Abstract

Three strains, Bacillus subtilis B1, B. subtilis B18 and Bacillus thuringiensis B12,were screened from wheat bran to produce milk-clotting enzyme. Among them, B. subtilis B1 exhibited considerable milk-clotting activity with low proteolytic activity. After response surface methodology optimization, milk-clotting activity wasimproved from 782 SU ml^-1 to 1129.05 ± 74.55 SU mL^-1 with a small inoculum size (0.130%, v/v). The optimized medium contained glucose (16.2 gL^-1), wheat bran (30 gL^-1), NaCl (5 gL^-1), MgSO₄·7H₂O (5 gL^-1), KH₂PO₄ (2 gL^-1) and CaCO₃ (3 gL^-1). The milk-clotting enzyme from B. subtilis B1 has the optimum pH 5.5 and CaCl₂concentration 50 mmol L^-1. It is completely inactivated after 5 min at 70°C. The result showed that B. subtilis B1 was a promising strain for industrial milk-clotting enzyme production.

Key words: Milk-clotting enzyme, Bacillus subtilis, submerged fermentation, wheat bran.

Abbreviation

MCA, Milk-clotting activity; MCE, milk-clotting enzyme; PA,proteolytic activity; SU, soxhlet unite.