Production and characterization of cellulase from mushroom (Pleurotus ostreatus) for effective degradation of cellulose

Abstract

Cellulases are a group of hydrolytic enzymes capable of hydrolyzing the most abundant organic polymer that means cellulose to smaller sugar components including glucose subunits. The aim of this study was to screen cellulase producing oyster mushroom collected from Eucalyptus tree bark to evaluate the in vitro production of cellulase by Pleurotus ostreatus using different lignocellulosic substrates, and to characterize the cellulase produced with respect to changes in pH, temperature, and concentration of substrates. A total of ten mushroom specimens were randomly collected from Eucalyptus tree bark in the premise of Holetta Agricultural Research Center campus. All of the collected mushroom specimens were identified morphologically and biochemically as Pleurotus ostreatus and also screened for their ability to produce cellulase by detecting and measuring zone of hydrolysis on commercial media containing Carbxymethyl Cellulose (CMC) as the sole carbon source. These mushroom specimens were cultivated using both solid state fermentation and submerged fermentation systems supplemented with different lignocellulosic substrates (wheat straw, teff straw, bean straw, wood fiber and Eucalyptus tree bark) to identify the most suitable medium for the production of cellulase. The highest enzyme production was obtained on bean straw and wheat straw which resulted in 0.191 U/ml, 0.868 U/ml and 0.389 U/ml; and 0.216 U/ml, 0.444 U/ml, and 0.245 U/ml of FPase, CMCase, and β-glucosidase in solid state fermentation. The lowest values were, however, obtained in media containing wood fiber in both solid state fermentation and submerged fermentation. Comparison of the lignocellulosic substrates revealed that wheat straw was selected for further growth parameter optimization. The production of cellulase was higher at the 5th day of incubation period, and the optimum pH and incubation temperature required for maximum cellulase production were 4 and 30°C, respectively. Sucrose and Yeast extract at 1% concentration were found to be the most preferred carbon and nitrogen sources for cellulase production by Pleurotus ostreatus. The optimum pH and temperature for cell_free cellulase activity on were found to be 4 and 50°C, respectively. Generally the cellulases produced by Pleurotus ostreatus were stable and active at temperatures ranging from 20-50°C. These characteristics hopefully would make this enzyme potentially attractive in a variety of industrial applications including animal feed treatments. There was a linear relationship between cellulase and its substrate concentration for there was an increase in activity with increase in substrate concentration. The relationship between rate of reaction and substrate concentration depended on the affinity of the enzyme for its substrate. Finally the cellulase was tested for its ability to saccharify agricultural wastes and the results showed the highest release of sugars from wheat straw.