Abstract: This study investigated the therapeutic mechanism of oyster polysaccharide (OPS) on type 2 diabetes mellitus (T2DM). A T2DM mouse model was established through high-fat diet combined with intraperitoneal injection of streptozotocin (STZ). The experimental mice were divided into six groups: normal control (NC), model control (MC), w-dose OPS (OPS-L), medium-dose OPS (OPS-M), high-dose OPS (OPS-H), and positive control (metformin, MET). Comprehensive evaluations were conducted, including blood glucose and lipid profiles, hepatic and renal histopathology, 16S rDNA-based gut microbiota analysis, and untargeted fecal metabolomics. Compared to the MC group, the OPS-L and OPS-M groups exhibited significant reductions in fasting blood glucose (FBG), fasting serum insulin (FINS), and homeostatic model assessment of insulin resistance (HOMA-IR), along with increased body weight. Serum levels of triglycerides (TG), total cholesterol (TC), and low-density lipoprotein cholesterol (LDL-C) levels were reduced to varying degrees, with alleviated hepatic and renal tissue damage. Notably, the OPS-M group demonstrated the most pronounced hypoglycemic effects. OPS treatment modulated gut microbiota composition in T2DM mice, restoring α- and β-diversity indices toward NC group levels. Serum metabolomics revealed alleviated metabolic pathways involving histidine metabolism, alpha-linolenic acid metabolism, and ketone body synthesis/degradation. Correlation analysis identified significant associations between differential bile acid metabolites and specific bacterial taxa, particularly g__Muribaculaceae and g__Rikenellaceae_RC9_gut_group. This study indicates OPS ameliorates T2DM in mice through mechanisms potentially linked to gut microbiota modulation, fecal metabolite regulation, and alterations in serum bile acid profiles.