Abstract | In this study, cellulase enzyme production was assessed by pretreating cotton stalk with dilute sulphuric acid which was optimized through response surface methodology. Pretreatment conditions were optimized using three variable with three levels like sulfuric acid concentrations (0.6%, 0.8%, 1% v/v), biomass loading (5%, 10%, 15% w/v), and residence time (4, 6 and 8h). After pretreatment process, cellulase production was achieved by solid substrate which was conducted in 250mL capacity Erlenmeyer flask in which incubation of Bacillus subtilis was carried out for 24 h of fermentation period at temperature of 50oC. Results showed that cellulase production was greatly affected by the thermochemical pretreatment as compared to the chemical pretreatment. At the pretreatment condition of 1% H2SO4 conc, 6h residence time, 15% substrate concentration maximum CMCase production (0.858 IU/ml/min) was obtained while at pretreatment conditions of 1% H2SO4 concentration, 8h residence time, 10% substrate concentration at room temperature followed by routine autoclaving, FPase production up to 0.876 IU/ml/min was recorded. The results of present study were found significant. The cellulase enzyme produced in this process affectively hydrolyzes the pretreated substrate at temperature of 50oC and 53 h of incubation period by releasing reducing sugars of 0.74 mg/ml. This study ensures the effective usage of lignocellulosic biomass at large scale biofuel production.
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