Lignocellulolytic characterization and comparative secretome analysis of a Trichoderma erinaceum strain isolated from decaying sugarcane straw

The fungus Trichoderma reesei is employed in the production of most enzyme cocktails used by the lignocellulosic biofuels industry today. Despite significant improvements, the cost of the required enzyme preparations remains high, representing a major obstacle for the industrial production of these alternative fuels. In this study, a new Trichoderma erinaceum strain was isolated from decaying sugarcane straw. The enzyme cocktail secreted by the new isolate during growth in pretreated sugarcane straw-containing medium presented higher specific activities of β-glucosidase, endoxylanase, β-xylosidase and α-galactosidase than the cocktail of a wild T. reesei strain and yielded more glucose in the hydrolysis of pretreated sugarcane straw. A proteomic analysis of the two strains' secretomes identified a total of 86 proteins, of which 48 were exclusive to T. erinaceum, 35 were exclusive to T. reesei and only 3 were common to both strains. The secretome of T. erinaceum also displayed a higher number of carbohydrate-active enzymes than that of T. reesei (37 and 27 enzymes, respectively). Altogether, these results reveal the significant potential of the T. erinaceum species for the production of lignocellulases, both as a possible source of enzymes for the supplementation of industrial cocktails and as a candidate chassis for enzyme production.     

Optimization of arylacetonitrilase production from Alcaligenes sp. MTCC 10675 and its application in mandelic acid synthesis

Alcaligenes sp. MTCC 10675 has been isolated from soil sample using enrichment method and has nitrilase catalytic system which is highly specific for the hydrolysis of arylaliphatic nitriles. Optimization of culture conditions using response surface methodology and inducer-mediated approach enhanced arylacetonitrilase production significantly (2.4-fold). Isobutyronitrile acted as an effective inducer for the induction of arylacetonitrilase, and it is highly specific for arylacetonitriles (phenyl acetonitrile and mandelonitrile). Arylacetonitrilase has no effect on its relative velocity (V _r) up to 20 mM substrate (mandelonitrile) concentration and at 30 mM mandelonitrile, 23.4 % degree of inhibition (I _d) was recorded. Half life of arylacetonitrilase of Alcaligenes sp. MTCC 10675 was 27.5 h at 25 °C. Hg²⁺, Ag⁺, Pb³⁺, and Co²⁺ were strong inhibitor of arylacetonitrilase activity which resulted into 100 %, 91 %, 84 %, and 83 % inhibition, respectively. Polar protic solvent (dichloromethane, dimethylsulphooxide, and n-butanol) reduce arylacetonitrilase activity up to 80–94 % at 10 % concentration. Alcaligenes sp. MTCC 10675 has higher biocatalytic activity, i.e., 3.9 gg^-1 dcw, which is highest in comparison to till reported organism. Arylacetonitrilase-mediated hydrolysis of racemic mandelonitrile resulted into R-(-) mandelic acid with 99.0 % enantiomeric excess (e.e.)     

Biodegradation of aflatoxin B1 in contaminated rice straw by Pleurotus ostreatus MTCC 142 and Pleurotus ostreatus GHBBF10 in the presence of metal salts and surfactants

Aflatoxin B1 (AFB1) is a highly toxic fungal metabolite having carcinogenic, mutagenic and teratogenic effects on human and animal health. Accidental feeding of aflatoxin-contaminated rice straw may be detrimental for ruminant livestock and can lead to transmission of this toxin or its metabolites into the milk of dairy cattle. White-rot basidiomycetous fungus Pleurotus ostreatus produces ligninolytic enzymes like laccase and manganese peroxidase (MnP). These extracellular enzymes have been reported to degrade many environmentally hazardous compounds. The present study examines the ability of P. ostreatus strains to degrade AFB1 in rice straw in the presence of metal salts and surfactants. Laccase and MnP activities were determined spectrophotometrically. The efficiency of AFB1 degradation was evaluated by high performance liquid chromatography. Highest degradation was recorded for both P. ostreatus MTCC 142 (89.14 %) and P. ostreatus GHBBF10 (91.76 %) at 0.5 µg mL^?1 initial concentration of AFB1. Enhanced degradation was noted for P. ostreatus MTCC 142 in the presence of Cu²⁺ and Triton X-100, at toxin concentration of 5 µg mL^?1. P. ostreatus GHBBF10 showed highest degradation in the presence of Zn²⁺ and Tween 80. Liquid chromatography-mass spectrometric analysis revealed the formation of hydrated, decarbonylated and O-dealkylated products. The present findings suggested that supplementation of AFB1-contaminated rice straw by certain metal salts and surfactants can improve the enzymatic degradation of this mycotoxin by P. ostreatus strains.     

