Levels Of Xylanase Research Articles

Xylanase production by Aspergillus awamori. Development of a medium and optimization of the fermentation parameters for the production of extracellular xylanase and β‐xylosidase while maintaining low protease production

A growth medium was developed for maximal production in batch culture of extracellular xylanase and beta-xylosidase by Aspergillus awamori CMI 142717 and a mutant (AANTG 43) derived from the wild-type strain. The optimum pH for the production of xylanase and beta-xylosidase was 4.0. The best temperature of xylanase production was 30 degrees C; 35 degrees C was optimal for beta-xylosidase. Protease production was never completely suppressed under any of the conditions tested. However, protease titre was 3.5-fold less than the control in medium in which proteose peptone and yeast extract were omitted: the level of xylanase was not affected (8.6 U mL(-1)) but beta-xylosidase titre was increased 4.7-fold to 1.5 U mL(-1). When corn steep liquor was used as the sole nitrogen source, xylanse and beta-xylosidase titres were further increased by 1.5- and 1.9-fold, respectively. Of the carbon sources investigated, ball-milled oat straw or oat spelt xylan produced the highest titres of xylanse and beta-xylosidase. None of the soluble carbon sources investigated produced the high titres of xylanase or beta-xylosidase induced by either oat straw for xylanse and beta-xylosidase was 2% and the optimum spore inoculum was between 10(6) and 10(7) spores/mL(-1) final concentration. The level of xylanse activity obtained in the culture filtrates of the mutant was a remarkable 820 U mL(-1) when the reducing sugar released was measured by the dinitrosalicylic acid method. This enzyme titre would appear to be the highest reported so far. The xylanases system contained the correct balance of enzymes to effect extensive hydrolysis of oat spelt xylan. The protease titre was very low.

Kinetics of wheat straw solid-state fermentation with Trametes versicolor and Pleurotus ostreatus ? lignin and polysaccharide alteration and production of related enzymatic activities

The kinetics of straw solid-state fermentation (SSF) with Trametes versicolor and Pleurotus ostreatus was investigated to characterize the delignification processes by these white-rot fungi. Two successive phases could be defined during straw transformation, characterized by changes in respiratory activity, changes in lignin and polysaccharide content and composition, increase in in-vitro digestibility, and enzymatic activities produced by the fungi. Lignin composition was analysed after CuO alkaline degradation, and decreases in syringyl/guaiacyl and syringyl/p-hydroxyphenyl ratios and cinnamic acid content were observed during the fungal treatment. An increase in the phenolic acid yield, revealing fungal degradation of side-chains in lignin, was produced by P. ostreatus. The highest xylanase level was produced by P. ostreatus, and exocellulase activity was nearly absent from straw treated with this fungus. Lactase activity was found in straw treated with both fungi, but lignin peroxidase was only detected during the initial phase of straw transformation with T. versicolor. High levels of H2O2-producing aryl-alcohol oxidase occurred throughout the straw SSF with P. ostreatus.

Xylan-hydrolysing enzymes from thermophilic and mesophilic fungi

Screening of 40 mesophilic and 13 thermophilic fungi indicated that enzyme activities capable of degrading oat spelt xylan extensively were produced by only a few of the mesophilic species investigated. The relatively low degree of hydrolysis effected by the enzymes from thermophilic organisms could be explained, in part, by their lack of β-xylosidase. Several strains of Aspergillus awamori and Aspergillus phoenicis were notable in producing high xylanase and β-xylosidase and low protease activities. Of the fungl tested, 13 produced activities capable of removing O-acetyl, arabinosyl, 4-O-methylglucuronyl, feruloyl and coumaroyl substituents from the backbone of xylan polysaccharides as well as endo-1,4-β-D-xylanase and β-1,4-xylosidase. When the growth medium contained oat spelt xylan as carbon source, higher levels of xylanase, β-xylosidase and acetyl xylan esterase were found than in cultures containing meadow fescue grass but the latter were richer in ferulic acid and coumaric acid esterases and 4-O-methylglucuronidase. No single organism or carbon source used was capabie of producing high levels of all the debranching enzymes as well as high levels of enzymes capable of cleaving the glycosidic linkages of the xylan backbone. The best ballnce of enzymes was obtained in cultures of A. awamori IMI 142717 and NRRL 2276 and A. phoenicis IMI 214827. Either of these would be suitable for strain improvement studies.

