High-temperature Daqu (HTD) is a crucial fermentation starter and plays a particularly important role in the production of sauce-flavor Baijiu. In this study, the original upper, middle, and lower layers of stacked HTD were labeled upon entry into the fermentation room, and the changes in their physicochemical properties, microbial communities, and volatile flavor compounds before and after flipping were investigated to reveal the effect of flipping on the quality of Daqu in different stacking layers. Flipping promoted pyrazine accumulation only when Daqu was in the upper layer, while the acidity gradient (lower > middle > upper) remained unchanged. Dominant bacteria shifted from Bacillus to Kroppenstedtia and Scopulibacillus after the first flipping, then to Lactobacillus and Virgibacillus after the second flipping. Fungi shifted from Thermoascus, Rhizopus, and Aspergillus to Pichia and Thermomyces after the first flipping, with Thermoascus becoming dominant after the second flipping. LEfSe identified bacterial and fungal genera differentially enriched across stacking layers, along with 30 differential volatiles (mainly esters). Flipping altered moisture and acidity, which were closely associated with microbial succession, and Lactobacillus abundance correlated with the acidity gradient. These findings provide new insights into how flipping influences different stacking layers, which can help improve the quality of HTD.
| [1] |
Cao D, Lv J, Liu C, Jin C, Zhang Y, Zhang Y, Zhang W. Metabolic power, volatile compounds, and flavor marker formation mechanisms of three fungi isolated from Fengxiangxing Daqu. Front Microbiol, 2025, 16: 1593638.
|
| [2] |
Chen X, Wu Y, Zhu H, Wang H, Lu H, Zhang C, Li X, Xu Y, Li W, Wang Y. Turning over fermented grains elevating heap temperature and driving microbial community succession during the heap fermentation of sauce-flavor baijiu. LWT, 2022, 172: 114173.
|
| [3] |
Chen C, Ni D, Yang Y, Tian J, Yang F, Ye X. Characterizing the microbial community constructure and the metabolites among different colour Moutai Daqu. Food Chem X, 2025, 26: 102223.
|
| [4] |
Chen P, Shen S, Lin L, Liu Q, Zhang C, Zhong C. Elucidating the impact of high-temperature Daqu on base Baijiu of Sauce-Flavor Baijiu: from key aroma compounds to microbial origins. Foods, 2026, 15: 1124.
|
| [5] |
Deng L, Mao X, Liu D, Ning XQ, Shen Y, Chen B, Nie HF, Huang D, Luo HB. Comparative analysis of physicochemical properties and microbial composition in high-temperature Daqu with different colors. Front Microbiol, 2020, 11: 588117.
|
| [6] |
Deng A, Tang J, Zhu C, Huang Y. Driving mechanisms for microbial community succession and change of volatile flavor compounds during the solid-state fermentation of black high-temperature Daqu. Food Sci, 2024, 45: 102-12.
|
| [7] |
Dong W, Yu X, Wang L, Zou M, Ma J, Liu J, Feng Y, Zhao S, Yang Q, Hu Y. Unveiling the microbiota of sauce-flavor Daqu and its relationships with flavors and color during maturation. Front Microbiol, 2024, 15: 1345772.
|
| [8] |
Du Y, Tang J, Liu D, Liu N, Peng K, Wang C, Huang D, Luo H. Microbial metabolism during the thermophilic phase promotes the generation of aroma substances in nongxiangxing Daqu. Food Chem X, 2023, 20: 101044.
|
| [9] |
He G, Huang J, Wu C, Jin Y, Zhou R. Bioturbation effect of fortified Daqu on microbial community and flavor metabolite in Chinese strong-flavor liquor brewing microecosystem. Food Res Int, 2020, 129: 108851.
|
| [10] |
He M, Jin Y, Zhou R, Zhao D, Zheng J, Wu C. Dynamic succession of microbial community in Nongxiangxing daqu and microbial roles involved in flavor formation. Food Res Int, 2022, 159: 111559.
|
| [11] |
Huang Y, Li D, Mu Y, Zhu Z, Wu Y, Qi Q, Mu Y, Su W. Exploring the heterogeneity of community and function and correspondence of species-enzymes among three types of Daqu with different fermentation peak-temperature via high-throughput sequencing and metagenomics. Food Res Int, 2024, 176: 113805.
|
| [12] |
Li P, Lin W, Liu X, Wang X, Luo L. Environmental factors affecting microbiota dynamics during traditional solid-state fermentation of Chinese Daqu starter. Front Microbiol, 2016, 7: 1237.
