Synergistic Effect of Multiple Saccharifying Enzymes on Alcoholic Fermentation for Chinese Baijiu Production

Chinese Jiuqu (fermentation starter) provides saccharifying enzymes for baijiu (Chinese liquor) fermentation, which undergoes a simultaneous saccharification and fermentation process. However, the key saccharifying enzymes associated with alcoholic fermentation from Jiuqu and their effects on ethanol production remain poorly understood. In this study, we identified 51 carbohydrate hydrolases in baijiu fermentation by metaproteomics analysis. Through source-tracking analysis, approximately 80% of carbohydrate hydrolases in the baijiu fermentation were provided by Jiuqu. Among these enzymes, alpha-amylase (EC 3.2.1.1) and glucoamylase (EC 3.2.1.3), from Aspergillus, Rhizomucor, and Rhizopus, were positively related to starch hydrolysis and ethanol production, indicating that they were the key saccharifying enzymes associated with alcoholic fermentation in the baijiu fermentation. Moreover, a combined mixture of alpha-amylase and glucoamylase (in a ratio of 1:6, wt/wt) enhanced ethanol production in a simulative baijiu fermentation under laboratory conditions. This result revealed a synergistic effect of multiple saccharifying enzymes on ethanol production in baijiu fermentation. Our study provides a potential approach to improve the efficiency of saccharification and alcoholic fermentation by optimizing the profile of saccharifying enzymes for fermentation of baijiu and other beverages. IMPORTANCE Jiuqu starter provides enzymes to the simultaneous saccharification and fermentation process of baijiu (Chinese liquor) production; however, the key saccharifying enzymes associated with alcoholic fermentation from Jiuqu and their effects on ethanol production remain unclear. We confirmed that Jiuqu was the main source of carbohydrate hydrolases for baijiu fermentation and identified two types of saccharifying enzymes from multiple microbes as the key enzymes associated with alcoholic fermentation. Moreover, a proper combination of multiple saccharifying enzymes could enhance ethanol production in baijiu fermentation. This combination provides an approach to optimize the profile of saccharifying enzymes for enhancing ethanol production in baijiu and other food fermentations.