Application of statistically-based experimental designs in medium optimization for spore production of <i>Bacillus subtilis</i> from distillery effluent

Spore production of Bacillus subtilis from distillery effluent was optimized using statistically-based experimental designs. The two-level Plackett–Burman design was applied to choose the nutrient supplements significantly influencing spore production. Among the seven variables we tested, the most significant variables influencing spore production were statistically elucidated for optimization, and included (NH 4 ) 2 SO 4 , corn flour and MgSO 4 . The optimum concentration of each significant variable was then predicted using Box–Behnken design. A second-order polynomial was determined by the multiple regression analysis of this experimental data. The optimum values for the critical nutrient supplements for the maximum were obtained as followed: (NH 4 ) 2 SO 4 , 4.54%; corn flour, 1.2%; MgSO 4 , 0.56% with the corresponding value of maximum spore production of 7.24 × 10 8 spores/ml. A verification experiment performed under the optimum conditions resulted in 6.95 × 10 8 spores/ml. The determination coefficient ( R 2 ) was 0.98, which ensure an adequate credibility of the model.