Production optimization and characterization of amylase from Bacillus subtilis strain 2i under submerged fermentation

Amylase holds significant importance in industrial biotechnology. In this study, amylase production by Bacillus subtilis strain 2i was statistically optimized using mango kernels as substrate in submerged fermentation. Nine nutritional variables (X1-X9) in the amylase production medium were screened through the Plackett-Burman design (PBD), out of which three variables were identified as significant. These include (NH₄)₂SO₄, MgSO₄ and FeSO₄. Further optimization of these variables by response surface methodology (RSM) through central composite design (CCD) was done for maximum amylase production. Characterization revealed that amylase displayed maximum activity at 70 °C and pH 8. Furthermore, the effect of metal ions, organic solvents, surfactants, and detergents was also evaluated. Enhanced activities were detected in the presence of Ca⁺⁺ and Mn⁺⁺ ions. Among solvents, methanol and acetone enhanced amylase activity. SDS displayed inhibitory effects. Amylase showed activation energy (Ea) of 13.7 kJ/mol, enthalpy change (∆H) of 10.63 kJ/mol, and entropy (∆S) of 0.079 kJ.mol^-1.K^-1. The enzyme remained stable in presence of most of surfactants and detergents which concludes that this enzyme can be incorporated in commercial detergent formulations and other industrial applications.