Utilization of pretreated coir pith for the optimized bioproduction of cellulase by Aspergillus nidulans

The present study is aimed at simultaneous cellulase synthesis and coir pith degradation by Aspergillus nidulans using coir pith as chief substrate. The lignocellulosic biomass, coir pith is known to be an excellent carbon source for microbial cellulase production under solid state fermentation. The alkali pretreatment with sodium hydroxide was seen to enhance enzymatic hydrolysis. The effect of coir pith weight, moisture content, initial pH and growth temperature on cellulase activity and yield were investigated by response surface methodology (RSM) employing a four-factor-five-level central composite design (CCD). The results of Fourier transform infrared spectroscopy (FTIR), X-Ray diffraction (XRD) and Scanning electron microscopy (SEM) of coir pith showed structural changes through pretreatment, in favor of enzymatic hydrolysis. Maximum carboxy methyl cellulase activity (CMCase) of 28.64 U/g and cellulase yield of 66.32% were achieved with 8 g coir pith at 70% moisture content and 40 °C temperature with pH 5 as evident from run numbers 25 and 30. Filter paper (FPase) and cellobiase (CBase) activities of 10.23 U/g and 4.31 U/g respectively were observed on the 11th day after the inoculation.     

Mechanistic studies on the binding of Acid Yellow 99 on coir pith

The interaction of Acid Yellow 99 (AY 99) with coir pith has been investigated in aqueous medium to understand the mechanism of adsorption and explore the potentiality of this biomass towards controlling pollution resulting from textile dyes. The obtained results establish that one gram of coir pith can adsorb 442.13 mg of AY 99. The adsorption process is found to be a function of pH of the solution, the optimum pH value being 2.0. The process follows Langmuir-Freundlich dual isotherm model. Scanning electron microscopic analysis demonstrates that on dye adsorption the biomass develops uneven and irregular surface. X-ray diffraction study indicates incorporation of the dye into the micropores and macropores of the adsorbent and thereby enhancing its degree of crystallinity. The results of Fourier transform infrared (FTIR) spectroscopy and chemical modification of the functional groups establish that binding of AY 99 on coir pith occurs through electrostatic and complexation reaction.     

Recycling spent larval food of Corcyra cephalonica Stainton for preparing spawn and sporophore of Pleurotus sajor-caju (Fr.) Singer

Large scale production of the rice moth, Corcyra cephalonica Stainton in pearl millet grain medium leads to a huge accumulation of spent larval medium in commercial insectaries. We attempted bioconversion of spent larval medium of C. cephalonica (CLM) for cultivation of the mushroom Pleurotus sajor-caju (Fr.) Singer, to increase the usage of these residues. Maximum efficiency limits of CLM for spawn run, sporophore cropping and as bed substrate were assessed with varying combinations of sorghum and rice straw. Sorghum grains and rice straw were the best substrates for spawn run and sporophore yield respectively. Having been crushed, macerated, heated and sterilized, CLM could also become a suitable substrate along with sorghum or rice straw. Sorghum and CLM at 16.7% + 83.3% and 33.3% + 66.7% combinations were very effective in supporting mycelial growth and quicker colonization of fungus, and mother spawn yield. The spawn that was obtained from these combinations yielded higher sporophore as well. The fungus did not rapidly colonize on other combinations (50% + 50%, 66.7% + 33.3% and 83.3% + 16.7%), and was completely unable to grow on CLM 100%. Combination of rice straw and CLM at 75% + 25% and 50% + 50% as bed substrate contributed higher sporophore yield. Analysis of the substrates indicated variation in their chemical and mineral composition, but they were good sources of N, P and Ca. The prospects of exploring CLM for the mushroom cultivation are discussed.     

