportion of the total cost of producing sugars and ultimately fuels from lignocellulosic biomass. Much of the high cost of enzymes is due to the low catalytic efficiency and stability of lignocellulolytic enzymes, especially cellulases, under conditions that include high temperatures and the presence of residual pretreatment chemicals, such as acids, organic solvents, bases, or ionic liquids. Improving the efficiency of the saccharification process on ionic liquid pretreated biomass will facilitate reduced enzyme loading and cost. Thermophilic cellulases have been shown to be stable and active in ionic liquids but their activity is typically at lower levels. Cel5A_Tma, a thermophilic endoglucanase from Thermotoga maritima, is highly active on cellulosic substrates and is stable in ionic liquid environments. Here, our motivation was to engineer mutants of Cel5A_Tma with higher activity on 1-ethyl-3-methylimidazolium acetate ([C2mim][OAc] pretreated biomass. We developed a robotic platform to screen a random mutagenesis library of Cel5A_Tma. Twelve mutants with 25-42% improvement in specific activity on carboxymethyl cellulose and up to 30% improvement on ionic-liquid pretreated switchgrass were successfully isolated and characterized from a library of twenty thousand variants. Interestingly, most of the mutations in the improved variants are located distally to the active site on the protein surface and are not directly involved with substrate binding.

  1. Efficient cellulase production by Trichoderma reesei in continuous cultivation on lactose medium with a computer-controlled feeding strategy.

    Science.gov (United States)

    Bailey, M J; Tähtiharju, J

    2003-08-01

    A low-foaming hydrophobin II deletant of the Trichoderma reesei strain Rut-C30 was used for production of cellulases by continuous cultivation on lactose medium in a laboratory fermenter. The control paradigm of the addition of new medium to the continuous process was based on the growth dynamics of the fungus. A decrease in the rate of base addition to the cultivation for pH-minimum control was used as an indicator of imminent exhaustion of carbon source for growth and enzyme induction. When the amount of base added per 5 min computation cycle decreased below a given value, new medium was added to the fermenter. When base addition for pH control thereafter increased above the criterion value, due to increased growth, the medium feed was discontinued or decreased. The medium feeding protocol employed was successful in locking the fungus in the stage of imminent, but not actual, exhaustion of carbon source. According to the results of a batch cultivation of the same strain on the same medium, this is the phase of maximal enzyme productivity. The medium addition protocol used in this work resulted in a very stable continuous process, in which cellulase productivity was maintained for several hundred hours at the maximum level observed in a batch cultivation for only about 10 h. Despite a major technical disturbance after about 420 h, the process was restored to stability. When the cultivation was terminated after 650 h, the level of enzyme production was still maximal, with no signs of instability of the process.

  2. A mitogen-activated protein kinase Tmk3 participates in high osmolarity resistance, cell wall integrity maintenance and cellulase production regulation in Trichoderma reesei.

    Directory of Open Access Journals (Sweden)

    Mingyu Wang

    Full Text Available The mitogen-activated protein kinase (MAPK pathways are important signal transduction pathways conserved in essentially all eukaryotes, but haven't been subjected to functional studies in the most important cellulase-producing filamentous fungus Trichoderma reesei. Previous reports suggested the presence of three MAPKs in T. reesei: Tmk1, Tmk2, and Tmk3. By exploring the phenotypic features of T. reesei Δtmk3, we first showed elevated NaCl sensitivity and repressed transcription of genes involved in glycerol/trehalose biosynthesis under higher osmolarity, suggesting Tmk3 participates in high osmolarity resistance via derepression of genes involved in osmotic stabilizer biosynthesis. We also showed significant downregulation of genes encoding chitin synthases and a β-1,3-glucan synthase, decreased chitin content, 'budded' hyphal appearance typical to cell wall defective strains, and increased sensitivity to calcofluor white/Congo red in the tmk3 deficient strain, suggesting Tmk3 is involved in cell wall integrity maintenance in T. reesei. We further observed the decrease of cellulase transcription and production in T. reesei Δtmk3 during submerged cultivation, as well as the presence of MAPK phosphorylation sites on known transcription factors involved in cellulase regulation, suggesting Tmk3 is also involved in the regulation of cellulase production. Finally, the expression of cell wall integrity related genes, the expression of cellulase coding genes, cellulase production and biomass accumulation were compared between T. reesei Δtmk3 grown in solid state media and submerged media, showing a strong restoration effect in solid state media from defects resulted from tmk3 deletion. These results showed novel physiological processes that fungal Hog1-type MAPKs are involved in, and present the first experimental investigation of MAPK signaling pathways in T. reesei. Our observations on the restoration effect during solid state cultivation suggest

  3. Fabrication of graphene oxide decorated with Fe{sub 3}O{sub 4}@SiO{sub 2} for immobilization of cellulase

    Energy Technology Data Exchange (ETDEWEB)

    Li, Yue; Wang, Xiang-Yu; Jiang, Xiao-Ping; Ye, Jing-Jing; Zhang, Ye-Wang, E-mail: zhangyewang@ujs.edu.cn [Jiangsu University, School of Pharmacy (China); Zhang, Xiao-Yun, E-mail: zhangxiaoyungu@126.com [Jiangsu University, School of Food and Biological Engineering (China)

    2015-01-15

    Fe{sub 3}O{sub 4}@SiO{sub 2}–graphene oxide (GO) composites were successfully fabricated by chemical binding of functional Fe{sub 3}O{sub 4}@SiO{sub 2} and GO and applied to immobilization of cellulase via covalent attachment. The prepared composites were further characterized by transmission electron microscopy and Fourier transform infrared spectroscopy. Fe{sub 3}O{sub 4} nanoparticles (NPs) were monodisperse spheres with a mean diameter of 17 ± 0.2 nm. The thickness of SiO{sub 2} layer was calculated as being 6.5 ± 0.2 nm. The size of Fe{sub 3}O{sub 4}@SiO{sub 2} NPs was 24 ± 0.3 nm, similar to that of Fe{sub 3}O{sub 4}@SiO{sub 2}–NH{sub 2}. Fe{sub 3}O{sub 4}@SiO{sub 2}–GO composites were synthesized by linking of Fe{sub 3}O{sub 4}@SiO{sub 2}–NH{sub 2} NPs to GO with the catalysis of EDC and NHS. The prepared composites were used for immobilization of cellulase. A high immobilization yield and efficiency of above 90 % were obtained after the optimization. The half-life of immobilized cellulase (722 min) was 3.34-fold higher than that of free enzyme (216 min) at 50 °C. Compared with the free cellulase, the optimal temperature of the immobilized enzyme was not changed; but the optimal pH was shifted from 5.0 to 4.0, and the thermal stability was enhanced. The immobilized cellulase could be easily separated and reused under magnetic field. These results strongly indicate that the cellulase immobilized onto the Fe{sub 3}O{sub 4}@SiO{sub 2}–GO composite has potential applications in the production of bioethanol.

  4. Effect of lignin structure and surfactant on cellulase adsorption by lignin%木质素结构以及表面活性剂对木质素吸附纤维素酶的影响

    Institute of Scientific and Technical Information of China (English)

    姚兰; 赵建; 谢益民; 杨海涛; 曲音波

    2012-01-01

    As substitute of petrol, bioethanol production from lignocelluloses is one of hot point of research in recent years. A main obstacle of bioethanol production in commercial scale is its cost.'One of the most important methods to solve the key problem is raising yield of cellulose hydrolysis by enzyme. However, content and type of lignin in lignocellulosic substrates has a significant effect on the hydrolysis process, as lignin has double actions: as a physical barrier that decreases accessibility of cellulase to cellulose, and as an attractant that adsorbs cellulases i. e. forming non-productive binding. In this paper, structure and adsorption for cellulase of three kinds of ligin, milled wood lignin, Klasson lignin and alkaline lignin were studied, and various surfactants were screened and used for improving enzymatic hydrolysis of lignocelluloses because they have a great impact on cellulase adsorption on lignin. The results showed that there are different structure characteristics for the three kinds of lignin: the highest ability of adsorption for cellulase is Klasson lignin, and then milled wood lignin, the lowest alkaline lignin. The surfactant that can significantly increase enzymatic hydrolysis yield of lignocelluloses is PEG-8000, because it can make cellulase desorption from lignin and reactivation for hydrolysis of cellulose.

  5. Effect of physical and chemical properties of oil palm empty fruit bunch, decanter cake and sago pith residue on cellulases production by Trichoderma asperellum UPM1 and Aspergillus fumigatus UPM2.

    Science.gov (United States)

    Zanirun, Zuraidah; Bahrin, Ezyana Kamal; Lai-Yee, Phang; Hassan, Mohd Ali; Abd-Aziz, Suraini

    2014-01-01

    The effect of cultivation condition of two locally isolated ascomycetes strains namely Trichoderma asperellum UPM1 and Aspergillus fumigatus UPM2 were compared in submerged and solid state fermentation. Physical evaluation on water absorption index, solubility index and chemical properties of lignin, hemicellulose and cellulose content as well as the cellulose structure on crystallinity and amorphous region of treated oil palm empty fruit bunch (OPEFB) (resulted in partial removal of lignin), sago pith residues (SPR) and oil palm decanter cake towards cellulases production were determined. Submerged fermentation shows significant cellulases production for both strains in all types of substrates. Crystallinity of cellulose and its chemical composition mainly holocellulose components was found to significantly affect the total cellulase synthesis in submerged fermentation as the higher crystallinity index, and holocellulose composition will increase cellulase production. Treated OPEFB apparently induced the total cellulases from T. asperellum UPM1 and A. fumigatus UPM2 with 0.66 U/mg FPase, 53.79 U/mg CMCase, 0.92 U/mg β-glucosidase and 0.67 U/mg FPase, 47.56 U/mg and 0.14 U/mg β-glucosidase, respectively. Physical properties of water absorption and solubility for OPEFB and SPR also had shown significant correlation on the cellulases production. PMID:24085387

  6. Cellulose hydrolysis by Trichoderma reesei cellulases: studies on adsorption, sugar production and synergism of cellobiohydrolase I,II and endoglucanase II

    Energy Technology Data Exchange (ETDEWEB)

    Medve, J.

    1997-02-01

    Three major cellulases have been purified by ion-exchange chromatography in an FPLC system. Microcrystalline cellulose (Avicel) was hydrolyzed by the single enzymes and by equimolar mixtures of CBH I-CBH II and CBH I-EG II. Enzyme adsorption was followed indirectly by selectively quantifying the enzymes in the supernatant by ion-exchange chromatography in an FPLC system. The (synergistic) production of small, soluble sugars (glucose, cellobiose and cellotriose) by the enzymes was followed by HPLC. 76 refs

  7. Enhanced alkaline cellulases production by the thermohalophilic Aspergillus terreus AUMC 10138 mutated by physical and chemical mutagens using corn stover as substrate.

    Science.gov (United States)

    Isaac, George Saad; Abu-Tahon, Medhat Ahmed

    2015-01-01

    A thermohalophilic fungus, Aspergillus terreus AUMC 10138, isolated from the Wadi El-Natrun soda lakes in northern Egypt was exposed successively to gamma and UV-radiation (physical mutagens) and ethyl methan-sulfonate (EMS; chemical mutagen) to enhance alkaline cellulase production under solid state fermentation (SSF) conditions. The effects of different carbon sources, initial moisture, incubation temperature, initial pH, incubation period, inoculum levels and different concentrations of NaCl on production of alkaline filter paper activity (FPase), carboxymethyl cellulase (CMCase) and β-glucosidase by the wild-type and mutant strains of A. terreus were evaluated under SSF. The optimum conditions for maximum production of FPase, CMCase and β-glucosidase were found to be the corn stover: moisture ratio of 1:3(w/v), temperature 45 °C, pH range, 9.0-11.0, and fermentation for 4, 4 and 7 day, respectively. Inoculum levels of 30% for β-glucosidase and 40% for FPase, CMCase gave the higher cellulase production by the wild-type and mutant strains, respectively. Higher production of all three enzymes was obtained at a 5% NaCl. Under the optimized conditions, the mutant strain A. terreus M-17 produced FPase (729 U/g), CMCase (1,783 U/g), and β-glucosidase (342 U/g), which is, 1.85, 1.97 and 2.31-fold higher than the wild-type strain. Our results confirmed that mutant strain M-17 could be a promising alkaline cellulase enzyme producer employing lignocellulosics especially corn stover.

  8. Efficacy of xylanase purified from Aspergillus niger DFR-5 alone and in combination with pectinase and cellulase to improve yield and clarity of pineapple juice.

    Science.gov (United States)

    Pal, Ajay; Khanum, Farhath

    2011-10-01

    Pineapple is one of the fruits having xylan rich hemicellulose content more than pectin. Therefore, the efficacy of absolutely purified xylanase from A. niger DFR-5 alone and in combination with pectinase and cellulase on juice yield and clarity was studied. Xylanase provided maximum yield (71.3%) and clarity (64.7%) of juice in comparison to control responses (61.8% yield and 57.8% clarity). When used together, a synergistic effect of xylanase, pectinase and cellulase on process responses was observed indicating the necessity of a cock-tail of hydrolytic enzymes for complete cell wall degradation. Overall, an increase in juice yield by 52.9% was observed. The process was numerically optimized with the constraint of 'minimum' pectinase and cellulase and 'maximum' xylanase and incubation time for 'maximum' juice yield and clarity. The closeness of observed response (90.2% yield and 80.9% clarity) to the predicted one (89.6% yield and 80.3% clarity) indicated the validity of developed model. PMID:23572788

  9. Isolation of Bacillus sp. strains capable of decomposing alkali lignin and their application in combination with lactic acid bacteria for enhancing cellulase performance.

    Science.gov (United States)

    Chang, Young-Cheol; Choi, Dubok; Takamizawa, Kazuhiro; Kikuchi, Shintaro

    2014-01-01

    Effective biological pretreatment method for enhancing cellulase performance was investigated. Two alkali lignin-degrading bacteria were isolated from forest soils in Japan and named CS-1 and CS-2. 16S rDNA sequence analysis indicated that CS-1 and CS-2 were Bacillus sp. Strains CS-1 and CS-2 displayed alkali lignin degradation capability. With initial concentrations of 0.05-2.0 g L(-1), at least 61% alkali lignin could be degraded within 48 h. High laccase activities were observed in crude enzyme extracts from the isolated strains. This result indicated that alkali lignin degradation was correlated with laccase activities. Judging from the net yields of sugars after enzymatic hydrolysis, the most effective pretreatment method for enhancing cellulase performance was a two-step processing procedure (pretreatment using Bacillus sp. CS-1 followed by lactic acid bacteria) at 68.6%. These results suggest that the two-step pretreatment procedure is effective at accelerating cellulase performance. PMID:24316485

  10. CELLULASES PRODUCTION UNDER SOLID STATE FERMENTATION USING AGRO WASTE AS A SUBSTRATE AND ITS APPLICATION IN SACCHARIFICATION BY TRAMETES HIRSUTA NCIM

    Directory of Open Access Journals (Sweden)

    Bhaumik R. Dave

    2014-12-01

    Full Text Available Food and energy crisis are the biggest constraint all over the world which has focused lights on need of utilizing renewable resources to meet the future demand. A promising strategy is efficient utilization of lignocellulosic waste and fermentation of the resulting sugars for production of desired metabolites or biofuel. Production of all the cellulase enzymes on wheat bran and different parameters regulating it like pH, moisture ratio (substrate: liquid, temperature and inoculum size has been optimized which found to be 4.5, 1:3, 30°C and 108 spores respectively. Salient feature of partially purified enzyme with stability in the range of 30-50°C under acidic pH range was found to be prominent for industrial applications, moreover in this study, Trametes hirsuta, an efficient cellulase producer, was observed to be an effective species for saccharification of wheat straw to enhance the sugar yield. Enzymatic hydrolysis of wheat straw with 15 FPU of cellulase from the species showed 73% yield in 20 hrs. It may prove to be a suitable choice for the industrial saccharification of lignocellulosic biomasses.

  11. Inexpensive, rapid procedure for bulk purification of cellulase-free. beta. -1,4-D-xylanase of high specific activity

    Energy Technology Data Exchange (ETDEWEB)

    Tan, L.U.L.; Yu, E.K.C.; Louis-Seize, G.W.; Saddler, J.N.

    1987-01-01

    A process has been developed for the bulk purification of cellulase-free ..beta..-14-D-xylanase from the fungus Tirchoderma harzianum E58. The process involved the primary step of ultrafiltering the culture filtrate via a 10,000-molecular-weight cut-off membrane to separate the cellulase (retentate) and xylanase (permeate) fractions. The cellulase component was concentrated by 40- to 60-fold, resulting in an enzyme complex that could effectively hydrolyze high concentrations of cellulose and xylan to glucose and xylose. The xylanase was concentrated and solvent exchanged by adsorption to a cationic exchanger, SP-ZetaPrep 250, followed by elution with a pH change in the buffer to give a purified and concentrated xylanase complex dissolved in a low-salt buffer. The resultant xylanase system was pure by the criteria of sodium dodecyl sulfate polyacrylamide electrophoresis, had a very high specific activity of 2400 IU/mg protein, was virtually free of filter paper activity, and had a ratio of contaminating filter paper activity of 2 x 10/sup -6/. Approximately 3.3 g protein, which contained in excess of 7 x 10/sup 6/ IU xylanase activity was obtained from 17 L original culture filtrate. The process scheme was designed to facilitate scale-up to an industrial level of production.

  12. Avocado cellulase: nucleotide sequence of a putative full-length cDNA clone and evidence for a small gene family.

    Science.gov (United States)

    Tucker, M L; Durbin, M L; Clegg, M T; Lewis, L N

    1987-05-01

    A cDNA library was prepared from ripe avocado fruit (Persea americana Mill. cv. Hass) and screened for clones hybridizing to a 600 bp cDNA clone (pAV5) coding for avocado fruit cellulase. This screening led to the isolation of a clone (pAV363) containing a 2021 nucleotide transcribed sequence and an approximately 150 nucleotide poly(A) tail. Hybridization of pAV363 to a northern blot shows that the length of the homologous message is approximately 2.2 kb. The nucleotide sequence of this putative full-length mRNA clone contains an open reading frame of 1482 nucleotides which codes for a polypeptide of 54.1 kD. The deduced amino acid composition compares favorably with the amino acid composition of native avocado cellulase determined by amino acid analysis. Southern blot analysis of Hind III and Eco RI endonuclease digested genomic DNA indicates a small family of cellulase genes.

  13. Novel Cellulase Profile of Trichoderma reesei Strains Constructed by cbh1 Gene Replacement with eg3 Gene Expression Cassette

    Institute of Scientific and Technical Information of China (English)

    Tian-Hong WANG; Ti LIU; Zhi-Hong WU; Shi-Li LIU; Yi LU; Yin-Bo QU

    2004-01-01

    To construct strains of the filamentous fungus Trichoderma reesei with low cellobiohydrolases while high endoglucanase activity, the Pcbh1-eg3-Tcbh1 cassette was constructed and the coding sequence of the cellobiohydrolase I (CBHI) gene was replaced with the coding sequence of the eg3 gene by homologous recombination. Disruption of the cbh1 gene was confirmed by PCR, Southern dot blot and Western hybridization analysis in two transforments denoted as L 13 and L29. The filter paper-hydrolyzing activity of strain L29 was 60% of the parent strain Rut C30, and the CMCase activity was increased by 33%. This relatively modest increase suggested that the eg3 cDNA under the control of the cbh1 promoter was not efficiently transcribed as the wild type cbhl gene. However our results confirmed that homologous recombination could be used to construct strains of the filamentous fungus Trichoderma reesei with novel cellulase profile. Such strains are of interest from the basic science perspective and also have potential industrial applications.

  14. Production, purification, and characterization of a cellulase-free thermostable endo-xylanase from Thermoanaerobacterium thermosaccharolyticum DSM 571.

    Science.gov (United States)

    Li, Xun; Shi, Hao; Ding, Huaihai; Zhang, Yu; Wang, Fei

    2014-12-01

    This is the first report describing the cloning, expression, and characterization of a putative thermostable, cellulase-free xylanase (XYN) from the thermophilic bacterium Thermoanaerobacterium thermosaccharolyticum DSM 571. The temperature and pH values for optimal enzyme activity of XYN were found to be 65 °C and pH 6.5, respectively. The XYN activity was apparently enhanced by Co(2+), Mn(2+), and Tween 60 and significantly inactivated by Al(3+), Cu(2+), Zn(2+), and SDS. The K m and V max values of XYN for the hydrolysis of beechwood xylan were 2.1 mg/ml and 222.1 U/mg, respectively. The k cat values of XYN for beechwood xylan at the optimal temperature and pH values were 481.4 s(-1). XYN represents an attractive candidate for use in the large-scale production of xylooligosaccharides (XOs) from forest residues because it is an endo-xylanase capable of degrading xylan. PMID:25261357

  15. ENHANCED PRODUCTION OF CELLULASE-FREE XYLANASE BY ALKALOPHILIC BACILLUS SUBTILIS ASH AND ITS APPLICATION IN BIOBLEACHING OF KRAFT PULP

    Directory of Open Access Journals (Sweden)

    Ashwani Sanghi

    2009-08-01

    Full Text Available This paper reports high level production of a cellulase-free xylanase using wheat bran, a cost-effective substrate, under submerged fermentation by alkalophilic Bacillus subtilis ASH. Production of xylanase was observed even at alkaline pH up to 11.0 and temperature 60 °C, although the highest enzyme titer was recorded at neutral pH and 37 °C. The enzyme production under optimized fermentation was 1.5-fold greater than under unoptimized conditions. Pre-treatment of unbleached pulp of 10% consistency with crude xylanase (6 IU/g o.d. pulp at 60 ºC for 2 h increased the final brightness by 4.9%. The enzyme treatment reduced the chlorine consumption by 28.6% with the same brightness as in the control. A reduction in kappa number and increase in viscosity was observed after enzyme pre-treatment. Scanning electron microscopy revealed loosening and swelling of pulp fibers. The strength properties viz. grammage, fiber thickness, beating degree, tensile index, breaking length, tear index and double fold of the treated pulp were improved as compared to the control pulp. This study reveals the potential of B. subtilis ASH xylanase as a biobleaching agent for the paper and pulp industry.

  16. Artificial Intelligence Techniques to Optimize the EDC/NHS-Mediated Immobilization of Cellulase on Eudragit L-100

    Directory of Open Access Journals (Sweden)

    Min-Chao He

    2012-06-01

    Full Text Available Two artificial intelligence techniques, namely artificial neural network (ANN and genetic algorithm (GA were combined to be used as a tool for optimizing the covalent immobilization of cellulase on a smart polymer, Eudragit L-100. 1-Ethyl-3-(3-dimethyllaminopropyl carbodiimide (EDC concentration, N-hydroxysuccinimide (NHS concentration and coupling time were taken as independent variables, and immobilization efficiency was taken as the response. The data of the central composite design were used to train ANN by back-propagation algorithm, and the result showed that the trained ANN fitted the data accurately (correlation coefficient R2 = 0.99. Then a maximum immobilization efficiency of 88.76% was searched by genetic algorithm at a EDC concentration of 0.44%, NHS concentration of 0.37% and a coupling time of 2.22 h, where the experimental value was 87.97 ± 6.45%. The application of ANN based optimization by GA is quite successful.

  17. Identification of a laccase from Ganoderma lucidum CBS 229.93 having potential for enhancing cellulase catalyzed lignocellulose degradation.

    Science.gov (United States)

    Sitarz, Anna K; Mikkelsen, Jørn D; Højrup, Peter; Meyer, Anne S

    2013-12-10

    Based on a differential pre-screening of 44 white-rot fungi on a lignocellulose-supplemented minimal medium, four basidiomycetes were selected for further study: Ganoderma lucidum, Polyporus brumalis, Polyporus ciliatus and Trametes versicolor. Only G. lucidum was able to grow vividly on malt extract or minimal media supplemented with alkali lignin. When grown on malt extract or minimal medium supplemented with lignocellulose (sugar cane bagasse), the crude G. lucidum protein extract exhibited high laccase activity, ∼3U/mL toward syringaldazine. This activity was 13-17 fold higher than the corresponding activities of the crude protein extracts of P. brumalis, P. ciliatus and T. versicolor. Native PAGE electrophoresis of the crude G. lucidum extract confirmed the presence of an active laccase. The G. lucidum laccase had a molecular weight of ∼62.5kDa, and a Km value of 0.107mM (determined on ABTS). A partial amino acid sequence analysis of four short de novo sequenced peptides, defined after trypsin digest analysis using MALDI-TOF MS/MS analysis, revealed 64-100% homology to sequences in related laccases in the UniProt database, but also indicated that certain sequence stretches had low homology. Addition of the laccase-rich G. lucidum broth to lignocellulosic biomass (pretreated sugar cane bagasse) together with a state-of-the-art cellulase enzyme preparation (Cellic™CTec1) produced significantly increased cellulolytic yields, which were also better than those obtained with a T. versicolor laccase addition, indicating that the laccase from G. lucidum has unique properties that may be momentous in lignocellulosic biomass conversion.

  18. Methane production of two roughage and total mixed ration as influenced by cellulase and xylanase enzyme addition

    Directory of Open Access Journals (Sweden)

    Belete Shenkute Gemeda

    2015-02-01

    Full Text Available In recent decades supplementation of animal feeds with exogenous fibrolytic enzymes has substantially improved digestibility and animal performance. However, information related to associated methane production is limited and inconsistent. This study evaluated the effect of cellulase and xylanase enzymes on in vitro methane production of Eragrostis curvula hay, maize (Zea mays stover and a total mixed ration (TMR at seven levels of the two enzymes. Feed samples were incubated for 2, 12, 24 and 48 h in an in vitro batch culture with buffer and rumen fluid, and fibrolytic enzymes. Gas production was measured using a pressure transducer connected to a data tracker, while methane gas was analysed using a gas chromatograph which was calibrated with standard CH4 and CO2. Increases in the level of enzyme application resulted in increases in gas volume, total volatile fatty acid (VFA production, dry matter (DM disappearance and associated increases in methane production. The linear increase in percentage and volume of methane production in tandem with increases in level of enzyme application might be due to increased fermentation, and organic matter degradability that resulted in a shift in VFA production towards acetate. Considering the efficiency of DM and neutral detergent fiber degradation and production of associated VFA with levels of enzymes, the use of 1 mg g−1 DM of enzyme can be a good option for the feeds tested. However, they cannot decrease methane production. It will be very important to consider other hydrogen sinks that can capture directly extra H+ produced by the addition of enzyme so that their supplementation could be very efficient and environmentally sound.

  19. Isolation and Characterization of Thermophilic Cellulase-Producing Bacteria from Empty Fruit Bunches-Palm Oil Mill Effluent Compost

    Directory of Open Access Journals (Sweden)

    Azhari S. Baharuddin

    2010-01-01

    Full Text Available Problems statement: Lack of information on locally isolated cellulase-producing bacterium in thermophilic compost using a mixture of Empty Fruit Bunch (EFB and Palm Oil Mill Effluent (POME as composting materials. Approach: The isolation of microbes from compost heap was conducted at day 7 of composting process where the mixture of composting materials consisted of 45.8% cellulose, 17.1% hemicellulose and 28.3% lignin content. The temperature, pH and moisture content of the composting pile at day 7 treatment were 58.3, 8.1 and 65.5°C, respectively. The morphological analysis of the isolated microbes was conducted using Scanning Electron Microscope (SEM and Gram stain method. The congo red test was conducted in order to detect 1% CMC agar degradation activities. Total genomic DNAs were extracted from approximately 1.0 g of mixed compost and amplified by using PCR primers. The PCR product was sequent to identify the nearest relatives of 16S rRNA genes. The localization of bacteria chromosomes was determined by Fluorescence In Situ Hybridization (FISH analysis. Results: Single isolated bacteria species was successfully isolated from Empty Fruit Bunch (EFB-Palm Oil Mill Effluent (POME compost at thermophilic stage. Restriction fragment length polymorphism profiles of the DNAs coding for the 16S rRNAs with the phylogenetic analysis showed that the isolated bacteria from EFB-POME thermophilic compost gave the highest homology (99% with similarity to Geobacillus pallidus. The strain was spore forming bacteria and able to grow at 60°C with pH 7. Conclusion: Thermophilic bacteria strain, Geobacillus pallidus was successfully isolated from Empty Fruit Bunch (EFB and Palm Oil Mil Effluent (POME compost and characterized.

  20. Cellulose accessibility limits the effectiveness of minimum cellulase loading on the efficient hydrolysis of pretreated lignocellulosic substrates

    Directory of Open Access Journals (Sweden)

    Saddler Jack N

    2011-02-01

    Full Text Available Abstract A range of lignocellulosic feedstocks (including agricultural, softwood and hardwood substrates were pretreated with either sulfur dioxide-catalyzed steam or an ethanol organosolv procedure to try to establish a reliable assessment of the factors governing the minimum protein loading that could be used to achieve efficient hydrolysis. A statistical design approach was first used to define what might constitute the minimum protein loading (cellulases and β-glucosidase that could be used to achieve efficient saccharification (defined as at least 70% glucan conversion of the pretreated substrates after 72 hours of hydrolysis. The likely substrate factors that limit cellulose availability/accessibility were assessed, and then compared with the optimized minimum amounts of protein used to obtain effective hydrolysis. The optimized minimum protein loadings to achieve efficient hydrolysis of seven pretreated substrates ranged between 18 and 63 mg protein per gram of glucan. Within the similarly pretreated group of lignocellulosic feedstocks, the agricultural residues (corn stover and corn fiber required significantly lower protein loadings to achieve efficient hydrolysis than did the pretreated woody biomass (poplar, douglas fir and lodgepole pine. Regardless of the substantial differences in the source, structure and chemical composition of the feedstocks, and the difference in the pretreatment technology used, the protein loading required to achieve efficient hydrolysis of lignocellulosic substrates was strongly dependent on the accessibility of the cellulosic component of each of the substrates. We found that cellulose-rich substrates with highly accessible cellulose, as assessed by the Simons' stain method, required a lower protein loading per gram of glucan to obtain efficient hydrolysis compared with substrates containing less accessible cellulose. These results suggest that the rate-limiting step during hydrolysis is not the catalytic

  1. Optimization and characterization of alkaline protease and carboxymethyl-cellulase produced by Bacillus pumillus grown on Ficus nitida wastes.

    Science.gov (United States)

    Gomaa, Eman Zakaria

    2013-01-01

    The potentiality of 23 bacterial isolates to produce alkaline protease and carboxymethyl-cellulase (CMCase) on Ficus nitida wastes was investigated. Bacillus pumillus ATCC7061 was selected as the most potent bacterial strain for the production of both enzymes. It was found that the optimum production of protease and CMCase were recorded at 30 °C, 5% Ficus nitida leaves and incubation period of 72 h. The best nitrogen sources for protease and CMCase production were yeast extract and casein, respectively. Also maximum protease and CMCase production were reported at pH 9 and pH 10, respectively. The enzymes possessed a good stability over a pH range of 8-10, expressed their maximum activities at pH10 and temperature range of 30-50 °C, expressed their maximum activities at 50 °C. Ions of Hg(2+), Fe2+ and Ag(+) showed a stimulatory effect on protease activity and ions of Fe(2+), Mg(2+), Ca(2+), Cu(2+) and Ag(+) caused enhancement of CMCase activity. The enzymes were stable not only towards the nonionic surfactants like Triton X-100 and Tween 80 but also the strong anionic surfactant, SDS. Moreover, the enzymes were not significantly inhibited by EDTA or cystein. Concerning biotechnological applications, the enzymes retained (51-97%) of their initial activities upon incubation in the presence of commercials detergents for 1 h. The potential use of the produced enzymes in the degradation of human hair and cotton fabric samples were also assessed. PMID:24294252

  2. Enzymatic saccharification of sugar cane bagasse by continuous xylanase and cellulase production from cellulomonas flavigena PR-22.

    Science.gov (United States)

    Rojas-Rejón, Óscar A; Poggi-Varaldo, Héctor M; Ramos-Valdivia, Ana C; Ponce-Noyola, Teresa; Cristiani-Urbina, Eliseo; Martínez, Alfredo; de la Torre, Mayra

    2016-03-01

    Cellulase (CMCase) and xylanase enzyme production and saccharification of sugar cane bagasse were coupled into two stages and named enzyme production and sugar cane bagasse saccharification. The performance of Cellulomonas flavigena (Cf) PR-22 cultured in a bubble column reactor (BCR) was compared to that in a stirred tank reactor (STR). Cells cultured in the BCR presented higher yields and productivity of both CMCase and xylanase activities than those grown in the STR configuration. A continuous culture with Cf PR-22 was run in the BCR using 1% alkali-pretreated sugar cane bagasse and mineral media, at dilution rates ranging from 0.04 to 0.22 1/h. The highest enzymatic productivity values were found at 0.08 1/h with 1846.4 ± 126.4 and 101.6 ± 5.6 U/L·h for xylanase and CMCase, respectively. Effluent from the BCR in steady state was transferred to an enzymatic reactor operated in fed-batch mode with an initial load of 75 g of pretreated sugar cane bagasse; saccharification was then performed in an STR at 55°C and 300 rpm for 90 h. The constant addition of fresh enzyme as well as the increase in time of contact with the substrate increased the total soluble sugar concentration 83% compared to the value obtained in a batch enzymatic reactor. This advantageous strategy may be used for industrial enzyme pretreatment and saccharification of lignocellulosic wastes to be used in bioethanol and chemicals production from lignocellulose. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:321-326, 2016. PMID:26701152

  3. Optimization and characterization of alkaline protease and carboxymethyl-cellulase produced by Bacillus pumillus grown on Ficus nitida wastes

    Directory of Open Access Journals (Sweden)

    Eman Zakaria Gomaa

    2013-01-01

    Full Text Available The potentiality of 23 bacterial isolates to produce alkaline protease and carboxymethyl-cellulase (CMCase on Ficus nitida wastes was investigated. Bacillus pumillus ATCC7061 was selected as the most potent bacterial strain for the production of both enzymes. It was found that the optimum production of protease and CMCase were recorded at 30 °C, 5% Ficus nitida leaves and incubation period of 72 h. The best nitrogen sources for protease and CMCase production were yeast extract and casein, respectively. Also maximum protease and CMCase production were reported at pH 9 and pH 10, respectively. The enzymes possessed a good stability over a pH range of 8-10, expressed their maximum activities at pH10 and temperature range of 30-50 °C, expressed their maximum activities at 50 °C. Ions of Hg2+, Fe2+ and Ag+ showed a stimulatory effect on protease activity and ions of Fe2+, Mg2+, Ca2+, Cu2+ and Ag+ caused enhancement of CMCase activity. The enzymes were stable not only towards the nonionic surfactants like Triton X-100 and Tween 80 but also the strong anionic surfactant, SDS. Moreover, the enzymes were not significantly inhibited by EDTA or cystein. Concerning biotechnological applications, the enzymes retained (51-97% of their initial activities upon incubation in the presence of commercials detergents for 1 h. The potential use of the produced enzymes in the degradation of human hair and cotton fabric samples were also assessed.

