Production of high activity thermostable phytase from thermotolerant Aspergillus niger in solid state fermentation

The thermotolerant fungus, Aspergillus niger NCIM 563, was used for production of extracellular phytase on agricultural residues: wheat bran, mustard cake, cowpea meal, groundnut cake, coconut cake, cotton cake and black bean flour in solid state fermentation (SSF). Maximum enzyme activity (108 U g⁻¹ dry mouldy bran, DMB) was obtained with cowpea meal. During the fermentation phytic acid was hydrolysed completely with a corresponding increase in biomass and phytase activity within 7 days. Phosphate in the form of KH₂PO₄ (10 mg per 100 g of agriculture residue) increased phytase activity. Among various surfactants added to SSF, Trition X-100 (0.5%) exhibited a 30% increase in phytase activity. The optimum pH and temperature of the crude enzyme were 5.0 and 50°C respectively. Phytase activity (86%) was retained in buffer of pH 3.5 for 24 h. The enzyme retained 75% of its activity on incubation at 55°C for 1 h. In the presence of 1 mM K⁺ and Zn²⁺, 95% and 55% of the activity were retained. Scanning electron microscopy showed a high density growth of fungal mycelia on wheat bran particles during SSF. Journal of Industrial Microbiology & Biotechnology (2000) 24, 237–243. Received 07 June 1999/ Accepted in revised form 18 December 1999     

Protease-conidia relationships of Aspergillus niger grown in solid state fermentation

Protease activity of Aspergillus niger growing on solid substrate correlated well with conidia formation (R: 0.91–0.96) for initial moisture contents of 38–48% (wet basis), initial pH 5.4 and 6 and temperature (29–37 °C ). However, conidia/protease ratio varied with most of these conditions and by NaCl addition indicating only a partial association between them.     

Cellulase production by solid state fermentation on lignocellulosic waste from the xylose industry

Cellulase production was carried out by solid state fermentation using corncob residue, a lignocellulosic waste from the xylose industry, as the substrate of Trichoderma reesei ZU-02. The effects of water content, dosage of wheat bran and initial pH value in solid substrate on cellulase synthesis were studied in shallow tray fermentors. The solid substrate could be reused in at least three batches and the highest cellulase activity (158 IFPU/g koji) was obtained in the second fermentation batch. To produce cellulase on a larger scale, a deep trough fermentor with forced aeration was used and 128 IFPU/g koji (305 IFPU/g cellulose) was reached after 5 days solid state fermentation. The enzyme koji produced in the present process can be used directly to hydrolyze corncob residue effectively, when the cellulase dosage was above 20 IFPU/g substrate, the saccharification yield could be over 84%.     

Production and characterization of a highly active cellobiase from Aspergillus niger grown in solid state fermentation

Summary Aspergillus niger produced extracellular cellobiase when grown on different lignocellulosic substrates in solid state fermentation. The enzyme activity and yield were variable according to the carbon source. In Vogel’s medium, the cellobiase productivity was significantly higher on wheat bran, followed by Leptochloa fusca (kallar grass) straw augmented with corn steep liquor. Maximum yield of cellobiase/g wheat bran was significantly higher than the values reported on other potent fungi, bacteria and recombinants, harboring heterologous gene for cellobiase. This enzyme in the presence and absence of Trichoderma reesei and celluloclast, saccharified the biomass and the percentage saccharification as well as glucose yield from lignocellulosic biomass was doubled in its presence. The partially purified enzyme was thermotolerant as evidenced by melting temperature, activation energy demand for active catalysis, enthalpy and entropy of activation for reversible or irreversible thermal inactivation.     

Enhanced rhizosphere competence of Trichoderma viride grown in solid state fermentation on corn cob residue

Abstract

A process has been disclosed in the present investigation for the production of a formulation of mycopesticide with enhanced shelf life of Trichoderma viride. A comparative evaluation of talc-based and corn cob formulations in the form of seed coat and soil treatment brought to light that the soil treatment significantly enhanced the plant growth performance of pea, green gram and pigeon pea. Among the two formulations, the corn cob formulation proved to be better as the rhizosphere competence of corn cob formulation was superior to that of talc formulation. Our results suggest that corn cob residue is a better substrate owing to high cellulase and spore production. Solid-state fermentation in batch cultures. The biocontrol ability of both formulations was tested against Fusarium in Cajanus species and it was found that the disease incidence was reduced by 86% when the corn cob formulation was given as soil application.     