Application of solid waste from anaerobic digestion of poultry litter in Agrocybe aegerita cultivation: mushroom production, lignocellulolytic enzymes activity and substrate utilization

The degradation and utilization of solid waste (SW) from anaerobic digestion of poultry litter by Agrocybe aegerita was evaluated through mushroom production, loss of organic matter (LOM), lignocellulolytic enzymes activity, lignocellulose degradation and mushroom nutrients content. Among the substrate combinations (SCs) tested, substrates composed of 10–20% SW, 70–80% wheat straw and 10% millet was found to produce the highest mushroom yield (770.5 and 642.9 g per 1.5 kg of substrate). LOM in all SCs tested varied between 8.8 and 48.2%. A. aegerita appears to degrade macromolecule components (0.6–21.8% lignin, 33.1–55.2% cellulose and 14–53.9% hemicellulose) during cultivation on the different SCs. Among the seven extracellular enzymes monitored, laccase, peroxidase and CMCase activities were higher before fruiting; while xylanase showed higher activities after fruiting. A source of carbohydrates (e.g., millet) in the substrate is needed in order to obtain yield and biological efficiency comparable to other commercially cultivated exotic mushrooms.     

Degradation dynamics of lignocellulose materials by wood-rottingPleurotus fungi

When grown on wheat straw,Pleurotus decomposes both the lignin and the cellulose components of the substrate. The course of degradation differs during growth and fructification. The losses of dry mass during growth were about 20 %. The absolute amount of hemicelluloses, cellulose and lignin was decreasing. Hemicelluloses and lignin were degraded at a higher rate than cellulose. The total mass losses of the substrate after fructification were 32 to 45 %. Cellulose was consumed at a higher rate than lignin.     

Oyster mushroom cultivation with rice and wheat straw

Cultivation of the oyster mushroom, Pleurotus sajor-caju, on rice and wheat straw without nutrient supplementation was investigated. The effects of straw size reduction method and particle size, spawn inoculation level, and type of substrate (rice straw versus wheat straw) on mushroom yield, biological efficiency, bioconversion efficiency, and substrate degradation were determined. Two size reduction methods, grinding and chopping, were compared. The ground straw yielded higher mushroom growth rate and yield than the chopped straw. The growth cycles of mushrooms with the ground substrate were five days shorter than with the chopped straw for a similar particle size. However, it was found that when the straw was ground into particles that were too small, the mushroom yield decreased. With the three spawn levels tested (12%, 16% and 18%), the 12% level resulted in significantly lower mushroom yield than the other two levels. Comparing rice straw with wheat straw, rice straw yielded about 10% more mushrooms than wheat straw under the same cultivation conditions. The dry matter loss of the substrate after mushroom growth varied from 30.1% to 44.3%. The straw fiber remaining after fungal utilization was not as degradable as the original straw fiber, indicating that the fungal fermentation did not improve the feed value of the straw.     

Experimental and kinetic studies on methylene blue adsorption by coir pith carbon

Varying the parameters such as agitation time, dye concentration, adsorbent dose, pH and temperature carried out the potential feasibility of thermally activated coir pith carbon prepared from coconut husk for removal of methylene blue. Greater percentage of dye was removed with decrease in the initial concentration of dye and increase in amount of adsorbent used. Kinetic study showed that the adsorption of dye on coir pith carbon was a gradual process. Lagergren first-order, second-order, intra particle diffusion model and Bangham were used to fit the experimental data. Equilibrium isotherms were analysed by Langmuir, Freundlich, Dubnin-Radushkevich, and Tempkin isotherm. The adsorption capacity was found to be 5.87 mg/g by Langmuir isotherm for the particle size 250-500 microm. The equilibrium time was found to be 30 and 60 min for 10 and 20 mg/L and 100 min for 30, 40 mg/L dye concentrations, respectively. A maximum removal of 97% was obtained at natural pH 6.9 for an adsorbent dose of 100 mg/50 mL and 100% removal was obtained for an adsorbent dose of 600 mg/50 mL of 10 mg/L dye concentration. The pH effect and desorption studies suggest that chemisorption might be the major mode of the adsorption process. The change in entropy (DeltaS0) and heat of adsorption (DeltaH0) of coir pith carbon was estimated as 117.20 J/mol/K and 30.88 kJ/mol, respectively. The high negative value of change in Gibbs free energy indicates the feasible and spontaneous adsorption of methylene blue on coir pith carbon.     

