Production of fructosyl transferase by <Emphasis Type="Italic">Aspergillus oryzae</Emphasis> CFR 202 in solid-state fermentation using agricultural by-products

Fructosyl transferase (FTase) production by Aspergillus oryzae CFR 202 was carried out by solid-state fermentation (SSF), using various agricultural by-products like cereal bran, corn products, sugarcane bagasse,cassava bagasse (tippi) and by-products of coffee and tea processing. The FTase produced was used for the production of fructo-oligosaccharides (FOS), using 60% sucrose as substrate. Among the cereal bran used, rice bran and wheat bran were good substrates for FTase production by A. oryzae CFR 202. Among the various corn products used, corn germ supported maximum FTase production, whereas among the by-products of coffee and tea processing used, spent coffee and spent tea were good substrates, with supplementation of yeast extract and complete synthetic media. FTase had maximum activity at 60°C and pH 6.0. FTase was stable up to 40°C and in the pH range 5.0–7.0. Maximum FOS production was obtained with FTase after 8 h of reaction with 60% sucrose. FTase produced by SSF using wheat bran was purified 107-fold by ammonium sulphate precipitation (30–80%), DEAE cellulose chromatography and Sephadex G-200 chromatography. The molecular mass of the purified FTase was 116.3 kDa by SDS-PAGE. This study indicates the potential for the use of agricultural by-products for the efficient production of FTase enzyme by A. oryzae CFR 202 in SSF, thereby resulting in value addition of those by-products.     

Mixture design as first step for improved glutaminase production in solid-state fermentation by isolated Bacillus

Aim: Investigation of mixture‐design impact on glutaminase production by isolated Bacillus sp. Methods and Results: An augmented simplex centroid design was used to optimize a three (wheat bran, Bengal gram husk and palm seed fibre) component mixture for glutaminase production. Selected substrate materials showed impact on glutaminase production values at individual level by Bengal gram husk [2789 U gds^?1 (gram dry substrate] and in two‐level combination with wheat bran and Bengal gram husk (maximum of 3300 U gds^?1). Conclusion: Bengal gram husk is the most suitable substrate medium for glutaminase production by Bacillus sp. Maximum glutaminase production is achieved using solid‐substrate mixture at two‐level combinations in the ratio of 66 : 34 for Bengal gram husk and wheat bran, respectively. Significance and Impact of the Study: The present study has significance in large‐scale production of glutaminase at commercial level with the use of multi‐substrate rather than single‐substrate/support material.     

Acid Protease Production by Fungi Used in Soybean Food Fermentation

Growth conditions for maximum protease production by Rhizopus oligosporus, Mucor dispersus, and Actinomucor elegans, used in Oriental food fermentations, were investigated. Enzyme yields by all three fungi were higher in solid substrate fermentations than in submerged culture. The level of moisture in solid substrate must be at about 50 to 60%. Very little growth of these fungi was noted when the moisture of substrate was below 35%, whereas many fungi including most storage fungi generally grow well on solid substrate with that level of moisture. Among the three substrates tested—wheat bran, wheat, and soybeans—wheat bran was the most satisfactory one for enzyme production. The optimal conditions for maximum enzyme production of the three fungi grown on wheat bran were: R. oligosporus, 50% moisture at 25 C for 3 to 4 days; M. dispersus, 50 to 63% moisture at 25 C for 3 to 4 days; A. elegans, 50 to 63% moisture at 20 C for 3 days. Because these fungi are fast growing and require high moisture for growth and for enzyme synthesis, the danger of contamination by toxin-producing fungi would be minimal.     

