• Title/Summary/Keyword: cellulase C

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Effect of Dilute Alkali on Structural Features and Enzymatic Hydrolysis of Barley Straw (Hordeum vulgare) at Boiling Temperature with Low Residence Time

  • Haque, Md. Azizul;Barman, Dhirendra Nath;Kang, Tae Ho;Kim, Min Keun;Kim, Jungho;Kim, Hoon;Yun, Han Dae
    • Journal of Microbiology and Biotechnology
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    • v.22 no.12
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    • pp.1681-1691
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    • 2012
  • This work was conducted to evaluate the effect of dilute sodium hydroxide (NaOH) on barley straw at boiling temperature and fractionation of its biomass components into lignin, hemicellulose, and reducing sugars. To this end, various concentrations of NaOH (0.5% to 2%) were applied for pretreatment of barley straw at $105^{\circ}C$ for 10 min. Scanning electron microscopy (SEM), atomic force microscopy (AFM), and Fourier transform infrared (FTIR) spectroscopy studies revealed that 2% NaOH-pretreated barley straw exposed cellulose fibers on which surface granules were abolished due to comprehensive removal of lignin and hemicellulose. The X-ray diffractometer (XRD) result showed that the crystalline index was increased with increased concentration of NaOH and found a maximum 71.5% for 2% NaOH-pretreated sample. The maximum removal of lignin and hemicellulose was 84.8% and 79.5% from 2% NaOH-pretreated liquor, respectively. Reducing sugar yield was 86.5% from 2% NaOH-pretreated sample using an enzyme dose containing 20 FPU of cellulase, 40 IU of ${\beta}$-glucosidase, and 4 FXU of xylanase/g substrate. The results of this study suggest that it is possible to produce the bioethanol precursor from barley straw using 2% NaOH at boiling temperature.

Effect of dietary supplementation of fermented spent mushroom substrates from Pleurotus eryngii on Hanwoo Steers (큰느타리버섯 수확후배지 발효사료 급여가 비육한우에 미치는 영향)

  • Moon, Yea Hwang;Kim, Hye Soo;Kim, Chul Hwan;Cho, Woong Gi;Yoo, Young Bok;Shin, Pyung Gyun;Cho, Soo Jeong
    • Journal of Mushroom
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    • v.13 no.2
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    • pp.108-113
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    • 2015
  • This study was carried out to investigate the feeding effects of dietary supplementation of fermented spent mushroom substrates (F-SMS) from Pleurotus eryngii with Bacillus subtilis CS21 and Saccharomyces cerevisiae on Hanwoo steers. The cellulase and xylanase producing bacteria, designated CS21, was isolated from freshly spent mushroom substrates from Pleurotus eryngii and used as probiotics to fermented spent mushroom substrates. Twenty Hanwoo steers were allocated into two feeding groups and assigned equally to two dietary treatments; Control (TMR) and TMR including 30% F-SMS (30% F-SMS TMR). Total gain and feed intake was significantly greater in the 30% F-SMS TMR than control (p<0.05), but carcass grades were not influenced by the experimental diets. Based on this study, fermented spent mushroom substrates from Pleurotus eryngii with B. subtilis CS21 and Saccharomyces cerevisiae is able to use as an ingredient feed in TMR for Hanwoo steers.

Purification and Characteriztion of an Antifungal Antibiotic from Bacillus megaterium KL 39, a Biocontrol Agent of Red-Papper Phytophtora Blight Disease. (고추역병균 Phytophthora capsici를 방제하는 길항균주 Bacillus megaterium KL39의 선발과 길항물질)