Profile of the extracellular lignocellulolytic enzymes activities as a tool to select the promising strains of Volvariella volvacea (Bull. ex Fr.) sing

Out of 26 strains of Volvariella volvacea used, 18 were of ‘typical’ type and possessed all the characteristics of a normal V. volvacea mycelium, while the rest 4 ‘atypical’ type strains showed completely distinct mycelial growth characteristics. The remaining 4 strains grew very slowly and exhibited growth characteristics of single spore isolates of V. volvacea. Strains varied in their extracellular lignocellulolytic activities and strains; OE-274, OE-272 and OE-210 with high ligninase enzymes (laccase and polyphenol oxidase) activities, gave highest mushroom yield on pasteurized paddy straw substrate. On the composted paddy straw substrate, additional two strains, OE-213 and OE-215 with lower activities of ligninases also gave higher mushroom yield. Mushrooms were harvested 3 to 4 d early from the composted substrate than on the pasteurized substrate. Activities of endoglucanase, laccase and polyphenol oxidase were found to be more crucial for mushroom yield on pasteurized substrate, while xylanase and β-glucosidase were more important for composted substrate. Strains also varied in their fruiting bodies quality and the substrate used for mushroom cultivation also affected the fruiting body quality. The superior yielding strains varied in shape, size, weight, colour and contents of sodium and potassium in their fruiting bodies; while contents of carbon, calcium and protein did not vary much with the strains.     

Effect of nutritional parameters on laccase production by the culinary and medicinal mushroom, Grifola frondosa

Summary Extracellular laccase in cultures of Grifola frondosa grown in liquid culture on a defined medium was first detectable in the early/middle stages of primary growth, and enzyme activity continued to increase even after fungal biomass production had peaked. Laccase production was significantly increased by supplementing cultures with 100–500 (M Cu over the basal level (1.6 mM Cu) and peak levels observed at 300 mM Cu were ∼ ∼7-fold higher than in unsupplemented controls. Decreased laccase activity similar to levels detected in unsupplemented controls, as well as an adverse effect on fungal growth, occurred with further supplementation up to and including 0.9 mM Cu, but higher enzyme titres (2- to 16-fold compared with controls) were induced in cultures supplemented with 1–2 mM Cu²⁺. SDS-PAGE combined with activity staining revealed the presence of a single protein band (M _r ∼ ∼70 kDa) exhibiting laccase activity in control culture fluids, whereas an additional distinct second laccase protein band (M _r␣∼ ∼45 kDa) was observed in cultures supplemented with 1–2 mM Cu. Increased levels of extracellular laccase activity, and both laccase isozymes, were also detected in cultures of G. frondosa supplemented with ferulic, vanillic, veratric and 4-hydroxybenzoic acids, and 4-hydroxybenzaldehyde. The optimal temperature and pH values for laccase activity were 65 °C and pH 2.2 (using 2,2′-azinobis(3-ethylbenzthiazoline-6-sulfonate) {ABTS} as substrate), respectively, and the enzyme was relatively heat stable. In solid-state cultures of G. frondosa grown under conditions adopted for industrial-scale mushroom production, extracellular laccase levels increased during the substrate colonization phase, peaked when the substrate was fully colonized, and then decreased sharply during fruit body development.     

Laccase, cellulase and xylanase activities during growth ofPleurotus sajor-caju on sagohampas

Sagohampas, the fibrous pith residue left after starch extraction from sago palm, is abundant at sago-processing factories and can be used as a substrate for the production of laccase by solid substrate fermentation (SSF) withPleurotus sajorcaju, an edible mushroom. The fungus grown onhampas with an adjusted carbon : nitrogen ratio of 35:1, exhibited high laccase activity together with variable cellulase (0.3-2.8 U/g) and xylanase (0.9-10.1 U/g) activity. The maximum amount of laccase produced was approximately 17.7 U/g after 6 days of SSF using 4-week-old inoculum at a density of 10%. With the mature four-week inoculum, laccase activity increased 12-fold compared to that achieved with two-week-old inoculum. The optimum pH and temperature of the crude laccase were 6.0 and 50‡C, respectively. The apparent K_m and V_max values obtained were 0.073 mM and 0.962 U/min, respectively. The maximum laccase activity could be almost doubled after 6 days of fermentation by addition of 0.2 mM vanillin or ferulic acid; the cellulose to lignin ratio increased significantly during the 12 days of SSF, from 2.74 in the control to 3.3, when 0.2 mM of either vanillin or ferulic acid was added to the substrate.     

