The variation of two extracellular enzymes and soybean meal bitterness during solid-state fermentation of Bacillus subtilis

The debittering effect of extracellular enzymes from Bacillus subtilis ACCC 01746 was studied using soybean meal as a substrate for solid-state fermentation (SSF). Results showed that B. subtilis produces proteases and carboxypeptidase in the early stage of SSF (0–8 h). Proteases are dominant and can hydrolyze the soybean protein into long-chain peptides with mild bitterness. Carboxypeptidase production is dominant at 8–16 h SSF, at which point soybean protein is further hydrolyzed and bitterness is enhanced. The strain then produces additional carboxypeptidase after 16 h, and bitterness is reduced. We compared the amino acid composition of the hydrolysates from soybean protein isolates to that of the fermented liquid of SSF. In the hydrolysates from soybean protein isolates that exhibit strong bitterness, 62.81% of amino acids are hydrophobic and occur in the form of peptides. In the fermented liquid from soybean meal, 16.22% of amino acids are hydrophobic and are mainly present in the form of free amino acids. The bitterness of fermented soybean hydrolysate is reduced from 5 to 0 when fermented for 24 h, suggesting that B. subtilis can effectively reduce bitterness, possibly due to the carboxypeptidase. Enzyme analysis shows that B. subtilis excretes carboxypeptidase during growth. The amino acids phenylalanine, alanine, tyrosine, and leucine at the C-terminal of the soybean bitter peptides in hydrolysates are cleaved in the presence of carboxypeptidase, resulting in complete debitterness.