  • 정희경;김상달
    • Microbiology and Biotechnology Letters
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    • v.31 no.3
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    • pp.235-241
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    • 2003
  • For the biological control of Phytophthora blight of red-pepper caused by Phytophthora capsici, an antibiotic-producing plant growth promoting rhizobacteria (PGPR) Bacillus sp. KL 39 was selected from a local soil of Kyongbuk, Korea. The strain KL 39 was identified as Bacillus megaterium by various cultural, biochemical test and API and Microlog system. B. megaterium KL 39 could produce the highest antifungal antibiotic after 40 h of incubation under the optimal medium which was 0.4% fructose, 0.3% yeast extract, and 5 mM KCl at 30 C with initial pH 8.0. The antifungal antibiotic KL 39 was purified by Diaion HP-20 column, silica gel column, Sephadex LH-20 column, and HPLC. Its RF value was confirmed 0.32 by thin-layer chromatography with Ethanol:Ammonia:Water = 8:1:1. The crude antibiotic KL39 was active against a broad range of plant pathogenic fungi, Rhizoctonia solani, Pyricularia oryzae, Monilinia fructicola, Botrytis cinenea, Alteranria kikuchiana, Fusarium oxysporum and Fusarium solani. The purified antifungal antibiotic KL39 had a powerful biocontrol activity against red-pepper phytophthora blight disease with in vivo pot test as well as the strain B. megaterium KL 39.

Characterization of a Multimodular Endo-β-1,4-Glucanase (Cel9K) from Paenibacillus sp. X4 with a Potential Additive for Saccharification

  • Lee, Jae Pil;Kim, Yoon A;Kim, Sung Kyum;Kim, Hoon
    • Journal of Microbiology and Biotechnology
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    • v.28 no.4
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    • pp.588-596
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    • 2018
  • An endo-${\beta}$-1,4-glucanase gene, cel9K, was cloned using the shot-gun method from Paenibacillus sp. X4, which was isolated from alpine soil. The gene was 2,994 bp in length, encoding a protein of 997 amino acid residues with a predicted signal peptide composed of 32 amino acid residues. Cel9K was a multimodular enzyme, and the molecular mass and theoretical pI of the mature Cel9K were 103.5 kDa and 4.81, respectively. Cel9K contains the GGxxDAGD, PHHR, GAxxGG, YxDDI, and EVxxDYN motifs found in most glycoside hydrolase family 9 (GH9) members. The protein sequence showed the highest similarity (88%) with the cellulase of Bacillus sp. BP23 in comparison with the enzymes with reported properties. The enzyme was purified by chromatography using HiTrap Q, CHT-II, and HiTrap Butyl HP. Using SDS-PAGE/activity staining, the molecular mass of Cel9K was estimated to be 93 kDa, which is a truncated form produced by the proteolytic cleavage of its C-terminus. Cel9K was optimally active at pH 5.5 and $50^{\circ}C$ and showed a half-life of 59.2 min at $50^{\circ}C$. The CMCase activity was increased to more than 150% in the presence of 2 mM $Na^+$, $K^+$, and $Ba^{2+}$, but decreased significantly to less than 50% by $Mn^{2+}$ and $Co^{2+}$. The addition of Cel9K to a commercial enzyme set (Celluclast 1.5L + Novozym 188) increased the saccharification of the pretreated reed and rice straw powders by 30.4% and 15.9%, respectively. The results suggest that Cel9K can be used to enhance the enzymatic conversion of lignocellulosic biomass to reducing sugars as an additive.

Production and Characterization of Ethanol- and Protease-Tolerant and Xylooligosaccharides-Producing Endoxylanase from Humicola sp. Ly01