Evaluation of lignocellulosic biomass from coconut palm as substrate for cultivation of Pleurotus sajor-caju (Fr.) Singer

The lignocellulosic biomass from coconut palm (Cocos nucifera Linn.) such as bunch waste (spathe+spadices), leafstalk (petiole), leaflets and coir pith (by-product from coir processing industry) were evaluated as substrates for cultivation of oyster mushroom, Pleurotus sajor-caju (Fr.) Singer. A low-cost mushroom shed built exclusively of coconut materials such as coconut wood and plaited coconut leaves inside a coconut plantation was used for spawn run and cropping. Leafstalk and bunch waste were superior to leaflets and coir pith in producing significantly more edible biomass of mushrooms. Biological efficiency of 58.9% was obtained in leafstalk, followed by bunch waste (56.9%), coir pith (39.7%) and leaflets (38.2%). The yield of sporophore was positively related to cellulose content and the cellulose : lignin ratio of the substrates.     

Effect of Rice Straw Incorporation on Phytotoxicity of Isoxaflutole to an Exotic Weed Phalaris minor Retz.

Phalaris minorRetz. is a major exotic annual weed in the wheat (Triticum aestivum L.) crop. Unharvested rice (Oryza sativa L.) straw, unburned and burned, is often incorporated in the field prior to cultivating wheat. Isoxaflutole (Balance), a pre-emergent systemic soil applied herbicide, has potential to control P. minor. Glasshouse experiments were conducted to determine the phytotoxicity of isoxaflutole defined by reductions in relation to shoot length of P. minor when grown in unamended soil or soil amended with unburned or burned rice straw. A 120 g soil was amended with 0, 1, 2 and 4 g of unburned or burned rice straw, and placed in 150 mL styrofoam pots. Appropriate amount of isoxaflutole (75% active ingredient, ai) was added to pots to get final concentration of 0, 7.5, 30, 60 and 120 μg ai/pot. Unamended soil and soil amended with unburned or burned rice straw were analyzed for pH and organic matter; two important determinants of isoxaflutole activity. Results indicate a significant reduction in shoot length of P. minor when grown in soil treated with isoxaflutole at 30, 60 or 120 μg ai/pot. Inhibition in the shoot length of P. minor was observed when soil amended with unburned straw was treated with isoxaflutole at 7.5 and 30 μg ai/pot compared with unamended soil treated with similar amounts of isoxaflutole. No significant change in isoxaflutole toxicity was observed when soil amended with unburned straw was treated with isoxaflutole at 60 and 120 lg ai/pot compared with unamended soil treated with similar amounts of isoxaflutole. Isoxaflutole phytotoxicity to P. minor shoot length was eliminated when soil amended with burned straw was treated with isoxaflutole at 7.5 and 30 μg ai/pot. P. minor shoot length was greater when soil amended with burned straw was treated with isoxaflutole at 60 and 120 μg ai/pot relative to herbicide-treated unamended soils. We conclude that incorporation of burned rice straw greatly reduces the phytotoxicity of isoxaflutole toP. minor.     

