• Asian-Australasian Journal of Animal Sciences
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• v.27 no.2
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• pp.290-301
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• 2014

The first purpose of this review is to outline some of the background information necessary to understand the mechanisms of action of fibre-degrading enzymes in non-ruminants. Secondly, the well-known and understood mechanisms are described, i) eliminating the nutrient encapsulating effect of the cell wall and ii) ameliorating viscosity problems associated with certain Non Starch Polysaccharides, particularly arabinoxylans and ${\beta}$-glucans. A third, indirect mechanism is then discussed: the activity of such enzymes in producing prebiotic oligosaccharides and promoting beneficial cecal fermentation. The literature contains a wealth of information on various non starch polysaccharide degrading enzyme (NSPase) preparations and this review aims to conclude by discussing this body of work, with reference to the above mechanisms. It is suggested that the way in which multi- versus single-component products are compared is often flawed and that some continuity should be employed in methods and terminology.

• Mycobiology
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• v.43 no.1
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• pp.81-86
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• 2015

To promote the selection of promising monokaryotic strains of button mushroom (Agaricus bisporus) during breeding, 61 progeny strains derived from basidiospores of two different lines of dikaryotic parental strains, ASI1038 and ASI1346, were analyzed by nucleotide sequencing of the intergenic spacer I (IGS I) region in their rDNA and by extracellular enzyme assays. Nineteen different sizes of IGS I, which ranged from 1,301 to 1,348 bp, were present among twenty ASI1346-derived progeny strains, while 15 different sizes of IGS I, which ranged from 700 to 1,347 bp, were present among twenty ASI1038-derived progeny strains. Phylogenetic analysis of the IGS sequences revealed that different clades were present in both the ASI10388- and ASI1346-derived progeny strains. Plating assays of seven kinds of extracellular enzymes (${\beta}$-glucosidase, avicelase, CM-cellulase, amylase, pectinase, xylanase, and protease) also revealed apparent variation in the ability to produce extracellular enzymes among the 40 tested progeny strains from both parental A. bisporus strains. Overall, this study demonstrates that characterization of IGS I regions and extracellular enzymes is useful for the assessment of the substrate-degrading ability and heterogenicity of A. bisporus monokaryotic strains.

• Journal of Animal Science and Technology
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• v.49 no.5
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• pp.667-676
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• 2007

This study was conducted to determine effects of cellulolytic microbes inoculation to sawdust-based spent mushroom substrate(SMS) during deepstacking on fermentation parameters, total microbial counts and cellulolytic enzyme activity and to on SMS nutrients utilization by sheep. For sheep metabolism trials, six sheep(ram, average 54.8kg) were fed a Control diet(70% concentrates, 15% rice straw and 15% SMS with no microbial treatment on a dry basis) and a Treatment diet(the same diet including SMS with a microbial treatment) for 2 trials. Spent mushroom substrates with or without a microbial(4 strains including 1 strain of Enterobacter ludwigii, 1 strain of Bacillus cereus and 2 strains of Bacillus subtillis) treatment (1% of SMS on wet basis) were deepstacked for 7 days. The internal temperatures in 1.2 M/T of SMS deepstacks reached to 50±5℃ within 7 days of storage. Total microbial counts remarkably decreased (P<0.05) with a deepstacking process and were not affected(P>0.05) by the microbial treatment. For fibrolytic enzyme activity, CMCase and xylanase activities were decreased(P<0.05) by a deepstacking process. After deepstacking, the microbial treatment showed about 2.5-times higher(P<0.05) for CMCase activity and about 4-times higher(P<0.05) for xylanase activity than those of the Control. Activities of ligninolytic enzymes such as laccase and MnP were not affected by the microbial treatment. The sheep fed the microbially treated SMS diet had a tendency of greater total tract digestibilities of ash(P=0.051), NFE (P=0.071), hemicellulose(P=0.087) and NDF(P=0.096) than those fed the untreated SMS diet. Nitrogen balance of sheep was not affected(P>0.05) by feeding of microbially treated SMS. Accordingly, these results indicate that cellulolytic microbes inoculation during deepstacking of SMS may improve the bio- utilization of SMS by sheep.