Solid‐state fermentation for the production of debittering enzyme naringinase using Aspergillus niger MTCC 1344

Aspergillus niger produced high levels of naringinase using easily available, inexpensive industrial waste residues such as rice bran, wheat bran, sugar cane bagasse, citrus peel, and press mud in solid‐state fermentation (SSF). Among these, rice bran was found to be the best substrate. Naringinase production was highest after 96 h of incubation at 27°C and at a substrate‐to‐moisture ratio of 1:1 w/v. Supplementation of the medium with 10% naringin caused maximum induction. An inoculum age of 72 h and an inoculum level of 15% resulted in maximum production of naringinase. Enzyme production was stimulated by the addition of nutrients such as naringin and peptone. Thus, A. niger produced a very high level of naringinase within a short time in solid‐state fermentation using inexpensive agro‐residues, a level that is much higher than reported for any other microbes.     

Cellulolytic enzymes on lignocellulosic substrates in solid state fermentation by <Emphasis Type="Italic">Aspergillus niger</Emphasis>

The production of cellulolytic enzymes by Aspergillus niger on lignocellulosic substrates groundnut fodder, wheat bran, rice bran and sawdust in solid state fermentation in a laboratory scale was compared. Czapek Dox liquid broth amended with cellulose (0.5%) was used to moisten lignocellulosic solid supports for cultivation of Aspergillus niger. The production of filter paperase, carboxymethyl cellulase and -glucosidase were monitored at daily intervals for 5 days. The peak production of the enzymes occurred within 3 days of incubation. Among solid supports used in the study, wheat bran was the best solid matrix followed by groundnut fodder in production of cellulolytic enzymes in solid state fermentation. Groundnut fodder supported significant production of FPase (2.09 FPU/g), CMCase (1.36 U/g) and -glucosidase activity (0.0117 U/g) in solid state fermentation. Considerable secretion of protein (5.10 mg/g) on groundnut fodder at peak time interval 1st day of incubation was recorded.     

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.     

Production of raw cassava starch-digestive glucoamylase by Rhizopus sp. in liquid culture

Among about 200 Rhizopus strains isolated in Thailand, Rhizopus sp. MB46 was selected as a producer of raw cassava starch-digestive glucoamylase. Rice bran was effective for the enzyme production in a solid culture as well as wheat bran. Addition of turpentine oil into the rice bran solid culture increased the productivity. Rhizopus sp. MB46 was found to produce glucoamylase in a liquid culture containing 1% rice bran but not in one consisting of 10% raw cassava starch of 2% glucose. The productivity per 1 g solids in the medium in liquid culture was finally improved 6-times by utilization of n-hexane-treated rice bran, supplement of 0.1% meat extract and addition of gauze as a support. The activity was superior to that in turpentine oil-supplemented solid culture.     

Kinetics of Enhanced Substrate Consumption and Endo-β-xylanase Production by a Mutant Derivative of Humicola lanuginosa in Solid-state Fermentation

Summary Industrial byproducts namely canola meal, rice bran, sunflower meal, and wheat straw were used as substrates for endo-xylanase production by Humicola lanuginosemutant TH1 through solid substrate fermentation. The enzyme was secreted extracellularly by both wild and mutant cultures. Rice bran supported the maximum production of endo-xylanase followed by wheat straw, canola meal and sunflower meal. The highest activity was achieved after 72 h of culture and the highest yields from the above substrates were 842, 840, 610 and 608 IU per g substrate consumed respectively. The highest productivity (281 IU flask⁻¹ h⁻¹ corresponding to 5620 l⁻¹ h^-1) of endo-xylanase by the mutant of H. lanuginosa was 1.6-fold more than that produced by the parental organism in solid-state fermentation of rice bran at 45 °C. Maximum specific activity (180 IU mg⁻¹ protein) and substrate consumption rates were significantly more than those reported by previous researchers on Humicola sp. The mutant possessed markedly low accompanying cellulase activity. Thermodynamic studies revealed that the mutant required significantly lower activation energy for enzyme production and higher for thermal inactivation which signified that the endogenous metabolic machinery of mutant cells exerted more protection against thermal inactivation during product formation than that needed by its parental cultures.     

