Screening of high yield tetramethylpyrazine Bacillus from high temperature Daqu and its application in the stacking of fermented grains

In this study, Bacillus spp were screened from high temperature Daqu. The strains were preliminarily screened through high temperature resistance test and protease activity determination, rescreened by V-P test and tetramethylpyrazine content analysis. The screened strains identified through morphological observation, physiological and biochemical tests and molecular biology techniques. The volatile flavor substances in the fermentation liquid of the strains were analyzed by GC-MS, and the strains were applied to the stacking of fermented grains. The results showed that a total of 18 proteinase- producing strains and 1 high yield tetramethylpyrazine strain D-13-11 were screened, and the high yield tetramethylpyrazine strain D-13-11 was identified as Bacillus velezensis with its content as high as 339.99 mg/L. The GC-MS results showed that a total of 42 substances were detected, including pyrazines 4, esters 15, alcohols 4, aldehydes and ketones 4, acids 6, phenols 2, alkanes and siloxanes 7. The relative content of pyrazine compounds was the highest, reaching 40.14%, and the tetramethylpyrazine was the main compound (24.84%). The strain D-13-11 was inoculated into the stacking of fermented grains with inoculum 7%, and the relative content of pyrazine compounds was significantly higher than that of the control group. Among them, the content of tetramethylpyrazine was 11.38 μg/g, which was 71.13 times that of the control group.