|
| [13] |
Li Y, Qin X, Zeng X, She M, Chen L, Lu J, Han Y, Yang F, Tu H, Du H. Comprehensive analysis of spatiotemporal heterogeneity reveals the effects of physicochemical and biological factors on temperature rise during the Moutai-flavor Baijiu stacking fermentation process. Chem Eng J, 2025, 508: 161049.
|
| [14] |
Li D, Zhang X, Guo H, Zhang H, Jiang S, Song Y, Mu Y. Linking microbial ecosystem to key flavor compounds in high-temperature Daqu: multi-omics insights into putatively bacillus-driven acetoin-pyrazine pathway. LWT, 2026.
|
| [15] |
Luo Y, Liao H, Luo Y, Gao L, Xia X. Temperature-driven microbial assembly and flavor compound dynamics in high-temperature Daqu: adaptive co-evolution for efficient tetramethylpyrazine production. Int J Food Microbiol, 2025, 441: 111325.
|
| [16] |
Ma S, Li Y, Dong Y, Zhou Y, Tu F, Zhang L, Wu C. Stochastic and deterministic processes shape microbial communities and flavor characteristics of strong-flavor daqu during storage. Int J Food Microbiol, 2025, 429: 111017.
|
| [17] |
Ministry of Industry and Information Technology of the People’s Republic of China. General methods of analysis for Daqu. QB/T 4257 – 2011; 2011.
|
| [18] |
Pang Z, Li W, Hao J, Xu Y, Du B, Zhang C, Wang K, Zhu H, Wang H, Li X. Correlational analysis of the physicochemical indexes, volatile flavor components, and microbial communities of high-temperature daqu in the northern region of China. Foods, 2023, 12: 326.
|
| [19] |
Peng Q, Wei C, Xie J, Yi Z, Ren Z. A high esterifying enzyme-producing rhizopus strain for fortified Daqu: screening, preparation, and microbial community characterization. Foods, 2026, 15: 1213.
|
| [20] |
Que P, Yao S, Hu J, Hu F, Wang D, Huang W. Associations of microbial communities with metabolites and enzyme activities in different colors of high-temperature Daqu. World J Microbiol Biotechnol, 2025, 41: 324.
|
| [21] |
Shi W, Chai LJ, Fang GY, Mei JL, Lu ZM, Zhang XJ, Xiao C, Wang ST, Shen CH, Shi JS. Spatial heterogeneity of the microbiome and metabolome profiles of high-temperature Daqu in the same workshop. Food Res Int, 2022, 156: 111298.
|
| [22] |
Shi G, Fang C, Xing S, Guo Y, Li X, Han X, Lin L, Zhang C. Heterogenetic mechanism in high-temperature Daqu fermentation by traditional craft and mechanical craft: from microbial assembly patterns to metabolism phenotypes. Food Res Int, 2024, 187: 114327.
|
| [23] |
Shi W, Chai LJ, Guo J, Huang T, Li FX, Wang YY, Zheng HM, He YX, Mei JL, Zhang XJ. Spatial location within the incubation room shapes the microbial succession and metabolic profiles of high-temperature Daqu. LWT. 2026;119065. https://doi.org/10.1016/j.lwt.2026.119065.
|
| [24] |
Tang P, Wang L, Lu J, Qiao M, Li C, Xiao D, Guo X. Characterization of key aroma compounds and relationship between aroma compounds and sensory attributes in different quality of high temperature Daqu. LWT, 2024, 194: 115801.
|
| [25] |
Tang P, Li Q, Shan Q, Qin L, Kang X, Li C, Xiao D, Guo X. Physicochemical properties, microbial community evolution, and flavor compounds during the mechanized production of Jiangxiangxing high-temperature daqu. LWT. 2025;118418. https://doi.org/10.1016/j.lwt.2025.118418.
|
| [26] |
Tie Y, Yang Q, Chen X, Wang L, Wu Z, Takamine K, Zhang W. Bottom-up metaproteomic profiling of active microorganisms producing ethyl ester synthase and investigation of their key driving factors during Daqu fermentation. Food Biosci. 2025;107906. https://doi.org/10.1016/j.fbio.2025.107906.
|
| [27] |
Wang H, Huang Y, Huang Y. Microbiome diversity and evolution in stacking fermentation during different rounds of Jiang-flavoured Baijiu brewing. LWT, 2021, 143: 111119.