Comparative study of mycelial growth and basidiomata formation in seven different species of the edible mushroom genus Hericium

The potential of using several agricultural by-products as supplements of sawdust substrate for the production of edible mushroom Hericium was evaluated using seven Hericium species. All the tested supplements (rice bran, wheat bran, barley bran, Chinese cabbage, egg shell, and soybean powder) were found to be suitable for the mycelial growth of all the tested species. In mycelial growth, soybean powder was the best supplement for Hericium americanum, Hericium coralloides, and Hericium erinaceum while barley bran was the best for Hericium alpestre, Hericium laciniatum, and Hericium erinaceus. For Hericium abietis, rice bran and Chinese cabbage was the best. The possibility of mushroom production on oak sawdust substrate with 20% rice bran supplement was demonstrated with H. coralloides, H. americanum, H. erinaceus, and H. erinaceum which showed 26-70% biological efficiency. Our results also showed that strain selection is important to improve biological efficiency and mushroom yield in Hericium cultivation.     

Production of cellulases and d-xylanase by some selected fungal isolates

Cellulase [see 1,4-(1,3;1,4)-β-d-glucan 4-glucanohydrolase, EC 3.2.1.4], β-d-glucosidase (β-d-glucoside glucanohydrolase, EC 3.2.1.21) and d-xylanase (1,4-β-d-xylan xylanohydrolase, EC 3.2.1.8) production by Aspergillus ustus, Sporotrichum pulverulentum, Trichoderma sp. (a), Trichoderma sp. (b) and Botrytis sp. in solid state fermentation on different compounded media containing wheat bran (WB), rice straw (RS) and minerals was studied. Toyama's mineral solution mixed with RS was found to be a better substrate for cellulase and d-xylanase while with WB it induced higher β-d-glucosidase production. A ratio of substrate to mineral solution (w/v) of 1:4 or 1:5 supported high d-xylanase and cellulase production whereas a ratio of 1:2 gave the highest β-d-glucosidase activity. Among the fungal isolates, Aspergillus ustus gave the highest β-d-glucosidase activity of 60 U g⁻¹WB and the highest d-xylanase activity of 740 U g⁻¹was obtained with RS. A mixture of seven parts of RS and three of WB, mixed with 40 parts of Toyama's mineral solution yielded 6 U filter paper activity, 40 U β-d-glucosidase, 12 U carboxymethylcellulase and 650 U d-xylanase g⁻¹substrate.     

Effect of culture substrates on virulence of Beauveria bassiana (Ascomycota: Cordycipitaceae) conidia against the browntail moth,Euproctis chrysorrhoea (Lepidoptera: Lymantriidae)

We studied the effect of different solid substrates on virulence of two Beauveria bassiana isolates against the browntail moth, Euproctis chrysorrhoea (L.) (Lep.: Lymantriidae). Conidia produced on wheat grains, wheat flour, wheat bran, rice flour, rice bran, rice paddy, corn flour, millet, and Sabouraud's dextrose agar with 1% yeast extract (SDAY) as control were compared. There were significant differences among these substrates for their effects on the virulence of produced conidia. Applying 10⁷ conidia/mL of B. bassiana EUT105, produced on rice bran caused the highest (84.9%) and on rice flour, the lowest (57.6%) mortalities. Bioassay on fifth-instar larvae using aerial conidia harvested from wheat grains, rice paddy, and SDAY indicated that conidia from wheat grains had the highest virulence while those from rice paddy, the lowest.     

Xylanase production by Ganoderma lucidum on liquid and solid state fermentation

Ganoderma lucidum, a white rot fungus, was exploited for its potentials to produce xylanase employing shake and solid-state culture conditions. Different culture conditions such as pH, temperature, carbon and nitrogen requirements for its growth and production of xylanase were optimized. The culture media pH 6.0-7.0 and temperatures 30 degrees-35 degrees C significantly promoted the growth as well as xylanase secretion into the media. Xylan and peptone were found to be the suitable carbon and nitrogen sources. Among the different agrowastes used, wheat bran was found to be the best substrate for the test fungus for the production of xylanase than sugarcane bagasse and rice bran in solid-state fermentation.     