Biological detoxification of coffee husk by filamentous fungi using a solid state fermentation system

Studies were carried out on detoxification of coffee husk in solid state fermentation using three different strains of Rhizopus, Phanerochaete, and Aspergillus sp. Fungal strains were selected by their ability to grow on a coffee husk extract-agar medium. Using R. arrizus LPB-79, the best results on the degradation of caffeine (87%) and tannins (65%) were obtained with pH 6.0 and moisture 60% in 6 days. When P. chrysosporium BK was used, maximum degradation of caffeine and tannins were 70.8 and 45%, respectively, with coffee husk having 65% moisture and pH 5.5 in 14 days. The Aspergillus strain, isolated from the coffee husk, showed best biomass formation on coffee husk extract-agar medium. Optimization assays were conducted using factorial design, and surface response experiments with Aspergillus sp. The best detoxification rates achieved were 92% for caffeine and 65% for tannins. The results showed good prospects of using these fungal strains, in particular Aspergillus sp., for the detoxification of coffee husk.     

Utilisation of fruits waste for citric acid production by solid state fermentation

A solid state fermentation method was used to utilise pineapple, mixed fruit and maosmi waste as substrates for citric acid production using Aspergillus niger DS 1. Experiments were carried out in the presence and absence of methanol at different moisture levels. In the absence of methanol the maximum citric acid was obtained at 60% moisture level whereas in the presence of methanol the maximum citric acid was obtained at 70% moisture level. The stimulating effect of methanol was less at lower moisture level. The inhibitory effect of metal ions was also not observed and maximum citric acid yield of 51.4, 46.5 and 50% (based on sugar consumed) was obtained from pineapple, mixed fruit and maosmi residues, respectively.     

Use of various coffee industry residues for the cultivation of Pleurotus ostreatus in solid state fermentation

Studies were carried out to evaluate the feasibility of using coffee industry residues, viz. coffee husk, coffee leaves and spent coffee ground as substrates in solid state fermentation (SSF) to cultivate edible mushrooms Pleurotus. Eight strains of Pleurotus ostreatus and two strains of Pleurotus sajor‐caju were screened on a medium prepared from aqueous extract of coffee husk and agar. Based on best mycelial growth (9.68 mm/day) and biomass production (43.4 mg/plate in 9 days at 24°C), the strain P. ostreatus LPB 09 was selected for detailed studies. SSF was carried out using these substrates under different moisture conditions (45–75%) and spawn rates (2.5–25%). In general, although a 25% spawn rate appeared superior, the 10% spawn rate was recommended for all the three substrates in view of the process economics, as there was not any significant difference in the increase with 10 to 15%. The ideal moisture content for mycelial growth was 60–65% for coffee husk and spent coffee ground, and 60–70% for coffee leaves. The biological efficiency (BE), which is defined as the ratio of the weight of fresh fruiting bodies to the weight of dry substrate, multiplied by 100, and which indicates the fructification ability of the fungus for utilizing the substrate, was best with coffee husk. With coffee husk as the substrate, the first fructification occurred after 20 days of inoculation, and the biological efficiency reached about 97% after 60 days. When coffee leaves were used as the substrate, no fructification was observed even upon prolonged cultivation. With spent ground as the substrate, the first fructification occurred 23 days after inoculation and the biological efficiency reached about 90% in 50 days. There was a significant decrease in the caffeine and tannin contents (61 and 79%, respectively) of coffee husk after 60 days. It was remarkable to observe that caffeine was adsorbed onto the fruiting body (0.157%), indicating that it was not completely degraded by the fungal culture. However, no tannins were found in the fruiting body, indicating that the fungal strain was capable of degrading them. The results showed the feasibility of using coffee husk and spent coffee ground as substrates without any pre‐treatment for the cultivation of edible fungi in SSF, and provided one of the first steps towards an economical utilization of these otherwise unutilized or poorly utilized residues.     