  • Zhou, Junpei;Wu, Qian;Zhang, Rui;Yang, Yuying;Tang, Xianghua;Li, Junjun;Ding, Junmei;Dong, Yanyan;Huang, Zunxi
    • Journal of Microbiology and Biotechnology
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    • v.23 no.6
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    • pp.794-801
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    • 2013
  • This paper reports the production and characterization of crude xylanase from the newly isolated Humicola sp. Ly01. The highest (41.8 U/ml) production of the crude xylanase was obtained under the optimized conditions (w/v): 0.5% wheat bran, 0.2% $KH_2PO_4$, and 0.5% peptone; initial pH 7.0; incubation time 72 h; $30^{\circ}C$; and 150 rpm. A considerable amount of the crude xylanase was induced using hulless barley bran or soybean meal as the carbon source, but a small amount of the enzyme was produced when supplementary urea was used as the nitrogen source to wheat bran. The crude xylanase showed apparent optimal cellulase-free xylanase activity at $60^{\circ}C$ and pH 6.0, more than 71.8% of the maximum xylanase activity in 3.0-30.0% (v/v) ethanol and more than 82.3% of the initial xylanase activity after incubation in 3.0-30.0% (v/v) ethanol at $30^{\circ}C$ for 2 h. The crude xylanase was moderately resistant to both acid and neutral protease digestion, and released 7.9 and 10.9 ${\mu}mol/ml$ reducing sugar from xylan in the simulated gastric and intestinal fluids, respectively. The xylooligosaccharides were the main products of the hydrolysis of xylan by the crude xylanase. These properties suggested the potential of the crude enzyme for being applied in the animal feed industry, xylooligosaccharides production, and high-alcohol conditions such as ethanol production and brewing.

STUDIES ON THE TISSUE CULTURE OF PANAX GINSENG

  • Harn C
    • Proceedings of the Ginseng society Conference
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    • 1974.09a
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    • pp.9-22
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    • 1974
  • Unlike the tissue culture in animals and human being, in higher plants various parts of the plant are cultured for varied purposes, and they are named variously depending on which parts are used as explants or what purposes they are cultured for. Followings are some of the names of culture used frequently: organ culture, tissue culture, callus culture, single cell culture, meristem culture, mericlone culture, ovary culture, ovule culture, embryo culture, endosperm culture, anther culture, pollen culture, protoplast culture, etc.. As the names of the culture indicate, in some kinds of culture the explants used for culture are actually not tissues, but organs, single cells, or protoplasts. It seems, however, convenient to call all of the above-mentioned cultures grossly as tissue culture. Several kinds of tissue culture were attempted using Panax ginseng as material and some of the results were summarized below. 1. Callus culture After dormancy of the sed was broken, whole embryo or parts (hypocotyl, cotyledon and epicotyl) of partly grown embryo were cultured in the media supplemented with growth regulators. Rapid swelling occurred in a few weeks, but most of the swelling was observed only in the basal part of epicotyl, changes in the other parts of embryo appearing in much later stages. The swelling or increase in size, however, was resulted not from the divisions of cells, but from the mere expansion of cell. Real calli were formed about two months after inoculation of explants. Callus tissues developed from cortex, pith, and vascular bundle in the cases of hypo- and epicotyl, from mesophyl tissue in the case of cotyledon. Shoots developed more easily from cotyledons regardless of whether they are detached from or attached to the embryo proper. 2. Culture in the Knudson C medium When cotyledons, detached from or attached to the embryo proper, were cultured in the growth regulator-free Knudson C medium comprision only several kinds of mineral compounds and sucrose, shoot primordium or callus developed profusely and finally plantlets were produced directly from shoot primordium or indirectly through callus. In this medium epidermal cells as well as mesophyl cells of the cotyledon became meristematic and divided, changing into multinucleate cells or multicellular bodies, developing eventually into either shoot primordia or calli. 3. Anther culture Anthers were cultured in the media supplemented with various growth regulators applied singly or in combinations. Callus was formed mostly in the connective tissue of anther. Cells of anther wall layers changed in appearance, but no division occurred. Microspores of all stages in development were not changed, ruling out the possibility that microspore-originated callus might be formed. 4. Isolation of protoplast Protoplasts were isolated from young root, leaf, and epicotyl, using 0.7M D-mannitols as osmoticum and using macerozyme and cellulase respectively for maceration and digestion of the cell wall. Production in large number of naked intact protoplast was rather difficult as compared with other plant species. Fusion of protoplasts occurred infrequently mainly due to the fewer number of naked protoplasts in the solution.

