Optimization of Laccase Production from Bacillus sp. PK4 through Statistical Design of Experiments

Statistical design of experiments was employed to optimize the media composition for the production of laccase from Bacillus sp. PK4. In order to find the key ingredients for the best yield of enzyme production from the selected eleven variables viz yeast extract, glucose, zinc sulphate, copper sulphate, potassium chloride, magnesium sulphate, calcium chloride, ferrous sulphate, sodium chloride, potassium dihydrogen phosphate (KH2PO4) and dipotassium hydrogen phosphate (K2HPO4), Plackett-Burman design was applied. The MgSO4, FeSO4, and CuSO4 showed positive estimate, and their concentration optimized further. The steepest ascent method and Box-Behnken method revealed that 1.5 mM MgSO4, 0.33 g/l FeSO4 and 1.41 mM CuSO4 were optimal for the laccase production by Bacillus sp. PK4. This optimization strategy leads to enhancement of laccase production from 2.13 U/ml to 40.79 U/ml. Agro-wastes residues replace the carbon source glucose in the optimized media namely sugarcane bagasse, wheat bran, rice husk, and groundnut shell, among these groundnut shells (117 U/ml) was found to enhance the laccase production significantly. The laccase produced by Bacillus sp. PK4 was found to have the potential to degrade persistent organic pollutant benzo[a]pyrene.