Endocellulase Production by <Emphasis Type="Italic">Cotylidia pannosa</Emphasis> and its Application in Saccharification of Wheat Bran to Bioethanol

For efficient bioconversion of lignocellulosic materials to bioethanol, the study screened 19 white-rot fungal strains for their endocellulolytic activity and saccharification potential. Preliminary qualitative and quantitative screening revealed Cotylidia pannosa to be the most efficient endocellulase producing fungal strain when compared to the standard strain of Trichoderma reesei MTCC 164. Ensuing initial screening, the production of endocellulase was further optimized using submerged fermentation to recognize process parameters such as temperature, time, agitation pH, and supplementation of salts in media required for achieving maximum production of endocellulase. The strain C. pannosa produced the maximum amount of endocellulase (8.48 U/mL) under submerged fermentation with wheat bran (2%) supplemented yeast extract peptone dextrose (YEPD) medium after an incubation time of 56 h at 30 °C and pH 5.0 at an agitation rate of 120 rpm with a saccharification value of 50.5%. The fermentation of wheat bran hydrolysate with Saccharomyces cerevisiae MTCC 174 produced 4.12 g/L of bioethanol after 56 h of incubation at 30 °C. The results obtained from the present investigation establish the potential of white-rot fungus C. pannosa for hydrolysis and saccharification of wheat bran to yield fermentable sugars for their subsequent conversion to bioethanol, suggesting its application in efficient bioprocessing of lignocellulosic wastes.     

Chemical and biological properties of wheat soil in response to paddy straw incorporation and its biodegradation by fungal inoculants

A field experiment was conducted to evaluate the relative contribution of organic fertilizers (paddy straw, microbial inoculants and vermicompost) and inorganic fertilizers (urea and superphosphate) in improving pH, C, N, humus, microbial biomass, dehydrogenase, phosphatase, cellulase, β-glucosidase and xylanase activities of soil under wheat crop. Vermicompost fertilization resulted in highest microbial biomass, available phosphorus, and nitrogen content of wheat soil. It was also found effective in minimizing the alkalinity of soil compared to other treatments as indicated by pH change. However incorporation of paddy straw in conjunction with N₆₀P₆₀ and T. reesei inoculation resulted in maximum dehydrogenase, alkaline phosphatase and highest humus content of soil. Mixed inoculation of A. awamori and T. reesei did not prove effective in improving the soil biochemical properties in comparison to single inoculation of T. reesei. Results showed that in situ incorporation of paddy straw in combination with N₆₀P₆₀ and T. reesei inoculation can be used as an effective measure for valuable disposal of paddy straw and to improve the soil health by reducing mineral fertilization.     

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

Summary 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.     

Bench scale production of benzohydroxamic acid using acyl transfer activity of amidase from Alcaligenes sp. MTCC 10674

The acyl transfer activity of the amidase of Alcaligenes sp. MTCC 10674 has been applied to the conversion of benzamide and hydroxylamine to benzohydroxamic acid. The unique features of the acyl transfer activity of this organism include its optimal activity at 50 °C and very high substrate (100 mM benzamide) and product (90 mM benzohydroxamic acid) tolerance among the hitherto reported enzymes. The bench scale production of benzohydroxamic acid was carried out in a fed-batch reaction (final volume 1 l) by adding 50 mM benzamide and 250 mM of hydroxylamine after every 20 min for 80 min in 0.1 M potassium phosphate buffer (pH 7.0) at 50 °C, using resting cells equal to 4.0 mg dcm/ml of reaction mixture. From 1 l of reaction mixture 33 g of benzohydroxamic acid was recovered with 24.6 g l^?1 h^?1 productivity. The acyl transfer activity of the amidase of Alcaligenes sp. MTCC 10674 and the process developed in the present study are of industrial significance for the enzyme-mediated production of benzohydroxamic acid.     

