Turning-Modulated Vertical CO2 Gradients Drive Microbial Stratification and Amadori Product Accumulation in Winter Daqu

High-temperature Daqu (HTD)'s quality determines the characteristics and yield of the Chinese sauce-aroma baijiu. However, winter production frequently encounters challenges such as fermentation instability and metabolic fluctuations, primarily stemming from complex, unmonitored microenvironmental changes within the HTD pile. This study established a closed-loop system linking the microenvironment, HTD quality, microbiome, and metabolome. Through continuous monitoring of the winter fermentation pile's microenvironmental conditions and integrating multi-omics analyses, we revealed that CO₂ concentration within fermentation piles is the core factor causing quality variations in HTD. By breaking the respiratory bottleneck formed by carbon dioxide (CO₂) accumulation through the turning anaerobic stress can be alleviated, thereby driving metabolic succession. The study found that vertical CO₂ concentration heterogeneity severely restricts the enrichment of aerobic core functional microbial communities such as the <i>Bacillus</i> species. This directly blocks key metabolic pathways including amino acid metabolism and energy supply via ABC transporters. Moreover, the specific accumulation of Amadori products further confirms that this low-temperature environment under CO₂ stress causes the Maillard reaction to stall at intermediate stages. Consequently, this study proposes a steady-state control strategy centered on oxygen and CO₂ gas characteristics. By actively regulating the gaseous microenvironment to eliminate metabolic heterogeneity, it provides theoretical support for standardizing traditional fermentation processes.