Cultivation of oyster mushrooms on wheat straw and bagasse substrate amended with distillery effluent

Summary Molasses-based distilleries produce large quantities of dark coloured effluent, which is a major cause of environmental pollution. An experiment was conducted to investigate the efficacy of distillery effluent amendment for edible mushroom production. Three species of oyster mushroom, namely Pleurotus florida Eger (EM 1303), Pleurotus pulmonarius (Fries) Quelet (EM 1302) and Pleurotus sajor-caju (Fries) Singer (EM 1304) were grown on wheat straw (variety UP 2338) and bagasse amended with post-anaerobic distillery effluent, a high organic load wastewater with high biochemical oxygen demand and chemical oxygen demand. Three different levels of effluent treatment were applied to bagasse and wheat straw. Wheat straw was found to be the preferred substrate and showed better results than bagasse in all treatments with respect to yield, biological efficiency (BE) and dry matter loss. P. florida (EM 1303) and P. pulmonarius (EM 1302) gave significantly enhanced yield with increasing levels of effluent, with BE reaching highest at 238.6% for P. florida (EM 1303). Using bagasse as a substrate, P. sajor-caju (EM 1304) and P.␣pulmonarius (EM 1302) exhibited a decreasing trend as compared to control. However, the effect of effluent concentrations did not influence yield and BE significantly in case of bagasse. The dry matter loss of the substrate varied from 9.4% to 53.4% in wheat straw and 17.5% to 45.2% in bagasse respectively.     

Effect of culture parameters on the production of the edible mushroom <Emphasis Type="Italic">Grifola frondosa</Emphasis> (maitake) in tropical weathers

Hitherto, little effort has been directed to improve culture conditions for commercial development of maitake (Grifola frondosa), an edible and medicinal fungus, due to the short history of cultivation, particularly in tropical weathers. The purpose of this research was analyzing the environmental factors required for successful basidiome development on synthetic substrates in Colombia. We evaluated different cereal grains (corn, barley, sorghum and rice) for spawn production; and industrial by-products (such as coffee spent-ground and oak-sawdust) as substrates for mushroom production. Exploiting these residues for G. frondosa solid culturing would primarily provide edible mushroom and simultaneously help in resolving their disposal problem. The use of corn grains as substrate for spawn production results an important factor for reducing crop cycle time. A cold shock to 10°C was requisite for basidiome formation. Coffee spent-ground was a good substrate for mycelial growth, but not for mushroom production. When using oak sawdust plus corn bran as substrate, we obtained consistent yields with combined high biological efficiency (BE) (35.3%), best quality mushrooms, and a crop cycle of 12–14 weeks. The results achieved in this investigation contribute to expand the knowledge on this fungus, and compare favorably with previous works in the northern hemisphere with respect to BE, mushroom quality and crop cycle time.     

Biosoftening of coir fiber using selected microorganisms

Coir fiber belongs to the group of hard structural fibers obtained from coconut husk. As lignin is the main constituent of coir responsible for its stiffness, microbes that selectively remove lignin without loss of appreciable amounts of cellulose are extremely attractive in biosoftening. Five isolated strains were compared with known strains of bacteria and fungi. The raw fiber treated with Pseudomonas putida and Phanerocheate chrysosporium produced better softened fiber at 30±2 °C and neutral pH. FeSO₄ and humic acid were found to be the best inducers for P. chrysosporium and P. putida, respectively, while sucrose and dextrose were the best C-sources for both. Biosoftening of unretted coir fibers was more advantageous than the retted fibers. Unlike the weak chemically softened fiber, microbial treatment produced soft, whiter fibers having better tensile strength and elongation (44.6–44.8%) properties. Scanning electron microscopy photos showed the mycelia penetrating the pores of the fiber, removing the tylose plug and degrading lignin.     

Comparative study on the growth and yield of<Emphasis Type="Italic"> Pleurotus ostreatus</Emphasis> mushroom on different lignocellulosic by-products

Eight lignocellulosic by-products were evaluated as substrates for cultivation of the oyster mushroom, Pleurotus ostreatus (Jacq. ex. fr) Kummer. The yields of mushroom on the different substrates were 183.1, 151.8, 111.5, 87.8, 49.5, 23.3, 13.0 and 0.0 g for composted sawdust of Triplochiton scleroxylon, rice straw, banana leaves, maize stover, corn husk, rice husk, fresh sawdust, and elephant grass, respectively. The biological efficiency (BE) followed the same pattern and ranged from 61.0% for composted sawdust to 0.0% for elephant grass. The yield of mushroom was positively correlated to cellulose (r ² =0.6), lignin (r ² =0.7) and fibre (r ² =0.7) contents of the substrates. Based on the yield and BE of the substrates tested, rice straw appeared to be the best alternate substrate for growing oyster mushrooms. Electronic Publication     