Cassava fibrous waste residue: A substitute to wheat bran in solid state fermentation

Summary The cassava fibrous waste residue, after enrichment with urea, gave similar yields of fungal pectinases when used as a substitute to wheat bran in solid state fermentation. Comparatively, it is much less expensive and can be used in existing solid state fermentation plants without any modification.     

A novel thermostable xylanase from Thermomonospora sp.: influence of additives on thermostability

An alkalothermophilic Thermomonospora sp. producing high levels of xylanase was isolated from self-heating compost. The culture produced 125 IU/ml of xylanase when grown in shake flasks at pH 9 and 50 degrees C for 96 h. The culture filtrate also contained cellulase (23 IU/ml), mannanase (1 IU/ml) and beta-xylosidase (0.1 IU/ml) activities. The xylanase was active at a broad range of pH (5-9) and temperature (40-90 degrees C). The optimum pH and temperature were 7 and 70 degrees C, respectively. The enzyme was stable in the pH range 5-8 and was thermostable with half-lives of 8 and 4 h at 60 degrees C and 70 degrees C, respectively, but only 9 min at 80 degrees C. The effects of a variety of compounds to enhance the stability of xylanase at 80 degrees C was studied. Addition of sorbitol, mannitol and glycerol increased the thermostability of xylanase in proportion to the number of hydroxyl groups per polyol molecule. Glycine also offered protection against thermoinactivation. Xylan, trehalose, gelatin and trehalose-gelatin mixture had marginal effect on the thermostability of xylanase at 80 degrees C.     

Nutritional value and proteases of Lentinus citrinus produced by solid state fermentation of lignocellulosic waste from tropical region

This paper examined the growth and yield performance of Lentinus citrinus on cupuaçu exocarp (Theobroma grandiflorum) mixed with litter (CE + LI) or rice bran (Oryza sativa) (CE + RB) in the ratio of 2:1 (800 g:200 g) to investigate the nutritional composition and proteolytic potential of the fruiting body produced. Significance values of yield were determined on substrate combinations. In CE + LI the biological efficiency of the mushrooms was 93.5% and the content of fat (4.5%), fiber (11.0%), protein (27.0%) and amino acids were higher when compared with CE + RB. Among the amino acids, the amount of glutamic acid, aspartic acid, alanine, arginine and leucine was high. The biological efficiency on CE + RB reduced to 84.2% and based on the nutritional value, carbohydrates (53.59%), energy (324.33 kcal) and minerals such as zinc, iron, copper, potassium and phosphorus were higher in this substrate combination. Protease activity from fruiting body was significant in CE + LI (463.55 U/mL). This protease showed an optimal activity at 50 °C in neutral and alkaline pH with maximum stability at 30 °C at alkaline pH. This is the first report of L. citrinus fruiting body nutritional composition with potential for human food and application in industrial processes.     

烟梗为原料固态发酵生产果胶酶

以烟梗为主要原料,采用单因素和正交实验对筛选到的丝状菌JXY-17固态发酵产果胶酶的培养基进行了优化,正交实验结果表明,影响该菌株产果胶酶的因素依次为含水量(料水比)(A)＞(NH4)2SO4(B)＞KH2PO4(D)＞吐温-80(C),产酶培养基组成为A3B2C2D1,即固液比1∶1.5,(NH4)2SO4 5.0％,吐温-80 0.10％,KH2 PO40.20％.采用该固态发酵培养基,自然pH,接种量25 mL,装料量为50 g(干基)/1000 mL三角瓶,30℃恒温培养6d,产酶最高达8171.35U/g干曲,为初始酶活的3.8倍.提取酶液后的残余烟梗还可用于提取烟梗纤维类物质.残余烟梗的化学成分检测结果表明,与原始烟梗(或对照)相比,其果胶质降低了45％左右,残余烟梗固形物回收率约50％.     

