Unveiling the synthesis of aromatic compounds in sauce-flavor Daqu from the functional microorganisms to enzymes.

Aromatic compounds serve as the primary source of floral and fruity aromas in sauce-flavor (Maotai flavor) baijiu, constituting the skeleton components of its flavor profile. Nevertheless, the formation mechanism of these compounds and key aroma-producing enzymes in sauce-flavor Daqu (fermentation agent, SFD) remain elusive. Here, we combined metagenomics, metaproteomics, metabolomics, and key enzyme activity to verify the biosynthesis pathway of aromatic compounds and to identify key enzymes, genes, and characteristic microorganisms in SFD. The results showed that the later period of fermentation was critical for the generation of aromatic compounds in SFD. In-situ verification was conducted on the potential key enzymes and profiles in various metabolites, providing comprehensive evidence for the main synthetic pathways of aromatic compounds in SFD. Notably, our results showed that primary amine oxidase (PrAO) and aldehyde dehydrogenase (ALDH) emerged as two key enzymes promoting aromatic compound synthesis. Additionally, two potential key functional genes regulating aromatics generation were identified during SFD fermentation through correlation analysis between proteins and relevant metabolites, coupled with in vitro amplification test. Furthermore, original functional strains (Aspergillus flavus-C10 and Aspergillus niger-IN2) exhibiting high PrAO and ALDH production were successfully isolated from SFD, thus validating the results of metagenomics and metaproteomics analyses. This study comprehensively elucidates the pathway of aromatic compound formation in SFD at the genetic, proteomic, enzymatic, and metabolomic levels, providing new ideas for the investigation of key flavor substances in baijiu. Additionally, these findings offer valuable insights into the regulatory mechanisms of aromatic compounds generation.

Analysis of fermentation control factors on volatile compounds of primary microorganisms in Jiang-flavor Daqu.

Chinese Jiang-flavor Baijiu is the most widely consumed liquor. Jiang-flavor Daqu, a fermentation starter, is important sources of key flavors of Jiang-flavor Baijiu. Some microbes play significant roles in flavor formation of Daqu. In order to clarify the microbial population that promotes the formation of Daqu flavor, we use high throughput sequencing technology combined with headspace solid-phase microextraction gas chromatography-mass spectrometry to investigate microbial population and volatile compounds in Jiang-flavor Daqu. In addition, the dynamic changes of physicochemical factors and enzyme activities in Jiang-flavor Daqu were investigated. Correlations between microbial population, volatile compounds, physicochemical factors, and enzyme activities of Jiang-flavor Daqu were disclosed by redundancy analysis and Spearman correlation analysis. A total of 66 volatile compounds were identified and 14 primary microorganisms were selected. Results showed that high temperature environment could promote the formation of acids, aldehydes and ketones, phenols, furans by affecting the growth of Monascus, Trichomonascus, Cutaneotrichosporon, Wallemia, Millerozyma, Nigrospora, Cladosporium, Bacillus, and Pediococcus in the early fermentation stage. While high nitrogen environment was more suitable for the growth of Virgibacillus and Kroppenstedtia, who could promote the formation of pyrazines in the late fermentation stage. PRACTICAL APPLICATIONS: This study has provided a scientific basis for the directed regulation of Daqu fermentation through physicochemical factors, developed scientific basis for artificially constructing Daqu microbial population and obtaining an easy-to-operate, reproducible fermentation system for Daqu production.