Isolation and screening of potential actinobacteria for rapid composting of rice straw

Rice straw is produced as a by-product from rice cultivation, which is composed largely of lignocellulosic materials amenable to general biodegradation. Lignocellulolytic actinobacteria can be used as a potential agent for rapid composting of bulky rice straw. Twenty-five actinobacteria isolates were isolated from various in situ and in vitro rice straw compost sources. Isolates A2, A4, A7, A9 and A24 were selected through enzymatic degradation of starch, cellulose and lignin followed by the screening for their adaptability on rice straw powder amended media. The best adapted isolate (A7) was identified as Micromonospora carbonacea. It was able to degrade cellulose, hemicelluloses and carbon significantly (P ≤ 0.05) over the control. C/N ratio was reduced to 18.1 from an initial value of 29.3 in 6 weeks of composting thus having the potential to be used in large scale composting of rice straw.     

Effect of carbon,nitrogen sources and inducers on ligninolytic enzyme production by Morchella crassipes

The effect of different carbon, nitrogen sources and inducers on growth and ligninolytic activity by Morel mushroom Morchella crassipes was investigated. The maximum growth was observed in mineral salts broth containing glucose as the carbon source and sodium nitrate as the nitrogen source. Among the inducers, chemical inducers inhibited the growth whereas in natural substrates, growth was not affected much. Manganese peroxidase and lignin peroxidase activity were not detected in the medium with different carbon and nitrogen sources, whereas laccase activity varied depending on carbon source (0.7–3.48 U/ml). Among the inducers, natural inducers resulted in an increase in the enzyme activities. Maximum laccase activity was observed in rice straw (12. 6 U/ml) followed by ABTS (11.6 U/ml); Manganese peroxidase activity was maximum in rice straw (14.32 U/l) wheat straw (12.16 U/l) and phenol red (15 U/l) as the inducers, whereas for Lignin peroxidase activity, rice straw (22 U/l), wheat straw (16 U/l) and veratrylalcohol (20 U/l) served as the best inducers.     

Biodegradation of gossypol by the white oyster mushroom,Pleurotus florida,during culturing on rice straw growth substrate,supplemented with cottonseed powder

The capacity of the white oyster mushroom, Pleurotus florida to biodegrade gossypol was studied, when grown on rice straw supplemented with cottonseed powder. The mushroom fruiting bodies did not contain any residues of gossypol at concentrations of cottonseed powder 0.15–0.60% nitrogen contents of rice straw at the end of mycelial ramification. However, the cottonseed supplementation (at 0.30% N level itself) caused a doubling in the mushroom yield and its protein content, per unit weight straw substrate. The mushroom mycelium when grown on synthetic medium in liquid cultures was able to biodegrade gossypol. A pre-incubation period of 5 days before the addition of gossypol into the culture medium, an inoculum load 10 mg and an incubation period of 10 days at 25 °C caused the biodegradation of 100 g gossypol. Increased concentrations of gossypol required increased duration and increased inoculum levels to effect biodegradation. However, the effect was more pronounced with an increase in inoculum density. The fungal monoculture when grown in rice straw (powder) (5%) + glucose (1%) liquid culture medium, showed an increase in hexosamine content and laccase activity that produced an increased degradation of gossypol over an incubation period from 5 to 25 days. Enzymic extracts of the mycelial monoculture raised on the chopped rice straw substrate when incubated with 100 g of gossypol demonstrated its biodegradability; the increase in enzyme concentration showed enhanced gossypol degradation. This study adds to the world list of organic compounds that Pleurotus is able to biodegrade, and explains the cause of non-yellowing of the white oyster mushroom (P. florida) fruiting bodies, during culture on rice straw with supplementation of cottonseed powder for enhancing the mushroom yields.