• Biotechnology and Bioprocess Engineering:BBE
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• v.10 no.6
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• pp.482-493
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• 2005

Biomass is originally photosynthesized from inorgainic compounds such as $CO_2$, minerals, water and solar energy. Recent studies have shown that anaerobic bacteria have the ability to convert recalcitrant biomass such as cellullosic or chitinoic materials to useful compounds. The biomass containing agricultural waste, unutilized wood and other garbage is expected to utilize as feed, food and fuel by microbial degradation and other metabolic functions. In this study we isolated several anaerobic, cellulolytic and chitinolytic bacteria from rumen fluid, compost and soil to study their related enzymes and genes. The anaerobic and cellulolytic bacteria, Clostridium thermocellum, Clostridium stercorarium, and Clostridium josui, were isolated from compost and the chitinolytic Clostridium paraputrificum from beach soil and Ruminococcus albus was isolated from cow rumen. After isolation, novel cellulase and xylanase genes from these anaerobes were cloned and expressed in Escherichia coli. The properties of the cloned enzymes showed that some of them were the components of the enzyme (cellulase) complex, i.e., cellulosome, which is known to form complexes by binding cohesin domains on the cellulase integrating protein (Cip: or core protein) and dockerin domains on the enzymes. Several dockerin and cohesin polypeptides were independently produced by E. coli and their binding properties were specified with BIAcore by measuring surface plasmon resonance. Three pairs of cohesin-dockerin with differing binding specificities were selected. Two of their genes encoding their respective cohesin polypeptides were combined to one gene and expressed in E. coli as a chimeric core protein, on which two dockerin-dehydrogenase chimeras, the dockerin-formaldehyde dehydrogenase and the dockerin-NADH dehydrogenase are planning to bind for catalyzing $CO_2$ reduction to formic acid by feeding NADH. This reaction may represent a novel strategy for the reduction of the green house gases. Enzymes from the anaerobes were also expressed in tobacco and rice plants. The activity of a xylanase from C. stercorarium was detected in leaves, stems, and rice grain under the control of CaMV35S promoter. The digestibility of transgenic rice leaves in goat rumen was slightly accelerated. C. paraputrificum was found to solubilize shrimp shells and chitin to generate hydrogen gas. Hydrogen productivity (1.7 mol $H_2/mol$ glucos) of the organism was improved up to 1.8 times by additional expression of the own hydrogenase gene in C. paraputrficum using a modified vector of Clostridiu, perfringens. The hydrygen producing microflora from soil, garbage and dried pelletted garbage, known as refuse derived fuel(RDF), were also found to be effective in converting biomass waste to hydrogen gas.

• Animal Bioscience
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• v.35 no.7
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• pp.1030-1038
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• 2022

Objective: Two experiments were conducted to evaluate the effect of enzyme complex (EC) on the standardized ileal digestibility (SID) of amino acids (AA) in corn gluten meal (60%) (CGM), soy protein concentrate (SPC), dried bovine plasma (DBP), and poultry offal meal (POM). Experiments I and II were conducted with broilers in the pre-starter (1 to 7 days of age) and starter (1 to 21 days of age) phases, respectively. Methods: The treatments consisted of a protein-free diet (PFD) containing feedstuffs either supplemented with EC (xylanase, amylase, and protease) or not. In Experiment I, a total of 360 one-day-old male Cobb-500 broiler chicks were randomly housed in 45 pens, resulting in five replicates with eight birds each, totalizing eight treatments and one PFD group. In Experiment II a total of 270 one-day-old male Cobb-500 broiler chicks were randomly housed in 45 pens, resulting in five replicates with six birds each, totalizing eight treatments and one PFD group. The PFD groups were used to assess the endogenous AA losses. The birds were slaughtered to collect the ileal content. Results: In the pre-starter phase, the SID of arginine, branched chain-aminoacids, glycine, serine, aspartate, and glutamic acid increased with EC addition. The EC improved the SID of arginine and glutamic acid of CGM; the SID of valine and cystine of SPC; the SID of leucine, glycine, and aspartate of POM and the SID of isoleucine of DBP. In the starter phase, the SID of isoleucine, phenylalanine and glycine increased in EC-supplemented diets. The EC improved the SID of isoleucine of DBP; the SID of phenylalanine of CGM and POM. The SID of AA of SPC was not influenced by the EC. Conclusion: The addition of an EC to broiler pre-starter and starter diets is efficient in increasing the SID of AA on SPC, POM, and DBP.

