Untargeted Metabolomics Reveals Metabolic Changes in Cyperus esculentus L. by Mixed Fermentation of Lacticaseibacillus paracasei and Saccharomyces cerevisiae

This study investigated the effects of single-strain fermentation (using Lactobacillus paracasei XY1-4 and Saccharomyces cerevisiae XX1-2) and 1:1 mixed fermentation on the differential metabolites and in vitro cholesterol-lowering activity of Cyperus esculentus L. tuber milk. A non-targeted metabolomics approach based on liquid chromatography-tandem mass spectrometry (LC-MS/MS) was applied, and the dynamic changes in physicochemical properties and in vitro cholesterol-lowering rates during fermentation were determined to systematically compare the metabolite profiles and functional characteristics of tuber milk samples subjected to single-strain fermentation and mixed fermentation for 12 h and 24 h, respectively. The results showed that a total of 1085 differential metabolites were screened across all fermentation groups under the criteria of VIP ≥ 1, p < 0.05, and FC ≥ 2 or FC ≤ 0.5, which were primarily classified as lipids and lipid-like molecules as well as organic acids and their derivatives. Enrichment analysis of these differential metabolites identified ten metabolic pathways closely associated with fermentation progression and functional activity, including arginine biosynthesis, the tricarboxylic acid (TCA) cycle, and glutathione metabolism. Further correlation analysis demonstrated that key metabolites such as succinic acid and L-glutamic acid were significantly and positively correlated with the in vitro cholesterol-lowering rate. This study clarifies the effects of different fermentation treatments on the metabolome and functional properties of tuber milk, laying a solid theoretical foundation for the development and quality optimization of functional fermented Cyperus esculentus L. products.