## 3 Results and Discussion
### Analysis of Flavor Compounds  
The results of the physicochemical factors showed that the fermentation process of Baijiu could be simulated under laboratory conditions, while the analysis of flavor compounds was significant to judge the quality of the fermentation products, as flavor compounds play an important role in the quality of Baijiu. The flavor compounds of fermented grains were determined by GC-MS. A total of 78 flavor compounds were identified, including forty-seven esters, eight alcohols, six phenols, five alkanes, four acids, two pyrazines, two aldehydes, two ketones, and one furan in all samples during the whole fermentation process (Table S1). There were 43 flavor compounds detected in sample 1, while samples 2, 3, and 4 contained 64, 56, and 62 flavor compounds, respectively. In addition, the relative contents of most major flavor compounds in samples 2, 3, and 4 were higher than in sample 1. For example, the relative contents of hexanoic acid, ethyl ester in samples 2, 3, and 4 were 311.18 \(\pm\) 299.40 \(\upmu\)g/kg, 545.34 \(\pm\) 269.44 \(\upmu\)g/kg, 290.05 \(\pm\) 37.99 \(\upmu\)g/kg, respectively, all being higher than that in sample 1 (124.51 \(\pm\) 36.76 \(\upmu\)g/kg); butanoic acid, ethyl ester in samples 1, 2, 3, and 4 were 131.70 \(\pm\) 51.34 \(\upmu\)g/kg, 1942.47 \(\pm\) 490.64 \(\upmu\)g/kg, 14109.75 \(\pm\) 1964.85 \(\upmu\)g/kg, 4633.89 \(\pm\) 788.53 \(\upmu\)g/kg, respectively; octanoic acid, ethyl ester in samples 1, 2, 3, and 4 were 92.82 \(\pm\) 16.18 \(\upmu\)g/kg, 110.63 \(\pm\) 14.00 \(\upmu\)g/kg, 304.11 \(\pm\) 38.05 \(\upmu\)g/kg, 289.13 \(\pm\) 25.31 \(\upmu\)g/kg, respectively (Figure 4 and Table S1). This indicated that the microbial combination could enhance the stable production of flavor compounds. The heatmap analysis of the changes of flavor compounds during the fermentation process showed that many flavor compounds in the samples increased significantly with prolongation of the fermentation time (Figures S1-S4). However, the changed tendencies of some flavor compounds were different, such as 2-heptanone, hexadecane, methyl caproate, and benzaldehyde in sample 1 were only detected on day 0 with very low contents, and were almost not identified with prolonged fermentation (Figure S1). These flavor compounds were possibly brought into the fermented grains by the raw materials, and no microorganisms could produce these flavor compounds during the fermentation process of the control group. 2,3,5,6-Tetramethylpyrazine is generally considered as a functional factor in Baijiu, with nutty, earth, cocoa, peanut, coffee, and asparagus aromas [30], and it was only identified in sample 2 (Figure S2) and sample 4 (Figure S4). Therefore, it is possible that 2,3,5,6-tetramethylpyrazine was introduced to the fermentation system by Daqu. The content of 2,3,5,6-tetramethylpyrazine gradually increased in the early and middle stages of fermentation, but decreased in the middle and late stages of fermentation. This indicates that the fermented samples contained microorganisms capable of the synthesis and degradation of 2,3,5,6-tetramethylpyrazine [27,30,31]. In addition, there were several middle-chain fatty acid esters, such as hexanoic acid, ethyl ester; hexanoic acid, butyl ester; octanoic acid, ethyl ester; and octanoic acid, methyl ester in sample 3 (Figure S3) and sample 4 (Figure S4), and their relative contents increased during the fermentation period (0-28 days), but decreased slightly on day 42. Studies have reported that one of the synthetic pathways of medium chain fatty acids during the fermentation of Baijiu is an esterification reaction using acid and alcohol as the substrates [2,4]. This suggests that the amount of precursors of the esters caproic acid and octanoic acid might be insufficient in the late fermentation stages, while the content of other acids was higher, and led to the reverse reaction of ester synthesis [2,32]. Figure 3: Changes of physicochemical factors in fermented grains.