PRODUCTION OF AMYLASE, CELLULASE AND PROTEASE BY SRAW MUSHROOM (VOLVARIELLA VOLVACEA) USING SELECTED' AGRO-INDUSTRIAL WASTES

Abstract

Fresh fruiting body of Volvariella volvacea was collected and identified using standard method. Active mycelium was produced by tissues culture techniques on potato dextrose agar. Proximate and mineral compositions of selected agro-industrial residues before and after use in fermentation were determined. Proximate analysis of the residues used revealed that the percentage protein and moisture contents increased while the fat, crude fibre, ash and carbohydrate contents decreased with fermentation time. Analysis of. the mineral contents of the agroindusrial wastes revealed a reduction with fermentation time. Volvariefla volvacea produced extracellular cellulase, amylases and proteases, when wheat bran, rice bran, palmkernel pcricarp and sawdust (white alfra) were used as carbon sources under submerged fermentation. The amount of these enzymes produced and the time required for the maximum production of enzymes differed with each substrate.Crude protein content and enzyme activity were determined at different period of incubation in culture filtrate (24hrs, 48hrs, 72hrs and 96hrs).p-amylase was selected for further studies because of its high activity enzyme. Wheat bran supported the highest beta amylase activity of 240 umol/min/ml at 72hrs fermentation.The enzyme was purified by ion-exchange chromatography (DEAE-Sephadex-A50) and gel filtration (Sephadex GJ 50). The molecular weight of p-amylase was estimated to be approximately 69kDa. The activity of the enzyme was affected by the pH and temperature of the reaction mixture. The optimum pH and temperatureforB-arnylase activity were 5.0 and 50°C respectively .. The apparent Km for the hydrolysis of soluble starch from the Lineweaver-Bulk plot was approximately 1.39mg/ml while the Ymax was 0.365 urnol/rnin/ml. In the presence of salts such as CUS04, NaCI, HgCI and metal chelator EDTA, the activity of the enzyme was inhibited at 0.04M. However, higher degree of inhibition was observed with HgCI and EDTA. The low Krn values and improved activities of enzymes from the mushroom, Volvariella volvacea makes them a more favoured alternative to enzymes from other microorganisms for industrial scale applications.