Citric Acid Production from Hydrolysate of Pretreated Straw Cellulose by Yarrowia lipolytica SWJ-1b Using Batch and Fed-Batch Cultivation

In this study, crude cellulase produced by Trichoderma reesei Rut-30 was used to hydrolyze pretreated straw. After the compositions of the hydrolysate of pretreated straw were optimized, the study showed that natural components of pretreated straw without addition of any other components such as (NH₄)₂SO₄, KH₂PO₄, or Mg²⁺ were suitable for citric acid production by Yarrowia lipolytica SWJ-1b, and the optimal ventilatory capacity was 10.0 L/min/L medium. Batch and fed-batch production of citric acid from the hydrolysate of pretreated straw by Yarrowia lipolytica SWJ-1b has been investigated. In the batch cultivation, 25.4 g/L and 26.7 g/L citric acid were yields from glucose and hydrolysate of straw cellulose, respectively, while the cultivation time was 120 hr. In the three-cycle fed-batch cultivation, citric acid (CA) production was increased to 42.4 g/L and the cultivation time was extended to 240 hr. However, iso-citric acid (ICA) yield in fed-batch cultivation (4.0 g/L) was similar to that during the batch cultivation (3.9 g/L), and only 1.6 g/L of reducing sugar was left in the medium at the end of fed-batch cultivation, suggesting that most of the added carbon was used in the cultivation.     

Effect of chemical treatments on the degradability of cotton straw by rumen microorganisms and by fungal cellulase

Three different chemical treatments—sulfur dioxide, ozone, and sodium hydroxide—were applied on cotton straw, and the effect on cell-wall degradability was assessed by using rumen microorganism and Trichoderma reesei cellulase. Sulfur dioxide (applied at 70°C for 72 h) did not change the lignin content of cotton straw but reduced the concentration of hemicellulose by 48%. Ozone exerted a dual effect, both on lignin (a 40% reduction) and hemicellulose (a 54% decrease). The treatment with NaOH did not solublize cell-wall components. The in vitro organic matter digestibility with rumen fluid of cotton straw was increased significantly by ozone and SO₂ treatments, by 120% and 50%, respectively, but not by NaOH. T. reesei cellulase was applied on the chemically pretreated cotton straw at a low level (6 filter paper U/g straw, organic matter), and the release of reducing sugars was determined. The highest level of reducing sugars (30.6 g/100 g organic matter) was obtained with the O₃-cellulase combination, which solubilized 64% of the cellulose and 88% of the hemicellulose. the SO₂- and the NaOH-pretreated cotton straw were hydrolyzed by T. reesei cellulase to the same extent (21 g reducing sugars/100 g organic matter). The rumen fluid digestibility of the enzymatic ally hydrolyzed straw was not increased further over the effect already obtained with the chemical pretreatments. However, the fermentability of the combined treatments was increased markedly. In the O₃-cellulase-treated cotton straw, 83% of the rumen fluid digestible material consisted of highly fermentable components. Although ozone proved to be the most potent pretreatment for enzymic saccharification in this study, the absolute result was modest. The limited effect of the combined O₃-cellulase treatment was probably associated with the pretreatment limitations, but not with the enzyme level. Based on the differential response of the chemically treated cotton straw to attack by rumen microorganisms on the one hand, and by T. reesei cellulase on the other hand, a hypothesis has been suggested as to the location of lignin and hemicellulose in the cellwall unit of cotton straw.     