Expression of xylanase and cellulase enzymes in a newly isolated Clostridium sp.SAIV

Clostridium sp. SAIV produces a high level of extracellular xylanase when grown on either xylan or xylose (520 and 480 mU ml −1). β-Xylosidase enzyme is cell-bound. Clostridium sp.SAIV showed growth on filter paper cellulose, but growth decreased with repeated transfer on this substrate. However, cellulase enzymes were expressed and continued degradation of cellulose occurred when the organism was grown on xylose plus cellulose.

Isolation of mutants of Aspergillus awamori with enhanced production of extracellular xylanase and ?-xylosidase

Plate screening tests were designed for the selection and isolation of mutant strains of the fungus Aspergillus awamori CMI 142717 showing over-production and constitutive synthesis of xylanase and β-xylosidase. Following mutation by N-methyl-N-nitro-N-nitrosoguanidine, nitrous acid and UV (254 nm), two generations of mutants were isolated and cultured in shake fiasks containing glucose, ball-milled oat straw or oat speit xylan as carbon source. Growth of a number of selected mutants in shake flask culture on medium containing oat spelt xylan produced the highest titres of xylanase and β-xylosidase. Thus, xylanase producton by mutant AANTG43 was 132 U/ml when the Somogyl-Nelson (alkaline copper) method of measuring reducing sugar released was used, or 1160 U/ml using the dinitrosalicylic acid method of reducing sugar analysis. These values were 8-fold higher than those produced by the wild type. A 20-fold improvement in β-xylosidase production was produced by mutant AANO19 (3.51 U/ml). The titres for these two enzyme activities are the highest recorded so far in the literature. Mutant AANTG43 also produced high levels of xylanase (49.8 U/ml) in submerged culture in a fermenter and showed a substantial improvement in the overall productivity of enzyme compared to the wild type strain.

Xylanase production by Trichoderma harzianum E58

Growth of Trichoderma harzianum E58 on hemicellulose-rich media, both in batch and fermentor cultures, resulted in independent profiles for the production of xylanase and endoglucanase enzymes. Dramatic differences in the ratio of xylanase to endoglucanase activities were observed among cultures grown on cellulose-rich Solka Floc and xylan. These results indicated that the induction of xylanases and cellulases was likely to be under separate regulatory control. The specific activity and amount of xylanases produced were found to be dependent on the concentration of xylan in the growth media. Growth on oat spelts xylan or the hemicellulose-rich, water-soluble fraction from steam-treated aspenwood (SEA-WS) greatly enhanced the production of xylanases and xylosidase in the culture filtrates. Constitutive levels of xylanase and endoglucanase enzymes were detected during growth of the fungus on glucose.

Xylanase production by Trichoderma longibrachiatum

The effects of carbon source, substrate concentration, culture pH, and spore inoculum concentration on production of extracellular xylanase and cellulase were examined. Very low enzyme activities were obtained with growth on glucose, xylose, and cellobiose, while significantly higher levels were produced from lactose and arabinose. Higher levels of both enzymes were generated from alpha cellulose, wood pulp, and fibrous paper waste than from purified xylan. However, the ratio of xylanase to cellulase activity was higher with pure xylan. High levels of both xylanase and cellulase activity were obtained when the culture was grown on lactose plus xylan. A factorial experiment showed that spore and substrate concentration had significant effects on xylanase yield, and that an interaction existed between these two variables. Highest levels of xylanase were generated with cultures grown on 1% wood pulp at pH 7.0 using an inoculum of 10 5 spores ml -1. Maximal xylanase activity was observed at pH 4.8-5.5 at 55°C using a 30-min assay. The type of xylan used as substrate and the method of reducing sugar detection significantly affected measured xylanase activity.

