Unraveling Qu-aroma variation between inner and outer layers of medium-temperature Daqu: A multi-omics and sensory approach

Solid-state fermentation heterogeneity causes microenvironmental differences, shaping diverse microbial communities and metabolite compositions. Here, we aimed to investigate the variations in Qu -aroma between the inner (I-) and outer (O-) layers of medium-temperature Daqu (MT- Daqu ) starter, and to analyze the differences in microbial communities as well as the associated metabolic pathways related to Qu -aroma volatile organic compounds (VOCs). Firstly, quantitative descriptive analysis revealed that the I-layer exhibited intensified rancid-roasted notes, whereas the O-layer showed stronger grain and woody notes. Secondly, a total of 225 VOCs were identified from 88 inner and outer layer samples using HS-SPME-GC–MS, among which 43 differential VOCs were confirmed between the layers (VIP > 1, P  < 0.05, and Fold change ≥1.5 or ≤0.67). Thin film (TF)-SPME-GC-O/MS revealed 52 and 47 odors with corresponding VOCs in the I- and O-layers, respectively. Subsequently, cross-referencing metagenomic and metabolic databases revealed that 37 aroma-active VOCs were functionally linked to six metabolic modules within the MT- Daqu microbial metabolic network. Finally, integrated metagenomic and amplicon sequencing identified potential functional microorganisms associated with specific metabolic modules. Five genera, including Kroppenstedtia and Thermomyces , were identified as characteristic of the I-layer, while 22 genera, such as Lactobacillus and Saccharomycopsis , were characteristic of the O-layer. Notably, Kroppenstedtia , Thermomyces , Lactobacillus , Saccharomycopsis , and Weissella were ubiquitously associated with all six metabolic modules across both layers of MT- Daqu . This study delivers a perspective for clarifying the spatial heterogeneity of Qu -aroma and its microbial drivers.