20(S/R)-ginsenoside Rh1 is a less-studied ginsenoside. This study investigated its role in restoring glycaemic homeostasis via the gut microbiota–bile acid axis in type 2 diabetes mellitus (T2DM). The antidiabetic mechanism was evaluated in a high-fat diet/streptozotocin (HFD/STZ)-induced T2DM mouse model treated with 20(S/R)-ginsenoside Rh1 (60 mg/kg/day) for 4 weeks. Fasting blood glucose, oral glucose tolerance test (OGTT), serum insulin and glucagon-like peptide-1 (GLP-1) levels, and lipid profiles were assessed. Gut microbiota composition was analyzed by 16S rRNA sequencing, and targeted metabolomics measured bile acid profiles. Western blotting, gene expression, and enzyme activity assays evaluated the effects of 20(S/R)-Ginsenoside Rh1 on hepatic bile acid synthesis enzymes, microbial bile salt hydrolase (BSH), and ileal FXR/TGR5 signaling pathways. 20(S/R)-Ginsenoside Rh1 treatment significantly improved glucose and lipid metabolism and was associated with increased hepatic expression of CYP27A1 and CYP7B1, the rate-limiting enzymes of the alternative bile acid synthesis pathway. It reshaped the gut microbiota, altering BSH and bile acid-inducible ( bai ) gene expression, which increased ileal accumulation of tauro-β-muricholic acid (T-β-MCA) and lithocholic acid (LCA). T-β-MCA suppressed farnesoid X receptor (FXR), while LCA activated Takeda G protein-coupled receptor 5 (TGR5). Consequently, FXR inhibition and TGR5 activation modulated the CREB (cAMP response element binding protein)/cAMP (cyclic AMP) pathway, upregulated GCG (proglucagon) and PCSK1 (prohormone convertase 1), promoted L -cell differentiation, and enhanced GLP-1 secretion. Thus, 20(S/R)-ginsenoside Rh1 alleviates T2DM by modulating microbiota–mediated BA metabolism and enhancing GLP-1 release via FXR/TGR5 signaling.