  4. Controlled production of cellulases in plants for biomass conversion. Annual report, March 11, 1997--March 14, 1998

    Energy Technology Data Exchange (ETDEWEB)

    Danna, K.J.

    1998-06-01

    The goal of this project is to facilitate conversion of plant biomass to usable energy by developing transgenic plants that express genes for microbial cellulases, which can be activated after harvest of the plants. In particular, the feasibility of targeting an endoglucanase and a cellobiohydrolase to the plant apoplast (cell wall milieu) is to be determined. To avoid detrimental effects of cellulose expression in plants, enzymes with high temperature optima were chosen; the genes for these enzymes are from thermophilic organisms that can use cellulose as a sole energy source. During the past year (year 2 of the grant), efforts have been focused on testing expression of endoglucanase E{sub 1}, from Acidothermus cellulolyticus, in the apoplast of both tobacco suspension cells and Arabidopsis thaliana plants. Using the plasmids constructed during the first year, transgenic cells and plants that contain the gene for the E{sub 1} catalytic domain fused to a signal peptide sequence were obtained. This gene was constructed so that the fusion protein will be secreted into the apoplast. The enzyme is made in large quantities and is secreted into the apoplast. More importantly, it is enzymatically active when placed under optimal reaction conditions (high temperature). Moreover, the plant cells and intact plants exhibit no obvious problems with growth and development under laboratory conditions. Work has also continued to improve binary vectors for Agrobacterium-mediated transformation, to determine activity of E{sub 1} at various temperatures, and to investigate the activity of the 35S Cauliflower Mosaic Virus promoter in E. coli. 9 figs.

  5. Optimization of covalent immobilization of Trichoderma reesei cellulase onto modified ReliZyme HA403 and Sepabeads EC-EP supports for cellulose hydrolysis, in buffer and ionic liquids/buffer media.

    Science.gov (United States)

    Bilgin, Ramazan; Yalcin, M Serkan; Yildirim, Deniz

    2016-08-01

    The covalent immobilization of Trichoderma reesei cellulase onto modified ReliZyme HA403 and Sepabeads EC-EP supports were carried out. The optimal immobilization conditions were determined using response surface methodology. The hydrolysis of cellulose using the free and immobilized cellulase preparations in ionic liquids (IL) using cosolvents was investigated. The hydrolytic activities in buffer medium containing 25% (v/v) of 1-butyl-3-methylimidazolium hexafluorophosphate were around 2.6-, 1.6-, and 5.5-fold higher than the activities in buffer medium. The retained initial activities were 32% and 57%, respectively for cellulase preparations immobilized onto Sepabeads EC-EP support and onto modified ReliZyme HA403 support after 5 reuses. PMID:25811997

  6. 纤维素酶产生菌的诱变与筛选%Mutation and Screening for Cellulase Production Strain

    Institute of Scientific and Technical Information of China (English)

    王美珠; 曹月坤; 陈文艺; 姚秀清; 李妍妍

    2012-01-01

    [目的]为提高纤维素酶产生菌的酶活力.[方法]以抚顺近郊堆积腐烂秸秆的土壤为分离源,利用CMC刚果红平板培养基筛选出纤维素水解圈与菌落直径比值(Hc值)较大的菌株Y-07.以Y-07为出发菌株,在最适诱变剂量条件下,对其进行紫外线诱变和亚硝酸诱变.[结果]经过3轮复筛,紫外线诱变后的U10菌株为试验中获得的高产纤维素酶菌株,其酶活最高达到2.499 IU/ml,其活性是Y-07的3.41倍.[结论]该方法为进一步提高纤维素酶产生菌的酶活力奠定了基础.%[Objective] The research aimed to improve enzyme activity of cellulase production strain. [Method] A strain named Y-07 was isolated from rotten straw soil nearby the suburb of Fushun with culture medium containing the carboxymethylcellulose sodium and Congo Red in accordance with its larger size of ratio of hydrolysis ring to colony diameter. With the strain of Y-07 as starting Strain, the strain of Y-07 was mutagenized by ultraviolet radiation and nitrous acid mutagenesis under the most appropriate amount of mutation. [Result] After three rounds of secondary screening, the strain of U10 was achieved with the biggest cellulase activity 2. 5 IU/ml which was 3.4 times of Y-07 strain. [ Conclusion] The method provided the basis for further improvements of enzyme activity of cellulase production strain.

  7. Design and characterizations of two novel cellulases through single-gene shuffling of Cel12A (EG3) gene from Trichoderma reseei.

    Science.gov (United States)

    Yenenler, Aslı; Sezerman, Osman Uğur

    2016-06-01

    Cellulases have great potential to be widely used for industrial applications. In general, naturally occurring cellulases are not optimized and limited to meet the industrial needs. These limitations lead to demand for novel cellulases with enhanced enzymatic properties. Here, we describe the enzymatic and structural properties of two novel enzymes, EG3_S1 and EG3_S2, obtained through the single-gene shuffling approach of Cel12A(EG3) gene from Trichoderma reseei EG3_S1 and EG3_S2 shuffled enzymes display 59 and 75% identity in protein sequence with respect to native, respectively. Toward 4-MUC, the minimum activity of EG3_S1 was reported as 5.9-fold decrease in native at 35°C, whereas the maximum activity of EG3_S2 was reported as 15.4-fold increase in native activity at 40°C. Also, the diminished enzyme activity of EG3_S1 was reported within range of 0.6- to 0.8-fold of native and within range of 0.5- to 0.7-fold of native toward CMC and Na-CMC, respectively. For EG3_S2 enzyme, the improved enzymatic activities within range of 1.1- to 1.4-fold of native and within range of 1.1- to 1.6-fold of native were reported toward CMC and Na-CMC, respectively. Moreover, we have reported 6.5-fold increase in the kcat/Km ratio of EG3_S2 with respect to native and suggested EG3_S2 enzyme as more efficient catalysis for hydrolysis reactions than its native counterpart. PMID:27129352

  8. Study of a High-Yield Cellulase System Created by Heavy-Ion Irradiation-Induced Mutagenesis of Aspergillus niger and Mixed Fermentation with Trichoderma reesei.

    Directory of Open Access Journals (Sweden)

    Shu-Yang Wang

    Full Text Available The aim of this study was to evaluate and validate the efficiency of 12C6+ irradiation of Aspergillus niger (A. niger or mutagenesis via mixed Trichoderma viride (T. viride culturing as well as a liquid cultivation method for cellulase production via mixed Trichoderma reesei (T. reesei and A. niger culture fermentation. The first mutagenesis approach was employed to optimize yield from a cellulase-producing strain via heavy-ion mutagenesis and high-throughput screening, and the second was to effectively achieve enzymatic hydrolysis of cellulase from a mixed culture of mutant T. viride and A. niger. We found that 12C6+-ion irradiation induced changes in cellulase biosynthesis in A. niger but had no effect on the time course of the synthesis. It is notable that the exoglucanases (CBH activities of A. niger strains H11-1 and H differed (6.71 U/mL vs. 6.01 U/mL and were significantly higher than that of A. niger mutant H3-1. Compared with strain H, the filter paper assay (FPA, endoglucanase (EG and β-glucosidase (BGL activities of mutant strain H11-1 were increased by 250.26%, 30.26% and 34.91%, respectively. A mixed culture system was successfully optimized, and the best ratio of T. reesei to A. niger was 5:1 for 96 h with simultaneous inoculation. The BGL activity of the mixed culture increased after 72 h. At 96 h, the FPA and BGL activities of the mixed culture were 689.00 and 797.15 U/mL, respectively, significantly higher than those of monocultures, which were 408.70 and 646.98 U/mL for T. reesei and 447.29 and 658.89 U/mL for A. niger, respectively. The EG activity of the mixed culture was 2342.81 U/mL, a value that was significantly higher than that of monocultures at 2206.57 U/mL for T. reesei and 1727.62 U/mL for A. niger. In summary, cellulose production and hydrolysis yields were significantly enhanced by the proposed combination scheme.

  9. Use of a Cellulase-Derepressed Mutant of Cellulomonas in the Production of a Single-Cell Protein Product from Cellulose †

    OpenAIRE

    Hitchner, E. V.; Leatherwood, J. M.

    1980-01-01

    A cellulase-derepressed mutant of a Cellulomonas species was used to produce single-cell protein from crystalline cellulose. In preliminary tests, maximum yield of single-cell protein was obtained at 30°C (pH 7.0) with urea as the nitrogen source. A continuous-flow foam flotation procedure was developed for rapid and efficient separation of bacteria from the culture liquid and cellulose residue. A pH of 4.5 was optimum for foam flotation of this organism. In preliminary trials, recovery was 8...

  10. OPTIMIZATION OF CELLULASE-FREE XYLANASE PRODUCED BY A POTENTIAL THERMOALKALOPHILIC PAENIBACILLUS SP.N1 ISOLATED FROM HOT SPRINGS OF NORTHERN HIMALAYAS IN INDIA

    Directory of Open Access Journals (Sweden)

    Sanjeev Kumar Verma

    2012-08-01

    Full Text Available Hot spring bacteria are found a novel source of highly active xylanase enzyme with significant activity at high temperature. Among bacteria, Paenibacillus sp.N1 isolated from hot water spring of Manikaran, H.P., India showed highest 24.60 IU.ml-1 of cellulase-free xylanase on Reese medium. Growth conditions including medium, incubation time, pH, temperature, inoculum size, aminoacids, carbon sources, nitrogen sources and additives that affect the xylanase production by Paenibacillus sp.N1 were studied sequentially using the classical “change-one factor at a time” method. The optimal cultivation conditions predicated from canonical analysis of this model were achieved by using basal salt medium on 3rd day, pH 9.0, temperature 50ºC with inoculum size of 12.5%, phenylalanine as aminoacid, xylose as carbon source, (NH42HPO4 as nitrogen source and Tween 20 as detergent added with an approximate yield of 52.30 IU.ml-1 escalating the over level of xylanase production by 113.38%. A rare combination of all characters i.e. thermoalkalophilic nature and high units of cellulase-free xylanase produced from a new Paenibacillus sp.N1 make it of special industrial interest.

  11. Clostridium thermocellum cellulase CelT, a family 9 endoglucanase without an Ig-like domain or family 3c carbohydrate-binding module.

    Science.gov (United States)

    Kurokawa, J; Hemjinda, E; Arai, T; Kimura, T; Sakka, K; Ohmiya, K

    2002-08-01

    The celT gene of Clostridium thermocellum strain F1 was found downstream of the mannanase gene man26B [Kurokawa J et al. (2001) Biosci Biotechnol Biochem 65:548-554] in pKS305. The open reading frame of celT consists of 1,833 nucleotides encoding a protein of 611 amino acids with a predicted molecular weight of 68,510. The mature form of CelT consists of a family 9 cellulase domain and a dockerin domain responsible for cellulosome assembly, but lacks a family 3c carbohydrate-binding module (CBM) and an immunoglobulin (Ig)-like domain, which are often found with family 9 catalytic domains. CelT devoid of the dockerin domain (CelTDeltadoc) was constructed and purified from a recombinant Escherichia coli, and its enzyme properties were examined. CelTDeltadoc showed strong activity toward carboxymethylcellulose (CMC) and barley beta-glucan, and low activity toward xylan. The V(max) and K(m) values were 137 micro mol min(-1) mg(-1) and 16.7 mg/ml, respectively, for CMC. Immunological analysis indicated that CelT is a catalytic component of the C. thermocellum F1 cellulosome. This is the first report describing the characterization of a family 9 cellulase without an Ig-like domain or family 3c CBM.

  12. Characterization and Strain Improvement of a Hypercellulytic Variant, Trichoderma reesei SN1, by Genetic Engineering for Optimized Cellulase Production in Biomass Conversion Improvement.

    Science.gov (United States)

    Qian, Yuanchao; Zhong, Lixia; Hou, Yunhua; Qu, Yinbo; Zhong, Yaohua

    2016-01-01

    The filamentous fungus Trichoderma reesei is a widely used strain for cellulolytic enzyme production. A hypercellulolytic T. reesei variant SN1 was identified in this study and found to be different from the well-known cellulase producers QM9414 and RUT-C30. The cellulose-degrading enzymes of T. reesei SN1 show higher endoglucanase (EG) activity but lower β-glucosidase (BGL) activity than those of the others. A uracil auxotroph strain, SP4, was constructed by pyr4 deletion in SN1 to improve transformation efficiency. The BGL1-encoding gene bgl1 under the control of a modified cbh1 promoter was overexpressed in SP4. A transformant, SPB2, with four additional copies of bgl1 exhibited a 17.1-fold increase in BGL activity and a 30.0% increase in filter paper activity. Saccharification of corncob residues with crude enzyme showed that the glucose yield of SPB2 is 65.0% higher than that of SP4. These results reveal the feasibility of strain improvement through the development of an efficient genetic transformation platform to construct a balanced cellulase system for biomass conversion. PMID:27621727

  13. Optimization of Enzyme Production Conditions for High-yield Cellulase Strains%纤维素酶高产菌株的产酶条件优化

    Institute of Scientific and Technical Information of China (English)

    张铁涛; 徐云升

    2016-01-01

    CMC-Congo red stain is used to screen the cellulase-degrading strain A2 with high cellulase activity and good stability from lodgy or felled banana plant near the humus soil sample.Then the strain mentioned above is identified as fungus by its morphology.The results of fermentation test show that the initial pH of medium is 5.5,the content of peptone is 0.60%,the content of carbon source is 2.40%,and the maximal activity of FPA is 2.069 IU/g.%采用刚果红鉴别培养基,从倒伏或被砍伐的香蕉株附近的腐质土样中,通过筛选得到一株产纤维素酶活力高且稳定性良好的菌株 A2,镜检初步断定菌株 A2为真菌,确定最佳产酶条件为培养基初始pH 5.5,蛋白胨含量0.60%,碳源含量2.40%,此时滤纸片酶活法(FPA)测定酶活力最大为2.069 IU/g。

  14. Use of a new Trichoderma harzianum strain isolated from the Amazon rainforest with pretreated sugar cane bagasse for on-site cellulase production.

    Science.gov (United States)

    Delabona, Priscila da Silva; Farinas, Cristiane Sanchez; da Silva, Mateus Ribeiro; Azzoni, Sindelia Freitas; Pradella, José Geraldo da Cruz

    2012-03-01

    The on-site production of cellulases is an important strategy for the development of sustainable second-generation ethanol production processes. This study concerns the use of a specific cellulolytic enzyme complex for hydrolysis of pretreated sugar cane bagasse. Glycosyl hydrolases (FPase, xylanase, and β-glucosidase) were produced using a new strain of Trichoderma harzianum, isolated from the Amazon rainforest and cultivated under different conditions. The influence of the carbon source was first investigated using shake-flask cultures. Selected carbon sources were then further studied under different pH conditions using a stirred tank bioreactor. Enzymatic activities up to 121 FPU/g, 8000 IU/g, and 1730 IU/g of delignified steam-exploded bagasse+sucrose were achieved for cellulase, xylanase and β-glucosidase, respectively. This enzymatic complex was used to hydrolyze pretreated sugar cane bagasse. A comparative evaluation, using an enzymatic extract from Trichoderma reesei RUTC30, indicated similar performance of the T. harzianum enzyme complex, being a potential candidate for on-site production of enzymes.

  15. BIOCONVERSION OF SUGARCANE BAGASSE INTO SECOND GENERATION BIOETHANOL AFTER ENZYMATIC HYDROLYSIS WITH IN-HOUSE PRODUCED CELLULASES FROM Aspergillus sp. S4B2F

    Directory of Open Access Journals (Sweden)

    Sanjeev Kumar Soni

    2010-05-01

    Full Text Available An integrated approach was studied for in-house cellulase production, pretreatment, and enzymatic conversion of sugarcane bagasse into glucose followed by the production of second generation bioethanol. Solid state cultures of Aspergillus sp. S4B2F produced significant levels of cellulase complex on wheat bran, supplemented with 1% (w/w soyabean meal, moistened with 1.5 parts of distilled water after 96 h of incubation at 30oC. The highest productivities of endoglucanase, exoglucanase, and β-glucosidase were 66, 60, and 26 IU/g of fermented dry bran, respectively. The enzyme components had a temperature and pH optima at 50oC and 4.0, respectively and revealed high thermostability at 50oC, retaining 66, 54, and 84% residual activities after 72 h. Pretreatment with 2% alkali in combination with steam was the most efficient pre-hydrolysis method for enzymatic bioconversion and fermentation of cellulosic residue of sugarcane bagasse, which produced the highest cellulose conversion (67%, with glucose and alcohol yields of 323 mg and 175 l respectively per dry gram of bagasse.

  16. Molecular characterization of SCO0765 as a cellotriose releasing endo-β-1,4-cellulase from Streptomyces coelicolor A(3).

    Science.gov (United States)

    Hong, Joo-Bin; Dhakshnamoorthy, Vijayalakshmi; Lee, Chang-Ro

    2016-09-01

    The sco0765 gene was annotated as a glycosyl hydrolase family 5 endoglucanase from the genomic sequence of Streptomyces coelicolor A3(2) and consisted of 2,241 bp encoding a polypeptide of 747 amino acids (molecular weight of 80.5 kDa) with a 29-amino acid signal peptide for secretion. The SCO0765 recombinant protein was heterogeneously over-expressed in Streptomyces lividans TK24 under the control of a strong ermE* promoter. The purified SCO0765 protein showed the expected molecular weight of the mature form (718 aa, 77.6 kDa) on sodium dodecyl sulfate-polyacryl amide gel electrophoresis. SCO0765 showed high activity toward β-glucan and carboxymethyl cellulose (CMC) and negligible activity to Avicel, xylan, and xyloglucan. The SCO0765 cellulase had a maximum activity at pH 6.0 and 40°C toward CMC and at pH 9.0 and 50-60°C toward β-glucan. Thin layer chromatography of the hydrolyzed products of CMC and β-glucan by SCO0765 gave cellotriose as the major product and cellotetraose, cellopentaose, and longer oligosaccharides as the minor products. These results clearly demonstrate that SCO0765 is an endo-β-1,4-cellulase, hydrolyzing the β-1,4 glycosidic bond of cellulose into cellotriose. PMID:27572512

  17. Partial purification and properties of cellulase-free alkaline xylanase produced by Rhizopus stolonifer in solid-state fermentation

    Directory of Open Access Journals (Sweden)

    Antonio José Goulart

    2005-05-01

    Full Text Available Rhizopus stolonifer was cultivated in wheat bran to produce a cellulase-free alkaline xylanase. The purified enzyme obtained after molecular exclusion chromatography in Sephacryl S-200 HR showed optimum temperature as 45º C and hydrolysis pHs optima as pH 6.0 and 9.0. Xylanase presented higher Vmax at pH 9.0 (0.87 µmol/mg protein than at pH 6.0 and minor Km at pH 6.0 (7.42 mg/mL than at pH 9.0.Rhizopus stolonifer foi cultivado em meio de farelo de trigo para produzir uma xilanase alcalina celulase-free. Uma amostra parcialmente purificada desta enzima foi obtida após cromatografia de exclusão molecular em Sephacryl S-200 HR. A temperatura ótima de hidrólise determinada (45º C está dentro do intervalo citado na literatura (45º C a 60º C para xilanases microbianas. Quanto ao pH ótimo, a amostra obtida apresentou atividades máximas em pH 6,0 e 9,0. Estes dados diferem da literatura, uma vez que o pH ótimo citado para a maioria das xilanases estudadas varia entre 4,0 e 5,5. De acordo com os estudos cinéticos realizados, a xilanase apresentou maior Vmax em pH 9,0 (0,87 µmol/mg proteína e menor Km em pH 6,0 (7,42 mg/mL. Os dois pHs ótimos determinados podem indicar a presença de isoformas desta enzima. Estes dados são interessantes pelo fato de que enzimas xilanolíticas alcalinas celulase-free podem ser utilizadas para o biobranqueamento da polpa na indústria de papel.

  18. 一株纤维素酶产生菌的抗阻遏选育%Anti-inhibit breeding of a cellulase-producing strain

    Institute of Scientific and Technical Information of China (English)

    贾奎艳; 段学辉; 郭炳其; 魏斌; 傅奇

    2011-01-01

    从沂蒙地区灌木枯枝中筛选到一株产纤维素酶菌株,初步鉴定为绿色木霉(T.viride Persex Fx NS90).该研究考察了不同浓度阻遏剂甘油、葡萄糖、纤维二糖对菌株生长和产酶的影响.结果显示:菌株生长的生物量随发酵培养基中甘油、葡萄糖、纤维二糖的浓度增加而提高,当培养基中分别含0.6%的甘油或0.4%葡萄糖时,菌株的相应发酵产酶活力较高,葡萄糖的浓度达2%时,菌株发酵产酶能力受到明显抑制,纤维二糖浓度对菌株发酵产酶没有明显的影响.实验采用紫外和微波对筛选菌株进行诱变处理,在含一定浓度阻遏剂的分离培养基上进行修复筛选,对选育出突变菌株的显微形态、抗阻遏性能和发酵产酶能力进行考察.结果表明:经紫外照射90s,微波辐射60s,结合2%葡萄糖平板修复选育,筛选到的突变株在生长过程中,菌落由黄色逐渐变为绿色,呈现黄色孢子,相对于出发菌株其抗阻遏性能明显提高,摇瓶发酵产CMCA和FPA酶活分别提高了41.0%和44.95%.%A cellulase -producing strain, which was identified as Trichoderma viride (T.viride Persex Fx NS90), was isolated from sapless shrubs in Yimeng area.Effects of different concentrations of glycerol, glucose, cellobiose on growth of T.viride and enzyme production were studied.The results indicated that biomass of strain growing positively correlated with concentrations of glycerol, glucose, cellobiose in the medium.The yield of cellulase was the highest when the media added 0.6% of glycerol or 0.4% glucose, and addition of 2% glucose in medium significantly inhibited cellulase production, and addition of cellobiose had no effect on yield of cellulase.The start strain was mutated with UV and microwave and the strain was repaired and screened in medium containing inhibitors.Colony morphology, anti - inhibit ability and cellulase - producing capacity of mutant strain were investigated.The results showed

  19. Engineering cellulase mixtures by varying the mole fraction of Thermomonospora fusca E[sub 5] and E[sub 3], Trichoderma reesei CBHI, and Caldocellum saccharolyticum [beta]-glucosidase

    Energy Technology Data Exchange (ETDEWEB)

    Walker, L.P.; Belair, C.D.; Wilson, D.B.; Irwin, D.C. (Cornell Univ., Ithaca, NY (United States))

    1993-11-05

    In this study, different mole fractions of pure Thermomonospora fusca E[sub 5] and E[sub 3], plus Trichoderma reesei CBHI were studied for reducing sugar production at 2 h, degree of synergism, and cellulose binding. In addition, the effects of introducing the Caldocellum saccharolyticum [Beta]-glucosidase into this cellulase system were investigated. The cellulases used were purified to homogeneity. Avicel PH 102 was the substrate. Reactions were run at 50 C for 2 h using total cellulase concentrations of 8.3 or 12.2 [mu]M. A bimixture of T. fusca E[sub 3] and T. reesei CBHI was very effective in hydrolyzing microcrystalline cellulose. The addition of endoglucanase E[sub 5] to the mixture only increased conversion to 9.8%. However, when both E[sub 5] and [Beta]-glucosidase were added, conversion increased to 14%. It was also observed that increasing total cellulase concentration beyond 8.3 [mu]M did little to increase percent conversion of cellulose into glucose. The results of the binding studies indicate no competition for binding sites between the endo- and exocellulases.

  20. 高产纤维素酶的绿色木霉菌种的诱变和筛选%Screening for a Novel Trichoderma vride Strain Highly Producing Cellulase via Ultraviolet Mutagenesis

    Institute of Scientific and Technical Information of China (English)

    屈二军; 谢展; 马孟星; 张亚飞

    2011-01-01

    [目的]诱变和筛选高产纤维素酶的绿色木霉菌种。[方法]利用紫外线对出发菌株进行诱变,经过初筛和发酵检测挑选出高产纤维素酶的菌株。[结果]筛选到高产纤维素酶的绿色木霉K6,酶活是出发菌株的1.39倍。[结论]诱变筛选得到的K6菌株高产纤维素酶,为秸秆纤维素的利用奠定了基础。%[Objective] The aim of this study was to find out a new Trichoderma vride K strain highly producing cellulase.[Method] Ultraviolet(UV) was used to induce mutagenesis on T.vride K and to select out a new Trichoderma vride strain highly producing cellulase from the first round and further selection.[Result] A new T.vride strain K6 with high yield of cellulase was obtained with the enzyme production amount of 1.39 times over that of starting strain K.This strain showed highest cellulase yield under the culture condition of 28 ℃ for 96 h.[Conclusion] The strain K6 selected out from induced mutation is endowed with better capacity of producing cellulase,which provides a new method for the utilization of straw.

  1. Structure and stability of metagenome-derived glycoside hydrolase family 12 cellulase (LC-CelA a homolog of Cel12A from Rhodothermus marinus

    Directory of Open Access Journals (Sweden)

    Hiroyuki Okano

    2014-01-01

    Full Text Available Ten genes encoding novel cellulases with putative signal peptides at the N-terminus, termed pre-LC-CelA–J, were isolated from a fosmid library of a leaf–branch compost metagenome by functional screening using agar plates containing carboxymethyl cellulose and trypan blue. All the cellulases except pre-LC-CelG have a 14–29 residue long flexible linker (FL between the signal peptide and the catalytic domain. LC-CelA without a signal peptide (residues 20–261, which shows 76% amino acid sequence identity to Cel12A from Rhodothermus marinus (RmCel12A, was overproduced in Escherichia coli, purified and characterized. LC-CelA exhibited its highest activity across a broad pH range (pH 5–9 and at 90 °C, indicating that LC-CelA is a highly thermostable cellulase, like RmCel12A. The crystal structure of LC-CelA was determined at 1.85 Å resolution and is nearly identical to that of RmCel12A determined in a form without the FL. Both proteins contain two disulfide bonds. LC-CelA has a 16-residue FL (residues 20–35, most of which is not visible in the electron density map, probably due to structural disorder. However, Glu34 and Pro35 form hydrogen bonds with the central region of the protein. ΔFL-LC-CelA (residues 36–261 and E34A-LC-CelA with a single Glu34 → Ala mutation were therefore constructed and characterized. ΔFL-LC-CelA and E34A-LC-CelA had lower melting temperatures (Tm than LC-CelA by 14.7 and 12.0 °C respectively. The Tm of LC-CelA was also decreased by 28.0 °C in the presence of dithiothreitol. These results suggest that Glu34-mediated hydrogen bonds and the two disulfide bonds contribute to the stabilization of LC-CelA.

  2. Cellulase, Clostridia, and Ethanol†

    OpenAIRE

    Demain, Arnold L.; Newcomb, Michael; Wu, J. H. David

    2005-01-01

    Biomass conversion to ethanol as a liquid fuel by the thermophilic and anaerobic clostridia offers a potential partial solution to the problem of the world's dependence on petroleum for energy. Coculture of a cellulolytic strain and a saccharolytic strain of Clostridium on agricultural resources, as well as on urban and industrial cellulosic wastes, is a promising approach to an alternate energy source from an economic viewpoint. This review discusses the need for such a process, the cellulas...

  3. Study on Isolation and Characterization of Bacteria Producing Cellulase%纤维素酶产生菌的筛选及酶学性质研究

    Institute of Scientific and Technical Information of China (English)

    孙玉宇

    2014-01-01

    Isolated Cellulose decomposing ability was the strongest from city life garbage, and extracellular enzyme production strains, tentatively identified as Bacillus ,and enzymatic properties of cellulase produced by the bacteria were investigated. The cellulase reaction, the optimum temperature is 50℃ ; the optimum pH7.0; enzyme at 40~50℃ has good thermal stability; the activity of CMCase in pH 6 ~ 7 can be maintained above 70%, the activity of FPA in pH 6~7 can be maintained more than 79%; Ca2+ and Mg2+ for the enzyme reaction were increased markedly, While Fe3+ and Mn2+ had inhibitory effect on the enzyme reaction, effect of Na+, Zn2+ and K+ on the enzyme is very small.%从城市生活垃圾中分离到一株分解纤维素能力最强,并产生胞外酶的菌株,初步确定为芽孢杆菌,并对该菌所产的纤维素酶的酶学性质进行了初步研究.该纤维素酶反应最佳温度为50℃;最适 pH7.0;酶在40~50℃热稳定性较好; CMCase活力在 pH 6.0~7.0处可保持70%以上, FPA酶活在pH 6.0~7.0处可保持79%以上; Ca2+和Mg2+对酶反应表现为明显的促进作用,而Fe3+和Mn2+对酶反应有抑制作用, Na+、Zn2+和K+对酶的影响很小.

  4. 产纤维素酶细菌的筛选及其紫外诱变%Screening and Ultraviolet Mutagenesis of Cellulase-Producing Bacteria

    Institute of Scientific and Technical Information of China (English)

    张学佳; 田云; 卢向阳

    2013-01-01

    采用刚果红脱色圈法初筛得到143株有纤维素酶活性的细菌,然后采用DNS法复筛得到纤维素酶酶活最高的6株细菌,进一步进行紫外诱变处理,获得酶活提高最大且具有遗传稳定性的菌株E140',酶活为0.90 IU· mL-1,较出发菌株(0.68 IU·mL-1)提高了32.35%.表明采用刚果红脱色圈法和DNS法联合筛选并结合紫外诱变,可以获得纤维素酶活性高的细菌.%Firstly, 143 strains with cellulase activity were obtained by primary screening with Congo red decolorizing ring method,and then 6 strains with the highest enzyme activity were obtained by screening with DNS method. After the furtherly UV mutagenesis treatment,a mutant strain E140' with the highest enzyme activity increasing and genetic stability was got, whose enzyme activity reached 0. 90 IU · mL-1,increasing by 32. 35% .than that of the starting strain(0. 68 IU · mL-1). It showed that cellulase-producing bacteria with high enzyme activity could be obtained by screening with both Congo red decolorizing ring method and DNS method, combining with UV mutagenesis treatment.

  5. 里氏木霉产纤维素酶的诱导和合成机理研究进展%Research progress of the mechanism of induction and synthesis of cellulase by Trichoderma reesei

    Institute of Scientific and Technical Information of China (English)

    李辉; 王义强; 陈介南; 张伟涛

    2011-01-01

    Cellulase is a kind of inducible enzyme.The inducer is required for cellulase expression.Cellulose, sophorose, lactose, cellobiose and sorbitol are common inducers.Trichoderma reesei mainly expresses three cellulases, endoglucanase, exoglucanase and β-1,4-glucosidase, in which the expression level of exoglucanase Ⅰ is highest, followed by endoglucanase and β-1,4-glucosidase.The synthesis and expression of cellulase is affected by active elements and inhibitory factors.In details, the combination of inducer and active element induces the synthesis and expression of cellulose.When the decomposition products of cellulase accumulate to a certain level, the intracellular inhibitory factor will combine with promoter to prevent the expression of cellulase.%纤维素酶是一类诱导酶,诱导剂对纤维素酶的表达是必需的.常用的诱导剂如纤维素、槐糖、乳糖、纤维二糖、山梨糖等.里氏木霉主要表达3种纤维素酶:内切葡聚糖酶、外切葡聚糖酶和β-1,4-葡萄糖苷酶,其中以外切葡聚糖苷酶Ⅰ的合成量最多,其次是内切葡聚糖苷酶和β-1,4-葡萄糖苷酶.纤维素酶的合成和表达是激活原件和抑制因子共同作用的结果:诱导物质和激活元件结合激活细胞内纤维素酶的合成和表达;当纤维素酶的分解产物达到一定水平时,细胞内的抑制因子和启动子结合,阻止纤维素酶的表达.

