Enhanced Cellulase Production from Bacillus subtilis by Optimizing Physical Parameters for Bioethanol Production

Effect of physical parameters such as initial pH, agitation (rpm), and temperature (°C) for cellulase production from Bacillus subtilis AS3 was investigated. Central composite design of experiments followed by multiple desirability function was applied for the optimization of cellulase activity and cell growth. The effect of the temperature and agitation was found to be significant among the three independent variables. The optimum levels of initial pH, temperature, and agitation for alkaline carboxymethylcellulase (CMCase) production predicted by the model were 7.2, 39°C, and 121 rpm, respectively. The CMCase activity with unoptimized physical parameters and previously optimized medium composition was 0.43 U/mL. The maximum activity (0.56 U/mL) and cell growth (2.01 mg/mL) predicted by the model were in consensus with values (0.57 U/mL, 2.1 mg/mL) obtained using optimized medium and optimal values of physical parameters. After optimization, 33% enhancement in CMCase activity (0.57 U/mL) was recorded. On scale-up of cellulase production process in bioreactor with all the optimized conditions, an activity of 0.75 U/mL was achieved. Consequently, the bacterial cellulase employed for bioethanol production expending (5%, w/v) NaOH-pretreated wild grass with Zymomonas mobilis yielded an utmost ethanol titre of 7.56 g/L and 11.65 g/L at shake flask and bioreactor level, respectively.