Thermogravimetric analysis of flax shives degraded by pleurotus ostreatus and ceraceomyces sublaevis

The polysaccharide-degrading and laccase enzymes released during the controlled degradation of flax shives by the fungal colonists Ceraceomyces sublaevis and Pleurotus ostreatus were assayed. Thermogravimetric analysis was used to identify the major components of the degraded flax shives. Changes in the peak patterns of derivative thermogravimetry were only observed 8 days after the start of fungal incubations, coinciding with an increased content of degrading enzymes. Two major peaks of thermogravimetric decomposition were observed; one at 280°C, believed to represent holocellulose, and one at 450°C representing lignin. The two fungal colonists degraded different components of the flax shive. P. ostreatus released higher levels of pectic, xylanase and cellulase enzymes into the substrate and, as a result, degraded the holocellulose present in shive more efficiently than lignin. C. sublaevis released higher levels of laccase in the substrate and, subsequently, degraded lignin more efficiently. The difference in the enzyme-release patterns of the two fungi was correlated with the changes in thermogravimetric weight loss of the different components of shive.

Role of β‐Xylosidase in Hemicellulose Hydrolysis by Xylanase from Penicillium funiculosum1

Penicillium funiculosum produces 16.7 and 0.2 Uml−1 of xylanase and xylosidase with traces of endoglucanase and filter paper activity when grown on a medium containing xylan. The rate of hydrolysis of xylan increased with the increasing levels of xylanase but was not strictly proportional to enzyme concentration. Fractions enriched in xylanase and xylosidase activity were obtained by fractional precipitation of culture broth with ammonium sulfate. Hydrolysis of xylan using varying amounts of xylosidase showed that the optimum ratio of xylanase to xylosidase for maximum hydrolysis is 7.14. The results on the decreased rate of hydrolysis, due to a further increase in xylosidase, are discussed.

Xylanase and β‐xylosidase production by Aspergillus ochraceus during growth on lignocelluloses

Xylan is a polysaccharide located in the cell wall of higher plants which is one of the major components of lignocellulosic materials. In some higher plants and agricultural wastes the xylan is 20-40% of the dry material.' It consists of p-1, 4-linked D-xylose chain. The breakdown of hemicellulose is accomplished by the synergistic action of xylanase (1,4+-~-xylan xylanohydrolase, EC 3.2.1.8) and P-xylosidase (P-D-xyloside xylohydrolase, EC 3.2.1.37). Such enzymes are useful in the degradation of biomaterials containing hemicellulose for the production of monosaccharides from which single-cell protein, single-cell oil, or ethanol could be produced. '-I1 The production and properties of enzymes capable of breaking down cellulose have been extensively st~died. '~-~' Although there are reports on the production of xylanase,'8-26 the behavior of xylanases and its involvement in the utilization of complex lignocellulosic material is still incomplete. Therefore, the present communication deals with the utilization of some lignocellulosic materials by A . ochraceus for xylanase and pxylosidase production. The use of purified xylan enhances the cost of enzyme production and is a major limitation to the economic feasibility of bioconversion and utilization of lignocellulosic materials. Therefore, in this work, easily available substrates like wheat straw, wheat bran, and sugarcane bagasse were used in the experiment to obtain optimum levels of xylanases by the fungal strain A . ochruceus.

The role of bacteria in retting of desiccated flax during damp weather

The bacteria present on samples of desiccated flax stems were Bacillus mycoides, B. subtilis, Erwinia carotovora, Pseudomonas fluorescens, P. putida and Micrococcus sp. and the fungi present were Cladosporium herbarum, Fusarium culmorum, Botrytis cineria, Epicoccum nigrum and yeast. When inoculated on autoclaved stems or in liquid culture, B. subtilis produced mainly pectin-lyase and xylanase. However, only pectin-lyase was detected in significant levels in autoclaved stem sections or in liquid cultures inoculated with E. carotovora. Enhanced pectin-lyase and xylanase levels were detected in field-retted stems sprayed with B. subtilis compared with enzyme levels in stems sprayed with E. carotovora or the control stem tissues. Increases in the fungal population coincided with a reduction in the bacterial population on treated stems at the later part of retting. Enhanced retting was observed in stems sprayed with B. subtilis and consequently the stems produced finer fibres than fibres from E. carotovora-sprayed or control stems.