  6. 里氏木霉Trichoderma reesei产纤维素酶的发酵培养基碳氮源优化%Culture medium optimization for producing cellulase by Trichoderma reesei

    Institute of Scientific and Technical Information of China (English)

    祖彩霞; 李志敏; 叶勤

    2011-01-01

    The influence of carbon and nitrogen sources on cellulase production was investigated. Single factor method and response surface methodology were used to optimize the medium for cellulase production by Trichoderma reesei. Firstly, it showed that soybean powder, corn cob and corn steep liquor had positive effects on cellulase production in one factor at a time. Then, the optimal concentrations of soybean powder (32. 21 g/L) , corn cob (42. 29 g/L) and corn steep liquor (4.45 g/L) were determined by central compostion design and response surface analysis. Under these optimizations conditions, the cellulase production reached (10. 65 ±0. 50) U/mL on the flask cultivation for 7 d.%研究C、N源对里氏木霉(Trichoderma reesei)生产纤维素酶的影响,采用单因素实验方法和中心复合方法对发酵培养基进行优化.单因素实验表明:黄豆饼粉、玉米芯、玉米浆对纤维素酶的影响显著.通过响应面优化,得到最优培养基C、N源的组成:黄豆饼粉32.21g/L,玉米芯42.29g/L,玉米浆4.45g/L.优化条件下,摇瓶发酵7 d的比酶活达到(10.65±0.50)U/mL.

  7. Production of Cellulases, Xylanase, Pectinase, alpha-amylase and Protease Enzymes Cocktail by Bacillus spp. and Their Mixed Cultures with Candida tropicalis and Rhodotorula glutinis under Solid State Fermentation

    International Nuclear Information System (INIS)

    A group of twelve locally isolated Bacillus species, B.megaterium (MAI and MA II), B.licheniformis (MLI and ML II); B. circulans, B. stearothermophilis, B.cereus, B.sphaericus, B. pumilus, B. laterosporus, B. coagulans and B. pantothenticus, were examined for the production of cellulases, xylanase, pectinase, alpha-amylase and protease enzymes cocktail on wheat bran under solid state fermentation (SSF). All species were found to be potent hydrolyzing enzymes producers and the superior producing species were B. megaterium MAI and B. licheniformis. On the other hand, both of them still produced highest enzyme titres when mixed with Candida tropicalis or Rhodotorula glutinis, yeast strains. The two superior bacterial strains produced the highest enzymatic activities when coculturing with C. tropicalis compared with coculturing with R. glutinis only or with both C. tropicalis and R. glutinis in combination. The inferior activities of cocultures (B. megaterinm MAI and R. glutinis) were enhanced in carboxymethyl cellulase (CMCase), filter paper cellulase (FPase), avecilase, xylanase, pectinase, -amylase and protease by gamma irradiation at dose 1.0 kGy with percent increase 8 %, 20 %, 10 %, 4 %, 31 %, 22 % and 34 %, respectively as compared with un-irradiated cocultures

  8. 硫化物对纤维素酶活及织物处理效果的影响%Effects of sulfide on cellulase activity and enzyme treatment performance of cotton fabric

    Institute of Scientific and Technical Information of China (English)

    姚继明; 窦春晓; 李倩

    2014-01-01

    1%(on the mass of acid cellulase solutions) of thiourea, sodium thiocyanate, sodium sulfite, so-dium bisulfite, sodium metabisulfite, sodium hyposulfite, sodium hydrosulfide and sodium sulfide were added to acid cellulase solutions, respectively. The filter paper enzymatic activities were measured according to cen-tral composite experiment design under 40, 50, 60 ℃ and pH 4.5, 5.0, 5.5. The effects of various sulfides on the enzymatic activities were analyzed by Minitab software. The cotton knitted fabrics were treated with cellu-lase under the same conditions, and the bursting strength, hairiness removal percentage and glucose content of residue liquid were measured to evaluate the practical performance on cotton fabrics. The results showed that the addition of sulfides could slightly enhance the cellulase activity, and widen cellulase working tempera-ture and pH range. However, except sodium sulfite and sodium hyposulfite, the addition of other six sulfides decreased the hairiness removal ability.%在酸性纤维素酶液中分别加入1%(对溶液质量)的硫脲、硫氰酸钠、亚硫酸钠、亚硫酸氢钠、焦亚硫酸钠、连二亚硫酸钠、硫氢化钠、硫化钠等8种不同的硫化物,在温度40、50、60℃,pH=4.5、5.0、5.5的条件下,根据中心复合试验设计方案测定滤纸酶活力,并用Minitab软件分析各种硫化物对纤维素酶活力的影响.在相同的条件下对纯棉针织物进行酶处理,测定处理前后织物的顶破强力、毛羽去除率、处理残液葡萄糖含量来评价对织物的实际作用效果.结果表明,硫化物的加入可以小幅提高纤维素酶活力、扩大酶的温度和pH作用区间,但除了亚硫酸钠和连二亚硫酸钠外,其他6种硫化物的加入反而降低了织物的除毛效果.

  9. 产纤维素酶食用菌筛选及其培养基的优化%Screening for Cellulase Producing Edible Fungi and Optimization of Medium

    Institute of Scientific and Technical Information of China (English)

    丁玉萍; 张康; 叶红强; 王晓娟; 杨莎; 梅竹; 任欢欢

    2012-01-01

    利用刚果红杯碟快速筛选法对大白菇、金针菇、香菇、竹荪、磷盖红菇、杏鲍菇6种食用菌产纤维素酶能力进行比较,同时对高产纤维素酶食用菌的培养基最佳碳源和配方进行筛选、优化。结果表明:磷盖红菇出现透明圈最大,直径为20mm;对比羧甲基纤维素钠(CMC-Na)、豆渣、秸秆粉、葡萄糖4种碳源,豆渣对磷盖红菇产纤维素酶有促进作用,优化培养基为豆渣60g/L、硫酸铵2.0g/L、吐温-80 1.5mL/L、pH5.4。%The ability of six edible fungi,Russula delica Fr,Flammulina velutipe,Lentinus javanicus,Bamboo-sun,Russula lepida and Abalone mushroom,to produce cellulase were compared using the Congo red saucer rapid screening method.Meanwhile,the optimum carbon source and medium formula for cellulase production were screened and optimized.The result showed that Russula lepida produced the largest transparent rings of 20 mm in diameter,showing its high yield of cellulase.Among the four kinds of carbon source,CMC-Na,bean dregs,straw powder and glucose,the bean dregs promoted Russula lepida to produce cellulase.The optimized medium for cellulase production by Russula lepida was bean dregs 60 g/L,ammonium sulfate 2.0 g/L,Tween-80 1.5 mL/L,and pH 5.4.

  10. 大伏革菌产纤维素酶条件优化及高效菌株筛选%Optimization of Submerged Culture Requirements for the Cellulase Activity by Phlebiopsis gigantea and Screening the Most Effective Strain

    Institute of Scientific and Technical Information of China (English)

    李杏春; 何双辉; 戴玉成

    2014-01-01

    The activity of extracellular cellulase could be used for screening the effective biocontrol strain. We could acquire the most effective Phlebiopsis gigantea isolate associated with biocontrol conifer root and butt rots by determining cellulase activity of them. Based on single factor and orthogonal experiment, the optimum medium compositions for the highest cellulase activity were(g/L):glucose 30.00, peptone 5.00, KH2PO4 3.00, MgSO4·7H2O 1.50, the optimal liquid medium volume was 120 mL/250 mL, the optimal inoculum volume was 5%(V/V), initial pH4.0. By determining cellulase activity of P. gigantea isolates, we concluded that P. gigantea 08077 had the highest cellulase activity and 13025 was the lowest. P. gigantea 08077 is better to control the Chinese Heterobasidion parviporum than Rotstop-F that had been registered in Finland.%真菌胞外纤维素酶活可作为高效生防菌株筛选的指标之一,通过测定野生大伏革菌(Phlebiopsis gigantea)菌株在最优条件下所产生纤维素酶活力高低来达到筛选防治针叶树根腐病高效菌株的目的。以大伏革菌菌株液体培养过程中纤维素酶活力为指标,通过单因素和正交试验结合的方法,筛选出大伏革菌最适产纤维素酶条件为(g/L):葡萄糖30.00,蛋白胨5.00,磷酸二氢钾3.00,硫酸镁1.50,装液量120 mL/250 mL,接种量5%(V/V),初始pH为4.0。测定野生大伏革菌菌株纤维素酶活力的大小显示:08077号菌株纤维素酶活力最大,13025号菌株最小。研究得出08077号菌株比芬兰商业化生物制剂Rotstop-F具有更好的防治中国小孔异担子菌的潜能。

  11. 一株产纤维素酶的甲醇利用细菌的鉴定及其纤维素降解条件优化%Identification and optimal degradation conditions for cellulase-degrading enzyme of a methanol-utilizing and cellulase-producing bacterium

    Institute of Scientific and Technical Information of China (English)

    高健; 肖丹青; 刘喜平; 王能强; 张大为

    2012-01-01

    采用刚果红染色法,从废弃矿山周边土壤中筛选出一株产纤维素酶的甲醇利用细菌,命名为xt - 04.形态特征、生理试验及16S rDNA序列和gyrB序列分析表明,该菌株属于Bacillus methylotrophicus.为提高该菌所产纤维素酶的降解能力,首先通过单因子实验考察了底物CMC -Na浓度、反应温度及缓冲液pH值对纤维素酶活力的影响;然后采用响应面分析法对影响纤维素酶活力的3个单因子进行了优化.结果表明,单因素实验得出的适宜反应温度、缓冲液pH和底物浓度分别为70℃、5.0和2% (20 mg/mL);响应面法得出的最高酶活力条件:反应温度、pH和底物浓度分别为66.1℃、4.81和19.01mg/mL.在最优条件下,酶活力达到17.85 U/mL,比优化前的酶活力12.84 U/mL提高了39.01%.因此,鉴于这种纤维素酶能耐受较高温度和酸性条件,该菌株所产纤维素酶可能在工业中具有良好的应用前景.%A methanol-utilizing and cellulase-producing bacteria, designated strain xt-04, was isolated from the soil of abandoned mine lands of Hunan Province by Congo red staining test. This strain was identified as the species Bacillus methylotrophicus based on the morphological, physiological characteristics and 16S rDNA and gyrB gene sequences analysis. In order to improve the catalytic ability of the cellulase produced by this strain, influences of the concentration of CMC-Na, temperature and pH on the activity of cellulose-decomposing enzyme were investigated by single factor experiment. Then, response surface analysis was used to optimize the influences of three factors on the cellulase activity. The results indicated that the appropriate concentrations of CMC-Na, temperature and pH, based on single-factor experiments, were 20 mg/mL, 70 ℃ , and 5.0, respectively. And the optimal concentrations of CMC-Na, temperature and pH of cellulase-producing, from response surface analysis, were 19.01 mg/mL, 66.1 ℃ and 4

  12. A highly thermostable alkaline cellulase-free xylanase from thermoalkalophilic Bacillus sp. JB 99 suitable for paper and pulp industry: purification and characterization.

    Science.gov (United States)

    Shrinivas, Dengeti; Savitha, Gunashekaran; Raviranjan, Kumar; Naik, Gajanan Ramchandra

    2010-11-01

    A highly thermostable alkaline xylanase was purified to homogeneity from culture supernatant of Bacillus sp. JB 99 using DEAE-Sepharose and Sephadex G-100 gel filtration with 25.7-fold increase in activity and 43.5% recovery. The molecular weight of the purified xylanase was found to be 20 kDA by SDS-PAGE and zymogram analysis. The enzyme was optimally active at 70 °C, pH 8.0 and stable over pH range of 6.0-10.0.The relative activity at 9.0 and 10.0 were 90% and 85% of that of pH 8.0, respectively. The enzyme showed high thermal stability at 60 °C with 95% of its activity after 5 h. The K (m) and V (max) of enzyme for oat spelt xylan were 4.8 mg/ml and 218.6 µM min(-1) mg(-1), respectively. Analysis of N-terminal amino acid sequence revealed that the xylanase belongs to glycosyl hydrolase family 11 from thermoalkalophilic Bacillus sp. with basic pI. Substrate specificity showed a high activity on xylan-containing substrate and cellulase-free nature. The hydrolyzed product pattern of oat spelt xylan on thin-layer chromatography suggested xylanase as an endoxylanase. Due to these properties, xylanase from Bacillus sp. JB 99 was found to be highly compatible for paper and pulp industry.

  13. The industrial applicability of purified cellulase complex indigenously produced by Trichoderma viride through solid-state bio-processing of agro-industrial and municipal paper wastes

    Directory of Open Access Journals (Sweden)

    Muhammad Irshad

    2013-02-01

    Full Text Available An indigenous strain of Trichoderma viride produced high titers of cellulase complex in solid-state bio-processing of agro-industrial orange peel waste, which was used as the growth-supporting substrate. When the conditions of the SSF medium containing 15 g orange peel (50% w/w moisture inoculated with 5 mL of inoculum were optimal, the maximum productions of endoglucanase (655 ± 5.5 U/mL, exoglucanase (412 ± 4.3 U/mL, and β-glucosidase (515 ± 3.7 U/mL were recorded after 4 days of incubation at pH 5 and 35 °C. The enzyme with maximum activity (endoglucanase was purified by ammonium sulfate fractionation and Sephadex G-100 column gel filtration chromatographic technique. Endoglucanase was 5.5-fold purified with specific activity of 498 U/mg in comparison to the crude enzyme. The enzyme was shown to have a molecular weight of 58 kDa by sodium dodecyl sulphate poly-acrylamide gel electrophoresis (SDS-PAGE. The shelf life profile revealed that the enzyme could be stored at room temperature (30 °C for up to 45 days without losing much of its activity.

  14. A 24.7-kDa copper-containing oxidase, secreted by Thermobifida fusca, significantly increasing the xylanase/cellulase-catalyzed hydrolysis of sugarcane bagasse.

    Science.gov (United States)

    Chen, Cheng-Yu; Hsieh, Zhi-Shen; Cheepudom, Jatuporn; Yang, Chao-Hsun; Meng, Menghsiao

    2013-10-01

    Thermobifida fusca is a moderately thermophilic soil bacterium belonging to Actinobacteria. It has been known for its capability to degrade plant cell wall polymers except lignin and pectin. To know whether it can produce enzymes to facilitate lignin degradation, the extracellular proteins bound to sugarcane bagasse were harvested and identified by liquid chromatography tandem mass spectrometry. Among the identified proteins, a putative copper-containing polyphenol oxidase of 241 amino acids, encoded by the locus Tfu_1114, was thought to presumably play a role in lignin degradation. This protein (Tfu1114) was thus expressed in E. coli and characterized. Similarly to common laccases, Tfu1114 is able to catalyze the oxidation reaction of phenolic and nonphenolic lignin related compounds such as 2,6-dimethoxyphenol and veratryl alcohol. More interestingly, it can significantly enhance the enzymatic hydrolysis of bagasse by xylanase and cellulase. Tfu1114 is stable against heat, with a half-life of 4.7 h at 90 °C, and organic solvents. It is sensitive to ethylenediaminetetraacetic acid and reducing agents but resistant to sodium azide, a potent inhibitor of laccases. Atomic absorption spectroscopy indicated that the ratio of copper to the protein monomer is 1, instead of 4, a feature of classical laccases. All these data suggest that Tfu1114 is a novel oxidase with laccase-like activity, potentially useful in biotechnology application. PMID:23377789

  15. Purification and characterization of cellulase-free low molecular weight endo β-1,4 xylanase from an alkalophilic Cellulosimicrobium cellulans CKMX1 isolated from mushroom compost.

    Science.gov (United States)

    Walia, Abhishek; Mehta, Preeti; Chauhan, Anjali; Kulshrestha, Saurabh; Shirkot, C K

    2014-10-01

    Alkalophilic Cellulosimicrobium cellulans CKMX1 isolated from mushroom compost is first report on actinomycetes that has the ability to produce thermostable cellulase-free xylanase, which is an important industrial enzyme used in the pulp and paper industry. Strain CKMX1 was characterized by metabolic fingerprinting, whole-cell fatty acids methyl ester analysis and 16Sr DNA and found to be C. cellulans CKMX1.The enzyme was purified by gel permeation and anion exchange chromatography and had a molecular mass of 29 kDa. Xylanase activity was optimum at pH 8.0 and 55 °C. The enzyme was somewhat thermostable, retaining 50 % of the original activity after incubation at 50 °C for 30 min. The xylanase had K m and V max values of 2.64 mg/ml and 2,000 µmol/min/mg protein in oat spelt xylan, respectively. All metal ions except HgCl2, CoCl2 as well as CdCl2 were well tolerated and did not adversely affect xylanase activity. The deduced internal amino acid sequence of C. cellulans CKMX1 xylanase by matrix assisted laser desorption ionization-time of flight mass spectrometry resembled the sequence of β-1,4-endoxylanase, which is a member of glycoside hydrolase family 11. Some of the novel characteristics that make this enzyme potentially effective in xylan biodegradation could be useful for pulp and paper biobleaching are discussed in this manuscript. PMID:24908422

  16. Extracellular production of novel halotolerant, thermostable, and alkali-stable carboxymethyl cellulase by marine bacterium Marinimicrobium sp. LS-A18.

    Science.gov (United States)

    Zhao, Kun; Guo, Li-Zhong; Lu, Wei-Dong

    2012-10-01

    Cellulases which are active and stable under extreme conditions have attracted considerable attention because of their potential industrial applications. Marinimicrobium sp. LS-A18 showed high extracellular carboxymethylcellulase (CMCase) activity when grown on mineral salt medium containing carboxymethylcellulose as the sole carbon source. Maximum CMCase activity was obtained at 55°C and pH 7.0 in the absence of NaCl. Under the optimized fermentation conditions, the yield of CMCase was increased up to 2.5 U/ml, which was 3.1-fold higher than that before optimization. The enzyme retained 84 % of residual activity after incubation at 60°C for 1 h and more than 88 % of residual activity after incubation for 72 h in the presence of different pH (5-11) and NaCl concentrations (0-25 %, w/v), indicating it was halotolerant, thermostable and alkali-stable. These characteristics made the CMCase from Marinimicrobium sp. LS-A18 as a potentially novel biocatalyst in biotechnological and industrial applications.

  17. Use of a cellulase-derepressed mutant of cellulomonas in the production of a single-cell protein product from cellulose.

    Science.gov (United States)

    Hitchner, E V; Leatherwood, J M

    1980-02-01

    A cellulase-derepressed mutant of a Cellulomonas species was used to produce single-cell protein from crystalline cellulose. In preliminary tests, maximum yield of single-cell protein was obtained at 30 degrees C (pH 7.0) with urea as the nitrogen source. A continuous-flow foam flotation procedure was developed for rapid and efficient separation of bacteria from the culture liquid and cellulose residue. A pH of 4.5 was optimum for foam flotation of this organism. In preliminary trials, recovery was 85% of the cells with the flotation procedure. Cellulomonas was 68% true protein and had an essential amino acid profile featuring a high lysine content (6.5% of protein). The Cellulomonas product was evaluated nutritionally with weanling rats. The net protein utilization value for the protein supplemented with methionine was 50.4% Weight gain of rats on the Cellulomonas diet was similar to that of rats fed a casein diet. PMID:16345511

  18. Isolation, Screening, and Identification of Cellulolytic Bacteria from Natural Reserves in the Subtropical Region of China and Optimization of Cellulase Production by Paenibacillus terrae ME27-1

    Directory of Open Access Journals (Sweden)

    Yan-Ling Liang

    2014-01-01

    Full Text Available From different natural reserves in the subtropical region of China, a total of 245 aerobic bacterial strains were isolated on agar plates containing sugarcane bagasse pulp as the sole carbon source. Of the 245 strains, 22 showed hydrolyzing zones on agar plates containing carboxymethyl cellulose after Congo-red staining. Molecular identification showed that the 22 strains belonged to 10 different genera, with the Burkholderia genus exhibiting the highest strain diversity and accounting for 36.36% of all the 22 strains. Three isolates among the 22 strains showed higher carboxymethyl cellulase (CMCase activity, and isolate ME27-1 exhibited the highest CMCase activity in liquid culture. The strain ME27-1 was identified as Paenibacillus terrae on the basis of 16S rRNA gene sequence analysis as well as physiological and biochemical properties. The optimum pH and temperature for CMCase activity produced by the strain ME27-1 were 5.5 and 50°C, respectively, and the enzyme was stable at a wide pH range of 5.0–9.5. A 12-fold improvement in the CMCase activity (2.08 U/mL of ME27-1 was obtained under optimal conditions for CMCase production. Thus, this study provided further information about the diversity of cellulose-degrading bacteria in the subtropical region of China and found P. terrae ME27-1 to be highly cellulolytic.

  19. 多菌混合固态发酵产纤维素酶研究%Cellulase production by solid-state fermentation with multi-strains

    Institute of Scientific and Technical Information of China (English)

    刘云云; 张宇; 许敬亮; 何敏超; 庄新姝; 袁振宏

    2014-01-01

    采用多菌混合发酵可以提高纤维素酶的活力,为获得高活力的纤维素酶制剂,文中以碱处理后的甘蔗渣和麸皮作为发酵产酶培养基,采用响应面法对2株纤维素酶生产菌里氏木霉CICC40359和斜卧青霉SMX固态混合发酵条件进行了优化。结果发现在发酵温度为28℃,料水比(质量体积比)1∶2.5(g/mL)的条件下,当V(青霉)∶V(木霉)为3∶1,总接种量8%(mL/g),培养基中 m(蔗渣)∶m(麸皮)为2∶1,发酵3 d 时,滤纸酶活有最大值达到101.825 FPU/g,这为后续优化工作的开展提供了依据,同时高酶活下发酵液中呈现高的糖质量分数为同步产酶发酵产乙醇提供了新思路。%Cellulase production efficiency can be improved via the mixed culture solid-state fermentation.In order to obtain high cellulase activity,alkali-treated sugarcane bagasse supplemented with wheat bran was used as enzyme fermentation substrate.The response surface method was introduced to optimize the solid-state mixed-fermentation with Trichoderma reesei CICC40359 and Penicillium decumbens SMX.The results indicate that the filter paper enzyme activity has an maximum value of 101.825 FPU/g under the following conditions:temperature is 28 ℃;mass and volume ratio of material to water is 1 ∶2.5 g/mL;V(Penicillium )∶V(Trichoderma) =3 ∶1, m(bagasse)∶m(wheat bran)=2∶1 ,inoculum dose is 0.08 g/mL and fermentation time is 3 d,which provides a basis for the next optimization work.The fermentation liquor showed high sugar content when the enzyme activity was high.These results indicate that simultaneous enzyme and ethanol production may be an effective approach for the economic production of bioethanol.

  20. Selection of Bacillus spp. for Cellulase and Xylanase Production as Direct-Fed Microbials to Reduce Digesta Viscosity and Clostridium perfringens Proliferation Using an in vitro Digestive Model in Different Poultry Diets

    Science.gov (United States)

    Latorre, Juan D.; Hernandez-Velasco, Xochitl; Kuttappan, Vivek A.; Wolfenden, Ross E.; Vicente, Jose L.; Wolfenden, Amanda D.; Bielke, Lisa R.; Prado-Rebolledo, Omar F.; Morales, Eduardo; Hargis, Billy M.; Tellez, Guillermo

    2015-01-01

    Previously, our laboratory has screened and identified Bacillus spp. isolates as direct-fed microbials (DFM). The purpose of the present study was to evaluate the cellulase and xylanase production of these isolates and select the most appropriate Bacillus spp. candidates for DFM. Furthermore, an in vitro digestive model, simulating different compartments of the gastrointestinal tract, was used to determine the effect of these selected candidates on digesta viscosity and Clostridium perfringens proliferation in different poultry diets. Production of cellulase and xylanase were based on their relative enzyme activity. Analysis of 16S rRNA sequence classified two strains as Bacillus amyloliquefaciens and one of the strains as Bacillus subtilis. The DFM was included at a concentration of 108 spores/g of feed in five different sterile soybean-based diets containing corn, wheat, rye, barley, or oat. After digestion time, supernatants from different diets were collected to measure viscosity, and C. perfringens proliferation. Additionally, from each in vitro simulated compartment, samples were taken to enumerate viable Bacillus spores using a plate count method after heat-treatment. Significant (P < 0.05) DFM-associated reductions in supernatant viscosity and C. perfringens proliferation were observed for all non-corn diets. These results suggest that antinutritional factors, such as non-starch polysaccharides from different cereals, can enhance viscosity and C. perfringens growth. Remarkably, dietary inclusion of the DFM that produce cellulase and xylanase reduced both viscosity and C. perfringens proliferation compared with control diets. Regardless of diet composition, 90% of the DFM spores germinated during the first 30 min in the crop compartment of the digestion model, followed by a noteworthy increased in the intestine compartment by ~2log10, suggesting a full-life cycle development. Further studies to evaluate in vivo necrotic enteritis effects are in

  1. Cellulase production by two mutant strain of Trichoderma longibrachiatum Qm9414 and Rut C30. Produccion de celulasas a partir de dos cepas hiperproductoras de trichoderma longibrachiatum Qm9414 y Rut C30

    Energy Technology Data Exchange (ETDEWEB)

    Blanco, M.J.

    1991-01-01

    Native or pretreated biomass from Onopordum nervosum boiss, has been examined as candidate feedstock for cellulase production by two mutant strain of trichoderma longibrachiatum QM9414 and Rut C30. Batch cultivation methods were evaluated and compared with previous experiments using ball-milled, crystalline cellulose (Solka floc). Batch cultivation of T. longibrachiatum Rut C30 on 55% (W/V) acid pretreated O. nervosum biomass yielded enzyme productivities and activities comparable to those obtained on Solka floc. However, the overall enzyme production performance was lower than on Solka floc at comparable cellulose concentrations. This fact may be due to the accumulation of pretreated by products and lignin in the fermentor.(author)

  2. Cellulase production by two mutant strain of Trichoderma longibrachiatum Qm9414 and Rut C30; Produccion de celulasas a partir de dos cepas hiperproductoras de trichoderma longibrachiatum Qm9414 y Rut C30

    Energy Technology Data Exchange (ETDEWEB)

    Blanco, M.J.

    1991-12-31

    Native or pretreated biomass from Onopordum nervosum boiss, has been examined as candidate feedstock for cellulase production by two mutant strain of trichoderma longibrachiatum QM9414 and Rut C30. Batch cultivation methods were evaluated and compared with previous experiments using ball-milled, crystalline cellulose (Solka floc). Batch cultivation of T. longibrachiatum Rut C30 on 55% (W/V) acid pretreated O. nervosum biomass yielded enzyme productivities and activities comparable to those obtained on Solka floc. However, the overall enzyme production performance was lower than on Solka floc at comparable cellulose concentrations. This fact may be due to the accumulation of pretreated by products and lignin in the fermentor.(author)

  3. EndB, a Multidomain Family 44 Cellulase from Ruminococcus flavefaciens 17, Binds to Cellulose via a Novel Cellulose-Binding Module and to Another R. flavefaciens Protein via a Dockerin Domain

    OpenAIRE

    Rincón, Marco T.; McCrae, Sheila I.; Kirby, James; Scott, Karen P.; Flint, Harry J.

    2001-01-01

    The mechanisms by which cellulolytic enzymes and enzyme complexes in Ruminococcus spp. bind to cellulose are not fully understood. The product of the newly isolated cellulase gene endB from Ruminococcus flavefaciens 17 was purified as a His-tagged product after expression in Escherichia coli and found to be able to bind directly to crystalline cellulose. The ability to bind cellulose is shown to be associated with a novel cellulose-binding module (CBM) located within a region of 200 amino aci...

  4. 酶解工艺对籽瓜浓缩汁出汁率的影响%Effect of Enzymatic Treatment with Pectinase and Cellulase on Juice Yield of Seed Melon

    Institute of Scientific and Technical Information of China (English)

    刘忆冬; 罗鹏; 刘娅; 陈国刚

    2015-01-01

    The seed melon was used as the raw material in the experiment. The application of pectinase, cellulase to increase juice yield in the production of seed melon concentrated juice was studied in this paper through a single factor and an orthogonal experiment. The results showed that the optimum enzymatic hydrolysis processing were as followed: 0.1% pectinase, cellulase 0.05%, Enzyme solution temperature 45℃, enzymolysis time 2h and enzyme solution pH=4.5. On this condition, the yield of juice was 86.3%.%以籽瓜为试验材料,利用果胶酶和纤维素酶进行酶解。通过单因素和正交试验确定籽瓜的最佳酶解工艺:果胶酶0.1%,纤维素酶0.05%,酶解温度45℃,酶解时间2h,酶解pH=4.5。在此最佳酶解条件下,出汁率达86.3%。

  5. High-throughput screening of cellulase F mutants from multiplexed plasmid sets using an automated plate assay on a functional proteomic robotic workcell

    Directory of Open Access Journals (Sweden)

    Qureshi Nasib

    2006-05-01

    Full Text Available Abstract Background The field of plasmid-based functional proteomics requires the rapid assay of proteins expressed from plasmid libraries. Automation is essential since large sets of mutant open reading frames are being cloned for evaluation. To date no integrated automated platform is available to carry out the entire process including production of plasmid libraries, expression of cloned genes, and functional testing of expressed proteins. Results We used a functional proteomic assay in a multiplexed setting on an integrated plasmid-based robotic workcell for high-throughput screening of mutants of cellulase F, an endoglucanase from the anaerobic fungus Orpinomyces PC-2. This allowed us to identify plasmids containing optimized clones expressing mutants with improved activity at lower pH. A plasmid library of mutagenized clones of the celF gene with targeted variations in the last four codons was constructed by site-directed PCR mutagenesis and transformed into Escherichia coli. A robotic picker integrated into the workcell was used to inoculate medium in a 96-well deep well plate, combining the transformants into a multiplexed set in each well, and the plate was incubated on the workcell. Plasmids were prepared from the multiplexed culture on the liquid handler component of the workcell and used for in vitro transcription/translation. The multiplexed expressed recombinant proteins were screened for improved activity and stability in an azo-carboxymethylcellulose plate assay. The multiplexed wells containing mutants with improved activity were identified and linked back to the corresponding multiplexed cultures stored in glycerol. Spread plates were prepared from the glycerol stocks and the workcell was used to pick single colonies from the spread plates, prepare plasmid, produce recombinant protein, and assay for activity. The screening assay and subsequent deconvolution of the multiplexed wells resulted in identification of improved Cel

  6. Use of Cellulases to Predict in vivo Digestible Organic Matter (D value in Pasture Silages Uso de Celulasas para Predecir el Contenido de Materia Orgánica Digestible (Valor D in vivo, en Ensilajes de Praderas

    Directory of Open Access Journals (Sweden)

    Claudia Barchiesi-Ferrari

    2011-06-01

    Full Text Available In pasture-based dairy herds where silage is a widely adopted supplement, optimized feeding requires reliable estimations of nutritional quality of this conserved forage. Metabolizable energy, an important nutritional fraction, can be predicted from digestibility-related traits, such as the digestible organic matter contained in the dry matter (D-value. The aim of the present study was to evaluate the prediction of D-value and dry matter digestibility (DMD of grass silages made from four different pastures and maturity stages, using the pepsin-cellulase method. Fungal cellulase was used, applying different enzyme concentrations, incubation times and types of final wash. The silages were prepared from permanent pasture (Dactylis glomerata L., Lolium perenne L., Bromus catharticus Vahl var. catharticus, Trifolium repens L. and Holcus lanatus L., rotation pasture (Lolium multiflorum Lam. cv. Tama, oats (Avena sativa L., and mixed pasture (L. perenne-T. repens. These were harvested at three different physiological stages (vegetative, ear emergence and dough grain. The treatment using an incubation time of 24 h, a cellulase concentration of 6.25 g L-1 and final wash with water (Treatment 3 presented the best prediction capacity of the in vivo D-value (R² = 0.78 and in vivo DMD (R² = 0.71. In vivo D-value prediction improved (R² = 0.8 when a chemical determination (crude fibre, gross energy, neutral detergent fibre, total ash or acid detergent fibre was included in addition (multiple regression to D-value obtained with cellulases (Treatment 3. Results of DMD obtained with cellulases show good precision, but underestimate in vivo values, and are closer to those obtained with ruminal fluid. Suitable equations could be used to improve accuracy.En sistemas lecheros pastoriles que utilizan ensilaje como suplemento, se requiere conocer el valor nutricional de éste para optimizar la alimentación del ganado. La energía metabolizable, importante fracci

  7. Isolation of a Cotton Stalk Cellulose Decomposing Fungus and Its cellulase Properties%一株棉秆纤维素分解真菌的分离筛选及酶学性质研究

    Institute of Scientific and Technical Information of China (English)

    白宝伟; 张琴; 滕立平; 杨瑛; 李艳宾

    2012-01-01

    [Objective] The purpose of this project was to study cellulase properties of a cellulose decomposing fungus SJ - 1 isolated from cotton stalk decomposing products were studied. [Method] Effects of temperature, pH value and metal ions on cellulase activity were studied, and Michaelis constant Km of enzymatic reaction and the maximum reaction rate Vmax were examined by Lineweaver - Burk method. [ Result]The best reaction temperature to CMCase and FPase was ranged from 50 to 60℃, and the best pH value was 7. 0, which showed the tolerance to high temperature and alkali. K+ and Fe2+ activated the cellulase activity significantly, while Cu2 + , Mg2 + , Ca2+ and Al3 + showed certain inhibition, the inhibition of Al3+ exhibited much more strongly, Cu2+ inhibited FPase strongly as well, Mn2+ showed activation to FPase but inhibition to CMCase, Zn2 + had no influence on cellulase activity. When CMC - Na was taken as substrate, Km and Vmax of the enzymatic reaction were 2.69 mg/mL and 0.53 mg/( mL·min) respectively. [Conclusion] The strain showed good properties of cellulase activity, and provided references for further breeding and alteration of this fungus.%[目的]对从棉秆腐解物中筛选得到的高效棉秆分解真菌SJ-1进行纤维素酶酶学性质的研究.[方法]研究温度、pH、金属离子对纤维素酶活力的影响,并以Lineweaver -Burk作图法测定酶促反应米氏常数Km及最大反应速率Vmax.[结果]该菌株CMCase和FPase的最适反应温度在50~60℃,最适pH为7.0,有较好的耐高温及耐碱能力.K+、Fe2对酶活有显著的激活作用,而Cu2+、Mg2+、Ca2+、Al3+等有一定的抑制作用,其中Al3的抑制作用较为强烈,Cu2+对FPase抑制较强,Mn2-对FPase有激活作用而对CMCase有抑制作用,Zn2对酶活性无明显影响.以CMC - Na做底物时酶反应的Km为2.69 mg/mL,Vmax为0.53 mg/( mL·min).[结论]菌株SJ-1的纤维素酶性质较为优良,为进一步进行菌株的选育与改造提供参考依据.