• Microbiology and Biotechnology Letters
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• v.22 no.5
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• pp.507-514
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• 1994

Acetyl xylan esterase was produced by E. coli HB101 harboring a recombinant plasmid pKMG6 which contained the estI gene of Bacillus stearothermophilus. The maximum production was observed when the E. coli strain was grown at 37$\circC for 12 hours in the medium containing 0.5% acetyl xylan, 1.0% tryptons, 1.0% sodium chloride, and 0.5% yeast extract. The esterase produced was purified to homogeneity using a combination of ammonium sulfate fractionation, DEAE Sepharose CL-6B ion exchange chromatography and Sephacryl S-200 gel filtration. The native enzyme had an apparent molecular mass of 60 kd and was composed of two identical subunits of 29 kd. The N-terminal amino acid sequence of the polypeptide was Ala-X-Leu-Gln- Ile-Gln-Phe-X-X-Gln. The acetyl esterase displayed a pH optimum of 6.5 and a temperature opti- mum of 45$\circC. The heavy metal ions such as Ag$^{++}$, Hg$^{++}$ and Cu$^{++}$ inhibited nearly completely the activity of the esterase, and no specific metal ion was found to be required for the enzyme activity. The enzyme readily cleaved MAS, $\beta$-D-glucose pentaacetate, $\alpha$-naphthyl acetate, $\rho$-nitrophenyl acetate as well as acetyl xylan, but had no activity on $\rho$-nitrophenyl propionate, $\beta$-nitrophenyl butyrate or $\beta$-nitrophenyl valerate. The Km and Vmax values for MAS were 2.87 mM and 11.55 $\mu$mole/min, respectively. Synergistic behavior was demonstrated with a combination of xylanase and esterase from B. stearothermophilus in hydrolyzing acetyl xylan.

• Applied Biological Chemistry
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• v.23 no.1
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• pp.64-72
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• 1980

Industrial wastes from pulp and food plants were treated with microorganisms to clarify organic waste-water and to produce cells as animal feed, and results were summarized as follows. (1) Waste-water from pulp, beer, bread yeast, and ethanol distillation plants contained $1.4{\sim}1.5%$ of total sugar, $0.25{\sim}0.35%$ nitrogen, and biological oxygen demand (BOD) was $400{\sim}25,000$, chemical oxygen demand (COD), $500{\sim}28,000$, and pH, $3.8{\sim}7.0$. The BOD and COD were highest in waste-water from ethanol distillation plants among others. (2) Bacterial and yeast counts were $4{\times}10^4-1{\times}10^9,\;2{\times}10^2-7{\times}10^4/ml$ in waste-water. (3) Bacteria grew better in pulp waste and yeasts in beer, bread yeast, and ethanol distillation waste. (4) Saccharomyces cerevisiae SAFM 1008 and Candida curvata SAFM 70 were the most suitable microorganisms for clarification of ethanol distillation waste. (5) When liquid and solid waste from ethanol distillation were treated with microbial cellulase, xylanase, and pectinase, solid waste was reduced by 36%, soluble waste was increased, and recuding sugar content was increased by 1.3 times which provided better medium than untreated waste for cultivation of yeasts. (6) Optimum growth conditions of the two species of yeast in ethanol distillation waste were pH 5.0, $30^{\circ}C$, and addition of 0.2% of urea, 0.1% of $KH_2PO_4$ and 0.02% of $MgSO_4$. (7) Minimum number of yeast for proper propagation was $1.8{\times}10^5/ml$. (8) C. curvata70 was better than cerevisae for the production of yeast cells from ethanol distillation waste treated with microbial enzymes. (9) S. cerevisiae produced 16 g of dried cell per 1,000ml of ethanol distillation waste and reduced BOD by 46%. C. curvata produced 17.6g of dried cell and reduced BOD by 52% at the same condition. (10) Yeast cells produced from the ethanol distillation waste contained 46-52% protein indicating suitability as a protein source for animal feed.

• ANNALS OF ANIMAL RESOURCE SCIENCES
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• v.28 no.3
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• pp.97-107
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• 2017

