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Jiuyao as a generally small saccharifying and spontaneously fermenting starter is indispensable for huangjiu brewing. However, jiuyao currently in use is facing a major challenge of standardized manufacturing because of its open operating environment and rather primitive operations. This study provided a stead manufacturing procedure for jiuyao by controlling the processing parameters. Saccharomycopsis fibuligera and Rhizopus microsporus were revealed as the core functional species related to starch-degrading enzyme activities with single-molecule real-time (SMRT) sequencing. Two strains with high starch-degrading abilities were successfully isolated and identified as Saccharomycopsis fibuligera CY2111 and Rhizopus microsporus SM4. Meanwhile, the environmental fermentation parameters of jiuyao were optimized to enhance enzyme activities. The glucoamylase and α-amylase activities of fortified jiuyao with strain CY2111 and SM4 were 3.07 and 3.15 times that of traditional jiuyao , respectively. Moreover, the alcohol content increased by approximately 7.5% when fortified jiuyao was used for huangjiu brewing. The flavor profiles determined by gas chromatography–mass spectrometry indicated there was no significant difference in the key aromas between the huangjiu fermented with fortified jiuyao and that of traditional jiuyao. Further, fortified jiuyao produced more pleasant esters in huangjiu such as ethyl isovalerate, isoamyl acetate, and ethyl caprylate. Combined with the sensory evaluation, huangjiu fermented with fortified jiuyao presented stronger fruity aroma. These results demonstrated that careful fermentation settings combined with biofortification technology were feasible to improve jiuyao quality, thus promoting huangjiu flavor. Taken together, it provided scientific guidance to improve traditional handcrafting and scale up the production of jiuyao under controllable fermentation. [Display omitted] • The core saccharifying microbiota of huangjiu jiuyao was revealed. • The fermentation environment of jiuyao was simulated and optimized. • The fortified jiuyao was developed for higher enzyme activities. • The fortified jiuyao could improve the flavor quality of huangjiu. [ABSTRACT FROM AUTHOR]

Wheat Qu plays the role of saccharification fermentation, providing microorganisms and flavor in the fermentation of huangjiu , and the use of functional microorganisms to fortify wheat Qu is becoming increasingly popular. Yet, the mechanisms promoting microbial successions of wheat Qu remain unclear. In this study, we first correlated microbial community succession with physicochemical factors (moisture, temperature, acidity, glucoamylase and amylase) in inoculated raw wheat Qu (IRWQ) with Saccharopolyspora rosea. The Mantel test was performed to investigate the significance and found that temperature (r = 0.759, P = 0.001), moisture (r = 0.732, P = 0.006), and acidity (r = 0.712, P = 0.017) correlated significantly with the bacterial community in phase 1 (0–40 h). Meanwhile, temperature correlated significantly with the fungal community in phases 1 and 2 (40–120 h). To confirm the effect of temperature on microbial communities, the artificial reduction of bio-heat (37°C) in IRWQ also reduced the relative abundance of heat-resistant microorganisms including Bacillus and Saccharopolyspora. A higher abundance of Saccharopolyspora (87%) in IRWQ was observed following biofortified inoculation of S. rosea , in which glucoamylase activity increased by 40% compared to non-inoculated raw wheat Qu (NIRWQ) (1086 U/g vs 776 U/g). Finally, the IRWQ was employed to mechanized huangjiu fermentation and it was found to reduce the bitter amino acid and higher alcohol content by 27% and 8%, respectively, improving the drinking comfort and quality of huangjiu. [ABSTRACT FROM AUTHOR]

[Display omitted] • P. pentosaceus , W. paramesenteroides , R. emersonii, B. spectabilis were dominant in Daqu. • Four Saccharopolyspora species were dominant in wheat Qu. • Flavor metabolic pathways in Qu microbiota were reconstructed. • The dominant microbes in Qu may not be the main functional microbes. Daqu and wheat Qu are saccharification and fermenting agents in Chinese huangjiu and baijiu production. This study aimed to investigate the difference between Daqu and wheat Qu in physicochemical indices, microbial communities, functional genes, and the metabolic network of key microbes responsible for flavor synthesis by whole-metagenome sequencing and metabolite analysis. Herein, physicochemical indices indicated that compared with wheat Qu , Daqu exhibited higher protease and cellulase activity and acidity, and lower glucoamylase and amylase enzyme activity. Metagenomic sequencing reveals that although Daqu and wheat Qu community composition have significant differences at species level, they have similar functional genes. Daqu were enriched in Pediococcus pentosaceus , Weissella paramesenteroides , Rasamsonia emersonii and Byssochlamys spectabilis (22.48% of the total abundance), while wheat Qu harbored greater abundances of Saccharopolyspora (54.78%, Saccharopolyspora rectivirgula , Saccharopolyspora shandongensis , Saccharopolyspora hirsuta , Saccharopolyspora spinose , and Saccharopolyspora erythraea). From a functional perspective, the important functions of Daqu and wheat Qu are both amino acid metabolism and carbohydrate metabolism. Meanwhile, a combined analysis among microbiota, functional genes, and dominant flavors indicated S. shandongensis , S. rectivirgula , and S. spinose might be the main contributor to the synthesis of flavor compounds in wheat Qu , while R. emersonii , W. paramesenteroides , Leuconostoc citreum , Leuconostoc mesenteroides , Weissella cibaria and P. pentosaceus may make the greatest contribution to flavor compounds synthesis in Daqu. This study reveals the microbial and functional dissimilarities of Daqu and wheat Qu , and helps elucidating different metabolic roles of microbes during flavor formation. [ABSTRACT FROM AUTHOR]