  8. MnO2纳米粒子固载纤维素酶用于高效水解农业废弃物制备生物乙醇%Immobilization of cellulase onto MnO2 nanoparticles for bioethanol production by enhanced hydrolysis of agricultural waste

    Institute of Scientific and Technical Information of China (English)

    Elsa Cherian; Mahendradas Dharmendirakumar; Gurunathan Baskar

    2015-01-01

    Cellulase is an efficient enzymatic catalyst that hydrolyses cellulosic substances. The high costs associated with using enzymes for industrial applications can be reduced by immobilizing the cellu-lase. In the current study, cellulase produced by Aspergillus fumigatus JCF was immobilized onto MnO2 nanoparticles, which improve the activity of cellulase and offer a superior support. The sur-face characteristics of synthesized MnO2 nanoparticles and cellulase-bound MnO2 nanoparticles were investigated by scanning electron microscopy, and Fourier transform infrared spectroscopy was used to analyze the functional characteristics of the immobilized cellulase. The maximum cellu-lase binding efficiency was 75%. The properties of the immobilized cellulase, including activity, operational pH, temperature, thermal stability, and reusability were investigated and were found to be more stable than for the free enzyme. It was found that cellulase immobilized on MnO2 nanopar-ticles could be used to hydrolyze cellulosic substances over a broad range of temperature and pH. The results confirmed that cellulase immobilized on MnO2 nanoparticles was very efficient in terms of cellulolytic activity.%纤维素酶是一种有效的纤维质类物质水解催化剂,工业应用时可通过固定化纤维素酶来降低其成本。本文将烟曲霉原变种JCF产生的纤维素酶固定在MnO2纳米颗粒上。 MnO2可提高纤维素酶的活性,并充当一个更好的载体。采用扫描电镜表征了所制MnO2纳米粒子及其负载纤维素酶的表面性质,以傅里叶变换红外光谱分析了固定在MnO2纳米粒子上纤维素酶的官能团性质。纤维素酶在MnO2纳米粒子上最大的固定化效率为75%。考察了固定化纤维素酶的活性、操作pH值、温度、热稳定性和重复使用性等性质。结果表明,所制固定化酶的稳定性比游离酶更高。固定于MnO2纳米粒子上的纤维素酶可用于纤维质类物质的水解反

  9. 纤维素酶降解秸秆特性及其基因工程研究进展%Advances in Research of Straw Degradation with Cellulase and Its Genetic Engineering

    Institute of Scientific and Technical Information of China (English)

    张森翔; 尹小燕; 龚志伟; 杨忠华; 侯亚利; 周卫

    2015-01-01

    Energy shortage and environmental pollution have become the public focusing issue. Straw biomass with its rich resources, non-polluting and renewable feature, has great application prospect in solving the energy crisis. Converting straw biomass to fermentable sugars by hydrolysis with cellulase and combing with fermentation may produce ethanol, hydrogen and other materials of energy, which has been a mature technology route. The crucial steps of utilizing straw biomass are the pretreatment of straw biomass and efficient obtaining of glycoside hydrolases. We summarize and analyze the current research from 3 aspects: the structural characteristics of straw and its biological pretreatment; the mechanism of cellulase in hydrolysis of straw biomass; and the gene engineering for cellulase. It has a guiding significance in the promotion of applying straw biomass for energy.%能源短缺和环境污染问题是人们关注的焦点.秸秆类生物质以其资源丰富、无污染及可再生等特性使其在解决能源危机方面具有极大应用前景.对秸秆类生物质通过纤维素酶的水解转化为可发酵性的糖,再结合发酵技术可进一步生产乙醇、氢气等能源物质,是一条成熟的能源化技术路线.其关键是秸秆生物质的预处理与高效的糖苷水解酶获得.将从对秸秆类生物质的预处理、纤维素酶的作用机理研究和纤维素酶基因工程3个方面对当前的研究进展进行综述与分析.这对于促进秸秆类生物质能源化应用具有指导意义.

  10. Effects of acids, pH and temperature on acid cellulase activity%酸的种类、pH和温度对酸性纤维素酶活力的影响

    Institute of Scientific and Technical Information of China (English)

    刘幸乐; 姚继明; 吴远明

    2012-01-01

    以8种酸(甲酸、冰醋酸、羟基乙酸、乳酸、柠檬酸、酒石酸、氨基乙酸和氨基磺酸)配制成pH=5.0的缓冲液,测定酸性纤维素酶在各缓冲液中的活力.同时以冰醋酸作为pH调节剂,采用中心合成设计法,分析和优化pH和温度对酶活力的影响,得出线性回归方程和优化值.结果表明,在pH=5.0的条件下,乳酸和酒石酸为酸剂的酶活力高于冰醋酸,羟基乙酸、甲酸和柠檬酸比冰醋酸略低,而氨基乙酸和氨基磺酸明显低于冰醋酸;采用冰醋酸调节pH,酶活力随着温度的升高和pH的降低而增加,pH影响的显著性要大于温度.酸性纤维素酶在49.8℃,pH=4.8可以实现最佳的活力.%Eight kinds of acids (formic acid, acetic acid, glycoiic acid, lactic acid, citric acid, tartaric acid; glycine and aminosulphonic acid) were used as buffer solutions with Ph=5.0. The acid cellulase activities were measured individually in these buffer solutions. Meanwhile using acetic acid as Ph regulator, the effects of Ph and temperature on cellulose activity were analyzed and optimized by central composite design to obtain linear regression equatian and optimal factors. The results showed that tartaric acid and lactic acid exhibited higher activity than acetic acid at Ph 5.0, glycoiic acid, formic acid and citric acid gave little lower activity compared with acetic acid, while the glycine, aminosulphonic acid gave much lower cellulase activity than acetic acid. Applying acetic acid to adjust Ph, cellulase activity increased with the rising of temperature and the decline of Ph. Ph showed more significance than temperature, the optimum performance of activity was at 49.8 ℃,Ph= 4.8.

  11. Molecular Identification and Characterization of a Thermostable Cellulase-Secreting Strain%产耐热纤维素酶菌株的分子鉴定及产酶条件优化

    Institute of Scientific and Technical Information of China (English)

    陈红漫; 孙玉辉; 阚国仕; 任大明; 杨佳颖; 蔡金涛

    2011-01-01

    A thermophilic strain NR615 possessing cellulase was isolated at 60℃ from hot spring samples. Based on the ITS sequence and phylogenetic tree analysis, strain NR615 was identified as Aspergillusfumigatus. The strain was capable of growing well at 28 - 60 ℃. Aeeording to optimization condition of cell growth and enzyme production, the initial pH is 6. 5. The supernatant of strain incubated medium showed efficient activity of cellulase with addition of crystalline cellulose as carbon and beef extract as nitrogen sources respectively. The highest enzyme activity at 2.41 IU/mL was reached after incubation for 5d. The optimal pH of the celullase for the reaction is 5.0. This enzyme shows stable aetivity over a broad range of temperature from 50 ℃ to 70 ℃. These characteristics of the eellulase demonstrated that it has potential application in the thermostable cellulase field.%从温泉热源地区采集的水样中筛选出1株60℃生长的纤维素酶产生菌NR615。通过ITS序列及系统发育进化树的分析表明该菌株为烟曲霉(Aspergillus fumigatus)。该菌株在28~60℃生长较好.对菌体及产酶条件进行优化得出:初始pH6.5,碳、氮源分别为结晶纤维素、牛肉膏时有利于产酶。其产生的纤维素酶的最适反应温度65℃,最适pH5.0,且在50~70℃有一定稳定的活性。经过响应面分析优化培养基,发酵5d时活力达到2.4113IU/mL。这些特征表明,该菌株是1株性能良好的耐热纤维素酶生产菌株。

  12. Atividade da celulase e β-galactosidase no estudo da firmeza da polpa de mamões 'golden' e 'gran golden' Cellulase and β-galactosidase activities in 'golden' and 'gran golden' papaya softening

    Directory of Open Access Journals (Sweden)

    Camilla Zanotti Gallon

    2009-12-01

    Full Text Available O objetivo desse trabalho foi avaliar a ação das enzimas celulase e β-galactosidase em relação à perda de firmeza dessas cultivares de mamões 'Gran Golden' e 'Golden' devido a relatos de uma perda de firmeza diferenciada entre as cvs. Os frutos foram armazenados a 25ºC e analisados diariamente quanto à firmeza da polpa e à atividade enzimática da celulase e β-galactosidase durante 8 dias. Os resultados de firmeza da polpa e atividade enzimática foram submetidos às análises de correlação e regressão. No 4º dia pós-colheita os mamões 'Golden' apresentaram firmeza média de 60,6 N e os 'Gran Golden' 31,1 N e a um aumento da atividade da celulase e da β-galactosidase. Os dados gerados neste trabalho sugerem que as enzimas celulase e β-galactosidase atuam diferentemente no processo de perda de firmeza dos frutos das cultivares Goldene Gran Golden. Aantecipaçãonaperdade firmezade 'Gran Golden' pode estar relacionada com a maior atividade dessas enzimas.It has been reported by orchards from the north of Espírito Santo state that 'Gran Golden' papaya loses firmness faster than 'Golden'. The goal of this work was to evaluate the action of cellulase and β-galactosidase related to the softening on papaya. The fruits have been stored at 25ºC and firmness and enzymes activities were daily analyzed during 8 days. The results were submitted to correlation and regression analysis. The activity of cellulase and β-galactosidase had increased for both cultivars. The 4th postharvest day showed that 'Golden' firmness was 6.18 while 'Gran Golden' was 31.1 N. Fruit softening in 'Gran Golden' was intense and the fruit was very soft at ripe stage. These works show that hydrolytic enzymes cellulase and β-galactosidase act differently in the softening process in 'Golden'and 'Gran Golden'papaya. The flesh firmness on 'Gran Golden' is related to the increased activity of these enzymes. These results can help to choose which cultivar to produce

  13. Engineering Cellulase Enzymes for Bioenergy

    OpenAIRE

    Atreya, Meera Elizabeth

    2015-01-01

    Sustainable energy sources, such as biofuels, offer increasingly important alternatives to fossil fuels that contribute less to global climate change. The energy contained within cellulosic biofuels derives from sunlight energy stored in the form of carbon-carbon bonds comprising sugars such as glucose. Second-generation biofuels are produced from lignocellulosic biomass feedstocks, including agricultural waste products and non-food crops like Miscanthus, that contain lignin and the polysacch...

  14. Optimization of Brown Rice Germination Process with Cellulase Treatment%纤维素酶预处理糙米发芽工艺优化

    Institute of Scientific and Technical Information of China (English)

    张强; 贾富国; 杨瑞雪; 付倩; 王吉泰; 韩珊

    2012-01-01

    In order to improve the taste of the cooked germinated brown rice, the research of the pre - treating technology for cellulose - immersing brown rice was explored to provide conditions for germination and appropriate degradation of the cortex crude fiber. Influences of the enzyme concentration, the enzyme temperature and enzymolys-is time on the germinating rate of brown rice, and the variation of hardness were studied by quadratic regression rotary combination experiments. The GAB A content of germinated brown rice was taken as the index for evaluation and the comparative test was carried out with traditional process of soaking and germination. The results showed that experimental factors had significant effects on the germinating rate of brown rice and hardness of germinated brown rice. And the optimum parameter combination in the germination rate of germinated brown rice was that the enzyme concentration being 0. 4 mg/mL, enzyme temperature being 33 ℃ , and enzyme time being 110 min. Under such conditions , the germination rate of brown rice can reach up to higher than 90% of that soaking in a traditional way, and the hardness of germinated brown rice can be reduced by 14. 1%. The GAB A content in germinated brown rice by the cellulase pretreatment is a little lower than that by traditional immersion. The cortex crude fiber degradation of germinated brown rice proved to be the reason for the decline of hardness with scanning electron microscope analysis.%为解决发芽糙米蒸煮后口感差的问题,提出酶溶液浸泡糙米提供发芽条件的同时适当降解皮层粗纤维预处理工艺.研究酶浓度、酶解温度以及酶解时间对糙米发芽率及发芽糙米硬度的影响规律,采用二次旋转组合试验方法设计试验.并以GABA含量为考核指标,将酶预处理工艺与传统浸泡工艺进行了对比试验.结果表明:试验因素对糙米发芽率及发芽糙米硬度变化影响显著;酶预处理工艺优化参

  15. Study on the Conditions for Producing Cellulase by Liquid Fermentation with Aspergillus niger HQ-1%黑曲霉HQ-1液体发酵产纤维素酶条件研究

    Institute of Scientific and Technical Information of China (English)

    张辉; 桑青

    2009-01-01

    [Objective] The study aimed to discuss the optimum condition for producing cellulase by liquid fermentation of Aspergillus niger HQ-1. [Method] With 1 strain of A. niger HQ-1 with higher activity of producing cellulase as the original srtain, the optimum conditions for liquid fermentation of A. niger HQ-1 to produce cellulase was studied primarily through the single factor experiments and orthogonal tests and the activities of CMCase and FPAase were measured. [Result] The factors influencing the production of CMCase and FPAase by liquid fermentation of A, niger HQ-1 were in order were the content of compound carbon origin, the ratio of 2 kinds of carbon, the content of nitrogen and the liquid volume in flask. The optimal medium of producing CMCase was as follows: 15‰ bran+corn stalk powder (1∶1), 2.0‰ NH4Cl, other components as same as Mandel's medium. The optimal medium of producing FPAase was as follows: 20‰ bran+corn stalk powder (1∶1), 1.5‰ NH4Cl, others were as same as Mandel's medium. For both medium, the optimum pH was 5.0 and medium volume was 50 ml/250 ml triangle flask. The activities of CMCase and FPAase reached 525.2 and 217.6 IU respectively after culturing for 4 d under 30 ℃ and 170 r/min. [Conclusion] This study provided some technique reference for the research and development of producing cellulase strain.%[目的] 探讨黑曲霉HQ-1液体发酵产纤维素酶的最适条件.[方法] 以1株产酶活力相对较高的黑曲霉HQ-1为出发菌株,采用单因子试验和正交试验对黑曲霉HQ-1液体发酵产纤维素酶的最适条件进行初步研究,并测定了CMC酶(CMCase)和滤纸酶(FPAase)的活力.[结果] 影响黑曲霉HQ-1液体发酵产CMCase和FPAase的因素依次是复合碳源的含量、2种碳源的比例、氮源含量和装液量.该菌产CMCase的最适培养基为:麸皮+玉米秸秆粉15‰(1∶1),NH4Cl 2.0‰,其他成分同Mandel's营养液;产FPAase的最适培养基为:麸皮+玉米秸秆粉20‰(1

  16. An Applied Research on Cellulase Pre-treatment in Anaerobic Fermentation of Municipal Solid Waste%城市生活垃圾厌氧发酵中纤维素酶预处理的应用研究

    Institute of Scientific and Technical Information of China (English)

    何娟; 孙可伟; 李建昌; 尚江涛

    2011-01-01

    为解决常规厌氧发酵存在发酵速率滞后,产气量、产甲烷量低等缺点,首先利用纤维素酶对城市生活垃圾进行预处理(水解),通过单因素实验研究了纤维素酶添加量、水解温度、水解时间、水解pH和不同底物浓度(VS)对生活垃圾中纤维素水解率的影响。实验结果表明,利用纤维素酶水解城市生活垃圾的适宜条件为:纤维素酶添加量120 U/g(VS)、水解时间24 h、水解温度60℃、底物浓度为8%、水解pH5.6,此时水解率可达35.2%。随后将生活垃圾在此最佳水解条件下经过纤维素酶预处理后用于中温厌氧发酵。结果表明,经过纤维素酶预处理后用于厌氧发酵与不经过纤维素酶预处理直接进行厌氧发酵相比较,平均日产气率、VS产气量、VS产气率、平均产甲烷率、VS产甲烷量、VS产甲烷率等均显著提高;且累积产气量提高62.38%,累积产甲烷量提高87.94%。因此,采用纤维素酶预处理城市生活垃圾后应用于中温厌氧发酵,对解决目前常规厌氧发酵存在的主要问题是可行的。%To resolve the defects in conventional anaerobic fermentation process such as fermenting rate lag,low biogas production rate and so on,municipal solid waste(MSW) was pre-treated with cellulase(as hydrolysis).By using single-factor experiment technique,the effects of different cellulase doses,hydrolysis temperature and duration,hydrolysis pH level and substrate concentration(volatile solide,VS) were tested on the degree of hydrolysis for cellulose in MSW.It was indicated that optimal conditions of the MSW pre-treatment were cellulase dosage 120 units per gram of VS,hydrolysis duration 24 hours,hydrolysis temperature 60℃,substrate concentration 8%and pH value 5.6 whilst the degree of hydrolysis could reach 35.2%.Such pre-treated MSW was then taken to carry on anaerobic fermentation at moderate temperature.The results have shown that the cellulase pre

  17. Kinetics of Cellulase Based on the Model of Second-order Enzymatic Deactivation%基于二级酶失活模型的纤维素酶反应动力学

    Institute of Scientific and Technical Information of China (English)

    张宇; 许敬亮; 余强; 袁振宏; 刘云云; 亓伟

    2011-01-01

    酶催化、失活机制的模糊以及影响异相体系因素的大量存在,使得纤维素水解的酶催化过程高度复杂,很难为之建立机理模型.假定纤维素酶失活模型为二级反应,由准稳态理论推导出最终产物葡萄糖浓度与时间,初始酶浓度关系的半经验半理论模型.该模型只含两个参数,能对试验数据很好的拟合,相关系数R2均在0.98以上.用Bailey改进的米式方程进一步拟合不同初始酶浓度下的初速度关系,相关系数R2=0.977 3,求得最大反应速度为2.7425 g/(L·h),半饱和常数为3.010 6 g/L.该模型还表明酶失活速率常数随着初始酶浓度的增加而减小,呈线性关系.失活速度随着初始酶浓度的增加而增加.%Enzymatic hydrolysis of cellulose was extremely complex because of the unclear enzymatic hydrolysis , deactivation mechanisms and many factors that affect the heterogeneous system. Therefore, it is difficult to build a mechanistic model to study cellulose hydrolysis by cellulase. Under some assumed conditions such as the second-order cellulase deactivation and quasi-steady state theory, a semi mechanistic and empirical model describing the relationship between product concentration and time, the initial enzyme concentration was deduced. The mathematic model was a simple mathematic function that contained only two parameters. The experimental result was in accordance with the deduced mathematic model, where the correlation coefficients (R2) were above 0. 98. The relationship between initial enzyme concentration and initial hydrolysis rate calculated from the mathematic model showed good agreement with another type of Henri-Michaelis-Menten equation proposed by Bailey, where the R2 was 0. 977 3. The maximum hydrolysis rate and half saturation constant was 2. 742 4 g/ (L · h) and 3. 013 0 g/L, respectively. It was shown from the model that the rate constant of cellulase deactivation decreased when initial enzyme concentration increased

  18. Cellulase production by two mutant strain of Trichoderma longibranchiatum QM9414 and Rut C30; Produccion de celulasas a partir de dos cepas hiperproductoras de trichoderma longibranchiatum Qm9-41 4 y Rut C30

    Energy Technology Data Exchange (ETDEWEB)

    Blanco, M. J.

    1991-07-01

    Native or pretreated biomass from Onopordum nervosum Boiss, has been examined as candidate feedstock for cellulase production by two mutant strain of Trichoderma Ionqibrachiatum QM9414 and Rut C30. Batch cultivation methods were evaluated and compared with previous experiments using ball-milled, crystalline cellulose (Solka floc). Batch cultivation of T. Ionqibrachiatum Rut C30 on 5% (w/v) acid pretreated O. nervosum biomass yielded enzyme productivities and activities comparable to those obtained on Solka floc. However, the overall enzyme production performance was lower than on Solka floc at comparable cellulose concentrations. This fact may be due to the accumulation of pretreated by products and lignin in the ferment. (Author) 40 refs.

  19. Study on Optimal Conditions of Degradation of Wheat Straw by Cellulase and Analysis of Kinetics%纤维素酶降解小麦秸秆最适条件的研究及其动力学分析

    Institute of Scientific and Technical Information of China (English)

    田萍; 王浩菊; 马齐; 陈坤奇; 周婷

    2012-01-01

    以小麦秸秆为原料,通过正交实验对纤维素酶降解秸秆纤维的影响因素进行了研究.结果表明,影响小麦秸秆降解的因素依次为:酶量>酶解时间>料液比>反应温度,其最适条件是:加酶量为40 u/g,酶解时间为10h,反应温度为40℃,料液比为1∶3,总糖含量达到43.24%.以米氏方程为基础,建立起最适酶解条件下总纤维素降解的动力学模型.%The influence factors of degradation of straw fiber by cellulase were studied using wheat straw as the raw material through the orthogonal experiments. The results showed that the factors of influence on enzymatic hydrolysis of wheat straw were successively: the amount of enzyme>the time of hydrolysis>the ratio of material and liquid> reaction temperature. The optimal conditions of enzymatic hydrolysis were that the amount of cellulase was 40 u/g, the time of hydrolysis was 10 h, the reaction temperature was 40 C and the ratio of material to liquid was 1:3. The total sugar contents could reach 43. 24%. Under the optimal conditions, the kinetics model of degradation of the total cellulose was established based on Michaelis-Menten equation.

  20. 短小芽孢杆菌产碱性纤维素酶的发酵工艺优化%Optimization of Fermentation Process of Bacillus pumilus Producing Alkalescence Cellulase

    Institute of Scientific and Technical Information of China (English)

    黄谚谚; 吴华珠; 许旭萍

    2012-01-01

    With the purpose of getting the optimum fermentation condition of alkalescence cellulase produced by B. Pumilus SI 2, the fermentation condition was optimized through the single factor and orthogonal design. The results showed that the bran and CMC-Na were the suitable carbon source and the best nitrogen source was yeast. The enzyme producing rate of organic nitrogen sources was higher than that of inorganic nitrogen sources. The optimal conditions were as follows: initial pH 8.5, culture temperature 32℃,70 mL media filled in the 250 mL triangle flask, the inoculation amount 3%(v/v), fermentation time 20 h. The alkaline cellulase activity reached 367. 9 g/mL under the optimal condition.%为了确定短小芽孢杆菌(Bacillus pumilus S12)产碱性纤维素酶的最适发酵条件,采用液体摇瓶的发酵方法,进行最适发酵条件的单因素试验和正交试验.结果表明,Bacillus pumilus S12菌株合成纤维素酶的最适碳源为CMC和麸皮,最适氮源为酵母膏,有机氮比无机氮更有利于产酶;最适产酶条件为起始pH8.5,培养温度32℃,装液量为250mL三角瓶装70mL培养液,接种量为3%,在此条件下发酵20h,酶活力可达367.9 μg/mL.

  1. Optimization of Solid State Fermentation(SSF) Conditions with Orange Peels for Production of Cellulase by Rhizopus oryzae%米根霉固态发酵橘皮产纤维素酶工艺的优化

    Institute of Scientific and Technical Information of China (English)

    张帅; 董基; 吴紫俊

    2012-01-01

    With orange peels as raw materials and Rhizopus oryzae as production strains, Cellulase was producted by solid state fermentation(SSF). Three important factors of water content of medium, inoculums size and culture time on cellulose activities were researched respectively by single factor test. Based on it, the enzyme production conditions were optimized by Box-Behnken Design, regression and analysis of variance of experimental data was made by Design Expert. The optimum conditions of enzyme production was determined finally, water content of SSF medium was 12.24 mL, inoculum size was 10.76%, culture time was 72.64 h. Activity of cellulase producted on the above conditions were 464.33 U/g.%以橘皮为原料,以米根霉为生产菌株,采用固态发酵法生产纤维素酶。通过单因子试验分别考察了发酵培养基水分含量、接种量及培养时间三个重要因子对纤维素酶活力的影响,在此基础上,采用Box-Behnken设计对产酶工艺进行优化,利用Design Expert软件对试验数据进行回归拟合和方差分析,最终确定产酶最优工艺条件为:发酵培养基水分含量12.24 mL,接种量10.76%,培养时间72.64 h,在最优条件下所得纤维素酶的酶活力为464.33 U/g。

  2. Hidrólise enzimática de casca de arroz utilizando-se celulases: efeito de tratamentos químicos e fotoquímicos Enzymatic hydrolysis of rice hull using cellulases: effect of chemical and photochemical treatments

    OpenAIRE

    Juan Reyes; Patricio Peralta-Zamora; Nelson Durán

    1998-01-01

    In the present work we reported the study of rice hull enzymatic hydrolysis using a commercial cellulase preparation. The results showed that previous treatment with light and sodium chlorite inhibits the enzymatic process (31.4 and 11.8%, respectively) while hydrogen peroxide and ozone favoured the enzymatic production of reducing sugars (5.9 and 54.9%, respectively). Studies performed by quimiluminescence showed that the chlorite treatment produced the most significant change in the structu...

  3. 绿液预处理玉米秸秆产纤维素酶的研究%The Research on Cellulase Production of Green Liquor Pretreated Corn Stover

    Institute of Scientific and Technical Information of China (English)

    赵士明; 陆青山; 谷峰; 余世袁

    2012-01-01

    Abstract:The effect of green liquid pretreated corn stover on cellulase production of Trichoderma reesei was investigated. Inspecting the influence of pretreatment conditions on cellulase production of corn stover at different cooking temperature, total alkali charge and sulfidity by orthogonal experiment, we could conclude from range and variance analysis that the extent of impact to cellucase production of green liquor pretreated corn stover was in turn total alkali charge, cooking temperature and sultidity. The optimum filter paper activity (FPA) reached 2.6 IU/mL. FPA productions from corn stover oretreated bv cooking temperature140℃ and 170 ℃, total alkali charge 4 %, sulfidity 0, 20 %, 30 % and 40 %, were comprehensively compared and evaluated. It was found that the optimum conditions, i. e. , cooking temperature 140℃, total alkali charge 4 %, sulfidity 0, might ensure the value of FPA on and avoid the loss of materials. The results showed that green liquid pretreatment for corn stoverwas feasible for cellulase production of Trichoderma reesei. Taking corn stover pretreated by cooking temperature 140 ℃, total alkali charge 4% and sulfidity 0 as carbon sources whose concentration was 12 g/L, FPA and β-glucosidase activity(β-GA) were up to 2.5 IU/mL and 1.3 IU/mL respectively. The enzyme production period was 5 days.%研究了绿液预处理玉米秸秆对里氏木霉产纤维素酶的影响。通过正交试验考察3个预处理条件对玉米秸秆产纤维素酶的影响,从极差和方差分析可知,对绿液预处理玉米秸秆产纤维素酶的影响程度由大到小依次是总碱量、蒸煮温度、硫化度,最大滤纸酶活(FPA)达到2.6 IU/mL。比较在蒸煮温度140℃和170℃,总碱量4%,硫化度0、20%、30%与40%下所产的FPA,经综合评定,在最优条件下蒸煮温度140℃,总碱量4%和硫化度0时可以尽量避免原料损失的前提下,保证FPA的大小,结果表明,里氏木霉利用绿液预处

  4. 纤维素酶法提取竹叶黄酮的传质动力学%Mass Transfer Kinetics of Flavonoids Extraction from Baantant Leaves with Cellulase-Assisted Aqueous Extractant

    Institute of Scientific and Technical Information of China (English)

    魏凤玉; 陈玮; 方菊; 宝呼和

    2013-01-01

    采用纤维素酶法提取竹叶中的总黄酮类化合物,讨论了温度、原料尺寸、转速及纤维素酶用量对传质速率的影响,并用扫描电镜观察了水提和酶提取后样品的细胞结构变化。结果表明,竹叶黄酮的提取动力学符合平板型Fick 第二定律,提取过程主要受内扩散控制;纤维素酶不改变传质动力学机理,它只破坏细胞壁,从而降低扩散阻力使溶质的传质速率和表观扩散系数D′提高。实验还得到了D′与提取温度、竹叶几何尺寸之间的关系,在30~60℃,酶提时的 D′为1.95×10-13~6.85×10-13 m2×s-1,水提时的 D′为1.73×10-13~6.05×10-13 m2×s-1;酶提时的活化能 Ea 为11.853 kJ×mol-1,比水提时降低了9.59%。该研究为酶法提取竹叶黄酮的工艺设计及工业化应用提供了理论依据。%In this study, cellulase-assisted aqueous extractant (CAE) and aqueous extractant (AE) were adopted separatively for the extraction of the total flavonoid compounds from bamboo leaves. The influence of temperature, raw material size, rotation speed of stirrer and cellulase dosage on the mass transfer rates of the flavonoid compounds was examined. In addition, the micro-structural changes of bamboo leaf cells after the CAE or AE process were observed by scanning electron microscopy (SEM). Results show that the Fick’s second law fits well with the experimental data and the extraction rate is controlled mainly by the inner diffusion process. The presence of cellulase has nearly no influence on the mass transfer mechanism, and can only damage the cell walls and reduce the resistance of diffusion. The mass transfer rate and apparent diffusion constant D′ are increased accordingly. The relationship of D′with the extraction temperature and the bamboo leaf size was deduced. D′ varies from 1.95×10-13 to 6.85×10-13 m2×s-1 for CAE and from 1.73×10-13 to 6.05×10-13 m2×s-1 for AE in a temperature range between 30

  5. 绿色木霉纤维素酶AS3.3032固态发酵的研究%Production of cellulase by Trichoderma viride AS3.3032 in sloid state fermentation

    Institute of Scientific and Technical Information of China (English)

    张德强; 黄镇亚; 张志毅

    2001-01-01

    In the research, using wheat bran and steam exploded bagasse asmain substrate, cellulase is produced by solid state fermentation with Trichoderma viride AS3.3032. The effect on cellulase production is studied of nitrogen source, carbon source, surface active agent, inoculation, temperature, wate content and initial pH value of solid substrate. The results are:①With NH4SO4 as nitrogen source, FPA,CMC and β-gase all are high, 122.5 FPA U/g dried substrate, 1470.0 CMC U/g dried substrate and 119.3 β-Gase U/g dried substrate respectively;②With wheat bran and bigasse(their ratio 3 to 2) as carbon source, FPA, CMC and β-Gase get highest, 138.2 FPA U/g dried substrate, 134.6 β-Gase U/g dried substrate and 1603.1 CMC U/g dried substrate respectively;③Adding 0.1% Tween-80 to culture medium, FPA, β-Gase and CMC can increase by 230%,280% and 230% respectively, but with addition of 0.5%~0.7% washing powder, the three above indices increase by 310%,370%,300% resepctively;④The activity of produced cellulase goes up to the highest when the water amount of medium, the temperature and the initial pH are 250%,28℃ and 3.5.%该研究以麦麸和汽爆蔗渣为主要原料,采用绿色木霉AS3.3032(Trichodermaviride)固态发酵生产纤维素酶,研究了氮源、碳源、表面活性剂、接种方式、培养基含水量、培养温度、培养基起始pH值对绿色木霉产酶活力的影响.研究结果表明:①以硫酸铵为氮源,其FPA,CMC,和β-Gase酶活力均较高,每克干曲分别高达122.5FPAU/g,1470.0CMCU/g和119.3β-GaseU/g;②碳源以麸蔗比为3∶2时,FPA,β-Gase和CMC酶活力均为最高,每克干曲分别高达138.2FPAU/g,134.6β-GaseU/g和1603.1CMCU/g;③添加0.1%的Tween-80和0.5%~0.7%的洗衣粉可分别提高FPA,β-Gase和CMC为2.3倍、2.8倍、2.3倍和3.1倍、3.7倍、3.0倍;④培养基含水量、培养温度、培养起始pH值分别为250%,28℃和pH3.5,产酶活力最高.

  6. Heterologously expressed Aspergillus aculeatus β-glucosidase in Saccharomyces cerevisiae is a cost-effective alternative to commercial supplementation of β-glucosidase in industrial ethanol production using Trichoderma reesei cellulases.

    Science.gov (United States)

    Treebupachatsakul, Treesukon; Nakazawa, Hikaru; Shinbo, Hideaki; Fujikawa, Hiroki; Nagaiwa, Asami; Ochiai, Nobuhiro; Kawaguchi, Takashi; Nikaido, Mitsuru; Totani, Kazuhide; Shioya, Koki; Shida, Yosuke; Morikawa, Yasushi; Ogasawara, Wataru; Okada, Hirofumi

    2016-01-01

    Trichoderma reesei is a filamentous organism that secretes enzymes capable of degrading cellulose to cellobiose. The culture supernatant of T. reesei, however, lacks sufficient activity to convert cellobiose to glucose using β-glucosidase (BGL1). In this study, we identified a BGL (Cel3B) from T. reesei (TrCel3B) and compared it with the active β-glucosidases from Aspergillus aculeatus (AaBGL1). AaBGL1 showed higher stability and conversion of sugars to ethanol compared to TrCel3B, and therefore we chose to express this recombinant protein for use in fermentation processes. We expressed the recombinant protein in the yeast Saccharomyces cerevisiae, combined it with the superb T. reesei cellulase machinery and used the combination in a simultaneous saccharification and fermentation (SSF) process, with the hope that the recombinant would supplement the BGL activity. As the sugars were processed, the yeast immediately converted them to ethanol, thereby eliminating the problem posed by end product inhibition. Recombinant AaBGL1 activity was compared with Novozyme 188, a commercially available supplement for BGL activity. Our results show that the recombinant protein is as effective as the commercial supplement and can process sugars with equal efficiency. Expression of AaBGL1 in S. cerevisiae increased ethanol production effectively. Thus, heterologous expression of AaBGL1 in S. cerevisiae is a cost-effective and efficient process for the bioconversion of ethanol from lignocellulosic biomass.

