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Broccoli extract mainly contains polyphenols and glucosinolates (GSLs). GSLs can be hydrolyzed by gut microorganisms into isothiocyanates (ITCs) and other active substances. These substances have anticancer, antiinflammatory, antimicrobial, and atherosclerosis-reducing functions. In this study, a high concentration (2000 μmol/L GSLs and 24 μmol/L polyphenols) and a low concentration (83 μmol/L GSLs and 1 μmol/L polyphenols) of broccoli extract were prepared. Gut microorganisms from fresh human feces were cultured to simulate the gut environment in vitro. The GSL content decreased and the types and content of ITCs increased with broccoli extract hydrolysis through cyclic condensation and gas chromatography−mass spectrometry (GC-MS) analyses. Broccoli extract significantly increased probiotics and inhibited harmful bacteria through 16S rDNA sequencing. Based on phylum level analysis, Firmicutes and Lachnospiraceae increased significantly (P < 0.05). At the genus level, both high-and low-concentration groups significantly inhibited Escherichia and increased Bilophila and Alistipes (P < 0.05). The high-concentration group significantly increased Bif idobacterium (P < 0.05). The broccoli extract improved the richness of gut microorganisms and regulated their structure. The GSL hydrolysis was significantly correlated with Bilophila, Lachnospiraceae, Alistipes, Bif idobacterium, Escherichia, and Streptococcus (P < 0.05). These study findings provide a theoretical foundation for further exploring a probiotic mechanism of broccoli extract in the intestine.

The milk-clotting enzyme (MCE) is a crucial ingredient in cheese manufacture. Due to the limits of traditional MCE, finding viable substitute is a pressing issue. This study aims to isolate and identify a wild strain with high milk-clotting activity (MCA) and low proteolytic activity (PA) and optimize the fermentation conditions for MCE production. A strain of Bacillus velezensis DB219 with high MCA/PA value (9.2) was isolated from dairy soil (Wuchang, Heilongjiang, China) and identified through 16S rRNA from 40 strains. The optimal wheat bran, carbon, nitrogen, inoculum size, volume and initial pH were 60 g/L, soluble starch 12.5 g/L, corn steep liquor 3 g/L, 5%, 40 mL and 6.15, respectively for improving DB219 MCE production through single factor experiment. The wheat bran concentration, corn steep liquor concentration and volume were the most critical factor and their changed range was determined through Plackett–Burman design and the steepest ascent/descent experiments. The response surface analysis experiment of three factors and three levels was conducted by Box–Behnken design. The theoretical optimal fermentation conditions for DB219 MCE were as follows: wheat bran concentration 60.14 g/L, soluble starch 12.5 g/L, corn steep liquor 3 g/L, inoculum size 5%, volume 40.08 mL and initial pH 6.15. DB219 MCE achieved the maximal MCA (3164.84 SU/mL) that was 101.9% of the predicted value (3104.49 SU/mL) and 4.3-fold higher than the control.