Isolation and characterization of Aspergillus sp. for the production of extracellular polysaccharides by response surface methodology

In this study, Aspergillus sp. was isolated for the production of extracellular polysaccharide. The process parameters were initially optimized by traditional methods. The cheap substrate, wheat bran was used for the production of extracellular polysaccharide in solid state fermentation. Supplementation of (1%, w/w) maltose, gelatin enhanced EPS production (5.36?mg/g). The salts such as, Cu²⁺ (4.9?mg/g), Ca²⁺ (3.5?mg/g), Zn²⁺ (2.9?mg/g), Mn²⁺ (3.4?mg/g) and Mg²⁺ (1.8?mg/g) stimulated EPS production. In two level full factorial experimental designs, the EPS yield varied from 3.18 to 11.65?mg/g wheat bran substrate with various combinations of the components supplemented with wheat bran substrate. Among these selected factors in central composite design, maltose significantly influenced on extracellular polysaccharide production.     

蜡蚧轮枝菌固体发酵基质的筛选与组分优化研究

对昆虫病原真菌一蜡蚧轮枝菌菌株Tri—BA81进行5种单一基质和4种合基质固体发酵,初筛出谷子单一基质和谷子＋麦麸＋磷酸盐组合基质为最佳发酵基质,后者的产孢量是前者的3倍多。选取谷子、麦麸、磷酸盐和稻壳为4个组分因子,每个因子分别设有不同质量比例的3个水平,按正交设计（L94^3）进行优化组合筛选研究。结果表明,产孢量最高的正交组合为5号配方,分生孢子产量达1．68×10^10个／g。     

L-glutaminase production by <Emphasis Type="Italic">Trichoderma koningii</Emphasis> under solid-state fermentation

Solid state fermentation was conducted for the production of L-glutaminase by Trichoderma koningii Oud.aggr. using different agro-industrial byproducts inlcuding wheat bran, groundnut residues, rice hulls, soya bean meal, corn steep, sesamum oil cake, cotton seed residues and lentil industrial residues as solid substrates. Wheat bran was the best substrate for induction of L-glutaminase (12.1 U/mg protein) by T. koningii. The maximum productivity (23.2 U/mg protein) and yield (45.0 U/gds) of L-glutaminase by T. koningii occurred using wheat bran of 70% initial moisture content, initial pH 7.0, supplemented with D-glucose (1.0%) and L-glutamine (2.0% w/v), inoculated with 3 ml of 6 day old fungal culture and incubated at 30°C for 7 days. After optimization, the productivity of L-glutaminase by the solid cultures of T. koningii was increased by 2.2 fold regarding to the submerged culture.     

The use of spent brewery grains for Pleurotus ostreatus cultivation and enzyme production

In the brewing industry, spent brewery grains (SBGs) are byproducts with a low economic value. The potential use of this leftover as a substrate ingredient for Pleurotus ostreatus fruiting body cultivation and enzyme production was evaluated. The best substrate mixture for P. ostreatus mycelium growth comprised 30% wheat bran (WB), 68% beech sawdust (BS) and 2% CaCO3. On the substrates containing SBG, the fastest mycelium growth was observed on the substrate composed of 10% SBG, 20% WB, 68% BS and 2% CaCO3. The highest biological efficiency (51%) of fruiting bodies was determined on the mixtures containing 20% WB, 10% SBG and 2% CaCO3. The SBGs with the addition of WB were also shown to be suitable as a substrate for enzyme production. However, the supplementation levels designate which enzymes are produced and in what amounts.     