Cloning of a thermostable alpha-amylase gene from Thermomonospora curvata and its expression in Streptomyces lividans

The gene from Thermomonospora curvata CCM 3312 coding for thermostable alpha-amylase (tam) has been cloned in Streptomyces lividans TK 24 and localized to a 2.6 kb HindIII-BamHI fragment of DNA. The data presented here show that the tam gene is expressed at a high level in S. lividans and that the protein is efficiently excreted.     

The production of plant cell wall polysaccharide-degrading enzymes by hemicellulolytic rumen bacterial isolates grown on a range of carbohydrate substrates

Several cultures of bacteria, isolated from the rumen, that were able to utilize plant cell wall structural polysaccharides were grown on a range of carbohydrate substrates and the activities of the principal polysaccharide-degrading enzymes determined. The esterase activity was also monitored. The extent of hemicellulose degradation and utilization by the isolates was comparable with that of the hemicellulolytic type strains. Enzyme activities in all of the cultures examined were affected by the carbon source in the growth medium. Many responses were strain specific, although growth on glucose (or cellobiose and maltose to a lesser extent) resulted in reduced activities in most of the organisms examined, whilst polysaccharidic substrates resulted in higher levels of the appropriate polysaccharidase. However, enzyme activity was detectable in some isolates after culture on mono- or disaccharides in the absence of the principal or related polysaccharide substrate.     

Growth of Candida utilis in solid state fermentation

In this work, the growth of the yeast Candida utilis on different solid substrate (wheat bran) and supports (sugarcane bagasse and Amberlite resin) imbibed with a liquid culture medium was studied. Growth was followed by sugars consumption, carbon dioxide production rate (CDPR) and cell count. The results showed the ability of the yeast to grow on the three solid media with fairly good viability and total dextrose consumption in the case of sugarcane bagasse and Amberlite, and partial consumption of wheat bran sugars. After two or three days of culture, a five hundred fold increase in cell population was observed.     

高产纤维素酶的拟康宁木霉菌株8985固态发酵条件优化

木霉是重要的产纤维素酶真菌,在其可利用性评价筛选过程中,获得了一株在实验室条件下高产纤维素酶的拟康宁木霉菌株8985.采用响应面法对8985产纤维素酶的固态发酵条件进行了研究,以滤纸酶活为响应值,通过Plackett-Burman设计对11个因素进行了筛选,包括温度、湿度、发酵时间、K2HPO4、(NH4)2SO4、T...     

樟绒枝霉(Malbranchea cinnamomea)固体发酵产木聚糖酶的发酵条件优化

[目的] 研究樟绒枝霉(Malbranchea cinnamomea) CAU521利用农业废弃物固体发酵产木聚糖酶的发酵条件.[方法]采用单因素试验法优化影响菌株产酶的各个条件,包括碳源种类、氮源种类、初始pH、初始水分含量、培养温度及发酵时间共6个因素.[结果]获得的最佳产酶条件为:稻草为发酵碳源、2％(W/W)的酵母提取物为氮源、初始pH 7.0、初始水分含量80％和发酵温度45℃.在此条件下发酵6d后木聚糖酶的酶活力达到13 120 U/g干基碳源.[结论]樟绒枝霉固体发酵产木聚糖酶的产酶水平高,生产成本低,具有潜在的工业化应用前景.     

黑曲霉固态发酵生产单宁酶的条件优化

研究采用响应面法优化黑曲霉固态发酵生产单宁酶的培养条件。应用Plackett—Burman试验筛选出重要影响因子：五倍子粉含量、（NH4）2SO4浓度以及接种孢子量,最陡爬坡试验逼近最大响应区域。应用Box．Behnken响应面试验对重要影响因子进一步优化。得到最佳培养条件：每250mL三角瓶中装入1．0g五倍子粉、4．4g稻壳和0．5g麸皮、液固比（mL／g）2：1且营养盐溶液组成为（NH4）2s0421g/L、MgSO4·7H2O1g／L、NaCl1g／L,培养基pH自然,接种5．7×10^7个孢子后在30℃温度下培养4d。在此条件下,单宁酶产量从40U／g提高到114U／g,3次重复验证性试验平均值为115U／g,验证了模型的可靠性。