|
| [28] |
Wang C, Li C, Bin Z, Zhu G, Tang S, Zhang J, Chen Y, Xiao D, Guo X. Workshop environment heterogeneity shaped the microbiome and metabolome profiles during Xiasha round of Jiangxiangxing Baijiu. Food Chem X, 2024, 22: 101264.
|
| [29] |
Wei YH, Wen Z, Yao SJ, Cheng PY, Huang W, Jiang L, Bai M, Han DY, Zhu HY, Bai FY. Tempo-spatial dynamics of physicochemical properties and microbial communities in high-temperature Daqu during the fermentation process. Food Chem X. 2025;102815. https://doi.org/10.1016/j.fochx.2025.102815.
|
| [30] |
Wen Z, Wei YH, Han DY, Song L, Zhu HY, Guo LC, Chen SX, Lin B, He CJ, Guo ZX. Deciphering the role of traditional flipping crafts in medium-temperature Daqu fermentation: microbial succession and metabolic phenotypes. Curr Res Food Sci, 2025, 10: 101063.
|
| [31] |
Wu X, Jiang Q, Wang Z, Xu Y, Chen W, Sun J, Liu Y. Diversity, enzyme production and antibacterial activity of Bacillus strains isolated from sesame-flavored liquor Daqu. Arch Microbiol, 2021, 203: 5831-9.
|
| [32] |
Wu L, Yan M, Huang X, Liao H, Bao D, Ge Y, Wang S, Xia X. Temperature-mediated shift from competitive to facilitative interactions between lactic acid bacteria and Bacillus species in Daqu fermentation: insights from metagenomics, dual RNA-seq, and coculture analysis. Int J Food Microbiol. 2025;111352. https://doi.org/10.1016/j.ijfoodmicro.2025.111352.
|
| [33] |
Xing S, Shi G, Lu J, Fang C, Li C, Yuan S, Shi F, Lin L, Zhang C. The discrepancy in amino acids within high-temperature Daqu: a novel metabolic marker for the quality evaluation of Daqu. Food Chem, 2025, 470: 142645.
|
| [34] |
Yang L, Fan W, Xu Y. Chameleon-like microbes promote microecological differentiation of Daqu. Food Microbiol, 2023, 109: 104144.
|
| [35] |
Yang L, Fan W, Xu Y. Qu-omics elucidates the formation and spatio-temporal differentiation mechanism underlying the microecology of high temperature Daqu. Food Chem, 2024, 438: 137988.
|
| [36] |
Yuan M, Lu T, Zhang H, Wang J, Cui M, Zhao W, Wu Y, Ren X, Yao P, Zhang X. Verification of the esters-producing properties of Thermoascus aurantiacus QH-1 derived from low-temperature Daqu by multiomics. Food Microbiol, 2025.
|
| [37] |
Yuan S, Tu R, Shan B, Liu Y, Jiang X, Zheng M, Yang L, Liu H, Zhao T, Yang P. Turning-modulated vertical CO2 gradients drive microbial stratification and amadori product accumulation in winter Daqu. Foods, 2026, 15: 799.
|
| [38] |
Zhang K, Zhang TT, Guo RR, Ye Q, Zhao HL, Huang XH. The regulation of key flavor of traditional fermented food by microbial metabolism: a review. Food Chem X, 2023, 19: 100871.
|
| [39] |
Zhang Y, Kang J, Han BZ, Chen X. Wheat-origin Bacillus community drives the formation of characteristic metabolic profile in high-temperature Daqu. LWT, 2024, 191: 115597.
|
| [40] |
Zhang Z, Ran X, Guo Z, Hou Q, Qu D, Wang C, Xu Y, Wang Y. Microbial diversity, functional properties, and flavor characteristics of high-temperature Daqu with different colors. Food Res Int, 2025.
|
| [41] |
Zhang Z, Zhang K, Hou Q, Yang C, Guo Z, Li Y, Wang C, Wang Y. Microbial ecology and flavor formation mechanisms of high-temperature Daqu in the Huang-Huai River basin and adjacent regions: a comparative study from eastern Henan, Jiaodong peninsula, and southern Anhui. Food Res Int, 2026.
|
| [42] |
Zhu C, Cheng Y, Shi Q, Ge X, Yang Y, Huang Y. Metagenomic analyses reveal microbial communities and functional differences between Daqu from seven provinces. Food Res Int, 2023, 172: 113076.
|
Funding
2024 "Suqian Talent" Qunying Program Youth Funding Project(Grant No. SQQN202416)
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Jiangnan University
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