## 3 Results and Discussion  
### Flavor Analysis of the Samples in the First Three Rounds of Sauce-Flavor Baijiu  
A total of 99 flavor chemicals were detected by GC-MS, including 53 esters, 16 alcohols, 2 pyrazines, 1 furan, 8 phenols, 5 ketones, 8 aldehydes, 3 benzene, 1 ether, 1 pyrrole, and 1 thiazole in all the samples of the three rounds (Tables S1-S3, and Figure 2). The comparative analysis of flavor chemicals in the grain samples at the end of each round showed that the concentrations of 11 flavor chemicals in the third round of fermented grains increased, including hexanoic acid, ethyl ester; octanoic acid, ethyl ester; decanoic acid, ethyl ester; dodecanoic acid, ethyl ester; phenylacetic acid, ethyl ester; 3-(methylthio)-propionic acid ethyl ester; acetic acid, phenylethyl ester; hexanoic acid, butyl ester; benzaldehyde, 2,3-butanediol, and tetramethylpyrazine. These chemicals have an important contribution to fruity, floral, and sweet flavors, as hexanoic acid, ethyl ester, showed floral and sweet flavors with an odor threshold value of 55 \(\upmu\)g/L, and octanoic acid, ethyl ester showed a fruity flavor with an odor threshold value of 13 \(\upmu\)g/L [2]. Decanoic acid, ethyl ester, and dodecanoic acid, ethyl ester, and other medium and long chain fatty acid esters contributed to the typical characteristics of sauce-flavor Baijiu, namely \"a lingering aroma in empty glass\" [15]. In addition, other important flavor chemicals, such as tetramethylpyrazine (with baking aroma and a sweet flavor), also contributed significantly to the flavor of the sauce-flavor Baijiu [16], and its concentration was significantly increased in round 3 of fermented grains. 2,3-Butanediol can provide the Baijiu product with a mellow feeling orally with a sticky and slightly sweet taste [17]. This served as the reason for the improved quality of base Baijiu in round 3.  
Figure 2: Flavor chemical analysis of the fermented grains from rounds 1b, 2, and 3 during the sauce-flavor Rajjia manufacturing process. (**a**), round 1b. (**b**), round 2. (**c**) round 3.  
The concentrations of two phenols decreased significantly in the fermented grains of round 3, which were 4-ethylphenol and 2-methoxy-4-ethyl-phenol (4-ethyl guaiacol, with clove and spicy flavor) [18]. 4-Ethyl guaiacol, a controversial chemical for baijiu flavor, was frequently detected in sauce-flavor Baijius in the Guizhou region of China [19; 20]. However, although it was detected in the fermented grains of round 2, it was not identified in the fermented grains at the end of round 1b and round 3. This indicated the divergences in flavors between sauce-flavor Baijiu produced in the southern and northern regions of China [3; 21]. The concentration of benzaldehyde (with a bitter almond flavor) [22] in fermented grains at the end of round 3 was also higher than that in round 2. A moderate amount of benzaldehyde can enrich the flavor of sauce-flavor Baijiu but also bring bitterness in taste [23]. Flavor chemicals in each fermentation round showed fluctuated concentrations, indicating the coexistence of synthesis and degradation metabolisms of these chemicals by microorganisms in the fermentation process (Figure 2). However, what were the dynamic microbial compositions during the fermentation process, and what kinds of microorganisms were related to the synthesis or degradation of the key flavor chemicals? These questions were investigated in the following experiments.