Untargeted metabolomics analysis of Mucor racemosus Douchi fermentation process by gas chromatography with time‐of‐flight mass spectrometry

Intensive study of the metabolome during the Douchi fermentation can provide new knowledge for optimizing the fermentation process. In this work, the metabolic characterization throughout the fermentation of Mucor racemosus Douchi was investigated using gas chromatography with time‐of‐flight mass spectrometry. A total of 511 peaks were found, and 114 metabolites were identified. The fermentation process was clearly distinguished into two main phases by principal components analysis and orthogonal partial least squares‐discriminant analysis. All the samples in the score plots were within the 95% Hotelling T (2) ellipse. Two separated clusters can be seen clearly in the score plot, which represents the two stages of fermentation: koji‐making (within 48 hr) and postfermentation (after 48 hr). Besides, clear separation and discrimination by both methods were found among different fermentation time within 15 days, while the discrimination cannot be found with more than 15 days of fermentation, indicating that the fermentation of Douchi was finished in 15 days. Due to the synergistic effect of protease and hydrolase accumulated in the early stage, proteins and other big molecular substances are rapidly hydrolyzed into a large number of small molecule components. However, the activity of enzymes decreased with the further fermentation, and some free amino acids were consumed in Maillard reaction. Therefore, there was no significant change in the content of small molecular substances after 15 days of fermentation. Furthermore, the levels of some metabolites such as alanine and lysine involved in the fermentation varied significantly throughout the processes. This study provides new insights for the metabolomics characteristics of Douchi fermentation.