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Isolation and biochemical characterization of acid tolerance xylanase producing Bacteria, Bacillus sp. GJY from city park soil (도심공원으로부터 산내성 xylanase를 생산하는 박테리아 분리 및 효소학적 특성)

  • Jang, Min-Young;Park, Hwa Rang;Lee, Chong Gyu;Choo, Gab-Chul;Cho, Hyun Seo;Park, Sam-Bong;Oh, Ki-Cheol;Kim, Bong-Gyu
    • Journal of Applied Biological Chemistry
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    • v.60 no.1
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    • pp.79-86
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    • 2017
  • Microbes in forest are very important due to not only to enhance soil fertility but also maintain a healthy ecosystem by supplying the energy available to living organisms by producing various kinds of enzymes related to degradation of lignocellulosic biomass. In order to isolate a lignocellulosic biomass degrading bacterial strain from the Jurassic park located in Gyeongnam National University of Science and Technology, We used the Luria-Bertani-Carboxymethyl cellulose (CMC) agar trypan blue method containing 0.4 % carboxymethyl cellulose and 0.01 % trypan blue. As a result, we isolated a bacterial strain showing both activity on the CMC and xylan. To identify the isolated strain, 16S rRNA sequencing and API kit analysis were used. The isolated strain turned out to belong to Bacillus species and then named Bacillus sp. GJY. In the CMC zymogram analysis, it showed that one active band of about 28kDa in size is present. Xylan zymogram analysis also showed to have one active band of about 25kDa in size. The optimal growth temperature of Bacillus sp. GJY was $37^{\circ}C$. The maximal activities of CMCase and xylanase were 12 hour after incubation. The optimal pH and temperature for CMCase were 5.0 and $40^{\circ}C$, respectively, whereas the optimal pH and temperature for xylanase was 4.0 and $40^{\circ}C$. Both activities for CMCase and xylanase showed to be thermally stable at 40and $50^{\circ}C$, while both activities rapidly decreased at over $60^{\circ}C$.

Physiological and Biochemical Characterization of Bacillus spp. from Polychaete, Perinereis aibuhitensis (갯지렁이(Perinereis aibuhitensis)에서 분리한 Bacillus spp.의 생리생화학적 특성 분석)

  • Shin, Seyeon;Yundendorj, Khorloo;Lee, Sang-Suk;Kang, Kyoung-Ho;Kahng, Hyung-Yeel
    • Journal of Life Science
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    • v.23 no.3
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    • pp.415-425
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    • 2013
  • This study compared the characteristics of five Bacillus strains capable of aerobic and anaerobic growth, CBW3, CBW4, CBW9, CBW14 and EBW10. They were isolated and selected from a polychaete, Perinereis aibuhitensis, which is known as a good degrader of organic compounds in marine wetland. Based on a 16S rRNA sequence, CBW3 and CBW14 were found to share more than 99.8% similarity with B. nanhaiensis, B. arsenicus and B. barbaricus. CBW4, CBW9 and EBW10 shared 92.7%, 99.8%, and 99.8% similarity with B. anthracis, B. algicoa and B. thuringiensis, respectively. The temperature, salinity, and pH ranges of the cell growth of the Bacillus strains were $4-45^{\circ}C$, 0-17%, and pH 5-pH 9, respectively. All Bacillus strains were found to exhibit enzyme activities for the degradation of casein and starch. Notably, strain EBW10 exhibited the enzyme activities for all the tested macromolecules, DNA, casein, starch, cellulose, and four kinds of Tweens, which suggests the possibility that it had protease, amylase, cellulose, and lipase. All five Bacillus strains had alkaline phosphatase activities, and the strains CBW3, CBW4, and EBW10 also had acid phospatase. Strains CBW3 and EBW10 exhibited the enzyme activities both for esterase (C4) and esterase lipase (C8). The analysis of fatty acids revealed that in all strains, major fatty acids were anteiso $C_{15:0}$ and iso $C_{15:0}$.