Evaluation of genetic and phenotypic consistency of <Emphasis Type="Italic">Bacillus coagulans</Emphasis> MTCC 5856: a commercial probiotic strain

Commercial probiotics preparation containing Bacillus coagulans have been sold in the market for several decades. Due to its high intra-species genomic diversity, it is very likely that B. coagulans strain may alter in different ways over multiple years of production. Therefore, the present study focuses to evaluate the genetic consistency and probiotic potential of B. coagulans MTCC 5856. Phenotypic and genotypic techniques including biochemical profiling, 16S rRNA sequencing, GTG 5″, BOX PCR fingerprinting, and Multi-Locus-Sequence typing (MLST) were carried out to evaluate the identity and consistency of the B. coagulans MTCC 5856. Further, in vitro probiotic potential, safety and stability at ambient temperature conditions of B. coagulans MTCC 5856 were evaluated. All the samples were identified as B. coagulans by biochemical profiling and 16S rRNA sequencing. GTG 5″, BOX PCR fingerprints and MLST studies revealed that the same strain was present over 3 years of commercial production. B. coagulans MTCC 5856 showed resistance to gastric acid, bile salt and exhibited antimicrobial activity in in-vitro studies. Additionally, B. coagulans MTCC 5856 was found to be non-mutagenic, non-cytotoxic, negative for enterotoxin genes and stable at ambient temperature (25 ± 2 °C) for 36 months. The data of the study verified that the same strain of B. coagulans MTCC 5856 was present in commercial preparation over multiple years of production.     

High expression of a neutral endo-β-glucanase gene from Humicola insolens in Trichoderma reesei

The neutral endo-β-glucanase gene cel5A from Humicola insolens was cloned and connected with the cellobiohydrolase 1 promoter from Trichoderma reesei to construct a recombinant plasmid pCB-hEG with the hygromycin B resistance marker. The plasmid was introduced into conidia of T. reesei using the Agrobacterium tumefaciens mediated transformation method. Eight transformants were obtained on screening plates with sodium carboxymethyl cellulose as the sole carbon source. Stable integration of the cel5A gene into the chromosomal DNA of T. reesei was confirmed by PCR. An obvious protein band (approximately 52 kDa) was detected by SDS-PAGE from fermentation broth, which showed that the cel5A gene in recombinant T. reesei successfully fulfilled efficient expression and extracellular secretion. After 96 h shaking-flask fermentation, the endo-β-glucanase activity at pH 6.5 from recombinant T. reesei reached 3,068 U/ml, which was 11 times higher than that of the host strain. In a 2 m³ fermenter, the endo-β-glucanase activity could be further increased to 8,012 U/ml after 96 h fermentation. The results showed a good prospect for application of neutral endo-β-glucanase in the textile industry.     

Assessing biodegradation of furfuryl alcohol using <Emphasis Type="Italic">Pseudomonas putida</Emphasis> MTCC 1194 and <Emphasis Type="Italic">Pseudomonas aeruginosa</Emphasis> MTCC 1034 and its kinetics

The biodegradation of furfuryl alcohol (FA) in shake flask experiments using a pure culture of Pseudomonas putida (MTCC 1194) and Pseudomonas aeruginosa (MTCC 1034) was studied at 30 °C and pH 7.0. Experiments were performed at different FA concentrations ranging from 50 to 500 mg/l. Before carrying out the biodegradation studies, the bacterial strains were acclimatized to the concentration of 500 mg/l of FA by gradually raising 100 mg/l of FA in each step. The well acclimatized culture of P. putida and P. aeruginosa degraded about 80 and 66% of 50 mg/l FA, respectively. At higher concentration of FA, the percentage of FA degradation decreased. The purpose of this study was to determine the kinetics of biodegradation of FA by measuring biomass growth rates and concentration of FA as a function of time. Substrate inhibition was calculated from experimental growth parameters using the Haldane equation. Data for P. putida were determined as µ _max ?=?0.23 h^?1, K _s ?=?23.93 mg/l and K _i ?=?217.1 mg/l and for P. aeruginosa were determined as µ _max ?=?0.13 h^?1, K _s ?=?21.3 mg/l and K _i ?=?284.9 mg/l. The experimental data were fitted in Haldane, Aiba and Edwards inhibition models.     

Impact of Proteolytic <Emphasis Type="Italic">Lactobacillus helveticus</Emphasis> MTCC5463 on Production of Bioactive Peptides Derived from Honey Based Fermented Milk

In this study, Lactobacillus helveticus MTCC5463 was evaluated for its proteolytic activity and production of bioactive peptides during fermentation of honey supplemented milk under specified growth conditions. Generally, lactic acid bacteria have a strong proteolytic system. However, L. helveticus MTCC5463 showed maximum proteolytic activity at 4 % level of honey supplementation compared to 6 % and control. Similarly, water soluble extract derived from fermented honey based milks exhibited different level of bioactive peptides productions during fermentation. L. helveticus MTCC5463 showed maximum peptides production at 4 % level of honey supplementation compared to control during HPLC analysis and LC–MS analysis.     