  7. Development of malfunction diagnosis system for fermentation based on fuzzy inference and its application to industrial scale production of alkaline cellulase; Fuzzy suiron ni motozuku baiyo ijo shindan shisutemu no kaihatsu to arukarisei seruraze kogyo seisan eno tekiyo

    Energy Technology Data Exchange (ETDEWEB)

    Tanaka, T. [Kao Corp., Tokyo (Japan). Processing Development Research Lab.; Osaka Univ., Osaka (Japan). Graduate School of Engineering Science; Takeuchi, K. [Kao Corp., Tokyo (Japan). Processing Development Research Lab.; Taya, M. [Osaka Univ., Osaka (Japan). Graduate School of Engineering Science

    2000-09-10

    A malfunction diagnosis system based on fuzzy inference was developed for industrial scale fermentation with an alkaliphilic cellulase-producing bacterium. The index J indicating the degree of malfunction in the fermentation process is calculated from the deviations of actual values from the standard values with respect to three kinds of process data obtained from on-line sensors. The diagnosis system proposed in this study is as follows. The fermentation process is automatically categorized into one of three states (normal, intermediate and abnormal) according to computation of the J value. Only when the fermentation process is categorized to an 'intermediate' state, an operator should supervise the process by means of direct observation of process data. The diagnosis system was applied to 100 runs of batch fermentations conducted on an industrial scale. In all cases, it was confirmed that this system worked to offer the correct diagnosis results. The introduction of the system resulted in the reduction of over 99 % in the operators' diagnosing labor because their supervising tasks could be limited to the intermediate cases. (author)

  8. Research on the process of extraction pectin from orange peel using the cellulase%纤维素酶提取柑桔皮果胶工艺条件的研究

    Institute of Scientific and Technical Information of China (English)

    戴余军; 丁文; 石会军

    2011-01-01

    The research aims to investigate the best conditions for extracting pectin from orange peel. Taking orange peel as material, pectin was extracted from it using the cellulase. Results show that the maximum extracting rate of pectin could be obtained under the following conditions: the concentration of cellulose solution is 0. 5 %, the stirring time is 45 minutes, the temperature is 45 ℃, pH value is 5.6. The extracting rate of pectin is 36.56 %.%为了探索提取果胶的最佳试验条件,以柑桔皮为试验原料,采用纤维素酶法提取果胶,结果表明:在纤维素酶溶液浓度为0.5%,浸提时间为45 min,浸提温度为45℃,浸提pH值为5.6的条件下,果胶的得率最大,果胶提取率达到36.56%.

  9. EndB, a Multidomain Family 44 Cellulase from Ruminococcus flavefaciens 17, Binds to Cellulose via a Novel Cellulose-Binding Module and to Another R. flavefaciens Protein via a Dockerin Domain

    Science.gov (United States)

    Rincón, Marco T.; McCrae, Sheila I.; Kirby, James; Scott, Karen P.; Flint, Harry J.

    2001-01-01

    The mechanisms by which cellulolytic enzymes and enzyme complexes in Ruminococcus spp. bind to cellulose are not fully understood. The product of the newly isolated cellulase gene endB from Ruminococcus flavefaciens 17 was purified as a His-tagged product after expression in Escherichia coli and found to be able to bind directly to crystalline cellulose. The ability to bind cellulose is shown to be associated with a novel cellulose-binding module (CBM) located within a region of 200 amino acids that is unrelated to known protein sequences. EndB (808 amino acids) also contains a catalytic domain belonging to glycoside hydrolase family 44 and a C-terminal dockerin-like domain. Purified EndB is also shown to bind specifically via its dockerin domain to a polypeptide of ca. 130 kDa present among supernatant proteins from Avicel-grown R. flavefaciens that attach to cellulose. The protein to which EndB attaches is a strong candidate for the scaffolding component of a cellulosome-like multienzyme complex recently identified in this species (S.-Y. Ding et al., J. Bacteriol. 183:1945–1953, 2001). It is concluded that binding of EndB to cellulose may occur both through its own CBM and potentially also through its involvement in a cellulosome complex. PMID:11571138

  10. Analysis of a Commensal Strain Producing Cellulase from Zingiber officinale Rhizosphere%生姜根际一株纤维素降解共生菌的分析

    Institute of Scientific and Technical Information of China (English)

    侯进慧; 王富威; 樊继强

    2012-01-01

    A Zingiber officinale rhizosphere strain SP1 with higher cellulose-decomposing activity was preliminarily analyzed. With PCR, a 1 406 bp 16S rDNA fragment of SP.I was amplified. The method of table concentrator fermentation was used to preliminarily analyze the characteristics of SP1 cellulose. The result shows, pH 7 was the best acidity, 37℃ was the best temperature, and 40 h of fermentation could reach the peak of enzyme activity. Analyze ginger commensal cellulose-decomposing strain for the first time and provided data to develop new nature cellulase.%对一株从生姜根际筛选到的纤维素降解活性较高的菌株SP1进行了初步分析。通过PCR方法获得菌株SP1的16S rDNA序列,长度为1406bp。使用摇床发酵法对菌株SP1发酵产纤维素酶的特征进行了初步分析。结果表明,该菌株产酶的最适pH为7,适宜温度为40℃,发酵40h能达到产酶高峰。试验首次分析了生姜根际纤维素降解共生菌的情况,并为开发新型天然纤维素酶提供了资料。

  11. 纤维素酶—乙醇法提取苦丁茶中总黄酮%Extraction of Total Flavonoids from the Leaves of Ilex latifolia by Cellulase-Ethanol Method

    Institute of Scientific and Technical Information of China (English)

    黄靖

    2012-01-01

    Total flavonoids were extracted from leaves of Ilex latifolia (Kuding tea) by cellulase-ethanol method. The effects of cellulose dose, enzymolysis time, temperature, pH, volume fraction of ethanol, and dose of ethanol on yield of total flavonoids were studied by single factor tests so as to optimize the extraction conditions. The results showed that the optimum extraction conditions were obtained as follows, dose of cellulose enzyme, 1.0 mg per 5.0 g Kuding tea flour; enzymolysis time, 2.0 h; enzymolysis temperature, 55℃; pH, 4.5; volume fraction of ethanol, 60%; dose of ethanol, 35 mL per 5.0 g Kuding tea flour. The yield of flavonoids was up to 7.80% under these conditions.%采用纤维素酶酶解预处理与乙醇浸提法相结合从苦丁茶中提取总黄酮.通过单因素试验考察酶用量、酶解时间、酶解温度、pH、乙醇体积分数和乙醇用量对总黄酮提取率的影响,优化提取工艺条件.结果表明,优化的提取工艺条件为0.5 g苦丁茶粉末中加入纤维素酶1.0mg、酶解时间2.0 h、酶解温度55℃、酶解pH 4.5、体积分数60%的乙醇用量35 mL,最佳提取工艺条件下苦丁茶中总黄酮提取率达7.80%.

  12. 产蛋白酶和纤维素酶纳豆芽孢杆菌益生菌株的筛选及其生长特性研究%Screening of protease and cellulase producino Bacillus subtilis Natto strain and its growth characteristics

    Institute of Scientific and Technical Information of China (English)

    孙妍; 王加启; 奚晓琦; 魏宏阳; 周凌云

    2011-01-01

    According to the biological qualification of hydrolysised casein and CMC of Bacillus subtilis Natto, 10 strains were screened by pour plate method from their parent strains of Bacillus subtilis natto NB-1 and NR-1. The strain named NY-3 was selected with higher protease activity and cellulase activity through the test of protease activity and cellulase activity in 48 h fermenting medium among 10 strains. Further research of its growth characteristics showed that the best time of inoculation and fermentation were 14 and 48 h, respectively.%根据纳豆芽孢杆菌(Bacillus subtilis natto)水解酪蛋白和羧甲基纤维素的生物学特性,以菌株NB-1和NR-l为出发菌株,采用稀释涂平板法获得10株初筛纳豆芽孢杆菌,通过测定初筛菌株48 h发酵液中蛋白酶活性和纤维素酶活性,确定NY-3产蛋白酶和纤维素酶活性均相对较高.同时对该菌株生长特性进行了研究.结果表明,NY-3的最佳接种时间和最佳发酵时间分别为14和48 h.

  13. 纤维素酶催化与三液相萃取偶联制备盾叶薯蓣皂苷元%Preparation of diosgenin by integrated bioprocess of cellulase catalysis and extration in three-liquid-phase system

    Institute of Scientific and Technical Information of China (English)

    魏夺; 董悦生; 韩松; 修志龙

    2012-01-01

    Using coupling biotech of cellulase-catalysis and extrusion in three-liquid-phase system that composed of n-hexane, 1,4-dioxane and ammonium sulfate to hydrolyze dioscin in Chinese yan and glucose connected to steroidal saponins, and extration separation to collect diosgenin. The distribution of several steroidal saponins, hydrolyzed glucose, enzyme and its activity in the three-phase system were studied, and catalytic conversion by cellulase-enzyme and formation of diosgenin in three-liquid, organic and aqueous phases were compared. The results showed that diosgenin yield in the three-liquid-phase system could compare with that in organic phase and aqueous phase. The three-liquid-phase system could be suitable for simultaneous enzymatic hydrolysis of steroidal saponins and extration of diosgenin and glucose. The substrate and enzyme were held in the middle phase of 1,4-dioxane, whiles the products, diosgenin and glucose, were partitioned into the top and bottom phase, respectively. The yields of diosgenin was 69. 4% at 96 h in the three-liquid-phase system, which was twice and 27. 6 folds obtained in organic phase and aqueous phase respectively.

  14. 纤维素酶酶解法提取怀山药多糖工艺研究%Study on Extraction of Polysaccharide from Dioscorea Opposita Thunb by Enzymatic Hydrolysis of Cellulase

    Institute of Scientific and Technical Information of China (English)

    李培

    2015-01-01

    The Polysaccharide was extracted from Dioscorea opposita Thunb by enzymatic hydrolysis of Cellulase .First,the effects of the ratio of material and water ,the amount of the enzyme ,the time of enzyme hydrolysis , and the temperature of enzyme hydrolysis on the extraction rate of Polysaccharide from Di-oscorea opposita Thunb were studied by single factor experiment .Then the extraction process of Polysac-charide from Dioscorea opposita Thunb was optimized by orthogonal test .The results showed that the order of the primary and secondary factors affecting the extraction rate of Polysaccharides from Dioscorea opposi -ta Thunb was in order:the amount of the enzyme ,the time of enzyme hydrolysis ,the temperature of en-zyme hydrolysis ,the ratio of material and water .The optimal extraction conditions were as follows:the a-mount of the enzyme 3%, the time of enzyme hydrolysis 75 min,the temperature of enzyme hydrolysis 60℃,the ratio of material and water 1:30 .The Polysaccharide yield of 15.78% was achieved under these conditions .%以怀山药为材料,采用纤维素酶酶解,提取其活性成分多糖,先通过单因素实验考察了料水比、酶添加量、酶解时间,酶解温度对怀山药多糖提取得率的影响;然后通过正交试验对怀山药多糖的酶解提取工艺进行了优化,结果表明各因素对怀山药多糖提取率影响的主次顺序依次为:酶添加量、酶解时间、酶解温度、料水比;其最优提取工艺条件为:酶添加量3%,酶解时间75 min,酶解温度60 ℃,料水比1:30,在此最优工艺条件下,怀山药多糖的提取率为15.78%.

  15. Efeito de enzimas de maceração na textura do palmito (Euterpe edulis Mart Influence of cellulase, pectinase and hemicellulase on the texture of hearts of palm (Euterpe edulis Mart.

    Directory of Open Access Journals (Sweden)

    Regina Kitagawa GRIZOTTO

    1997-12-01

    Full Text Available Com a finalidade de ampliar o aproveitamento da palmeira produtora do palmito estudou-se a influência da poligalacturonase e de enzimas maceradoras na textura das partes semi-rígidas do vegetal não-comestíveis, incubando-se preparados comerciais de celulase, hemicelulase e poligalacturonase com o palmito preparado na forma de pequenos toletes (1-3 cm de comprimento e em porções de 2cm do raquis do vegetal. Embora os tratamentos com hemicelulase e mistura de hemicelulase e poligalacturonase tenham promovido ligeiro amaciamento do palmito, os resultados mostraram ,de modo geral, acréscimo na textura do palmito cortado em porções de 3,0 cm e em fatias de 1,0 cm indicando solubilização intensa das regiões suscetíveis a hidrólise com a permanência das regiões duras mais ricas em lignina. Como nos outros tecidos do palmito, no raquis fibroso, não foi comprovada estatiscamente a ação das enzimas na textura do vegetal.With the aim of amplifying the utilization of the palm tree the influence of poligalacturonase and macerating enzymes on the texture of the hard tips of hearts of palm was studied, incubating commercial preparations of cellulase , hemicellulase and poligalacturonase with the hearts of palm, prepared as small stalks of 1-3 cm length and with 2 cm portions of the rachis of the plant. Although the treatments containing hemicellulase and a mixture of hemicellulase and poligalacturonase promoted a slight softening of the hearts of palm, the results generally showed a hardening of the texture of the hearts of palm prepared as stalks of 1 to 3 cm indicating great solubilization of the parts susceptible to hydrolysis, the hard lignin rich parts remaining intact. As in the other tissue of the plant, in the rachis of the fibrous hearts of palm, the action of the enzymes was not statiscally proven.

  16. Analysis of Cellulase Production withTrichoderma reeseiRut C30 Induced by Corn Cob%玉米芯诱导里氏木霉Rut C30产纤维素酶的分析

    Institute of Scientific and Technical Information of China (English)

    马立娟; 蔡瑞; 崔有志; 赫荣琳; 陈树林; 肖冬光

    2015-01-01

    分别以不同木质纤维素类为碳源(质量浓度均为10,g/L)诱导里氏木霉Rut C30产酶,通过测定诱导过程的产酶曲线,结果表明:以玉米芯为诱导碳源时,各酶组分的产酶水平最高,诱导120,h 后纤维素酶滤纸酶活、β–葡萄糖苷酶和木聚糖酶酶活分别为1.53,FPU/mL、0.61,IU/mL 和72.86,IU/mL.采用高效液相色谱(HPLC)法分析了玉米芯水解过程中糖的组成及变化,可检测到的单糖包括葡萄糖、木糖、阿拉伯糖、甘露糖和半乳糖以及纤维二糖等寡糖.诱导产酶实验表明单糖中只有半乳糖可诱导纤维素酶的合成.%Trichoderma reesei Rut C30 was induced by different lignocellulosic inducers with the same concentration of 10,g/L to produce enzymes,and the enzyme-producing curves were determined. Results indicated that the production of cel-lulase,β-glucosidase and xylanase were the highest when corn cob was used as the inducing carbon source,and the produc-tion was up to 1.53,FPU/mL,0.61 IU/mL and 72.86 IU/mL respectively after inducing 120 hours. The sugar composition and variation during the enzymatic hydrolysis of corn cob were analyzed with high performance liquid chromatography(HPLC). Monosaccharide detected included glucose,xylose,arabinose,mannose and galactose. The results of induction experiments indicated that only galactose among monosaccharides could induce cellulase.

  17. Inductive effects of fungal pathogens of American Ginseng and Ginseng on chitinase and cellulase of antigonistic actinomycetes%西洋参和人参病原真菌菌体对放线菌2种水解酶的诱导

    Institute of Scientific and Technical Information of China (English)

    于妍华; 薛泉宏; 唐明

    2011-01-01

    【目的】研究特定拮抗放线菌对西洋参人参土传病害病原真菌的接触抗菌机理。【方法】以5株西洋参、人参土传病害病原真菌菌体为惟一碳源,用液体培养及3,5二硝基水杨酸(DNS)法研究5株供试病原真菌对9株拮抗放线菌几丁质酶和纤维素酶合成的诱导作用;采用搭片法,观察9株拮抗放线菌与5株供试病原真菌菌丝间的相互作用。【结果】①以5株病原真菌菌体为惟一碳源时,可诱导9株拮抗放线菌合成几丁质酶和纤维素酶;9株放线菌的几丁质酶和纤维素酶活性分别为7.17~11.58和6.14~21.20 U,其中西洋参恶疫霉菌体对9株放线%【Objective】 Inhibitory mechanism of antigonistic actimomycetes fighting against fungal pathogens of soil-borne disease in American Ginseng and Ginseng was studied.【Method】 The inductiveness was assessed by the activity of chitinase and cellulase,which were induced from 9 strains of antigonistic actinomycete by using 5 dried strains of fungal pathogens of American Ginseng and Ginseng as C-source in liquid culture medium and DNS measurement,and the mutual effects were observed between antigonistic actinomycetes and fungal pathogens of American Ginseng and Ginseng through the method of building pieces on plate.【Result】 ①The activity of chitinase and cellulase,which was mainly distributed among 7.17-11.58 and 6.14-21.20 U,respectively,differed from 9 strains of antigonistic antinomycete induced from 5 strains of fungal pathogen of American Ginseng and Ginseng.The powder of Phytophthora cactorum could induce antigonistic actinomycetes to produce much more chitinase and cellulase.②The mutural effects between mycelia of Act11,Act13,Act24 and Cylindrocarpon destructans,Cylindrocarpon sp.,such as winding and decomposition,were observed distinctively.【Conclusion】 The mycelia of 5 strains of fungal pathogen of American Ginseng and Ginseng could induce 9 strains of antigonistic actinomycete

  18. 木薯纤维素乙醇发酵的纤维素酶成本评价%Evaluation of the cellulase cost during the cassava cellulose ethanol fermentation process

    Institute of Scientific and Technical Information of China (English)

    方镇宏; 邓红波; 张小希; 张建; 鲍杰

    2013-01-01

    木薯中的纤维素成分约占木薯干重的10%(W/W).文中以木薯燃料乙醇生产的木薯纤维素酒渣为原料,从纤维素酶成本角度评估了三种利用木薯纤维素组分发酵生产乙醇的方法,包括木薯纤维素酒渣的直接糖化和乙醇发酵、木薯纤维素酒渣预处理后的糖化与乙醇发酵、木薯乙醇发酵中同步淀粉与纤维素糖化以及乙醇发酵.结果表明,前两种方法的纤维素利用效率不高,酶成本分别达到13602、11659元/吨乙醇.第三种方法,即在木薯乙醇发酵过程同时加入糖化酶和纤维素酶,进行同步淀粉与纤维素糖化,进而进行乙醇发酵,木薯纤维素乙醇的收益最高.发酵结束时的乙醇浓度从101.5g/L提高到107.0g/L,纤维素酶成本为3 589元/吨乙醇.此方法利用木薯纤维素与木薯淀粉同时进行,不会带来额外的设备及操作投入,酶成本低于产品乙醇价格,可实现盈利,因此第三种方法为木薯纤维用于乙醇发酵的最适方法,本研究结果将为木薯乙醇产业深度利用木薯纤维提供依据.%Cellulose takes nearly 10% (W/W) dry weight of cassava tubers. In this study, the cellulase cost of different ethanol fermentation from cassava cellulose was evaluated. The processes include the direct saccharification and fermentation of original cassava cellulose residues, the direct saccharification and fermentation of pretreated cassava cellulose residues, and the simultaneous co-saccharification and fermentation of cassava starch and cassava cellulose. The results show that the cassava cellulose utilization in the first two processes were low with the enzyme cost of 13 602 and 11 659 RMB Yuan per tone of ethanol, respectively. In the third process, the final ethanol concentration increased from 101.5 g/L to 107.0 g/L when cassava cellulose and cassava starch were saccharified simultaneously. Comparing to the first two processes, the third one demonstrated the lowest enzyme cost at 3

  19. Comparation of Gut Cellulase Activity in Four Herbivorous Beetles%4种植食性甲虫肠道内纤维素酶活性的比较

    Institute of Scientific and Technical Information of China (English)

    李燕利; 薛怀君; 胡春祥; 杨星科

    2013-01-01

    To compare and estimate the cellulose enzyme activity in different insects, the paper presented a comprehensive analysis of enzyme activity from the gut juices of four herbivorous insect species ( larvae and adults of three leaf-eating beetles, Chrysomela populi, Ambrostoma quadriimpressum and Gallerucida bifasciata, and larva of a trunk-borer , Semanotus bifasciatus). The results showed that there were complete cellulase systems in these four beetles, and that both temperature and pH had a significant impact on the enzyme activity in the gut contents. For both larvae and adults of three leaf beetles, the optimal conditions of enzyme activity were 40 - 60℃ and pH 4 - 6, and the enzyme activity could not be detected under high temperature and high pH conditions. However, in trunk-borer of 5. bifasciatus the highest enzyme activity of exoglucanase (C1) was observed at pH 9 and 70 ℃. In addition, the enzyme activities of endoglucanase (Cx) and β-glucosidase in S. bifasciatus were lower than that of the leaf beetles, while the enzyme activity of C1 was much higher than that of these leaf beetles. The results indicated that temperature and pH had significant influence on insect cellulose enzyme activity. The different feeding habits and the phylogenetic relationship may result in different enzyme activity.%为比较和客观评价昆虫体内纤维素酶的活性,在不同温度及pH值下测定杨叶甲、二纹柱萤叶甲、紫榆叶甲成、幼虫及双条杉天牛幼虫消化道内3类纤维素酶的活性.结果表明:4种甲虫体内均有消化纤维素的完整酶系,3种叶甲成、幼虫体内各纤维素酶的最适温区为40 ~ 60℃、最适pH值范围为4~6,在高温和强碱条件下均未能检测出酶活性;双条杉天牛幼虫C1酶活性在强碱pH =9和70℃条件下活性最高,Cx酶和β-葡萄糖苷酶活性低于3种叶甲的成虫和幼虫,C1酶活性显著高于3种叶甲的成虫和幼虫.温度和pH值对昆虫体内纤维素酶

  20. Production and properties of the cellulase-free xylanase from Thermomyces lanuginosus IOC-4145 Produção e propriedades de xilanase livre de celulase de Thermomyces lanuginosus IOC-4145

    Directory of Open Access Journals (Sweden)

    Mônica Caramez Triches Damaso

    2002-12-01

    Full Text Available In recent years, xylanases have expanded their use in many processing industries, such as pulp and paper, food and textile. Thermomyces lanuginosus IOC-4145 was able to produce a very high level of cellulase-free xylanase in shaken cultures using corncob as substrate (500 U/mL. An optimization of the medium composition in submerged fermentation was carried out aiming at a low cost medium composition for enzyme production. Statistical experiment design was employed for this purpose, pointing out corncob as the most important parameter, which affects enzyme production. Additionally, the influence of several chemicals on xylanase activity was investigated in the crude extract. A slight stimulation of the enzyme (5-15% was achieved with NaCl and urea, both at 3 and 5 mM of concentration. On the other hand, dithiothreitol and beta-mercaptoethanol at a molarity of 5mM have caused a strong stimulation of the enzyme (40-53%. The crude xylanase displayed appreciable thermostability, retaining almost 50% of activity during 24 hours of incubation at 50ºC; about 50% of activity was present at 60ºC even after 4 hours of incubation. The enzyme also exhibited good storage stability at -20ºC without any stabilizing agent.Nos últimos anos tem crescido o uso de xilanases em muitas indústrias, tais como polpa e papel, alimentos e têxtil. Thermomyces lanuginosus IOC-4145 foi capaz de produzir um alto nível de xilanase livre de celulase em culturas agitadas usando sabugo de milho como substrato (500 U/mL. Procedeu-se, inicialmente, à otimização da composição do meio de produção em fermentação submersa, com o intuito de alcançar uma composição de meio de produção de baixo custo para produção da enzima. Para este propósito, utilizou-se planejamento estatístico de experimentos. O sabugo de milho revelou-se como a mais importante variável que afeta a produção enzimática. Adicionalmente, a influência de vários reagentes na atividade xilan

  1. Effects of Cellulase and Propylene Glycol on Negative Energy Balance Related Serum Indexes of Postpartum Dairy Cows%纤维素酶和丙二醇对产后奶牛能量负平衡相关血清指标的影响

    Institute of Scientific and Technical Information of China (English)

    陈连民; 陈前岭; 王梦芝; 高健; 蔡青和; 张婵娟; 王洪荣

    2014-01-01

    本文旨在比较饲粮中添加纤维素酶和丙二醇对产后奶牛能量负平衡相关血清指标、尿液酮体浓度的影响。试验选择24头处于围产前期奶牛随机分成3组,对照组饲喂基础饲粮,另设置丙二醇组(基础饲粮+0.5%丙二醇制剂)和纤维素酶组(基础饲粮+0.1%纤维素酶制剂),测定产后第1、20、40、60和100天奶牛能量负平衡相关血清指标、尿液酮体的浓度。结果表明:在产后第1天丙二醇组血清葡萄糖和胰岛素浓度显著高于其余2组( P<0.05);在产后第1、20、40和60天,丙二醇组血清游离脂肪酸和尿液酮体浓度显著低于对照组(P<0.05),第20天显著低于纤维素酶组( P<0.05);甘油三酯等与肝脏脂肪代谢相关的血清指标试验组优于对照组,且以丙二醇组更好。综上,饲粮添加丙二醇及纤维素酶对奶牛产后能量负平衡有一定的改善作用,且以添加0.5%丙二醇制剂效果相对更好。%The objective of this study was to compare the effects of cellulase and propylene glycol on negative energy balance ( NEB) related serum indexes and urine ketone ( KET) concentration of postpartum dairy cows. Twenty four early perinatal dairy cows were randomly divided into 3 groups with 8 cows per group. Three groups were control group (basal diet), propylene glycol group (basal diet+0.5% propylene glycol prepara-tion) and cellulase group (basal diet+0.1% cellulose preparation). Serum indexes related to NEB and urine KET concentration were detected respectively on days 1, 20, 40, 60 and 100 after calving. The results showed as follows: on day 1 after calving, serum glucose and insulin concentrations of propylene glycol group were significantly higher than those of the other 2 groups ( P<0.05) . On days 1, 20, 40 and 60 after calving, serum non-esterified fatty acid and urine KET concentrations in propylene glycol group were significantly lower than those in control group (P<0.05), moreover

  2. 金属离子及表面活性剂对纤维素酶水解预处理玉米秸秆的影响%Influence of Metal Ions and Surfactants on the Hydrolysis of Pretreated Corn Stover by Cellulase

    Institute of Scientific and Technical Information of China (English)

    王娜娜; 姚秀清; 张全; 关浩

    2011-01-01

    Influences of metal ions and surfactants on the hydrolysis of pretreated corn stover by cellulase were researched. It was showed that some metal ions and surfactants loading in hydrolysis system, such as Cu2+ and Tween80, could enhance the activity of cellulase. In a 50 g hydrolysis system at pH 5.0, temperature 50 degrees centigrade , pretreated corn stover ( PCS) 10 wt% , enzyme loading 1 g ( 1300 from Zensun Sci & Tech Co. , Ltd) ,hydrolysis time 3 hours, the reducing sugar concentration with Tween80 (20 mg/L) loading is 21.3 percent of no Tween80 loading, and reducing sugar concentration with Cu2+ (0.13 mmol/L) loading is 138. 2 percent of no Cu2+loading.%研究了不同的金属离子及表面活性剂对纤维素酶水解的影响,试验表明添加某些金属离子及表面活性剂能够促进纤维素酶的水解,其中Cu2+和Tween80促进效果最佳.50g反应体系在pH 5.0、温度50℃、预处理玉米秸秆(PCS)10 wt%、1300纤维素酶(泽生科技)加入量1g条件水解3 h,添加20 mg/L Tween80的水解液中还原糖浓度比不加Tween80时提高21.3%;添加0.13 mmoL/LCu2+的水解液中还原糖浓度比不加金属离子时提高38.2%.

  3. Avaliação da celulase e pectinase como enzimas complementares, no processo de hidrólise-sacarificação do farelo de mandioca para produção de etanol Evaluation of the cellulase and pectinase by complementary enzymes in the process of hydrolysis-saccharification of cassava fibrous waste for alcohol production

    Directory of Open Access Journals (Sweden)

    Magali LEONEL

    1999-01-01

    Full Text Available Neste trabalho objetivou-se avaliar o uso de enzimas complementares no processo enzimático de hidrólise e sacarificação para a produção de etanol a partir do resíduo fibroso das fecularias. Os resultados obtidos demonstraram que 63,42% do amido foram hidrolisados no tratamento em que não se utilizaram enzimas complementares. No tratamento com as duas enzimas complementares foram hidrolisados 89,55%, no tratamento com celulase 65,42% e no tratamento com pectinase 88,73%. A prensagem do resíduo após o processo de hidrólise e sacarificação mostrou-se eficiente, ficando 10,43% do total de açúcares obtidos retidos no resíduo fibroso final. Portanto, o tratamento em que se utilizou a pectinase como enzima complementar na hidrólise foi o melhor. A celulase não apresentou efeito significativo no rendimento do processo.This work it was proposed to evaluate the use of complementary enzymes (cellulase and pectinase in the enzymatic process of hydrolysis-saccharification of the cassava fibrous waste for alcohol production. The results indicated that 63,42% of starch was hydrolyzed in the treatment without complementary enzymes, 89,55% in the treatment with the enzymes, 65,42% with the cellulase by complementary and 88,73% in the pectinase treatment. The pressing was efficacious for sugar recuperation and 10% of total sugar was retaining in the final fibrous residue. The pectinase was the better complementary enzyme enhance the yield.