The present study was conducted to investigate the effects on growth performance, nutrient digestibility, and gut health of broiler chickens when a dietary supplementation of multienzymes was added to diets, containing different energy levels. A total of 480 broiler chickens of similar body weight (Ross 308, 1-day-old) were randomly subjected to four treatments. The dietary treatments included a corn-soybean meal-based diet supplemented with: multienzyme (amylase+protease+ mannanase+xylanase+phytase), 0.05% enzyme, and different energy levels (3010 and 3060 kcal/kg). The experimental diets were fed to the chicks in a mash form for 35 days in two phases (1-21 d, phase I; and 22-35 d, phase II). During the overall period, chicks fed with diets supplemented with multienzymes had a better weight gain (p<0.05) and feed conversion ratio (FCR) than those fed with diets without enzymes. There was no difference in the growth rate and FCR among the chicks fed with diets supplemented with enzymes, even though the dietary energy levels were different. The apparent fecal and ileal digestibility of dry matter, gross, crude protein, calcium, and phosphorus were significantly enhanced (p<0.05). The population of cecal and ileal Lactobacillus spp. was significantly increased (p<0.05), and Clostridium spp. and coliforms were significantly decreased (p<0.05) in diets supplemented with enzymes. Villus height and villus height to crypt depth ratio in the small intestine was also significantly enhanced (p<0.05) in diets supplemented with enzymes. In conclusion, multienzyme supplementation had positive effects on the weight gain of broilers, FCR, digestibility of nutrients, and on the growth of intestinal microbiota.

• Journal of Mushroom
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• v.10 no.2
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• pp.74-82
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• 2012

Sixty strains of Pleurotus ostreatus, white-rot fungi, were screened for production ability of their lignocellulolytic enzymes to selectively wood degradation. That results were shown that all of screened strains were produced lignocellulolytic enzymes on 2nd screening liquid culture medium. However, cellulase activity of selected six strains of P. ostreatus was low in avicel-yeast-peptone liquid culture medium. In the case of xylan degrading enzyme, No. 6 and No. 38 strains produced a xylanase(above 1.0U/ml) and a 1,4-${\beta}$-xylosidase (above 0.15 U/ml). Examination of the ligninolytic enzyme profiles of selected thirteen strains of the P. ostreatus, in the presence of Remazol Brilliant Blue R(RBBR), were observed that laccase(Lac) activity were earlier reached maximum level(0.8-2.0 U/ml) and then Mn-dependent peroxidase (MnP) were reached maximum level(0.5-1.5 U/ml) in glucose-yeast-peptone(GYP) medium. On the other hand, activity of lignin peroxidase(LiP) was not detected in this medium. I selected the No. 42 strain of P. ostreatus produced high levels of Mn-dependent peroxidase and laccase based on the screening method.

• Asian-Australasian Journal of Animal Sciences
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• v.24 no.1
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• pp.56-64
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• 2011

This study was conducted to evaluate the use of exogenous enzymes as a potential means of improving the ruminal digestion (i.e., degradability) of alfalfa hay and rice straw. Twenty six enzyme-additives were examined in terms of protein concentration and enzymic activities on model substrates. The exogenous enzymes contained ranges of endoglucanase, xylanase, ${\beta}$-glucanase, ${\alpha}$-amylase, and protease activities. Six of the enzyme additives were chosen for further investigation. The enzyme additives and a control without enzyme were applied to mature quality alfalfa hay substrate and subsequently incubated in rumen batch cultures. Five of the enzyme additives (CE2, CE13, CE14, CE19, and CE24) increased total gas production (GP) at 48 h of incubation compared to the control (p<0.05). The two additives (CE14 and CE24) having the greatest positive effects on alfalfa hay dry matter, neutral detergent fibre (NDF) and acid detergent fibre (ADF) degradability were further characterized for their ability to enhance degradation of low quality forages. The treatments CE14, CE24, a 50:50 combination of CE14 and CE24 (CE14+24), and control (no enzyme) were applied to mature alfalfa hay and rice straw. For alfalfa hay, application of the two enzyme additives, alone and in combination, increased GP compared to the control at 48 h fermentation (p<0.05), whereas only CE14 and CE14+24 treatments improved GP from rice straw (p<0.05). Rumen fluid volatile fatty acid concentrations throughout the incubation of rice straw were analyzed. Acetate concentration was slightly lower (p<0.05) for CE14${\times}$CE24 compared to the control, although individually, CE14 and CE24 acetate concentrations were not different from the control. Increases (p<0.05) in alfalfa hay NDF degradability measured at 12 and 48 h of incubation occurred only for CE14 (at 12 h) and for CE14+24 (at 12 and 48 h). Similarly, ADF degradability increased (p<0.05) with CE14 and CE14+24. As for rice straw, increased DM degradability was observed at 12 and 48 h of incubation for all enzyme treatments with an exception for CE14 at 12 h. The degradability of NDF was improved by all the enzyme treatments at either incubation time, while ADF degradability was only enhanced at 48 h. Overall, the enzymes led to enhanced digestion of mature alfalfa and there was evidence of improved digestibility of rice straw, an even lower quality forage.