Effect of Different Carbon Sources on Decolourisation of an Industrial Textile Dye Under Alkaline–Saline Conditions

White-rot fungal strains of Trametes versicolor and Phanerochaete chrysosporium were selected to study the decolourisation of the textile dye, Reactive Black 5, under alkaline–saline conditions. Free and immobilised T. versicolor cells showed 100 % decolourisation in the growth medium supplemented with 15 g l^?1 NaCl, pH 9.5 at 30 °C in liquid batch culture. Continuous culture experiments were performed in a fixed-bed reactor using free and immobilised T. versicolor cells and allowed 85–100 % dye decolourisation. The immobilisation conditions for the biomass and the additional supply of carbon sources improved the decolourisation performance during a long-term trial of 40 days. Lignin peroxidase, laccase and glyoxal oxidase activities were detected during the experiments. The laccase activity varied depending on carbon source utilized and glycerol-enhanced laccase activity compared to sucrose during extended growth.     

Purification and characterization of pectin lyase secreted by Aspergillus flavus MTCC 10938

An indigenously isolated fungal strain Aspergillus flavus MTCC 10938 was subjected to pectin lyase (PNL) production under submerged fermentation conditions. The enzyme was purified to homogeneity from the culture filtrate of the fungus involving concentration by ultrafiltration, anion exchange chromatography on DEAE cellulose and gel filtration chromatography on Sephadex G-100. The purified PNL gave a single protein band in SDS-PAGE analysis with a relative molecular mass corresponding to 50 kDa. Using citrus pectin as the substrate the K _m and k _cat values of the enzyme were obtained as 1.7 mg/ml and 66 s^?1, respectively. The optimum pH of the purified PNL from A. flavus MTCC 10938 was 8.0 and up to 90% of its activity retained in the pH range from 3.0 to 11.0 after 24 h incubation. The optimum temperature of the purified enzyme was revealed at 55°C and it was completely stable up to 40°C when exposed for 30 min. The purified A. flavus MTCC 10938 PNL showed efficient retting of Crotalaria juncea fibres.     

Improvement of cellulase activity in Trichoderma reesei by heterologous expression of a beta-glucosidase gene from Penicillium decumbens

Trichoderma reesei is a well-known cellulase producer and widely applied in enzyme industry. To increase its ability to efficiently decompose cellulose, the beta-glucosidase activity of its enzyme cocktail needs to be enhanced. In this study, a beta-glucosidase I coding sequence from Penicillium decumbens was ligated with the cellobiohydrolase I (cbh1) promoter of T. reesei and introduced into the genome of T. reesei strain Rut-C30 by Agrobacterium-mediated transformation. In comparison to that from the parent strain, the beta-glucosidase activity of the enzyme complexes from two selected transformants increased 6- to 8-fold and their filter paper activity (FPAs) was enhanced by 30% on average. The transformant's saccharifying ability towards pretreated cornstalk was also significantly enhanced. To further confirm the effect of heterologous beta-glucosidase on the cellulase activity of T. reesei, the heterologously expressed pBGL1 was purified and added to the enzyme complex produced by T. reesei Rut-C30. Supplementation of the Rut-C30 enzyme complex with pBGL1 brought about 80% increase of glucose yield during the saccharification of pretreated cornstalk. Our results indicated that the heterologous expression of a beta-glucosidase gene in T. reesei might produce balanced cellulase preparation.     