  4. The Characteristics of Cellulase and Amylase from B.amy lolique f aciens FAJT-8754 and Optimization of Fermentation Conditions%解淀粉芽胞杆菌FJAT-8754产纤维素酶和淀粉酶特性及发酵条件优化

    Institute of Scientific and Technical Information of China (English)

    王凯; 蓝江林; 刘波; 刘程程; 李安琪

    2014-01-01

    采用淀粉平板和羧甲基纤维素钠(CMC-Na)平板从选取的140株芽胞杆菌中初筛出8株具有产纤维素酶和淀粉酶复合酶芽胞杆菌,经酶活力测定解淀粉芽胞杆菌FJAT-8754(Bacillus amyloliquef aciens)具有较高的淀粉酶、纤维素酶活力。通过研究解淀粉芽胞杆菌FJAT-8754的生长、产酶曲线以及酶学特性,确定在发酵28 h后菌体生长进入稳定期,培养44 h时发酵液中活菌数达到最大为4.41×109 cf u · mL -1,在36 h时纤维素酶、淀粉酶均达到酶活最高峰,酶活分别为135.8、1543.3 U · mL -1;纤维素酶反应最适p H值为5.5、最适温度为50℃,Vmax为5.14×10-3 mg · mL -1· min-1、 Km值为7.71×10-1 mg · mL -1;淀粉酶反应最适pH值为5.5、最适温度为55℃,Vmax为3.35×10-2 mg · mL -1· min-1、 Km值为6.03×10-3 mg · mL -1。采用3因素7水平,即U 7(73)均匀设计法优化解淀粉芽胞杆菌产酶条件,确定产纤维素酶、淀粉酶的最优条件均为:初始pH值6.2、培养温度37.5℃、转速180 r · min-1,优化后解淀粉芽胞杆菌 FJAT-8754纤维素酶活力为202.9 U·mL -1、淀粉酶活力为2392.9U·mL -1。%Eight Bacillus strains which produce cellulase and amylase were screened from 140 standard Bacillus strains by using starch medium and carboxymethylcellulose sodium (CMC-Na) plate screening method .FJAT-8754 had the highest cellulase and amylase producing activities and the biological characterizes (e .g .growth curves , dynamic changes of enzyme activity and enzymatic properties ) were tested .Our results suggested that the stationary phase of FJAT-8754 was at 28 h ,the colony count was 4.41 × 109 cfu · mL -1 at 44 h and the activity of cellulase and amylase peaked at 36 h simultaneously .Meanwhile ,the optimum reaction conditions for cellulase were at pH 5.5 ,55℃ ,Vmax was 5.14 × 10-3 mg · mL -1 · min-1 and Km was 7.71 × 10-1 mg · mL -1 ;the optimal reaction conditions of

  5. 甲酸与纤维素酶和木聚糖酶对多花黑麦草与白三叶混合青贮料发酵品质的影响%Effects of formic acid, cellulase and xylanase on fermentation quality of Lolium multiflorum and Trifolium repens mixture silage during ensiling

    Institute of Scientific and Technical Information of China (English)

    庄苏; 丁立人; 周建国; 王恬

    2013-01-01

    To evaluate the effects of formic acid, cellulase and xylanase on fermentation quality of mixture silage during ensiling, 2 000 g chopped Lolium multijlorum(80% ) and Trifolium repens(20% ) mixtures were ensiled in laboratory plastic bag either untreated (control) or treated with formic acid, cellulase, formic acid plus cellulase, xylanase, formic acid plus xylanase, cellulase plus xylanase and formic acid plus cellulase plus xylanase, respectively. Triplicate bags were opened at 0 d,2d,4d,6d,8d and 30 d of ensiling for chemical analyses. The pH value in all treated silages was lower (P<0. 05) than control at the end of ensiling. The formic acid, enzyme or formic acid plus enzyme treatments enhanced (P<0. 05) water soluble carbohydrate content significantly compared with control at all ensiling periods. The lactic acid content and acetic acid content were higher (P<0. 05) in the enzyme treatment than those in the formic acid-contained treatments and control, respectively. However, the acetic acid content was lower ( P<0. 05 ) in formic acid-contained treatments than that in enzymes treated silages. Relative to control, all treatments had lower (P< 0. 05 ) ammonia-N concentrations during ensiling. The enzyme treatments effectively (P<0. 05) decreased neutral detergent fiber and acid detergent fiber contents in the silages. The results suggested that the addition of formic acid and enzymes improved the Lolium multiflorum and Trifolium repens mixture silage quality, and the enzyme treatments were better than formic acid treatments during ensiling.%为评价甲酸与纤维素酶和木聚糖酶处理后多花黑麦草与白三叶混合青贮料发酵品质的变化,试验将混合青贮料分为对照组(未处理)、甲酸添加组、纤维素酶添加组、甲酸+纤维素酶添加组、木聚糖酶添加组、甲酸+木聚糖酶添加组、纤维素酶+木聚糖酶添加组、甲酸+纤维素酶+木聚糖酶添加组共8组,每组3个重复.青贮原料按80

  6. BÚSQUEDA DE LAS MEJORES CONDICIONES PARA LA EXTRACCIÓN Y MEDIDA DE ACTIVIDAD DE CELULASA Y XILANASA EXTRAÍDAS DE LA CORTEZA DE PITAYA AMARILLA (Acanthocereus pitajaya Searching the Best Conditions for the Extraction and Activity Measurement of Cellulase and Xylanase Extracted from the Yellow Pitaya Fruit Peel (Acanthocereus pitajaya

    Directory of Open Access Journals (Sweden)

    YENNY MARITZA DUEÑAS GÓMEZ

    Full Text Available Para pitaya amarilla (Acanthocereus pitajaya se ha encontrado que el ablandamiento excesivo de la cáscara contribuye al deterioro del fruto, al aplicar diferentes técnicas de conservación en fresco. Dado que tanto la celulasa como la xilanasa se han vinculado con el ablandamiento de la cáscara de frutos, este trabajo se basó en la búsqueda de las mejores condiciones de extracción y medida de actividad de celulasa y xilanasa. El mejor sistema de extracción fue buffer fosfato 20 mM, NaCl 0,5 M, pH 7,0. Para la medida de actividad de celulasa es necesario incubar durante 60 min a 37 ºC, con un volumen de extracto enzimático crudo de 30 µL, empleando buffer acetato 100 mM a pH 5,0; los valores de constante aparente de Michaelis Menten (K M aparente y velocidad máxima (V MÁX fueron 0,279 mg/mL y 0,00014 nmol glucosa/min, respectivamente. Para determinar la actividad de xilanasa se establecieron 15 min de tiempo de incuba-ción, a 50 ºC, empleando 30 µL de extracto enzimático crudo a pH 4,0 (buffer acetato 100 mM; los valores de K M aparente y V MÁX para xilanasa fueron 0,073 mg/mL y 0,0011 nmol glucosa/min, respectivamente.By applying different conservation techniques on yellow pitaya fruit (Acanthocereus pitajaya it has been found that excessive softening of the peel contributes to the deterioration of the fruit. Due to that both cellulase and xylanase have been related to the softening of the fruit's peel; this work was based on the search of the best conditions not only for the extraction, but also for the activity measurement of both cellulase and xylanase. The best extraction system for both enzymes was 20 mM buffer phosphate, 0.5 M NaCl, pH 7.0. For the cellulase activity measurement it was necessary to incubate during 60 min at 37 ºC, with a volume of raw enzymatic extract of 30 µL, using buffer acetate 100 mM at pH 5,0; the values of apparent K M and V MÁX were 0.279 mg/mL and 0.00014 nmol glucose/min, respectively. To

  7. Screening of a Cellulase-producing Strain from the Larval Gut of Bombyx mori and Optimum the Fermentation Condition%家蚕肠道纤维素酶菌株的筛选及产酶条件的优化研究

    Institute of Scientific and Technical Information of China (English)

    马如箭; 贾俊强; 刘艳伟; 任子旭; 桂仲争

    2013-01-01

    A novel cellulase-producing fungus named BMC-2 was isolated from the fifth-instar larval gut of Bom-byx mori. Fermentation conditions for the cellulase-producing, in which CMCase specific activity was taken as re-sponse value, were optimized with Plackett-Burman design, steepest ascent test and Box-Benhken design. The re-sults showed that fermentation time, fermentation temperature, initial pH of medium and rotational speed had influ-ence on cellulase-producing significantly. The affecting order was fermentation time, initial pH of medium, fermen-tation temperature and rotational speed, in turn. The optimized fermentation conditions were as follows:fermenta-tion time 94.35 h, fermentation temperature 30.3℃, initial pH of medium 7.01 and rotational speed 179 r/min. Under the optimal conditions, the CMCase specific activity was 25.801 U/mg in theory and 25.526 U/mg in actual value that was increased by 1 times compared with the initial fermentation condition. The result indicated that the established mathematical model by response surface methodology was feasible for practical prediction.%  从5龄家蚕肠道分离筛选得到一株产纤维素酶菌株BMC-2,以羧甲基纤维素酶(CMCase)比活力为响应值,通过Plackett-Burman试验设计、最陡爬坡试验和Box-Behnken试验设计对菌株BMC-2产纤维素酶发酵条件进行优化,结果表明,发酵时间、发酵温度、培养基初始pH和转速对CMCase比活力具有显著影响,其影响程度由大到小依次为发酵时间、培养基初始pH、发酵温度、转速.确定菌株BMC-2产纤维素酶最优发酵条件为:发酵时间94.35 h,发酵温度30.3℃,培养基初始pH 7.01,转速179 r/min.在此条件下, CMCase比活力理论值为25.801 U/mg,验证值为25.526 U/mg,较产酶条件优化前提高了1倍,预测模型可靠性高,可应用于菌株BMC-2产纤维素酶条件的优化.

  8. A Novel Approach to Degrading Plant Cellulose: Continual Adding Materials and Cycling Utilization of Acids and Cellulase%原料连续添加和酸酶循环利用法(CACU法)降解植物纤维素新技术

    Institute of Scientific and Technical Information of China (English)

    王卫国; 赵永亮

    2002-01-01

    A novel approach to degrading plant cellulose--Continual Adding Materials and Cycling Utilization of Acids and Cellulase(CACU) is developed on the basis of the optimum results of degrading cellulose with single acid, double or multiple acids and cellulase-lyses for short time by orthogonal experiments. The schematic flow diagram for continual adding materials and cycling utilization of acids and enzymes for the production of glucose from cellulose was designed, drawn and described. The experimental results show that the CACU method is an effective way of degrading cellulose, which possesses the properties of common conditions, simple process, lower cost and a short period. The whole system consists of five or six sets of equipment, including three sets of reaction equipment and two sets of plant containers. There are totally twelve steps of operation in the whole process. The final transformation ratio of glucose to cellulose and the final concentration of glucose in the end-residue can increase up to 95.34% and 3.21%, respectively, with CH3COOH, HCl and cellulase at 100℃, 1 kg/cm2, and for 15 h by the CACU method. It consumes a quarter of acetic acid and half of HCl compared with the traditional way. The CACU method can decrease the cost of production of glucose from cellulose greatly. Thus, the CACU method is worthy to be developed and spread because of its excellent properties.%以乙酸、草酸、盐酸、硫酸等单酸、二酸、三酸混和及纤维素酶降解植物纤维素的正交试验得出的最佳工艺条件为基础,进一步研究出一种原料连续添加和酸酶循环利用法降解植物纤维素新技术(CACU法).该技术的整个过程只需要12步操作,5或6套设备,包括3~4套反应釜和2套贮罐.在常压、温度100℃、反应15 h的条件下,以CH3COOH,HCl和纤维素酶为反应剂,按照该工艺技术能使纤维素转化成葡萄糖的转化率达95.34%,反应终液中的葡萄糖浓度达3.21%.与常规

  9. Analysis of the microflora and identification of the protease and cellulase producing strains from the traditional fermentative Douzhajun%传统豆渣菌的菌相分析及蛋白酶和纤维素酶主要产生菌株的鉴定

    Institute of Scientific and Technical Information of China (English)

    张燕鹏; 杨瑞金; 王贺; 蒋孝燕

    2012-01-01

    对江西瑞金的特色发酵食品-豆渣菌进行了菌相分析,结果表明豆渣菌是以真菌和细菌为主的混合型发酵食品。真菌主要为串珠霉属(Monilia)、根霉属(Rhizopus)和酵母属(Saccharomyces);细菌主要为微球菌属(Micrococcus)葡萄球菌属(Staphylococcus)、片球菌属(Pediococcus)、异常球菌属(Deinococcus)、肠杆菌属(Enterobacterspp),非乳杆菌属和三株枯草芽孢杆菌(B.subtilis)。串珠霉属真菌具有纤维素酶和蛋白酶活力,枯草芽孢杆菌和根霉属真菌则具有蛋白酶活力。%The microflora of Douzhajun,a traditional okara-fermented food,was analysed.The result indicated that Douzhajun was a food fermented mainly by fungi and bacteria.The fungi included the Monili.Rhizopu.and Saccharomyce,and the bacteria included Staphylococcus,Pediococcus,Deinococcus,Enterobacterspp,non-Lactobacillus and three strains of B.subtilis.Monilia.was able to secrete the cellulase and protease,the B.subtilis and Rhizopus were able to secrete the protease.

  10. Pepsin-Cellulase Digestibility of Pasture Silages: Effects of Pasture Type, Maturity Stage, and Variations in the Enzymatic Method Digestibilidad mediante Pepsina-Celulasa de Ensilajes de Pradera: Efectos del Tipo de Pradera, Estado de Madurez y Variaciones en el Método Enzimático

    Directory of Open Access Journals (Sweden)

    Claudia Barchiesi-Ferrari

    2011-06-01

    Full Text Available Enzymatic in vitro digestibility has been studied as a method to predict energy values of forages for ruminants, although results have been affected by type of forage and methodological details of the technique. This work was performed to evaluate the effects of cellulase concentration (0.75, 1.0 and 6.25 g L-1, incubation time (24 or 48 h and type of final washing of the residue (water or acetone on the in vitro digestibility of the dry matter (DMD, organic matter (OMD and content of digestible organic matter in the DM (D value of silages made at three maturity stages from three types of pastures: a permanent pasture (Dactylis glomerata, Lolium perenne, Bromus catharticus Vahl var. catharticus, Trifolium repens and Holcus lanatus; b Italian ryegrass ley (Lolium multiflorum Lam. cv. Tama; c oats (Avena sativa L. and d mixed pasture (L. perenne-T. repens. Regression equations among cellulase results and in vitro values obtained with rumen fluid were also developed. Higher enzyme concentration, longer incubation time and final washing with acetone, resulted in a significant (P La digestibilidad enzimática in vitro ha sido estudiada para predecir el valor energético de forrajes para rumiantes, aunque el tipo de forraje y los detalles metodológicos han afectado los resultados obtenidos. Este trabajo pretende evaluar los efectos de la concentración de celulasa (0,75; 1,0 y 6,25 g L-1, tiempo de incubación (24 o 48 h y tipo de lavado final (agua o acetona del residuo, sobre la digestibilidad in vitro de la materia seca (DMD, materia orgánica (OMD y el contenido de materia orgánica digestible en la materia seca, o valor D (D value de ensilajes hechos con tres estados de madurez, de diferentes praderas: a pradera permanente (Dactylis glomerata, Lolium perenne, Bromus catharticus Vahl var. catharticus, Trifolium repens y Holcus lanatus; b ballica Italiana (Lolium multiflorum Lam. cv. Tama; c avena (Avena sativa L.; y d pradera mixta (L. perenne

  11. 限饲与营养补偿对小尾寒羊生长性能、消化代谢和瘤胃液纤维素酶活性的影响%Effects of Dietary Restriction and Realimentation on Growth Performance, Digestion, Metabolism and Cellulase Activity in Ruminal Fluid of Small Tail Han Sheep

    Institute of Scientific and Technical Information of China (English)

    陈军强; 丁路明; 高强; 龙瑞军; 安吾; 刘培培; 张丽莉

    2015-01-01

    本试验旨在研究不同限饲水平与营养补偿对小尾寒羊生长性能、消化代谢以和瘤胃液纤维素酶活性的影响. 选择体重相近的3月龄小尾寒羊公羊40只,采用单因素4水平随机设计,每组10只,正试期90 d. 对照(Ⅰ)组全期自由采食;30 d限饲期内,Ⅱ、Ⅲ和Ⅳ组分别按照NRC(2007)推荐的日增重500、400和300 g/d所需要的代谢能(ME)和粗蛋白质(CP)的量配制饲粮,限饲结束后,试验组自由采食60 d为营养补偿期. 在24~30 d和84~90 d进行消化代谢试验. 结果表明:1 )限饲期,限饲水平对结束体重和平均日增重( ADG )有显著影响( P0.05);Ⅱ、Ⅲ和Ⅳ组体重补偿百分比分别为对照组的104.36%、101.71%、99.61%;限饲水平对ADG有显著影响( P0.05). 营养补偿期,组间营养物质的表观消化率差异不显著( P>0.05). 3)限饲期,限饲水平对食入氮、氮沉积、氮沉积率有显著影响( P0.05);粪氮和尿氮的变化不显著(P>0.05). 营养补偿期,组间氮代谢指标差异不显著(P>0.05). 4)1~30 d,试验组瘤胃液纤维素酶活性呈降低趋势,31~90 d,呈升高趋势,在90 d,试验组均高于对照组. 综上所述,对小尾寒羊采用短期适度限饲,经过营养补偿可以使其生长性能、消化代谢不受影响.%The objective of this study was to explore the effects of dietary restriction level and realimentation on growth performance, digestion, metabolism and cellulase activity in ruminal fluid of small tail Han sheep. For-ty 3-month old small tail Han sheep with average body weight (BW) of (19.77±1.34) kg were randomly split into four groups with ten sheep in each group by a single factor four levels design. The study lasted for 90 days. Control group ( group Ⅰ) was fed ad libitum during the whole period; during 30-day dietary restriction peri-od, three experimental groups were fed diets with different metabolizable energy ( ME ) and crude protein ( CP) levels, which were designed to meet

  12. Characterization of Thermostable Cellulases Produced by Bacillus and Geobacillus Strains

    Science.gov (United States)

    Bacterial community composition of thermophilic (60 deg C) mixed cellulose-enrichment cultures was examined by constructing a 16S rDNA clone library which demonstrated major lineages affiliated to Actinobacteria, Bacteroidetes, Chloroflexi, Deinococcus-Thermus, Firmicutes, and Proteobacteria. A tot...

  13. A kinetic model for the burst phase of processive cellulases

    DEFF Research Database (Denmark)

    Præstgaard, Eigil; Olsen, Jens Elmerdahl; Murphy, Leigh;

    2011-01-01

    is negligible. Here, we propose an explicit kinetic model for this behavior, which uses classical burst phase theory as the starting point. The model is tested against calorimetric measurements of the activity of the cellobiohydrolase Cel7A from Trichoderma reesei on amorphous cellulose. A simple version...

  14. Superactive cellulase formulation using cellobiohydrolase-1 from Penicillium funiculosum

    Energy Technology Data Exchange (ETDEWEB)

    Adney, William S.; Baker, John O.; Decker, Stephen R.; Chou, Yat-Chen; Himmel, Michael E.; Ding, Shi-You

    2012-10-09

    Purified cellobiohydrolase I (glycosyl hydrolase family 7 (Cel7A)) enzymes from Penicillium funiculosum demonstrate a high level of specific performance in comparison to other Cel7 family member enzymes when formulated with purified EIcd endoglucanase from A. cellulolyticus and tested on pretreated corn stover. This result is true of the purified native enzyme, as well as recombinantly expressed enzyme, for example, that enzyme expressed in a non-native Aspergillus host. In a specific example, the specific performance of the formulation using purified recombinant Cel7A from Penicillium funiculosum expressed in A. awamori is increased by more than 200% when compared to a formulation using purified Cel7A from Trichoderma reesei.

  15. Enzymatic lignocellulose hydrolysis: Improved cellulase productivity by insoluble solids recycling

    DEFF Research Database (Denmark)

    Weiss, Noah Daniel; Börjesson, Johan; Pedersen, Lars Saaby;

    2013-01-01

    To take advantage of this effect, the amount of solids recycled should be maximized, based on a given processes ability to deal with higher solids concentrations and volumes. Recycling of enzymes by recycling the insoluble solids fraction was thus shown to be an effective method to decrease enzyme...

  16. Soluble inhibitors/deactivators of cellulase enzymes from lignocellulosic biomass.

    Science.gov (United States)

    Kim, Youngmi; Ximenes, Eduardo; Mosier, Nathan S; Ladisch, Michael R

    2011-04-01

    Liquid hot water, steam explosion, and dilute acid pretreatments of lignocellulose generate soluble inhibitors which hamper enzymatic hydrolysis as well as fermentation of sugars to ethanol. Toxic and inhibitory compounds will vary with pretreatment and include soluble sugars, furan derivatives (hydroxymethyl fulfural, furfural), organic acids (acetic, formic and, levulinic acid), and phenolic compounds. Their effect is seen when an increase in the concentration of pretreated biomass in a hydrolysis slurry results in decreased cellulose conversion, even though the ratio of enzyme to cellulose is kept constant. We used lignin-free cellulose, Solka Floc, combined with mixtures of soluble components released during pretreatment of wood, to prove that the decrease in the rate and extent of cellulose hydrolysis is due to a combination of enzyme inhibition and deactivation. The causative agents were extracted from wood pretreatment liquid using PEG surfactant, activated charcoal or ethyl acetate and then desorbed, recovered, and added back to a mixture of enzyme and cellulose. At enzyme loadings of either 1 or 25mg protein/g glucan, the most inhibitory components, later identified as phenolics, decreased the rate and extent of cellulose hydrolysis by half due to both inhibition and precipitation of the enzymes. Full enzyme activity occurred when the phenols were removed. Hence detoxification of pretreated woods through phenol removal is expected to reduce enzyme loadings, and therefore reduce enzyme costs, for a given level of cellulose conversion.

  17. The Cellulase KORRIGAN Is Part of the Cellulose Synthase Complex

    NARCIS (Netherlands)

    Vain, T.; Crowell, E.F.; Timpano, H.; Biot, E.; Desprez, T.; Mansoori Zangir, N.; Trindade, L.M.; Pagant, S.; Robert, S.; Hofte, H.; Gonneau, M.; Vernhettes, S.

    2014-01-01

    Plant growth and organ formation depend on the oriented deposition of load-bearing cellulose microfibrils in the cell wall. Cellulose is synthesized by a large relative molecular weight cellulose synthase complex (CSC), which comprises at least three distinct cellulose synthases. Cellulose synthesis

  18. Single Molecule Study of Cellulase Hydrolysis of Crystalline Cellulose

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Y.-S.; Luo, Y.; Baker, J. O.; Zeng, Y.; Himmel, M. E.; Smith, S.; Ding, S.-Y.

    2009-12-01

    This report seeks to elucidate the role of cellobiohydrolase-I (CBH I) in the hydrolysis of crystalline cellulose. A single-molecule approach uses various imaging techniques to investigate the surface structure of crystalline cellulose and changes made in the structure by CBH I.

  19. Isolation, Purification and Characterization of Cellulases from Streptomyces Strains in the Gut of the Termite Odontotermesformosanus%白蚁肠道二株链霉菌株所产纤维素酶的分离、纯化及其特性

    Institute of Scientific and Technical Information of China (English)

    霍光华; 黄小晖; Shannon DILLON; 何刚强; 陈明辉; 龙昊知

    2012-01-01

    by Fe2+ and Ca2+, but inhibited by Mn2+, Cu2+, Zn2+ and Co2+. These results indicate that these cellulases originated from Streptomyce strains in O. formosanus gut are acidic, heat-tolerant enzymes and that their prosthetic groups may be Fe2+ and Ca2+. Therefore, these enzymes has potential in industrial applications for the decomposition of insoluble cellulose.

  20. Effect of commercial cellulases and refining on kraft pulp properties Correlations between treatment impacts and enzymatic activity components%商业纤维素酶和磨浆对硫酸盐浆性能的影响:机械处理和酶活性成分影响之间的相互关系

    Institute of Scientific and Technical Information of China (English)

    彭金勇

    2015-01-01

    In the papermaking industry, the significance of enzymes as biotechnological catalysts is now recognized. In this study, five cellulase formulations were used for fibre modification. The number of PFI revolutions decreased by about 50% while achieving the same freeness value (decrease in CSF by 200 ml) with the enzymatic pretreatment. The physical properties of handsheets were modified after enzymatic pretreatment followed by PFI refining. A slight decrease in tear strength was observed with enzymes C1 and C4 at pH 7 while the most decrease in tear was observed after C2, C3, C5 treatments. C1 and C4 which had xylanase activity improved paper properties, while other enzymes had a negative impact. Therefore, the intricate balance between cellulolytic and hemicellulolytic activity is the key to optimizing biorefining and paper properties. It was also observed that C1 impact was dependent on pH, which supports the importance of pH in developing an enzymatic strategy for refining energy reduction.%在造纸工业中,酶在生物技术催化剂中的重要性已经得到公认.此次研究选取了五种商业酶来改性纤维.结果显示,经过酶预处理后,浆料达到相同的游离度(下降200ml)所消耗的P F I磨浆机转数大约是未经处理的一半.并且,经过酶预处理及PFI磨浆后,手抄片的性能也会发生改变.最终得出结论,纤维经过酶C1和酶C4在pH=7下预处理后,其手抄片的撕裂强度略微下降.而经过酶C2、酶C3和酶C5预处理后,撕裂强度下降最明显.具有木聚糖酶活性的酶C1和酶C4预处理浆料后能够改善纸张性能,但是其他酶对纸的性能产生了负面影响.因此,水解纤维素活性和水解半纤维素活性之间复杂的平衡是优化生物质精炼和纸张性能的关键.同时发现,pH对酶C1的影响很大,这表明控制pH在开发减少磨浆能耗的酶策略中的重要性.

  1. Hypolipidemic effect of the polysaccharides extracted from pumpkin by cellulase-assisted method on mice.

    Science.gov (United States)

    Zhao, Xin-Hua; Qian, Li; Yin, De-Lu; Zhou, Yi

    2014-03-01

    The fruit of pumpkin (Cucurbita moschata) is one of the most important vegetables in the world. This study was conducted to investigate the hypolipidemic effect of the polysaccharide isolated from pumpkin (PP). Male Sprague-Dawley rats were divided into three groups and treated with diets containing either high fat, PP, or normal fat. Oral administration of PP could significantly decrease the levels of plasma triacylglycerol (TG), total cholesterol (TC), and plasma low-density lipoprotein cholesterol and increase the levels of fecal fat, cholesterol, and plasma high-density lipoprotein cholesterol. Therefore, results suggest that PP had a high hypolipidemic activity and could be explored as a possible agent for hyperlipidemia.

  2. Immobilization of cellulases on magnetic particles to enable enzyme recycling during hydrolysis of lignocellulose

    OpenAIRE

    Alftrén, Johan; Hobley, Timothy John

    2014-01-01

    Der er vigtigt at erstatte oliebaserede brændstoffer og kemikalier med mere miljøvenlige alternativer, da olie bidrager til en netto produktion af drivhusgasser og er en begrænset resurse. Biomasse fra lignocellulose er et meget interessant råmateriale for produktion af biokemikalier og biobrændstof på grund af den store overflod af råmaterialet og det faktum at det er spildmateriale og ikke direkte konkurrerer med fødevareproduktion. Lignocellulose består blandt andet af cellulose og hemicel...

  3. [Lignocellulose degrading bacteria and their genes encoding cellulase/hemicellulase in rumen--a review].

    Science.gov (United States)

    Chen, Furong; Zhu, Yaxin; Dong, Xiuzhu; Liu, Lihua; Huang, Li; Dai, Xin

    2010-08-01

    Rumen of ruminant animals is known as a natural reactor involved in highly efficient lignocelluloses degradation. Rumen fibrolytic microbes have attracted an increasing attention for their potential value in biofuel research. Studies on rumen microbes have traditionally entailed the isolation of fibrolytic bacteria and subsequent analysis of fibrolytic enzymes. Developments in genomic and metagenomic approaches have made it possible to isolate directly genes and gene clusters encoding fibrolytic activities from rumen samples, permitting a global analysis of mechanisms of degradation of lignocellulose in rumen. Research in this field shows that lignocellulose degradation in rumen is a complex process involving a number of different microbes and is effected by a huge array of hydrolytic enzymes in a concerted fashion. This review briefly summarizes results from recent studies, especially metagenomic studies, on lignocellulose degradation in rumen.

  4. IONIC LIQUID SALT-INDUCED INACTIVATION AND UNFOLDING OF CELLULASE FROM TRICHODERMA REESEI. (R828257)

    Science.gov (United States)

    The perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Concl...

  5. Biobleaching application of cellulase poor and alkali stable xylanase from Bacillus pumilus SV-85S

    OpenAIRE

    Nagar, Sushil; Jain, R. K.; Thakur, Vasanta Vadde; Gupta, Vijay Kumar

    2012-01-01

    The potential of extracellular alkali stable and thermo tolerant xylanase produced by Bacillus pumilus SV-85S through solid state fermentation was investigated in pulp bleaching in association with conventional bleaching using chlorine and chlorine dioxide. The biobleaching of kraft pulp with xylanase was the most effective at an enzyme dose of 10 IU/g oven dried pulp, pH 9.0 and 120 min incubation at 55 °C. Under the optimized conditions, xylanase pretreatment reduced Kappa number by 1.6 poi...

  6. Hydrolysis behavior of various crystalline celluloses treated by cellulase of Tricoderma viride

    OpenAIRE

    Abdullah, Rosnah; Saka, Shiro

    2014-01-01

    Cellobiose and glucose are valuable products that can be obtained from enzymatic hydrolysis of cellulose. This study discusses changes in the crystalline form of celluloses to enhance the production of sugars and examines the effect on structural properties during enzymatic hydrolysis. Various crystalline celluloses consisting of group I (cell I, cell III[I], cell IV[I]) and group II (cell II, cell III[II], cell IV[II]) of similar DPs were prepared as starting materials. The similar DP values...

  7. Application of the 2-cyanoacetamide method for spectrophotometric assay of cellulase enzyme activity

    Science.gov (United States)

    Cellulose is the most abundant form of carbon on the planet. Breakdown of cellulose microfibrils in the plant cell wall is a means by which microbes gain ingress into their respective hosts. Cellulose degradation is also important for global carbon recycling and is the primary substrate for producti...

  8. Hypolipidemic effect of the polysaccharides extracted from pumpkin by cellulase-assisted method on mice.

    Science.gov (United States)

    Zhao, Xin-Hua; Qian, Li; Yin, De-Lu; Zhou, Yi

    2014-03-01

    The fruit of pumpkin (Cucurbita moschata) is one of the most important vegetables in the world. This study was conducted to investigate the hypolipidemic effect of the polysaccharide isolated from pumpkin (PP). Male Sprague-Dawley rats were divided into three groups and treated with diets containing either high fat, PP, or normal fat. Oral administration of PP could significantly decrease the levels of plasma triacylglycerol (TG), total cholesterol (TC), and plasma low-density lipoprotein cholesterol and increase the levels of fecal fat, cholesterol, and plasma high-density lipoprotein cholesterol. Therefore, results suggest that PP had a high hypolipidemic activity and could be explored as a possible agent for hyperlipidemia. PMID:24325857

  9. Development of Specific Substrates for Hypocrea jecorina Cellulases

    DEFF Research Database (Denmark)

    Rasmussen, Tina Secher

    task since cellulose exists in different morphologies that can only be degraded efficiently by a battery of enzymes working together.3 One of the most efficient cellulosic systems is secreted by the fungus Hypocrea jecorina, which also has been employed in the industrial production of ethanol. However...... that they exhibit different substrate specificities. Therefore, a small library of derivatives of 2,4-dinitrophenyl cellobioside (2,4-DNPC) and 3,4-dinitrophenyl cellobioside (3,4-DNPC) was prepared. These derivatives contained a series of substituents (X and Y) located at the O4' and O6' position. Inspection...... chemistry or by amine coupling. The kinetic parameters of the library of compounds are to be determined....

  10. Biotechnological valorisation of agro-industrial wastes for the production of cellulases

    OpenAIRE

    DAMATO, GIUSEPPE

    2012-01-01

    This experimental work is embedded in a wider European-funded project named Etoile (FP7/2007-2013, Project n° 222331). The aim of Etoile project was to develop a new integrated process where the two main wastes coming from olive oil traditional three-phase production process, the solid lignocellulosic olive pomace (OP) and the liquid olive oil mill waste water (OOMW), are exploited for the production of cellulolytic enzymes and bioethanol.More specifically, this experimental work was focus...

  11. Investigation of adsorption kinetics and isotherm of cellulase and B-Glucosidase on lignocellulosic substrates

    Science.gov (United States)

    Clear understanding of enzyme adsorption during enzymatic hydrolysis of lignocellulosic biomass is essential to enhance the cost-efficiency of hydrolysis. However, conclusions from literatures often contradicted each other because enzyme adsorption is enzyme, biomass/pretreatment and experimental co...

  12. Recycling of cellulases in a continuous process for production of bioethanol

    DEFF Research Database (Denmark)

    Haven, Mai Østergaard

    The focus of the work presented in this thesis is recycling of commercial enzymes in a continuous process for production of bioethanol from biomass. To get a deeper understanding of the factors affecting the potential for enzyme recycling, the interactions between enzymes and biomass, the adsorpt......The focus of the work presented in this thesis is recycling of commercial enzymes in a continuous process for production of bioethanol from biomass. To get a deeper understanding of the factors affecting the potential for enzyme recycling, the interactions between enzymes and biomass......, the adsorption and desorption as well as stability and recovery of activity was investigated. More knowledge on these factors have enabled a process adapted for enzyme recycling. The driver being that enzyme consumption remains a major cost when producing bioethanol from lignocellulosic biomass. Unlike previous...

  13. Densities, cellulases, alginate and pectin lyases of luminous and other heterotrophic bacteria associated with marine algae

    Digital Repository Service at National Institute of Oceanography (India)

    Ramaiah, N.; Chandramohan, D.

    Epiphytic luminous and non-luminous bacteria were determined quantitatively for eight intertidal algal species from rocky beaches of Goa and Lakshadweep coral reef lagoon. Luminous bacteria were present on all eight algal species and contributed 2...

  14. A Metagenomic Advance for the Cloning and Characterization of a Cellulase from Red Rice Crop Residues.

    Science.gov (United States)

    Meneses, Carlos; Silva, Bruna; Medeiros, Betsy; Serrato, Rodrigo; Johnston-Monje, David

    2016-01-01

    Many naturally-occurring cellulolytic microorganisms are not readily cultivable, demanding a culture-independent approach in order to study their cellulolytic genes. Metagenomics involves the isolation of DNA from environmental sources and can be used to identify enzymes with biotechnological potential from uncultured microbes. In this study, a gene encoding an endoglucanase was cloned from red rice crop residues using a metagenomic strategy. The amino acid identity between this gene and its closest published counterparts is lower than 70%. The endoglucanase was named EglaRR01 and was biochemically characterized. This recombinant protein showed activity on carboxymethylcellulose, indicating that EglaRR01 is an endoactive lytic enzyme. The enzymatic activity was optimal at a pH of 6.8 and at a temperature of 30 °C. Ethanol production from this recombinant enzyme was also analyzed on EglaRR01 crop residues, and resulted in conversion of cellulose from red rice into simple sugars which were further fermented by Saccharomyces cerevisiae to produce ethanol after seven days. Ethanol yield in this study was approximately 8 g/L. The gene found herein shows strong potential for use in ethanol production from cellulosic biomass (second generation ethanol). PMID:27347917

  15. The use of cellulase and filter bag technique to predict digestibility of forages

    DEFF Research Database (Denmark)

    Kowalski, Z. M.; Ludwin, J.; Górka, P.;

    2014-01-01

    calculated for whole crop corn herbages, grass silages and clover silages. It can be concluded that the in vitro method presented in this study is a simple alternative for existing methods in which buffered rumen fluid is used. Using a standard enzyme available commercially worldwide may decrease variation...... between laboratories. Further, using filter bags and Daisy Incubator decreases labour costs and use of animals. Abbreviations AIC, akaike information criterion; ADF, acid detergent fibre; aNDF, neutral detergent fibre; AFBT, ANKOM filter bag technique; CP, crude protein; CS, subset ‘clover silages’; DK...

  16. Development of Activity-based Cost Functions for Cellulase, Invertase, and Other Enzymes

    Science.gov (United States)

    Stowers, Chris C.; Ferguson, Elizabeth M.; Tanner, Robert D.

    As enzyme chemistry plays an increasingly important role in the chemical industry, cost analysis of these enzymes becomes a necessity. In this paper, we examine the aspects that affect the cost of enzymes based upon enzyme activity. The basis for this study stems from a previously developed objective function that quantifies the tradeoffs in enzyme purification via the foam fractionation process (Cherry et al., Braz J Chem Eng 17:233-238, 2000). A generalized cost function is developed from our results that could be used to aid in both industrial and lab scale chemical processing. The generalized cost function shows several nonobvious results that could lead to significant savings. Additionally, the parameters involved in the operation and scaling up of enzyme processing could be optimized to minimize costs. We show that there are typically three regimes in the enzyme cost analysis function: the low activity prelinear region, the moderate activity linear region, and high activity power-law region. The overall form of the cost analysis function appears to robustly fit the power law form.

