Metagenomics-based insights into the microbial community succession and metabolic potential of flavor in Qingke high-temperature Daqu.

Raw grain selection profoundly influences the flavor and quality of high-temperature Daqu (HTD). However, the microbial and flavor profiles of Daqu made with Qingke (highland barley) remain unclear. This study explored how varying Qingke addition affects physicochemical indicators, volatile organic compounds (VOCs), and microbial succession, ultimately impacting the formation of characteristic flavor compounds. Results showed that Qingke high-temperature Daqu (QHTD) underwent notable physicochemical changes, with the 30%and 50% addition groups exhibiting higher moisture and total acid levels (P < 0.01). Eleven characteristic VOCs were identified in QHTD, primarily represented by tetramethyl-pyrazine, phenylethyl alcohol, benzaldehyde, and specific alcohols, aldehydes, and acids. Dominant taxa in mature QHTD included Pseudonocardiales (mainly Saccharopolyspora), Bacillales (mainly Kroppenstedtia and Lentibacillus), and Eurotiales (mainly Paecilomyces). Temperature, moisture, and total acid were the main drivers of microbial succession in QHTD, and Qingke supplementation shaped microbial co-occurrence patterns by altering the fermentation environment. Key genera such as Kroppenstedtia and Paecilomyces were positively correlated with pyrazines and aldehydes. Metabolic network analysis based on metagenomic prediction revealed that functional bacteria (mainly Bacillales and Lactobacillales) were extensively involved in macromolecular degradation and flavor synthesis, exhibiting higher enzyme abundances in QHTD. Fungi primarily drove macromolecule degradation, phenylethyl alcohol synthesis, and metabolism of acetate and ethanol. In summary, the addition of Qingke modified the physicochemical indicators, altered functional microbial abundance, and enriched the flavor compounds in HTD. These insights offer theoretical and practical guidance for enhancing multi-grain Daqu production and the quality of Sauce-flavor Baijiu.