<p>In this study, we determined whether gypenosides exert a therapeutic effect on colitis. We then explored the underlying mechanism of gypenosides in the treatment of ulcerative colitis (UC) via multiomics analyses. Dextran sulfate sodium (DSS) was used to establish a UC model in mice, and a subgroup of colitis model mice was subjected to gypenosides intervention. The inflammatory manifestations of the model and drug groups were determined via histological and molecular experiments. In addition, tissue and blood samples were collected for transcriptome and metabolomics analysis, respectively. Immunohistochemistry was used to detect the key proteins involved in UC pathogenesis and drug intervention. By combining transcriptomics and metabolomics assays, we explored one of the possible mechanisms by which gypenosides ameliorate UC in mice. Gypenosides treatment significantly alleviated clinical symptoms, prevented colon shortening, and decreased the disease activity index (DAI) in an 3% DSS-induced UC mouse model. Transcriptome sequencing revealed that the intestinal stem genes Ascl2 and Lgr5 ranked forefront in terms of differential expression before and after intervention with gypenosides. The metabolomics results suggest that the tricarboxylic acid cycle (TCA) and amino acid metabolism (AM) were the main metabolic pathways associated with UC and gypenosides treatment. Taken together, these results suggest that gypenosides may affect the biological activity of stem cells by regulating the tricarboxylic acid cycle and glutamine metabolism, promoting the repair of the damaged mucosa. In this study, we demonstrated that gypenosides alleviate colitis, likely by regulating the expression of genes associated with stemness and modulating the tricarboxylic acid cycle and amino acid metabolism, in a DSS-induced colitis mouse model.</p>