  17. Construction of a cellulase hyper-expression system in Trichoderma reesei by promoter and enzyme engineering

    NARCIS (Netherlands)

    Zou, G.; Shi, S.; Jiang, Y.; van den Brink, J.; de Vries, R.P.; Chen, L.; Zhang, J.; Ma, L.; Wang, C.; Zhou, Z.

    2012-01-01

    Background A bacterial strain previously isolated from pyrite mine drainage and named BAS-10 was tentatively identified as Klebsiella oxytoca. Unlikely other enterobacteria, BAS-10 is able to grow on Fe(III)-citrate as sole carbon and energy source, yielding acetic acid and CO2 coupled with Fe(III)

  18. Recovery of cellulase activity after ethanol stripping in a novel pilot-scale unit

    DEFF Research Database (Denmark)

    Skovgaard, Pernille Anastasia; Christensen, Børge Holm; Felby, Claus;

    2014-01-01

    Recycling of enzymes has a potential interest during cellulosic bioethanol production as purchasing enzymes is one of the largest expenses in the process. By recycling enzymes after distillation, loss of sugars and ethanol are avoided, but depending on the distillation temperature, there is a pot......Recycling of enzymes has a potential interest during cellulosic bioethanol production as purchasing enzymes is one of the largest expenses in the process. By recycling enzymes after distillation, loss of sugars and ethanol are avoided, but depending on the distillation temperature...

  19. Enzymatic hydrolysis of lignocelluloses: Identification of novel cellulase genes from filamentous fungi

    DEFF Research Database (Denmark)

    Kolasa, Marta; Ahring, Birgitte Kiær; Lübeck, Peter Stephensen;

    2010-01-01

    bonds. Cellulose can be degraded to simple sugar components by means of enzymatic hydrolysis. However, due to its complex, crystalline structure it is difficult to break it down and the cooperative action of a variety of cellulolytic enzymes is necessary. Fungi are known to have potential in production...... genes which subsequently will be cloned and expressed in a relevant fungal host for further characterization of the expressed enzymes. The goal is to introduce new enzymes to industrial processes....

  20. 耐热纤维素酶产生菌的筛选、鉴定及产酶条件优化%Screening and Identification of a Thermostable Cellulase-Secreting Bacteria and Its Optimization for Cellulase Production

    Institute of Scientific and Technical Information of China (English)

    罗颖; 欧阳嘉; 许婧; 何冰芳

    2007-01-01

    从温泉热源地区采集的大量泥土和水样中,筛选出一株在60 ℃生长的纤维素酶产生菌SH2.结合菌株的生理生化特性分析与Biolog微生物自动鉴定仪的鉴定结果,命名为热葡糖苷酶地芽孢杆菌(Geobacillus thermoglucosidasius) SH2,该菌株兼性好氧,在45~60 ℃能较好地生长.对菌体生长与产酶培养条件优化表明:初始pH值为5.5,碳、氮源分别为蔗糖和玉米浆时有利于产酶,经48 h发酵后纤维素酶酶活达0.36 IU/mL.纤维素酶反应条件研究表明,该纤维素酶的最适pH值为6.0,在pH 4.0~10.0范围具有较强的耐受性;在45~65 ℃间酶活差异仅在5%之内,显示了很好的温度耐受性.

  1. Study on Mutation of Cellulase-Decomposing Bacteria Induced by UV to Increase Cellulase Activity%紫外线诱变提高细菌产纤维素酶活力的研究

    Institute of Scientific and Technical Information of China (English)

    陈香; 蒋立建; 韩刚; 韩冲

    2008-01-01

    从南京红山动物园土样及食草动物的粪便中分离出14种产纤维素酶菌株,其中F1表现出较高的酶活力.以F1为出发菌株,通过紫外线诱变处理,采用透明圈法初筛和摇瓶培养复筛,获得了10株高产纤维素酶的突变株Q1~Q10.经紫外线诱变处理的Q3突变株产酶活力最高,与出发菌株相比酶活力提高了15.8倍.

  2. Produção de celulases por Aspergillus niger e cinética da desativação celulásica=Cellulases production by Aspergillus niger and cellulase deactivation kinetic

    OpenAIRE

    Caroline Mariana de Aguiar; Sérgio Luiz de Lucena

    2011-01-01

    O presente trabalho teve como objetivo a avaliação da cinética de produção de enzimas celulases pelo fungo Aspergillus niger e da cinética de desativação das celulases. Foi utilizado bagaço de cana-de-açúcar pré-tratado como fonte de carbono na fermentação para a produção do complexo celulásico e também como substrato da hidrólise enzimática. A. niger foi cultivado em três bateladas, cada uma contendo 10, 50 e 100 g L-1 de bagaço pré-tratado com NaOH 4% (m v-1). A cinética da produção das cel...

  3. Cellulase stability, adsorption/desorption profiles and recycling during successive cycles of hydrolysis and fermentation of wheat straw

    DEFF Research Database (Denmark)

    Rodrigues, Ana Cristina; Felby, Claus; Gama, Miguel

    2014-01-01

    The potential of enzymes recycling after hydrolysis and fermentation of wheat straw under a variety of conditions was investigated, monitoring the activity of the enzymes in the solid and liquid fractions, using low molecular weight substrates. A significant amount of active enzymes could be......) significantly undergo thermal deactivation. The hydrolysis yield and enzyme recycling efficiency in consecutive recycling rounds can be increased by using high enzyme loadings and moderate temperatures. Indeed, the amount of enzymes in the liquid phase increased with its thermostability and hydrolytic...... efficiency. This study contributes towards developing effective enzymes recycling strategies and helping to reduce the enzyme costs on bioethanol production....

  4. Optimization of alkaline cellulase production by the marine-derived fungus Chaetomium sp. using agricultural and industrial wastes as substrates

    Digital Repository Service at National Institute of Oceanography (India)

    Ravindran, C.; Naveenan, T.; Varatharajan, G.R.

    .0 for exoglucanase (Yazadi et al. 1990). In several other fungi, especially Trichoderma and Aspergillus sp., pH optima are variable but in acidic (3.4–6.3) range (Soni et al. 1999). In conclusion, the results of this study demonstrate that the Chaetomium sp... in case of exo- and endoglucanase, cellobiase in case of β-glucosidase) in 1 min under assay conditions (pH 5.0; 50°C for exo- and endoglucanase, 50°C for β-glucosidase). The released reducing sugar (glucose) was estimated by dinitrosalicylic acid (DNS...

  5. Influence of the carbon source on production of cellulases, hemicellulases and pectinases by Trichoderma reesei Rut C-30

    DEFF Research Database (Denmark)

    Olsson, Lisbeth; Christensen, T.M.I.E.; Hansen, K.P.;

    2003-01-01

    The growth and enzyme production by Trichoderma reesei Rut C-30 using different lignocellulosic materials as carbon source were investigated. Cellulose, sugar beet pulp and alkaline extracted sugar beet pulp (resulting in partial removal of hemicellulose, lignin and pectin) or mixtures thereof were...

  6. Untreated Chlorella homosphaera biomass allows for high rates of cell wall glucan enzymatic hydrolysis when using exoglucanase-free cellulases

    OpenAIRE

    Rodrigues, Marcoaurélio Almenara; Teixeira, Ricardo Sposina Sobral; Ferreira-Leitão, Viridiana Santana; da Silva Bon, Elba Pinto

    2015-01-01

    Background Chlorophyte microalgae have a cell wall containing a large quantity of cellulose Iα with a triclinic unit cell hydrogen-bonding pattern that is more susceptible to hydrolysis than that of the cellulose Iβ polymorphic form that is predominant in higher plants. This study addressed the enzymatic hydrolysis of untreated Chlorella homosphaera biomass using selected enzyme preparations, aiming to identify the relevant activity profile for the microalgae cellulose hydrolysis. Enzymes fro...

  7. Deconstruction of ionic liquid pretreated lignocellulosic biomass using mono-component cellulases and hemicellulases and commercial mixtures

    Science.gov (United States)

    Lignocellulosic biomass is comprised of cellulose and hemicellulose, sources of polysaccharides, and lignin, a macromolecule with extensive aromaticity. Lignocellulose requires pretreatment before biochemical conversion to its monomeric sugars which can provide a renewable carbon based feedstock for...

  8. Role of alkaline-tolerant fungal cellulases in release of total antioxidants from agro-wastes under solid state fermentation

    Digital Repository Service at National Institute of Oceanography (India)

    Ravindran, C.; Varatharajan, G. R.; Karthikeyan, A.

    -wastes and total antioxidant property. The increased antioxidant activity on free radical scavenging was also observed with the increase in pH. Thus, the present study makes it possible to produce nutraceutical ingredients cost-effectively from agricultural wastes....

  9. Cellulase activity of a Lentinula edodes (Berk. Pegl. strain grown in media containing Carboximetilcellulose or microcrystalline cellulose

    Directory of Open Access Journals (Sweden)

    José Antônio de Sousa Pereira Júnior

    2003-06-01

    Full Text Available Endoglucanase and exocellobiohydrolase produced b Lentinula edodes (Berk. Pegl. strain thatt was cultivated in carboxymetilcellulose (CMC or microcrystalline cellulose (Avicel liquid media. The concentration and type of cellulose influenced the enzyme activity and production. Extra-cellular cellobiase activity was not detected in CMC or Avicel media. This enzyme was detected in mycelial extracts only. With 1.7% Avicel liquid medium, the strain did not produce exocellobiohydrolase, but 74 µmol RBBR/mg protein/min was detected with 0.5% Avicel. The substitution of Avicel by 0.5% CMC reduced this activity. Endoglucanase also had maximum activity in 0.5% Avicel medium (approximately 820 UI/mg protein after 96 h incubation. In supernatants from 0.5% CMC, the maximum activity attained was 200 UI/mg protein only.Neste trabalho foram estudadas as atividades de endoglucanase, exocelobiohidrolase e celobiase em uma linhagem de Lentinula edodes (Berk. Pegl. cultivada em meio líquido contendo carboximetilcelulose (CMC ou celulose microcristalina (Avicel. Foram detectadas as atividades de endoglucanase e exocelobiohidrolase no sobrenadante das culturas crescidas tanto em meios contendo CMC como nos meios contendo Avicel, sendo observada a influência da concentração e do tipo de celulose. Não foi detectada atividade de celobiase nos sobrenadantes, sendo a mesma detectada somente no extrato micelial. Com uma concentração de 1,7% de Avicel, a linhagem estudada não demonstrou atividade de exocelobiohidrolase. Porém, à concentração de 0,5% obteve-se uma atividade de 74 µmol de RBBR/mg de proteína/min. Com a substituição de Avicel por CMC a 0,5%, a atividade de exocelobiohidrolase foi reduzida a menos de 50%. A máxima atividade de endoglucanase em sobrenadantes obtidos em meio com Avicel a 0,5% foi em torno de 800 UI/mg de proteína, após 96 horas de cultivo. Em sobrenadantes obtidos de meio com CMC, a atividade desta enzima foi de apenas 200 UI/mg de proteína.

  10. Glycoside Hydrolase (GH) 45 and 5 Candidate Cellulases in Aphelenchoides besseyi Isolated from Bird’s-Nest Fern

    Science.gov (United States)

    Tsay, Tung-Tsuan; Tsai, Isheng J.; Chen, Peichen J.

    2016-01-01

    Five Aphelenchoides besseyi isolates collected from bird’s-nest ferns or rice possess different parasitic capacities in bird’s-nest fern. Two different glycoside hydrolase (GH) 45 genes were identified in the fern isolates, and only one was found in the rice isolates. A Abe GH5-1 gene containing an SCP-like family domain was found only in the fern isolates. Abe GH5-1 gene has five introns suggesting a eukaryotic origin. A maximum likelihood phylogeny revealed that Abe GH5-1 is part of the nematode monophyletic group that can be clearly distinguished from those of other eukaryotic and bacterial GH5 sequences with high bootstrap support values. The fern A. besseyi isolates were the first parasitic plant nematode found to possess both GH5 and GH45 genes. Surveying the genome of the five A. besseyi isolates by Southern blotting using an 834 bp probe targeting the GH5 domain suggests the presence of at least two copies in the fern-origin isolates but none in the rice-origin isolates. The in situ hybridization shows that the Abe GH5-1 gene is expressed in the nematode ovary and testis. Our study provides insights into the diversity of GH in isolates of plant parasitic nematodes of different host origins. PMID:27391812

  11. Identification of a laccase from Ganoderma lucidum CBS 229.93 having potential for enhancing cellulase catalyzed lignocellulose degradation

    DEFF Research Database (Denmark)

    Sitarz, Anna Katarzyna; Mikkelsen, Jørn Dalgaard; Højrup, Peter;

    2013-01-01

    Based on a differential pre-screening of 44 white-rot fungi on a lignocellulose-supplemented minimal medium, four basidiomycetes were selected for further study: Ganoderma lucidum, Polyporus brumalis, Polyporus ciliatus and Trametes versicolor. Only G. lucidum was able to grow vividly on malt...... extract or minimal media supplemented with alkali lignin. When grown on malt extract or minimal medium supplemented with lignocellulose (sugar cane bagasse), the crude G. lucidum protein extract exhibited high laccase activity, ∼3U/mL toward syringaldazine. This activity was 13–17 fold higher than...... the corresponding activities of the crude protein extracts of P. brumalis, P. ciliatus and T. versicolor. Native PAGE electrophoresis of the crude G. lucidum extract confirmed the presence of an active laccase. The G. lucidum laccase had a molecular weight of ∼62.5kDa, and a Km value of 0.107mM (determined on ABTS...

  12. A graphene screen-printed carbon electrode for real-time measurements of unoccupied active sites in a cellulase

    DEFF Research Database (Denmark)

    Cruys-Bagger, Nicolaj; Tatsumi, Hirosuke; Borch, Kim;

    2014-01-01

    . In this work we describe a new electrochemical approach to the quantification of the populations of enzyme that are respectively free in the aqueous bulk, adsorbed to the insoluble substrate with an unoccupied active site or threaded with the cellulose strand in the active tunnel. Distinction of these three...... states appears essential to the idetification of the rate limiting step. The method is based on disposable graphene modified screen-printed carbon electrodes, and we show how the temporal development in the concentrations of the three enzyme forms can be derived from a combination of the electrochemical...... data and adsorption measuments. The approach was tested for the cellobiohydrolase Cel7A from Hypocrea jecorina acting on microcrystalline cellulose, and it was found that the threaded enzyme form dominates for this system while adsorbed enzyme with an unoccupied active site constitutes less than 5...

  13. Effect of Different Cellulase and Pectinase Enzyme Treatments on Protoplast Isolation and Viability in Lilium ledebeourii Bioss.

    OpenAIRE

    Esmaeil CHAMANI; Seyyed Karim TAHAMI; ZARE, Nasser; Rasool Asghari-ZAKARIA; Mehdi MOHEBODINI; Joyce, Daryl

    2012-01-01

    For overcoming interspecific incompatibility, protoplast combination method is a proper procedure for making a new plant withdesired traits. For this purpose, protoplast preparation is a first and important step. Hence, experiments were conducted to evaluatevarious combinations of cellulose, pectinase and their treatment times on protoplast production and protoplast viability in Liliumledebeourii Bioss. The results of experiment revealed that the protoplast yield was significantly affected by...

  14. PEA PEEL WASTE: A LIGNOCELLULOSIC WASTE AND ITS UTILITY IN CELLULASE PRODUCTION BY Trichoderma reesei UNDER SOLID STATE CULTIVATION

    OpenAIRE

    Nitin Verma; Mukesh C. Bansal; Vivek Kumar

    2011-01-01

    A wide variety of waste bioresources are available on our planet for conversion into bioproducts. In the biological systems, microorganisms are used to utilize waste as an energy source for the synthesis of valuable products such as biomass proteins and enzymes. The large quantities of byproducts generated during the processing of plant food involve an economic and environmental problem due to their high volumes and elimination costs. After isolation of the main constituent, there are abundan...

  15. The binding of cellulase variants to dislocations: a semi-quantitative analysis based on CLSM (confocal laser scanning microscopy) images

    DEFF Research Database (Denmark)

    Hidayat, Budi J.; Weisskopf, Carmen; Felby, Claus;

    2015-01-01

    Binding of enzymes to the substrate is the first step in enzymatic hydrolysis of lignocellulose, a key process within biorefining. During this process elongated plant cells such as fibers and tracheids have been found to break into segments at irregular cell wall regions known as dislocations or ...

  16. A graphene screen-printed carbon electrode for real-time measurements of unoccupied active sites in a cellulase

    DEFF Research Database (Denmark)

    Cruys-Bagger, Nicolaj; Tatsumi, Hirosuke; Borch, Kim;

    2014-01-01

    states appears essential to the idetification of the rate limiting step. The method is based on disposable graphene modified screen-printed carbon electrodes, and we show how the temporal development in the concentrations of the three enzyme forms can be derived from a combination of the electrochemical...

  17. Screening and characterization of a novel ruminal cellulase gene (Umcel-1) from a metagenomic library of gayal (Bos frontalis)

    Institute of Scientific and Technical Information of China (English)

    LI Bi-feng; MAO Hua-ming; YANG Shu-Li; ZHU Ya-xin; GU Zhao-bing; CHEN Yuan; LENG Jing; GOU Xiao; FENG Li; LI Qing; XI Dong-mei

    2016-01-01

    Gayal is a rare semi-wild bovine species found in the Indo-China. They can graze grasses, including bamboo leaves, as wel as reeds and other plant species, and grow to higher mature live weights than Yunnan Yelow cattle maintained in similar harsh environments. The aim of this study was to identify speciifc celulase in the gayal rumen. A metagenomic fosmid library was constructed using genomic DNA isolated from the ruminal contents of four adult gayals. This library contained 38400 clones with an average insert size of 35.5 kb. TheUmcel-1 gene was isolated from this library. Investigation of the celulase activity of 24 random clones led to the identiifcation of theUmcel-1 gene, which exhibited the most potent celulase activity. Sequencing theUmcel-1 gene revealed that it contained an open reading frame of 942 base pairs that encoded a product of 313 amino acids. The putativegene Umcel-1 product belonged to the glycosyl hydrolase family 5 and showed the highest homology to the celulase (GenBank accession no. YP_004310852.1) fromClostridium lentocelum DSM 5427, with 44% identity and 62% similarity. TheUmcel-1 gene was heterologously expressed inEscherichia coli BL21, and recombinant Umcel-1 was puriifed. The activity of puriifed recombinant Umcel-1 was assessed, and the results revealed that it hydrolyzed carboxymethyl celulose with optimal activity at pH 5.5 and 45°C. To our knowledge, this study provides the ifrst evidence for a celulase produced by bacteria in gayal rumen.

  18. Glycoside Hydrolase (GH) 45 and 5 Candidate Cellulases in Aphelenchoides besseyi Isolated from Bird's-Nest Fern.

    Science.gov (United States)

    Wu, Guan-Long; Kuo, Tzu-Hao; Tsay, Tung-Tsuan; Tsai, Isheng J; Chen, Peichen J

    2016-01-01

    Five Aphelenchoides besseyi isolates collected from bird's-nest ferns or rice possess different parasitic capacities in bird's-nest fern. Two different glycoside hydrolase (GH) 45 genes were identified in the fern isolates, and only one was found in the rice isolates. A Abe GH5-1 gene containing an SCP-like family domain was found only in the fern isolates. Abe GH5-1 gene has five introns suggesting a eukaryotic origin. A maximum likelihood phylogeny revealed that Abe GH5-1 is part of the nematode monophyletic group that can be clearly distinguished from those of other eukaryotic and bacterial GH5 sequences with high bootstrap support values. The fern A. besseyi isolates were the first parasitic plant nematode found to possess both GH5 and GH45 genes. Surveying the genome of the five A. besseyi isolates by Southern blotting using an 834 bp probe targeting the GH5 domain suggests the presence of at least two copies in the fern-origin isolates but none in the rice-origin isolates. The in situ hybridization shows that the Abe GH5-1 gene is expressed in the nematode ovary and testis. Our study provides insights into the diversity of GH in isolates of plant parasitic nematodes of different host origins. PMID:27391812

  19. DOWNREGULATION OF CINNAMYL-ALCOHOL DEHYDROGENASE IN SWITCHGRASS BY RNA SILENCING RESULTS IN ENHANCED GLUCOSE RELEASE AFTER CELLULASE TREATMENT

    Science.gov (United States)

    Cinnamyl alcohol dehydrogenase (CAD), catalyzes the last step in monolignol biosynthesis and genetic evidence indicates CAD deficiency in grasses both decreases overall lignin, alters lignin structure and increases enzymatic recovery of sugars. To ascertain the effect of CAD downregulation in switch...

  20. 黑胸散白蚁纤维素酶的体外酶学特性%In vitro Characteristics of the Cellulases from Reticulitermes chinensis Snyder

    Institute of Scientific and Technical Information of China (English)

    许利霞; 徐荣; 赵焕玉; 杨红

    2012-01-01

    Reticulitermes chinensis Snyder (Isoptera: Rhinotermitidae) is a wood-feeding lower termite widely distributed in China. For understanding the mechanisms of cellulose degradation in this termite, the in vitro enzyme activities of endo-β-l,4-glucanase (EG), exo-β-1,4-glucanase (namely cellobiohydrolase, CBH), and β-l,4-glucosidase (BG) from the worker termites of R. Chinensis were studied with DNS method. The results showed that for the three enzymes, the optimum reaction time, substrate concentration, pH and temperature were 15 min, 1%, 5.6 and 35 t, respectively. Under the above conditions, the activities of EG, CBH, and BG was 71.3(±13.9) U/mg, 5.8(±0.8) U/mg and 4.1(±0.7) U/mg, respectively. Endo-β-l,4-glucanase was not stable under high temperature in vitro. The incubation of crude enzyme at above 50 X. Would result in low activity or inactivation of the enzyme. However, this enzyme was relatively stable in the pH range from 3.2 to 8.0. Native-PAGE electrophoreses showed that there were at least eight different enzyme activity bands corresponding to endo-/M,4-glucanase distributing in different gut segments in R. Chinensis, which indicated the complexity of cellulose degradation process in wood-feeding lower termites. Fig 8, Ref 17%采用DNS法研究了我国广泛分布的一种低等木食性白蚁——黑胸散白蚁纤维素酶的体外酶活特性以了解其纤维素降解机制.结果表明,内切β-1,4-葡聚糖酶(EG)、纤维二糖水解酶(CBH)和β-葡萄糖苷酶(BG)这3种酶的最佳反应时间均为15 min,最佳底物浓度为1%,最适反应pH为5.6,最适反应温度为35℃.在最适反应条件下,EG、CBH和BG的活性分别达到71.3(±13.9) U/mg、5.8(±0.8) U/mg和4.1(±0.7) U/mg.EG在体外的热稳定性较差,在50℃及更高温度酶活很低或完全失活,但该酶对pH稳定性较好,在pH 3.2~8.0范围内酶活力变化不大.Native-PAGE电泳检测到该白蚁体内至少有8种不同的EG活性条带,肠道不同部位纤维素酶活性条带种类不同.这些研究表明,木食性白蚁降解纤维素是一个复杂的过程,需要多种纤维素酶的共同作用.

  1. Effects of cellulase and xylanase enzymes mixed with increasing doses ofSalix babylonicaextract onin vitrorumengas production kinetics of a mixture of corn silage with concentrate

    Institute of Scientific and Technical Information of China (English)

    Abdelfattah Z M Salem; German Buenda-Rodrguez; Mona M M Elghandour; Mara A Mariezcurrena Berasain; Francisco J Pea Jimnez; Alberto B Pliego; Juan C V Chagoyn; Mara A Cerrillo; Miguel A Rodrguez

    2015-01-01

    Anin vitro gas production (GP) technique was used to investigate the effects of combining different doses ofSalix babylonicaextract (SB) with exogenous ifbrolytic enzymes (EZ) based on xylanase (X) and celulase (C), or their mixture (XC; 1:1 v/v) onin vitrofermentation characteristics of a total mixed ration of corn silage and concentrate mixture (50:50, w/w)as substrate. Four levels of SB (0, 0.6, 1.2 and 1.8 mL g–1 dry matter (DM))andfour supplemental styles of EZ (1 µL g–1 DM; control (no enzymes), X, C and XC (1:1, v/v) were used in a 4×4 factorial arrangement.In vitro GP (mL g–1 DM) were recorded at 2, 4, 6, 8, 10, 12, 24, 36, 48 and 72 h of incubation. After 72 h, the incubation process was stopped and supernatant pH was determined, and then ifltered to determine dry matter degradability (DMD). Fermentation parameters, such as the 24 h gas yield (GY24),in vitro organic matter digestibility (OMD), metabolizable energy (ME), short chain fatty acid concentrations (SCFA), and microbial crude protein production (MCP) were also estimated. Results indicated that there was a SB´EZ interaction (P0.05) on OMD, pH, ME, GY24, SCFA and MP. The combination of SB with EZ increased (P<0.001) OMD, ME, SCFA, PF72 and GP24, whereas there was no impact on pH. It could be concluded that addition of SB extract, C, and X effectively improved thein vitro rumen fermentation, and the combination of enzyme with SB extract at the level of 1.2 mL g–1 was more effective than the other treatments.

  2. Production of ethanol from lepthochloa fusca L. (kallar grass) and panicum maximum using cellulases from trichoderma SSP and cultures of saccharomyces carlsbergensis

    International Nuclear Information System (INIS)

    Saline sodic soils have been used for production of biomass using salt tolerant grass, kallar grass followed by lesser tolerant plants, namely, sasbania aculeata or Panicum maximum or to provide biomass throughout the year for it utilization for microbial conversion. These substrates have been utilized to produce single cell protein and cellases or hemicellulases. The enzyme titer obtained after growth of cellulomonas biazotea on kallar grass could saccharify wheat straw ad bagasse for subsequent conversion to ethanol but kallar grass itself was saccharified to lesser extent, however, enzyme titres from different fungi could saccherify the biomass produced on saline lands to monomeric sugars. In these studies, the enzyme titre from Trichoderma spp. were used for saccharification purpose. (author)

  3. Influence of gamma-rays and some cultural conditions on the enhancement of cellulase production by some fungal strains isolated from cellulosic wastes

    International Nuclear Information System (INIS)

    In the present study, out of 51 fungal strains isolated from the cellulosic wastes, only 19 were CMCase-producers. Aspergillus, Fusarium and Penicillium were the most common fungal genera isolated from the cellulosic wastes. Fusarium neoceras, Aspergillus fumigatus and Fusarium oxysporium produced CMCase activity than Trichoderma viride. Out of 23 gamma-irradiated survivors from A.fumigatus and F. neoceras showing CMCase production, only two mutant strains A.fumigatus 8G-2 and F. neoceras 4G-2 produced the highest levels of CMCase than the parent strains. The results indicated that the maximum level of of CMCase activity was produced by A.fumigatus and F. neoceras strains under optiminizing conditions.

  4. Screening and Identifying of a Cellulase-Producing Strain%一株纤维素酶产生菌的筛选鉴定

    Institute of Scientific and Technical Information of China (English)

    张强; 杨岩

    2008-01-01

    用CMC平板筛选方法,从草丛土样中筛选到一株产纤维素酶的菌株,该菌在20℃~50℃、pH4.0~10.0.NaC1 1.0%~10.0%的条件下均能生长,在26℃~39℃、pH6.0~8.0、NaC1 4.0%~6.0%条件下生长快速.经采用Biolog进行碳源利用鉴定判定为黄曲霉,命名为AF-1.AF-1的CMCase为19.26U/mL,FPA为6.22U/mL,纤维分解率为20.6%.采用单因素法对AF-1产酶条件进行了初步测定.

  5. Real-Time Imaging of Plant Cell Wall Structure at Nanometer Scale, with Respect to Cellulase Accessibility and Degradation Kinetics (Presentation)

    Energy Technology Data Exchange (ETDEWEB)

    Ding, S. Y.

    2012-05-01

    Presentation on real-time imaging of plant cell wall structure at nanometer scale. Objectives are to develop tools to measure biomass at the nanometer scale; elucidate the molecular bases of biomass deconstruction; and identify factors that affect the conversion efficiency of biomass-to-biofuels.

  6. 纤维素酶促进菠萝皮果胶提取的研究%Extraction of Pectin from Pineapple Peel Promoted by Cellulase

    Institute of Scientific and Technical Information of China (English)

    李胜; 黄兰

    2011-01-01

    为了促进菠萝皮中果胶的提取,对采用酸解法处理后的菠萝皮酸解液作进一步的纤维素酶酶解处理,且初步确定此工艺的最佳酶解条件:酶解温度为55℃,菠萝皮酸解液pH值为4.2,单位菠萝皮酸解液中纤维素酶用量为15.5 U·mL-1,酶解时间为140 min。在此工艺条件下,酶解液中果胶含量可达7.4 g·L-1,比菠萝皮酸解液中的果胶含量提高了40.7%。%In order to promote the extraction of pectin from pineapple peel,the acid hyclrolyzed suspending solution of pineapple peel was treated with cellulose,and the conditions of enzymatic hydrolysis process were screened.The results indicated that the optimal conditions were as follows: enzymatic hydrolysis temperature was 55℃;enzymatic hydrolysis pH volume was pH 4.2;appending quantity of cellulose was 15.5 U·mL-1;and enzymatic hydrolysis time was 140 min,and under these conditions,the concentration of pectin in the enzymatic hyclrolyzed solution reached 7.4 g·L-1,which was 40.7% higher than that of the acid hyclrolyzed solution.

  7. 纤维素酶在可再生资源利用中的作用%The role of cellulase in utilization of renewable resource

    Institute of Scientific and Technical Information of China (English)

    安登第

    2003-01-01

    @@ 纤维素是组成植物体的主要物质,是地球上最丰富的可再生资源,但这类物质大多以秸秆的形式存在,难以被降解,人类对其利用极其有限.如能将纤维类物质转化为人类食品,由于其资源量极大,因而有非常重要的意义.当前转化的主要途径是通过反刍动物.反刍动物在严酷的自然进化中获得的前胃缓慢发酵系统,使其成为哺乳动物中最有效的粗饲料消化者.但秸秆饲料中,大量结构碳水化合物被木质素保护,瘤胃微生物不能降解木质素致使这些碳水化合物不能被利用(McSweeney等,1994).

  8. Las celulasas y su aplicación en la degradación de desechos agroindustriales The cellulases and their application in degrading agro-industrial waste

    Directory of Open Access Journals (Sweden)

    Schwarz Wolfgang H.

    2002-06-01

    Full Text Available Una cantidad de biomasa lignocelulósica está disponible y puede ser usada para producir energía almacenable, material básico de la industria química. Su uso es especialmente benéfico para un país, si esta biomasa hace parte de material de desecho que pude ser obtenido casi sin ningún costo y está presente en la carga ambiental. A pesar de esto, los polisacáridos presentes en ese tipo de biomasas son difíciles de degradar debido a su heterogeneidad y a su estructura cristalina. Este artículo está dirigido a la hidrólisis enzimática de la celulosa realizada por microorganismos que la degradan, las bacterias anaeróbicas. Se explican las dificultades para la digestión de la celulosa, así como las estrategias usadas por hidrolasas y complejos enzima ticos que permiten una degradación eficiente. La especificidad química de todas estas enzimas es idéntica, pero el modo de acción de cada uno de sus componentes es diferente. Sólo cuando se combinan con módulos de unión, se realiza una hidrólisis lenta pero eficiente. La variación de las estructuras modulares entre cada familia son un ejemplo de la diversificación evolutiva de la actividad enzimática. Se analiza un modelo para la hidrólisis de la celulosa en bruto (como está presente en la naturaleza; sin embargo, no se han realizado muchas investigaciones para explicar y describir el proceso a nivel molecular ni para optimizar la hidrólisis de la celulosa en el ámbito industrial. A huge amount of lignocellulosic biomass is available which can be used to produce storable energy and basic material for the chemical industry. Its use is especially beneficial for a country's economy if it is waste material, which can be obtained at almost no cost and which presents an environmental burden. However, the polysaccharides present in biomass are difficult to degrade due to their heterogeneity and crystalline structure. This article addresses the enzymatic hydrolysis of cellulose by its natural degraders, the anaerobic bacteria. The difficulties of cellulose digestion are explained and the strategies used by the hydrolytic enzymes and enzyme systems, allowing for efficient degradation. The multitude of enzymes is uniform in having an identical chemical specificity, but differs in each component's action mode. Only by combining this with binding modules can efficient hydrolysis be performed. The variation of modular structures within a single enzyme family is an example of enzymatic activity's evolutionary diversification. A model for hydrolytically degrading natural cellulose is presented, but much more research has to be done to explain and describe the process on the molecular level, and to optimize an industrial enzymatic cellulose hydrolysis process.

  9. Root cap specific expression of an endo-beta-1,4-D-glucanase (cellulase): a new marker to study root development in Arabidopsis.

    Science.gov (United States)

    del Campillo, Elena; Abdel-Aziz, Amal; Crawford, Damian; Patterson, Sara E

    2004-09-01

    The sloughing of root cap cells from the root tip is important because it assists the growing root in penetrating the soil. Using a promoter-reporter (GUS) and RT-PCR analysis, we identified an endo-beta-1,4-glucanase (AtCel5) of Arabidopsis thaliana that is expressed exclusively in root cap cells of both primary and secondary roots. Expression is inhibited by high concentrations of IAA, both exogenous and internal, as well as by ABA. AtCel5 expression begins once the mature tissue pattern is established and continues for 3 weeks. GUS staining is observed in both root cap cells that are still attached and cells that have already been shed. Using AtCel5-GUS as a marker, we observed that the root cap cells begin to separate at the sides of the tip while the cells of the central region of the tip separate last. Separation involves sequential tiers of intact cells that separate from the periphery of the root tip. A homozygous T-DNA insertion mutant that does not express AtCel5 forms the root cap and sheds root cap cells but sloughing is less efficient compared to wild type. The reduction in sloughing in the mutant does not affect the overall growth performance of the plant in loose media. The modest effect of abolishing AtCel5 expression suggests that there are multiple redundant genes regulating the process of sloughing of the root cap, including AtCel3/At1g71380, the paralog of the AtCel5 gene that is also expressed in the root cap cells. Thus, these two endo-1,4-beta-D-glucanases may have a role in the sloughing of border cells from the root tip. We propose that AtCel5, provides a new molecular marker to further analyze the process of root cap cell separation and a root cap specific promoter for targeting to the environment genes with beneficial properties for plant growth.

