Abstract: Objective: We optimized the sulfation modification process of Moutan cortex polysaccharide (MCP) and systematically evaluated the structural characteristics, antioxidant activity, and hypoglycemic potency of sulfated MCP (S-MCP). Methods: MCP were sulfated using concentrated sulfuric acid method. The process parameters were optimized using single-factor experiments, response surface methodology, and using degree of substitution (DS) as an index. S-MCP structure was characterized by FTIR, DLS, and SEM. Its antioxidant and hypoglycemic activities were evaluated in vitro and in vivo. Results: Optimal sulfation conditions for MCP included 39 min esterification time, 1.5 mL concentrated sulfuric acid, and 30 mg of ammonium sulfate. Under these conditions, the DS of S-MCP was 0.07391±0.0020. Structural characterization confirmed successful introduction of sulfate groups onto S-MCP. The S-MCP particle size (294.44±40.62 nm) was smaller than that of MCP (356.32±26.07 nm), and its microscopic characteristics changed from loose and porous to fragmented and layered. In vitro activity experiments showed respective scavenging rates of S-MCP against DPPH, ABTS⁺, and hydroxyl radicals of 64.85%, 82.42%, and 71.34% at 3.0 mg/mL significantly higher than those of MCP. Scavenging was also shown to be concentration-dependent. S-MCP displayed more potent inhibition of α-glucosidase (43.92% at 1.0 mg/mL) than MCP (31.31%). The mouse glucose tolerance test results showed that S-MCP significantly reduced the blood glucose levels after starch and sucrose loading, and decreased the area under the curve. Conclusion: Sulfation can effectively enhance MCP antioxidant and hypoglycemic activities, providing a scientific basis for their development and application in functional foods and pharmaceuticals.