Statistical Optimization of Medium Components for Milk-Clotting Enzyme Production by Bacillus amyloliquefaciens D4 Using Wheat Bran-an Agro-Industry Waste

Abstract

In this paper, two statistical methods were applied to optimize medium components to improve the production of the milk-clotting enzyme by Bacillus amyloliquefaciens D4. First, wheat bran juice, skim milk powder, and Na2HPO4 were shown to have significant effects on D4 enzyme production using the Plackett-Burman experimental design. Subsequently, an optimal medium was obtained using the Box-Behnken method, which consisted of 3.31 g/l of skim milk powder, 5.0 g/l of sucrose, 0.1 g/l of FeSO4·7H2O, 0.1 g/l of MgSO4·7H2O, 0.1 g/l of MnSO4·2H2O, 0.1 g/l of ZnSO4·7H2O, 1.52 g/l of Na2HPO4, and 172.45 g/l of wheat bran juice. With this optimal medium, the milk-clotting enzyme production was remarkably enhanced. The milk-clotting enzyme activity reached 3,326.7 SU/ml after incubation of 48 h, which was 1.76-fold higher than that of the basic medium, showing that the Plackett-Burman design and Box-Behnken response surface method are effective to optimize medium components, and B. amyloliquefaciens D4 possessed a high rennet-producing capacity in the optimal medium.

Newly Effective Milk-Clotting Enzyme from Bacillus subtilis and Its Application in Cheese Making

“…Although these MCEs could constitute the most common needs, the search for more efficient, lower cost MCEs is continuous . Besides, the MCEs from Bacillus attracted people’s interests ,, because it had a comparable ratio of milk-clotting activity to proteolytic activity with commercial calf Rennet but a high thermos-stability. It could produce less bitterness in the cheese ripening process .…”

The catalytic and kinetic characterization of Bacillus subtilis MK775302 milk clotting enzyme: comparison with calf rennet as a coagulant in white soft cheese manufacture

“…Most plant coagulants are unsuitable for cheese making due to the low yield of cheese, the bitter taste, and defects in the flavor of the produced cheese [ 8 ]. Milk clotting enzymes from microbial sources have gained great attention due to their various properties, ease of preparation, and reduced cost of production [ 9 ]. Bacterial milk clotting enzymes are more preferable compared to fungal enzymes due to lower production costs, greater biochemical diversity, faster cultivation, and easier genetic modification [ 10 ].…”