Characterization of novel phospholipase C from Bacillus licheniformis MTCC 7445 and its application in degumming of vegetable oils

A novel microbial phospholipase C (PLC) from Bacillus licheniformis MTCC 7445 was purified to homogeneity by ammonium sulphate fractionation, dialysis, anion exchange chromatography and gel exclusion chromatography. The bacteria growing on vegetable oils secreted significantly high amount of PLC. The enzyme was purified to 23.4-fold with 46% recovery and specific activity 398 U/mg. It exhibited optimum activity at 70°C and pH 10.0. Using diphosphatidylglycerol as substrate the PLC of B. licheniformis MTCC 7445 had a V _max and K _m of 0.68 mM/min and 32 mM, respectively. It hydrolyzed phosphatidylinositol and phosphatidylserine as well as phosphatidylcholine but not other glycerophospholipids. Its activity was enhanced by 113% with Mn²⁺ and 110% with Mg²⁺. During degumming of vegetable oils with this enzyme preparation, the phosphorus content of the oil became lower than 4 mg/kg after 5 h of enzyme treatment at 40°C. The novel PLC from B. licheniformis MTCC 7445 is potentially useful for the refining of high quality oils with 95% removal of phospholipids with attractive yield.     

Simultaneous saccharification and fermentation of enzyme pretreated Lantana camara using S. cerevisiae

Lantana camara, an abundantly available non-edible lignocellulosic biomass has been found to be a potential feedstock for ethanol production. The substrate was first pretreated with laccase followed by simultaneous saccharification and fermentation using cellulase and Saccharomyces cerevisiae, respectively. Laccase was produced from Pleurotus sp. and carbohydratases (cellulase and xylanase) were produced from Trichoderma reesei Rut C30. Using pretreated substrate simultaneous saccharification and fermentation was optimized through central composite design-based response surface methodology. Maximum bioethanol concentration of 5.14 % (v/v) was obtained at optimum process conditions of substrate concentration 17 % (w/v), inoculum volume 9 % (v/v), inoculum age 60 and 144 h of incubation time. To enhance ethanol yield, S. cerevisiae was treated with ethyl methane sulfonate, a chemical mutagenic agent which induced mutagenesis. A maximum bioethanol concentration of 6.01 % (v/v) was obtained using the mutated strain of S. cerevisiae (CM5).     

Kinetic studies of laccase enzyme of Coriolus versicolor MTCC 138 in an inexpensive culture medium

An attempt was made to use cyanobacterial biomass of water bloom, groundnut shell (GNS) and dye effluent as culture medium for laccase enzyme production by Coriolus versicolor. Laccase production was found to be 10.15 ± 2.21 U/ml in the medium containing groundnut shell and cyanobacterial bloom in a ratio of 9:1 (dry weight basis) in submerged fermentation at initial pH 5.0 and 28 ± 2 °C temperature. Half life of enzyme was found to be 74 min at 60 °C. Kinetic analysis of laccase when made with substrate ABTS, K_m and V_max were found to be 0.29 mM and 9.49 μmol/min respectively. Azide and hydroxylamine were found to exert significant inhibition on thermostable laccase. Inhibitor constant (k_i) for azide and hydroxylamine were 1.33 and 0.18 mM respectively. This study forms the first report on the potential application of waste water cyanobacterial bloom and dyeing effluent as a medium for laccase production by C. versicolor MTCC138.     

Utilization of agro-industrial waste for xylanase production by Aspergillus foetidus MTCC 4898 under solid state fermentation and its application in saccharification

Xylanase production by Aspergillus foetidus MTCC 4898 was carried out under solid state fermentation using wheat bran and anaerobically treated distillery spent wash. Response surface methodology involving Box–Behnken design was employed for optimizing xylanase production. The interactions among various fermentation parameters viz. moisture to substrate ratio, inoculum size, initial pH, effluent concentration and incubation time were investigated and modeled. The predicted xylanase activity under optimized parameters was 8200–8400 U/g and validated xylanase activity was 8450 U/g with very poor cellulase activity. Crude xylanase was used for enzymatic saccharification of agroresidues like wheat straw, rice straw and corncobs. Dilute NaOH and ammonia pretreatments were found to be beneficial for the efficient enzymatic hydrolysis of all the three substrates. Dilute NaOH pretreated wheat straw, rice straw and corncobs yielded 4, 4.2, 4.6 g/l reducing sugars, respectively whereas ammonia treated wheat straw, rice straw and corncobs yielded 4.9, 4.7, 4.6 g/l reducing sugars, respectively. The hydrolyzates were analysed by HPTLC. Xylose was found to be the major end product with traces of glucose in the enzymatic hydrolyzates of all the substrates.