Expression of Clostridium thermocellum Endoglucanase Gene in Lactobacillus bulgaricus and Lactobacillus plantarum and in vitro Survival Characteristics of the Transformed Lactobacilli (Lactobacillus bulgaricus와 Lactobacillus plantarum 균주에서 Clostridium thermocellum 유래 endoglucanase의 발현과 발현 유산균의 in vitro 생존 특성)

  • Cho, J.S.;Kang, S.H.;Lee, H.G.;Lee, H.J.;Woo, J.H.;Moon, Y.S.;Yang, C.J.;Choi, Y.J.
    • Journal of Animal Science and Technology
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    • v.45 no.4
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    • pp.659-666
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    • 2003
  • Endoglucanase A from Clostridium thermocellum which is resistant to pancreatic proteinase was selected out of numbers cellulases then were expressed in lactobacilli. Recombinant lactobacilli expression vector, pSD1, harboring the endoglucanase gene from C. thermocellum under the control of its own promoter, was constructed. Both L. bulgaricus and L. plantarum were electrotransformed with pSD1. The endoglucanase activities of 0.120 and 0.144 U/ml were found in culture media of L. bulgaricus and L. plantarum containing pSD1, respectively. In vitro survival characteristics of the transformed lactobacilli were tested. Both L. bulgaricus and L. plantarum showed a similar resistance to low pH 3. Moreover, L. plantarum was bile-salt resistant in the presence of 0.3 and 1% oxgall. L. bulgaricus and L. plantarum showed a rather homogenous resistant pattern against the tested antibiotics. Both of the strains were resistant to amikacin, gentamicin, streptomycin, kanamycin, and colistin.

Saccharification of Foodwastes Using Cellulolytic and Amylolytic Enzymes from Trichoderma harzianum FJ1 and Its Kinetics

  • Kim Kyoung-Cheol;Kim Si-Wouk;Kim Myong-Jun;Kim Seong-Jun
    • Biotechnology and Bioprocess Engineering:BBE
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    • v.10 no.1
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    • pp.52-59
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    • 2005
  • The study was targeted to saccharify foodwastes with the cellulolytic and amylolytic enzymes obtained from culture supernatant of Trichoderma harzianum FJ1 and analyze the kinetics of the saccharification in order to enlarge the utilization in industrial application. T. harzianum FJ1 highly produced various cellulolytic (filter paperase 0.9, carboxymethyl cellulase 22.0, ${\beta}$-glucosidase 1.2, Avicelase 0.4, xylanase 30.8, as U/mL-supernatant) and amylolytic (${alpha}$-amylase 5.6, ${\beta}$-amylase 3.1, glucoamylase 2.6, as U/mL-supernatant) enzymes. The $23{\sim}98\;g/L$ of reducing sugars were obtained under various experimental conditions by changing FPase to between $0.2{\sim}0.6\;U/mL$ and foodwastes between $5{\sim}20\%$ (w/v), with fixed conditions at $50^{\circ}C$, pH 5.0, and 100 rpm for 24 h. As the enzymatic hydrolysis of foodwastes were performed in a heterogeneous solid-liquid reaction system, it was significantly influenced by enzyme and substrate concentrations used, where the pH and temperature were fixed at their experimental optima of 5.0 and $50^{\circ}C$, respectively. An empirical model was employed to simplify the kinetics of the saccharification reaction. The reducing sugars concentration (X, g/L) in the saccharification reaction was expressed by a power curve ($X=K{\cdot}t^n$) for the reaction time (t), where the coefficient, K and n. were related to functions of the enzymes concentrations (E) and foodwastes concentrations (S), as follow: $K=10.894{\cdot}Ln(E{\cdot}S^2)-56.768,\;n=0.0608{\cdot}(E/S)^{-0.2130}$. The kinetic developed to analyze the effective saccharification of foodwastes composed of complex organic compounds could adequately explain the cases under various saccharification conditions. The kinetics results would be available for reducing sugars production processes, with the reducing sugars obtained at a lower cost can be used as carbon and energy sources in various fermentation industries.