  10. Cloning, Sequencing and Expression Analysis of the First Cellulase Gene Encoding Cellobiohydrolase 1 from a Cold-adaptive Penicillium chrysogenum FS010

    Institute of Scientific and Technical Information of China (English)

    Yunhua HOU; Tianhong WANG; Hao LONG; Huiyuan ZHU

    2007-01-01

    A cellobiohydrolase 1 gene (cbh1) was cloned from Penicillium chrysogenum FS010 by a modified thermal asymmetric interlaced polymerase chain reaction (TAIL-PCR). DNA sequencing shows that cbh1 has an open reading frame of 1590 bp, encoding a putative protein of 529 amino acid residues. The deduced amino acid sequence revealed that CBHI has a modular structure with a predicted molecular mass of 56 kDa and consists of a fungal type carbohydrate binding module separated from a catalytic domain by a threonine rich linker region. The putative gene product is homologous to fungal cellobiohydrolases in Family 7 of the glycosyl hydrolases. A novel cbh1 promoter (1.3 kb) was also cloned and sequenced, which contains seven putative binding sites (5'-SYGGRG-3') for the carbon catabolite repressor CRE1. Effect of various carbon sources to the cbh1 transcription of P. chrysogenum was examined by Northern analysis,suggesting that the expression of cbh1 is regulated at transcriptional level. The cbh1 gene in cold-adaptive fungus P. chysogenum was expressed as an active enzyme in Saccharomyces cerevisiae H158. The recombinant CBHI accumulated intracellularly and could not be secreted into the medium.

  11. Direct ethanol production from cassava pulp using a surface-engineered yeast strain co-displaying two amylases, two cellulases, and {beta}-glucosidase

    Energy Technology Data Exchange (ETDEWEB)

    Apiwatanapiwat, Waraporn; Rugthaworn, Prapassorn [Japan International Research Center for Agricultural Sciences (JIRCAS), Tsukuba, Ibaraki (Japan). Post-Harvest Science and Technology Div.; Kasetsart Univ., Bangkok (Thailand). Nanotechnology and Biotechnology Div.; Murata, Yoshinori; Kosugi, Akihiko; Arai, Takamitsu; Mori, Yutaka [Japan International Research Center for Agricultural Sciences (JIRCAS), Tsukuba, Ibaraki (Japan). Post-Harvest Science and Technology Div.; Yamada, Ryosuke; Kondo, Akihiko [Kobe Univ. (Japan). Dept. of Chemical Science and Engineering

    2011-04-15

    In order to develop a method for producing fuel ethanol from cassava pulp using cell surface engineering (arming) technology, an arming yeast co-displaying {alpha}-amylase ({alpha}-AM), glucoamylase, endoglucanase, cellobiohydrase, and {beta}-glucosidase on the surface of the yeast cells was constructed. The novel yeast strain, possessing the activities of all enzymes, was able to produce ethanol directly from soluble starch, barley {beta}-glucan, and acid-treated Avicel. Cassava is a major crop in Southeast Asia and used mainly for starch production. In the starch manufacturing process, large amounts of solid wastes, called cassava pulp, are produced. The major components of cassava pulp are starch (approximately 60%) and cellulose fiber (approximately 30%). We attempted simultaneous saccharification and ethanol fermentation of cassava pulp with this arming yeast. During fermentation, ethanol concentration increased as the starch and cellulose fiber substrates contained in the cassava pulp decreased. The results clearly showed that the arming yeast was able to produce ethanol directly from cassava pulp without addition of any hydrolytic enzymes. (orig.)

  12. Effects of Exogenous NSP Enzymes(Xylanase, β-glucanase and Cellulase) on Morphology and Functions of Digestive Tract in Growing Pigs Fed with Paddy-Based Diets

    Institute of Scientific and Technical Information of China (English)

    XU Zi-rong; LU Jian-jun

    2003-01-01

    Ninety Landrace X Jia 35±0.40 kg weight growing pigs were randomly allotted to three treatments, each of which was replicated three times with ten pigs per replicate. The pigs were reared on either a conventional corn-based diet (control Ⅰ ) or a paddy-based diet (control Ⅱ ) or a paddy diet supplemented with 0.2% NSP enzymes (test group). All pigs were given ad libitum access to both feed and water. The results of feeding trial showed that supplementation of NSP enzymes significantly increased ADG by 8.78 % (P<0.05) and decreased F/G by 9.42% (P<0.05) over the control group Ⅱ. No significant differences were found in ADG and F/G between control group I and the test group. The digestive trial showed that adding NSP enzymes significantly improved apparent digestibility of CP, EE and CF by 18.76 (P<0.01), 16.04 (P <0.05) and 108.57%(P<0.05), respectively, compared to control Ⅱ. The activities of proteolytic enzyme and α-amylase in duodenal contents were increased by 99.07 (P<0.01) and 18.41% (P<0. 05) with the addition of NSP enzymes. No significant differences between test and control Ⅱ group were found in activities of the pepsin in the gastric content, the trypsin and lipase in duodenal contents, the disaccharidase and γ-glutany transferase (γ-GT) in intestinal mucosa, but there was a tendency towards higher activities associated with the NSP enzymes diet (P>0. 05). The lengths of the villi within the duodenal, jejunal and ileal sections of the small intestine of pigs receiving the NSP enzymes diet were increased by 23.68 (P<0. 05), 56.00 (P<0. 01)and 76.90%(P<0.01) respectively, relative to the pigs in control Ⅱ.

  13. Nucleotide polymorphisms in a pine ortholog of the Arabidopsis degrading enzyme cellulase KORRIGAN are associated with early growth performance in Pinus pinaster.

    Science.gov (United States)

    Cabezas, José Antonio; González-Martínez, Santiago C; Collada, Carmen; Guevara, María Angeles; Boury, Christophe; de María, Nuria; Eveno, Emmanuelle; Aranda, Ismael; Garnier-Géré, Pauline H; Brach, Jean; Alía, Ricardo; Plomion, Christophe; Cervera, María Teresa

    2015-09-01

    We have carried out a candidate-gene-based association genetic study in Pinus pinaster Aiton and evaluated the predictive performance for genetic merit gain of the most significantly associated genes and single nucleotide polymorphisms (SNPs). We used a second generation 384-SNP array enriched with candidate genes for growth and wood properties to genotype mother trees collected in 20 natural populations covering most of the European distribution of the species. Phenotypic data for total height, polycyclism, root-collar diameter and biomass were obtained from a replicated provenance-progeny trial located in two sites with contrasting environments (Atlantic vs Mediterranean climate). General linear models identified strong associations between growth traits (total height and polycyclism) and four SNPs from the korrigan candidate gene, after multiple testing corrections using false discovery rate. The combined genomic breeding value predictions assessed for the four associated korrigan SNPs by ridge regression-best linear unbiased prediction (RR-BLUP) and cross-validation accounted for up to 8 and 15% of the phenotypic variance for height and polycyclic growth, respectively, and did not improve adding SNPs from other growth-related candidate genes. For root-collar diameter and total biomass, they accounted for 1.6 and 1.1% of the phenotypic variance, respectively, but increased to 15 and 4.1% when other SNPs from lp3.1, lp3.3 and cad were included in RR-BLUP models. These results point towards a desirable integration of candidate-gene studies as a means to pre-select relevant markers, and aid genomic selection in maritime pine breeding programs. PMID:26093373

  14. The Cellulases Endoglucanase I and Cellobiohydrolase II of Trichoderma reesei Act Synergistically To Solubilize Native Cotton Cellulose but Not To Decrease Its Molecular Size

    OpenAIRE

    Kleman-Leyer, K. M.; Siika-Aho, M.; Teeri, T. T.; Kirk, T K

    1996-01-01

    Degradation of cotton cellulose by Trichoderma reesei endoglucanase I (EGI) and cellobiohydrolase II (CBHII) was investigated by analyzing the insoluble cellulose fragments remaining after enzymatic hydrolysis. Changes in the molecular-size distribution of cellulose after attack by EGI, alone and in combination with CBHII, were determined by size exclusion chromatography of the tricarbanilate derivatives. Cotton cellulose incubated with EGI exhibited a single major peak, which with time shift...

  15. Las celulasas y su aplicación en la degradación de desechos agroindustriales The cellulases and their application in degrading agro-industrial waste

    OpenAIRE

    Schwarz Wolfgang H.

    2002-01-01

    Una cantidad de biomasa lignocelulósica está disponible y puede ser usada para producir energía almacenable, material básico de la industria química. Su uso es especialmente benéfico para un país, si esta biomasa hace parte de material de desecho que pude ser obtenido casi sin ningún costo y está presente en la carga ambiental. A pesar de esto, los polisacáridos presentes en ese tipo de biomasas son difíciles de degradar debido a su heterogeneidad y a su estructura cristalina. Este artículo e...

  16. The cellulases and their application in degrading agro-industrial waste Las celulasas y su aplicación en la degradación de desechos agroindustriales

    OpenAIRE

    Schwarz Wolfgang H.

    2002-01-01

    A huge amount of lignocellulosic biomass is available which can be used to produce storable energy and basic material for the chemical industry. Its use is especially beneficial for a country's economy if it is waste material, which can be obtained at almost no cost and which presents an environmental burden. However, the polysaccharides present in biomass are difficult to degrade due to their heterogeneity and crystalline structure. This article addresses the enzymatic hydrolysis of cellulos...

  17. Trichoderma viride 菌生物量测定及其纤维素酶合成特征%Trichoderma viride Biomass Assay and Its Cellulase Synthesis Characteristic

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    利用HPLC法测定Trichoderma viride菌固态发酵曲中的麦角固醇含量.研究了麦角固醇与菌丝体间的关系.该菌固态曲中麦角固醇分离条件以1∶25(m/v)的丙酮抽提1.5 h为最佳.当固态发酵培养至69 h时,曲中的生物量达到最大值,为每克干曲中含有0.575 g菌丝体.此时该菌所产生CMC酶和FP酶活力均达到最大值,呈现正相关性,说明这2种酶的合成特征均为同步合成型,而C1酶活力高峰滞后,出现在72 h.

  18. Defining the Interactions of Cellobiohydrolase with Substrate through Structure Function Studies: Cooperative Research and Development Final Report, CRADA Number CRD-10-409

    Energy Technology Data Exchange (ETDEWEB)

    Beckham, G. T.; Himmel, M. E.

    2013-07-01

    NREL researchers will use their expertise and skilled resources in numerical computational modeling to generate structure-function relationships for improved cellulase variant enzymes to support the development of cellulases with improved performance in biomass conversion.

  19. Vertical Integration of Biomass Saccharification of Enzymes for Sustainable Cellulosic Biofuel Production in a Biorefinery

    Energy Technology Data Exchange (ETDEWEB)

    Manoj Kumar, PhD

    2011-05-09

    Lignocellulosic biomass is the most abundant, least expensive renewable natural biological resource for the production of biobased products and bioenergy is important for the sustainable development of human civilization in 21st century. For making the fermentable sugars from lignocellulosic biomass, a reduction in cellulase production cost, an improvement in cellulase performance, and an increase in sugar yields are all vital to reduce the processing costs of biorefineries. Improvements in specific cellulase activities for non-complexed cellulase mixtures can be implemented through cellulase engineering based on rational design or directed evolution for each cellulase component enzyme, as well as on the reconstitution of cellulase components. In this paper, we will provide DSM's efforts in cellulase research and developments and focus on limitations. Cellulase improvement strategies based on directed evolution using screening on relevant substrates, screening for higher thermal tolerance based on activity screening approaches such as continuous culture using insoluble cellulosic substrates as a powerful selection tool for enriching beneficial cellulase mutants from the large library. We will illustrate why and how thermostable cellulases are vital for economic delivery of bioproducts from cellulosic biomass using biochemical conversion approach.

  20. Integrated cellulosic enzymes hydrolysis and fermentative advanced yeast bioconversion solution ready for biomass biorefineries

    Energy Technology Data Exchange (ETDEWEB)

    Manoj Kumar, PhD

    2011-05-04

    Lignocellulosic biomass is the most abundant, least expensive renewable natural biological resource for the production of biobased products and bioenergy is important for the sustainable development of human civilization in 21st century. For making the fermentable sugars from lignocellulosic biomass, a reduction in cellulase production cost, an improvement in cellulase performance, and an increase in sugar yields are all vital to reduce the processing costs of biorefineries. Improvements in specific cellulase activities for non-complexed cellulase mixtures can be implemented through cellulase engineering based on rational design or directed evolution for each cellulase component enzyme, as well as on the reconstitution of cellulase components. In this paper, we will provide DSM's efforts in cellulase research and developments and focus on limitations. Cellulase improvement strategies based on directed evolution using screening on relevant substrates, screening for higher thermal tolerance based on activity screening approaches such as continuous culture using insoluble cellulosic substrates as a powerful selection tool for enriching beneficial cellulase mutants from the large library. We will illustrate why and how thermostable cellulases are vital for economic delivery of bioproducts from cellulosic biomass using biochemical conversion approach.

  1. Fully Integrated Lignocellulosic Biorefinery with Onsite Production of Enzymes and Yeast

    Energy Technology Data Exchange (ETDEWEB)

    Manoj Kumar, PhD

    2010-06-14

    Lignocellulosic biomass is the most abundant, least expensive renewable natural biological resource for the production of biobased products and bioenergy is important for the sustainable development of human civilization in 21st century. For making the fermentable sugars from lignocellulosic biomass, a reduction in cellulase production cost, an improvement in cellulase performance, and an increase in sugar yields are all vital to reduce the processing costs of biorefineries. Improvements in specific cellulase activities for non-complexed cellulase mixtures can be implemented through cellulase engineering based on rational design or directed evolution for each cellulase component enzyme, as well as on the reconstitution of cellulase components. In this paper, we will provide DSM's efforts in cellulase research and developments and focus on limitations. Cellulase improvement strategies based on directed evolution using screening on relevant substrates, screening for higher thermal tolerance based on activity screening approaches such as continuous culture using insoluble cellulosic substrates as a powerful selection tool for enriching beneficial cellulase mutants from the large library. We will illustrate why and how thermostable cellulases are vital for economic delivery of bioproducts from cellulosic biomass using biochemical conversion approach.

  2. Development of a Commerical Enzyme System for Lignocellulosic Biomass Saccharification

    Energy Technology Data Exchange (ETDEWEB)

    Manoj Kumar, PhD

    2011-02-14

    Lignocellulosic biomass is the most abundant, least expensive renewable natural biological resource for the production of biobased products and bioenergy is important for the sustainable development of human civilization in 21st century. For making the fermentable sugars from lignocellulosic biomass, a reduction in cellulase production cost, an improvement in cellulase performance, and an increase in sugar yields are all vital to reduce the processing costs of biorefineries. Improvements in specific cellulase activities for non-complexed cellulase mixtures can be implemented through cellulase engineering based on rational design or directed evolution for each cellulase component enzyme, as well as on the reconstitution of cellulase components. In this paper, we will provide DSM's efforts in cellulase research and developments and focus on limitations. Cellulase improvement strategies based on directed evolution using screening on relevant substrates, screening for higher thermal tolerance based on activity screening approaches such as continuous culture using insoluble cellulosic substrates as a powerful selection tool for enriching beneficial cellulase mutants from the large library. We will illustrate why and how thermostable cellulases are vital for economic delivery of bioproducts from cellulosic biomass using biochemical conversion approach.

  3. 纤维素酶提取冬枣叶中总黄酮工艺的研究%Study on extraction technology of total flavones using cellulase from leaves of winter-jujube

    Institute of Scientific and Technical Information of China (English)

    张圣燕; 张成

    2013-01-01

    Total flavones in leaves of winter-jujube were extracted using cellulose.The total flavones extraction rate was determined by spectrophotometry method with rutin as reference substance.In the single factor analysis,the effect of extraction temperature, extraction time, enzyme amount and solid - liquid ratio were measured. The orthogonal test was used to optimize the conditions of extraction.The results showed that the best conditions of extraction total flavones from leaves of winter - jujube were the solid - liquid ratio 1 : 50 ( g/mL), extraction temperature 55?, extraction time 90min, enzyme amount 4mg/g. Under this optimal condition, extraction rate was 2.45%.The results showed that the order of extraction rate of total flavones was as follows;extraction temperature > extraction time > enzyme amount > solid-liquid ratio.%采用纤维素酶提取冬枣叶中的总黄酮.以芦丁为标准品,采用分光光度法测定冬枣叶中总黄酮的含量,通过单因素实验研究提取温度、提取时间、酶用量和料液比对提取率的影响,再利用正交实验优化最佳提取工艺条件.结果表明,纤维素酶提取冬枣叶中总黄酮的最佳工艺条件为:料液比1:50(g/mL),提取温度55℃,提取时间90min,酶用量4mg/g.在最佳提取工艺条件下,提取率可达2.45%.同时得到各影响因素对总黄酮提取率的显著性影响顺序为:提取温度>提取时间>酶用量>料液比.

  4. Isolation of cellulose-degradation thermophilic bacteria and character of cellulase%降解纤维素嗜热菌的分离及纤维素酶性质分析

    Institute of Scientific and Technical Information of China (English)

    陈路劼; 刘斌; 林白雪; 何柳; 张宁; 吴祖建; 谢联辉

    2010-01-01

    从福州永泰温泉分离到1株降解纤维素嗜热菌,通过形态特征和16S rDNA序列鉴定,将其命名为地芽孢杆菌(Geobacillus sp.)TC-S8.采用3,5-二硝基水杨酸显色法(DNS法)对菌株TC-S8产胞外纤维素酶的发酵条件及酶性质进行研究.在培养基的初始pH为6.5,温度为60 ℃,装液量20%,接种量2%,摇床转速为120 r·min~(-1)的条件下培养36 h,菌株TC-S8产酶量达到最大(229 mU·mL~(-1)).其分泌的纤维素酶最适作用温度为60-70 ℃,最适pH为7.0.纤维素酶降解产物主要为葡萄糖,并有少量纤维二糖.

  5. Optimization on Solid Fermentation of Corn Stover for the Production of Cellulase%固体发酵玉米秸秆产纤维素酶条件的优化

    Institute of Scientific and Technical Information of China (English)

    袁丽环

    2011-01-01

    以玉米秸秆为原料,利用菌株发酵生产纤维素酶,通过单因素试验和正交试验,考察了培养温度、接种量、发酵时间对纤维素酶活力的影响,确定了固体发酵玉米秸秆产纤维素酶的最佳工艺参数.试验结果表明,各因素对纤维素酶活力的影响程度由大到小依次为培养温度、接种量、发酵时间;最佳发酵条件为接种量8%,培养温度30℃的条件下发酵66 h,具有最大产酶量142.55 U/ml.

  6. 基于Taguchi试验设计稻草发酵产纤维素酶的优化%Optimization of the Fermentation of Straw to Produce Cellulase Based on Taguchi Design

    Institute of Scientific and Technical Information of China (English)

    辛婷; 谢逸萍; 张兴; 马代夫; 王慧

    2009-01-01

    采用Taguchi设计法综合考察接种量、硫酸铵、磷酸二氢钾、七水硫酸镁等因素对实验室筛选出的1株木霉菌种进行产纤维素酶的影响.结果表明,接种量时产酶的影响最大,其次是硫酸铵、七水硫酸镁,而磷酸二氢钾对产酶酶活的影响最小;Taguchi设计优化后得到产纤维素酶的最优条件为:接种量为8%,硫酸铵、磷酸二氢钾、七水硫酸镁添加量分别为0.6%、0.25%和0.03%.在最优条件下预期纤维素酶活能达到6.276μmol/g·min,而验证性试验证明在优化条件下,纤维素酶活达到了5.222 μmol/g·min,比优化前的纤维素酶活平均值3.128 μmol/g·min增加了2.094μmol/g·min,表明Taguchi方法对于产纤维素酶优化具有较好的应用价值.

  7. Cloning and Sequence Analysis of Cellulase Genes from Trichoderma longibrachiatum%长梗木霉纤维素酶基因的克隆及序列分析

    Institute of Scientific and Technical Information of China (English)

    石贤爱; 刘月; 陈飞; 杨锦

    2010-01-01

    从富含纤维素环境筛选获得一株纤维素降解菌株FU05,通过形态学特征及ITS序列分析确定其为长梗木霉(Tnchoderma longibrachiatum).PCR扩增获得该菌株的bgl2,cbh2和egl.序列分析表明,这3种纤维素酶基因与GenBank上其他木霉同种纤维素酶基因具有较高同源性:bgl2基因与里氏木霉bgl2基因(AB003110)同源性达91%;cbh2基因与康宁木霉cbh2基因(DQ504304)同源性达99%;egl基因与长梗木霉egl基因(X60652)同源性达95%.3种纤维素酶基因编码的相应氨基酸序列与其他木霉纤维素酶的氨基酸序列相似性也非常高.对上述纤维素酶基因编码的相应蛋白进行PROSITE motif search,对其N端糖基化位点、纤维素结合区、糖基水解酶家族特征结构区等进行了定位.

  8. 纤维素酶提取湘西椪柑皮中总黄酮的工艺优化研究%Extraction of Total Flavonoids from the Peel of 'Ponkan' Mandarin (Citrus reticulata Blanco) by Optimized Cellulase Hydrolysis

    Institute of Scientific and Technical Information of China (English)

    欧阳辉; 余佶; 张永康

    2010-01-01

    采用响应面法对纤维素酶提取椪柑皮中总黄酮的工艺进行优化.结果表明,酶法提取椪柑皮总黄酮的最铑佳工艺条件为酶用量120U/mL、酶解温度52℃、酶解时间140min、酶解pH4.35,总黄酮得率可达1.85%.

  9. 枝状枝孢菌(Cladosporium cladosporioides)发酵产酶培养基的响应面法优化%Response surface optimization of medium components for producing cellulase by Cladosporium cladosporioides F4-1

    Institute of Scientific and Technical Information of China (English)

    韩立荣; 王永宏; 何军; 安凤秋; 张兴

    2010-01-01

    采用响应面法对枝状枝孢菌(Cladosporium cladosporioides)产葡聚糖内切酶(CMCase)的液体发酵培养基进行优化.用单因素试验确定发酵培养基的最佳碳源、氮源和无机盐;通过中心组合试验和响应面法优化以上因素,得到的优化发酵培养基为:麦秸23.18 g·L~(-1)、麦麸30.00 g·L~(-1)、酵母膏13.62 g·L~(-1)、KH_2PO_4 4.60 g·L~(-1)、NaCl 4.60 g·L~(-1)、MgSO_4·7H_2O 0.46 g·L~(-1).在此条件下,该菌发酵液中CMCase活力达2.80 U·mL~(-1),较优化前提高了2.18倍.

  10. The process study of cellulase assisted acid extraction of pineapple peel pectin%纤维素酶辅助酸法提取菠萝皮果胶的工艺研究

    Institute of Scientific and Technical Information of China (English)

    姚春波; 来丽丽

    2013-01-01

    目的 考察影响酸法提取菠萝皮果胶的主要因素,并确定酸法提取果胶的最佳工艺条件.方法 通过正交试验,考察料液比、提取温度、提取时间和提取酸度对菠萝皮果胶提取的影响,从而得到最佳的提取工艺.结果 纤维素酶辅助酸法提取菠萝皮果胶的最佳工艺务件为:料液比1:20,提取温度80℃,提取时间60 min,提取pH 2.0.结论 纤维素酶辅助酸法具有提取率高、速度快、溶剂用量少、安全、节能、设备简单等优点,试验为优化菠萝皮果胶的提取工艺奠定了基础.

  11. DIFFERENT APPROACHES OF USING MULTIPLE CULTURE ISOLATES- FUSARIUM OXYSPORUM F8, PENICILLIUM NOTATUM 101 AND ASPERGILLIUS NIGER F7 FOR HIGHER PRODUCTION OF CELLULASE AND XYLANASE FROM PINUS ROXBURHII NEEDLES

    OpenAIRE

    DIVYA TANDON; NIVEDITA SHARMA; RICHA KAUSHAL

    2013-01-01

    Diminishing fossil fuel reserve and increasing cost of fossil products have rekindled effort on conversion of lignocelluloses to renewable fuel. Among various lignocelluloses' biomass, Pinus roxburghii is a predominant forest species which is scattered throughout the world. The continous shedding of pine needles and its exceptionally slow biodegradation in nature leads to huge accumulation of needles on earth by posing a serious threat to our environment which affects th...

  12. SAS 优化紫外诱变菌发酵产酶培养基及酶性质探讨%Study on optimization of UV-induced mutation fermentation medium for cellulase by SAS and its crude properties

    Institute of Scientific and Technical Information of China (English)

    米芳; 马英辉; 高锦明

    2010-01-01

    利用SAS中的Plackett-Burman二水平设计法成功地选取了对宇佐美曲霉固态发酵小麦秸秆产纤维素酶影响显著的4个因素(初始pH、尿素浓度、含水量和C源比例),并依据Box-Benhnken设计原则进行响应面回归模型分析(RSA),得出最佳的水平为:培养基初始pH为6.2、尿素浓度为1.54%、含水量为1.25:1、秸秆与麸皮的比例为4.6:5.4,在此务件下CMC酶和FPU酶的酶活分别达到1224.7u/g和254.5u/g;对粗酶液的酶学性质研究发现,CMC酶和FPU酶的最适作用pH分别为4.5和6.0,最适温度为65℃和50℃.

  13. Effects of Cellulase Origination on in vitro Fermentation Characteristics and Methane Production of Forage%纤维素酶来源对粗饲料体外发酵特性及甲烷产量的影响

    Institute of Scientific and Technical Information of China (English)

    邹瑶; 汤少勋; 谭支良; 刘勇; 冉涛

    2013-01-01

    采用体外发酵技术研究了瘤胃真菌、长柄木霉和李氏木霉三种来源纤维素酶在4种活性水平(0 IU/kg DM、1200IU/kgDM、3700 IU/kg DM、6700 IU/kg DM)下对粗饲料体外发酵特性及甲烷产量的影响.结果表明:粗饲料添加来源于李氏木霉的纤维素酶时,48h、72h产气量及最大产气量显著低于来源于瘤胃真菌和长柄木霉组及对照组.粗饲料添加来源于瘤胃真菌,李氏木霉和长柄木霉来源的纤维酶时各时间点干物质平均消失率比对照组分别高15%(P<0.001),20%(P<0.001)和6.8%(P<0.05)以上.干物质消失率随纤维素酶添加水平的增加呈三次曲线(P<0.01)增加,酶的添加量为1200 IU/kg DM时48h及72h干物质平均消失率达到最高.添加瘤胃真菌及李氏木霉来源纤维素酶时,粗饲料甲烷产量显著(P<0.001)低于对照组和长柄木霉组.各时间点粗饲料甲烷产量随酶添加水平的增加呈三次曲线降低(P<0.001).结论认为:添加纤维素酶虽然对产气量的影响不明显,但可提高粗饲料干物质消失率及降低甲烷产量,纤维素酶的来源可影响粗饲料的发酵特性.

  14. 纤维素酶法提取胡芦巴甾体总皂苷工艺条件的优化%Studies on Extraction Technology of Total Steroidal Saponins from Fenugreek Seeds by Cellulase

    Institute of Scientific and Technical Information of China (English)

    张黎明; 李霞; 徐玮

    2006-01-01

    以甾体皂苷元的提取率为指标,应用L9(34)正交试验设计优化了胡芦巴种子中甾体皂苷的酶法提取工艺,实验结果表明,影响酶法提取甾体皂苷的主次因素为:酶用量>提取温度>固液比>水解时间.纤维素酶处理能显著提高甾体皂苷的提取率,加酶后甾体皂苷元提取率的平均值为83.6%,而未加酶的结果则为58.4%.酶法提取新工艺能显著提高胡芦巴甾体皂苷的提取率.

  15. 没药中挥发性成分的酶提取及GC/MS分析%Analysis of Cellulase Extraction from the Volatile Components of Myrrha by GC/MS

    Institute of Scientific and Technical Information of China (English)

    陈华; 辛广; 张兰杰; 张博

    2007-01-01

    采用纤维素酶提取法(CE)和水蒸气蒸馏提取法(DSE)对没药挥发油成分进行比较研究.利用气相色谱-质谱联用技术对其化学成分进行分析,从中分别鉴定出34种和38种化学成分, 用峰面积归一法通过数据处理系统得出各化学成分在挥发油中的百分含量, 分别占挥发油总成分的70.05%和67.95%,共同组分中含量最多的是乙酸辛酯和辛醇.

  16. Analysis of essential oil from Myrrha by GC/MS with pretreatment of cellulase%纤维素酶前处理没药挥发油成分的GC/MS分析

    Institute of Scientific and Technical Information of China (English)

    陈华; 辛广; 张兰杰; 张博

    2008-01-01

    采用水蒸汽蒸馏法分别从未处理和纤维素酶前处理的没药中提取挥发油利用气相色谱-质谱(GC/MS)联用技术对其挥发油成分进行分析。从中分别鉴定出38种和34种化学成分,用峰面积归一法通过数据处理系统得出各化学成分在挥发油中的质量分数,占挥发油总成分的67.95%和70.05%。

  17. 提高草菇中纤维降解酶系mRNA丰度的最佳诱导条件初探%Inducing conditions for mRNA of cellulase and hemicellulase by Volvariella volvacea

    Institute of Scientific and Technical Information of China (English)

    李迅; 邵蔚蓝

    2007-01-01

    草菇具有完备的纤维降解酶系基因,在诱导条件下能产生完整的纤维降解酶系.为了提高纤维降解酶系的mRNA丰度,通过正交的方法确定提高草菇中纤维降解酶系mRNA丰度的最佳诱导条件为:培养基配方为xylan和CMC各0.2%、(NH4)2SO4 0.2%、KH2PO4 0.2%、VB 12.5mg/L、MgSO40.1%,调至pH为7.5,以150r/min的转速培养.

  18. 福寿螺多功能纤维素酶EGXA在酿酒酵母中的表达%Expression of a Multi-Functional Endogenous Cellulase Gene from Mollusc, Ampullaria Crossean in Saccharomyces Cerevisiae

    Institute of Scientific and Technical Information of China (English)

    高中华; 许根俊; 赵辅昆

    2007-01-01

    研究在酿酒酵母中表达福寿螺内源性纤维素酶基因egxa cDNA.结果显示:重组酶水解对-硝基酚纤维二糖苷(pNPC)、微晶纤维素(sigmacell)、羧甲基纤维素钠(CMC-Na)以及Oat spelt的木聚糖(xylan from oat spelt,麦木聚糖)等4种底物的活力分别为1.25,84,70和196U/L,并发现了天然信号肽,酵母性因子信号肽(MFα1)对重组酶表达的影响.

  19. 添加乳酸菌和纤维素酶对苜蓿青贮品质的影响%The effects of adding lactic acid bacteria and cellulase on alfalfa silage quality

    Institute of Scientific and Technical Information of China (English)

    万里强; 李向林; 何峰

    2011-01-01

    Alfalfa in initial flowering stage was harvested and wilted for 0, 8, or 32 hours 27.15%, 38.45%, and 50.87%, respectively), and was ensiled with lactic acid bacteria of 10s cfu/g+0.1 g/kg, 106 cfu/g+0.05 g/kg and 107 cfu/g+0. 025 g/kg. This study in fa (DM 50.87%) without additives had lower ammonia nitrogen content, conserved m produced more lactic acid. The addition of lactic (DM content was q-enzyme at rates dicated that alfal- ore DM, CP and acid bacteria and enzyme greatly improved alfalfa silage fermentation. Alfalfa (DM 38.45%) ensiled with lactic acid bact in reduction of ammonia nitrogen content and increases of CP and falfa (DM 38.45%) with the addi.tion of 104 cfu/g lactic acid ba than other treats. eria +enzyme performed well, resulting more lactic acid. The cteria and 0.05 g/kg silage quality of al- enzyme was better%初花期收获的苜蓿(Medicagosativa)经过0、8、32h的晾晒,添加乳酸菌和纤维素酶进行青贮。结果表明,苜蓿凋萎(干物质含量为38.45%)青贮可以使青贮料的氨态氮含量显著降低,并保存有更多的粗蛋白质,生成更多的乳酸。苜蓿青贮时添加乳酸菌和纤维素酶能明显改善苜蓿青贮料的发酵品质,即降低青贮料的氨态氮含量和保存更多的粗蛋白质以及生成更多的乳酸。晾晒8h的苜蓿添加乳酸菌(10^6cfu/g)和纤维素酶(0.05g/kg)的青贮效果最好;而晾晒32h的苜蓿直接青贮的效果最好。

  20. Study on relationship between reductive sugar amounts and fabric properties in treatment with cellulase%纤维素酶处理还原糖量与棉织物性能关系的研究

    Institute of Scientific and Technical Information of China (English)

    吴赞敏; 李昕; 戴晓红; 吕轶君

    2005-01-01

    在不同处理条件下,织物的减量率与酶解液中还原糖量的相关性.结果表明,纤维素酶处理后棉织物的减量率与酶解液中的还原糖量之间具有稳定良好的线性关系.