Key Laboratory of Industrial Biotechnology of Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi 214122, PR China; School of Chemistry and Life Sciences, Suzhou University of Science and Technology, Suzhou 215009, PR China; National Engineering Laboratory for Cereal Fermentation Technology, Jiangnan University, Wuxi 214122, PR China.
National Engineering Laboratory for Cereal Fermentation Technology, Jiangnan University, Wuxi 214122, PR China; School of Pharmaceutical Science, Jiangnan University, Wuxi 214122, PR China.
Food Res Int. 2021 Sep;147:110449. doi: 10.1016/j.foodres.2021.110449. Epub 2021 Jun 3.
Mud cellars have long been used as anaerobic bioreactors for the fermentation of Chinese strong-flavor Baijiu, where starchy raw materials (mainly sorghum) are metabolized to ethanol and various flavor compounds by multi-species microorganisms. Jiupei (fermented grains) and pit mud are two spatially linked microbial habitats in the mud cellar, yet their metabolic division of labor remains unclear. Here, we investigated the changes in environmental variables (e.g., temperature, oxygen, pH), key metabolites (e.g., ethanol, organic acids) and microbial communities in jiupei and pit mud during fermentation. Jiupei (low pH, high ethanol) and pit mud (neutral pH) provided two habitats with distinctly different environmental conditions for microbial growth. Lactic acid accumulated in jiupei, while butyric and hexanoic acids were mainly produced by microbes inhabiting the pit mud. Biomass analysis using quantitative real-time PCR showed that bacteria dominated the microbial consortia during fermentation, moreover cluster and principal coordinate analysis (PCoA) analysis showed that the bacterial communities of jiupei and pit mud were significantly divergent. The bacterial community diversity of jiupei decreased significantly during the fermentation process, and was relatively stable in pit mud. Lactobacillus dominated the jiupei bacterial community, and its relative abundance reached 98.0% at the end of fermentation. Clostridia (relative abundance: 42.9-85.5%) was the most abundant bacteria in pit mud, mainly distributed in the genus Hydrogenispora (5.3-68.4%). Fungal communities of jiupei and pit mud showed a similar succession pattern, and Kazachstania, Aspergillus and Thermoascus were the predominant genera. PICRUSt analysis demonstrated that enzymes participating in the biosynthesis of acetic and lactic acid were mainly enriched in jiupei samples, while the bacterial community in the pit mud displayed greater potential for butyric and hexanoic acid synthesis. Assays from an in vitro simulated fermentation further validated the roles of jiupei microbiota in acetic and lactic acid production, and these acids were subsequently metabolized to butyric and hexanoic acid by the pit mud microbiota. This work has demonstrated the synergistic cooperation between the microbial communities of jiupei and pit mud for the representative flavor formation of strong-flavor Baijiu.
泥窖长期以来一直被用作中国浓香型白酒发酵的厌氧生物反应器，在该反应器中，多种微生物将淀粉质原料（主要是高粱）代谢为乙醇和各种风味化合物。酒醅（发酵谷物）和窖泥是泥窖中两个空间上相连的微生物栖息地，但它们的代谢分工尚不清楚。在这里，我们研究了发酵过程中酒醅和窖泥中环境变量（例如温度、氧气、pH 值）、关键代谢物（例如乙醇、有机酸）和微生物群落的变化。酒醅（低 pH 值、高乙醇）和窖泥（中性 pH 值）为微生物生长提供了两个具有明显不同环境条件的栖息地。酒醅中积累了乳酸，而丁酸和己酸主要由栖息在窖泥中的微生物产生。使用定量实时 PCR 的生物量分析表明，细菌在发酵过程中主导了微生物群落，此外，聚类和主坐标分析（PCoA）分析表明，酒醅和窖泥的细菌群落差异显著。酒醅的细菌群落多样性在发酵过程中显著下降，而在窖泥中相对稳定。乳杆菌主导了酒醅的细菌群落，其相对丰度在发酵结束时达到 98.0%。梭菌（相对丰度：42.9-85.5%）是窖泥中最丰富的细菌，主要分布在氢化孢菌属（5.3-68.4%）中。酒醅和窖泥的真菌群落表现出相似的演替模式，其中卡氏酵母属、曲霉属和嗜热丝孢菌属是主要属。PICRUSt 分析表明，参与乙酸和乳酸生物合成的酶主要富集在酒醅样品中，而窖泥中的细菌群落显示出更大的丁酸和己酸合成潜力。体外模拟发酵实验进一步验证了酒醅微生物群在乙酸和乳酸生成中的作用，这些酸随后被窖泥微生物群代谢为丁酸和己酸。这项工作证明了酒醅和窖泥微生物群落之间的协同合作对浓香型白酒代表性风味的形成具有重要作用。
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