Synthesis of xylanase byTrichoderma reesei

Xylanase is synthesized byTrichoderma reesei when grown in the presence of xylan, cellulose or phosphocellulose. Xylan is the best inducer of xylanase. Other cellulolytic enzymes such as endoglucanase and FPase are synthesized in small quantities, but the level of β-glucosidase is very high. Mutants with altered levels of cellulolytic enzymes showed an unaltered level of xylanase, suggesting its independent control in this organism.

Interactive Effect of Temperature and Salinity on Growth and Activity of a Salt Marsh Isolate of Aureobasidium pullulans

The effect of various combinations of temperature and salinity on growth and xylanase production by a salt marsh isolate of Aureobasidium pullulans was investigated. At 10, 15, and 25 C growth was maximum at low salinities whereas the highest levels of xylanase were produced at intermediate salinities within this temperature range. Incubation at 35 C resulted in an upward shift in salinity optima such that maximal growth and xylanase production occurred at the highest salinity examined (45‰). The interactive effect of temperature and salinity on both growth and xylanase production was supported by highly significant F-values (P < 0.001) for the temperature/salinity interaction terms when the data were subjected to two-way analyses of variance.

High Cellobiase and Xylanase Production by Sclerotium rolfsii UV-8 Mutant in Submerged Culture

Sclerotium rolfsii UV-8 mutant secretes high levels of cellobiase and xylanase in addition to having high cellulase production. The apparent K(m) and V(max) of cellobiase (grown in NM-2 + 2% corn steep liquor medium) with cellobiose as a substrate were 5.6 mM and 22.2 mumol of glucose liberated per min per ml of culture filtrate, respectively. The addition of 2% corn steep liquor to NM-2 medium increased endo-beta-glucanase, cellobiase, and xylanase yields by approximately 1.5-fold.

Pleiotropic mutants of the wood-rotting fungus Polyporus adustus lacking cellulase, mannanase, and xylanase.

By screening for mutants which could not degrade cellulose, several cellulase-less mutants were isolated from the wood-rotting fungus Polyporus adustus. Most of the mutants lacked mannanase and xylanase as well. In wild type, the level of cellulase, mannanase, and xylanase was higher when the fungus was grown in a medium containing cellulose than in a medium lacking cellulose. It is proposed that in P. adustus, the induction of this group of enzymes is under the control of a single common regulator gene.

Effect Of Laboratory Produced Xylanase From Aspergillus niger On Fibre Digestibility Of Rice Husk And Guinea Grass

The efficacy of laboratory produced xylanase enzyme from Aspergillus niger on highly fibrous feeds and feedstuffs was investigated. Guinea grass (Panicum maximum) from grass hay and rice husk (Oryza sativa) from crop by-product characterised with high fibre content served as substrates (Feeds and Feedstuffs) for the experiment. The enzyme xylanase was prepared from Aspergillus niger (fungal extracts). The two substrates were treated with three levels of aqueous enzymes xylanase (0.02 ml/g, 0.04 ml/g, 0.08 ml/g). Substrate with no enzyme treatment (0 ml/g) served as control. Substrates were subjected to xylanase enzyme treatment at different time interval to determine the optimum incubation period for maximum fibre digestibility. Rice husk and guinea grass were analysed to determine the level of digestibility of fibre fractions such as crude fibre, hemicellulose, acid detergent fibre, neutral detergent fibre and lignin. The results indicate an improvement in fibre digestibility for rice husk and guinea grass treated with enzyme when compared with the control. There was no significant different in the fibre digestibility of substrate treated different levels of xylanase enzyme. Results obtained on the period of application showed no significant different across the time intervals (0 hr, 24 hr, 48 hr and 72 hr). The results therefore, suggest that optimum fibre digestibility could be obtained at level 0.02 ml/g of xylanase treatment at any time interval.Key Words: Aspergillus niger, Xylanase, Rice husk, Guinea grass, Laboratory