Optimization of xylanase production using inexpensive agro-residues by alkalophilic Bacillus subtilis ASH in solid-state fermentation

Alkalophilic Bacillus subtilis ASH produced high levels of xylanase using easily available inexpensive agricultural waste residues such as wheat bran, wheat straw, rice husk, sawdust, gram bran, groundnut and maize bran in solid-state fermentation (SSF). Among these, wheat bran was found to be best substrate. Xylanase production was highest after 72 h of incubation at 37 °C and at a substrate to moisture ratio of 1:2 (w/v). The inoculum level of 15% resulted in maximum production of xylanase. The enzyme production was stimulated by the addition of nutrients such as yeast extract, peptone and beef extract. In contrast, addition of glucose and xylose repressed the production of xylanase. The extent of repression by glucose (10%, w/v) was 81% and it was concentration-dependent. Supplementation of the medium with 4% xylose caused 59% repression. Under optimized conditions, xylanase production in SSF (8,964 U of xylanase/g dry wheat bran) was about twofold greater than in submerged fermentation. Thus, B. subtilis produced a very high level of xylanase in SSF using inexpensive agro-residues, a level which is much higher than that reported by any other bacterial isolate. Furthermore, the enzyme was produced at room temperature and with tap water without the addition of any mineral salt in SSF, leading to a marked decrease in the cost of xylanase production, which enhances its industrial potential.     

绿色木霉ZY-1固态发酵产纤维素酶

利用筛选的绿色木霉ZY-1（Trichoderma viride ZY-1）固态发酵产纤维素酶,采用稻草和麸皮为底物,考察稻草与麸皮比例随发酵时间对产酶的影响。结果表明：底物中,在m（稻草）：m（麸皮）为0：5和1：4时,发酵48h,pH保持4．5左右,还原糖量急剧上升,胞外蛋白产量最低;仅以稻草作底物时,整个发酵过程中pH约为7,还原糖量最低,胞外蛋白产量较高而滤纸酶活、羧甲基纤维素酶（CMCase）和β-葡萄糖苷酶（β-Gase）酶活均较低;在m（稻草）：m（麸皮）为3：2时,发酵96h,滤纸酶活达最大值5．01U／g干曲;m（稻草）：m（麸皮）为1：4时,发酵96h,β-Gase酶活达最大值4．6U／g干曲;m（稻草）：m（麸皮）为4：1时,发酵72h,CMCase酶活达最大值6．01U／g干曲。因此,底物中存在适量的稻草和麸皮有利于Trichoderma viride ZY—1产纤维素酶。     

Influence of the carbon source on the growth and lignocellulolytic enzyme production by Morchella esculenta strains

Growth and lignocellulolytic enzymes production by two Morchella esculenta strains (BAFC 1728 and BEL 124) growing in solid state fermentation using different lignocellulosic materials along 58 days was characterized. Both strains were able to grow on the three substrates: wheat bran, wheat bran plus corn starch, and rolled oat. The growth was characterized by measuring chitin content, reducing sugars, pH, dry weight loss, and extractable proteins, such parameters varied substantially with substrate and strain used. The maximum rate of growth in both strains was observed between 5 and 28 days. Regarding enzyme production, as a general trend strain BAFC 1728 produced the highest titres. The most evident difference was observed in laccase production by this strain on wheat bran, which exceeded that observed in strain BEL 124 by tenfold (7.45 U g⁻¹).     

Cloning of a novel feruloyl esterase gene from rumen microbial metagenome and enzyme characterization in synergism with endoxylanases

A feruloyl esterase (FAE) gene was isolated from a rumen microbial metagenome, cloned into E. coli, and expressed in active form. The enzyme (RuFae2) was identified as a type C feruloyl esterase. The RuFae2 alone released ferulic acid from rice bran, wheat bran, wheat-insoluble arabinoxylan, corn fiber, switchgrass, and corn bran in the order of decreasing activity. Using a saturating amount of RuFae2 for 100 mg substrate, a maximum of 18.7 and 80.0 μg FA was released from 100 mg corn fiber and wheat-insoluble arabinoxylan, respectively. Addition of GH10 endoxylanase (EX) synergistically increased the release of FA with the highest level of 6.7-fold for wheat bran. The synergistic effect of adding GH11 EX was significantly smaller with all the substrates tested. The difference in the effect of the two EXs was further analyzed by comparing the rate in the release of FA with increasing EX concentration using wheat-insoluble arabinoxylan as the substrate.