Opuntia dillenii (Ker Gawl.) Haw, which has long been prized for its therapeutic virtues, has shown promise in treating hyperglycemic conditions. This study looks at the chemical composition and antihyperglycemic capabilities of aqueous extracts from O. dillenii's pulp and peel, as well as their effects on major carbohydrate metabolism enzymes. Significant changes in the composition of bioactive chemicals between pulp and peel were discovered using high-performance liquid chromatography with photodiode array detection (HPLC-DAD), with high amounts of p-coumaric acid, flavone, quercetin, and kaempferol. Key compounds included gallic acid, vanillic acid, p-coumaric acid, 3-hydroxy flavone, quercetin, cinnamic acid, kaempferol, and flavone. P-coumaric acid was highest in the pulp (298.71±0.43 mg/100g) and peel (38.18±1.08 mg/100g), while flavone was higher in the peel (120.03±0.26 mg/100g). In vitro enzyme inhibition tests showed that the extracts successfully inhibited pancreatic α-amylase, lipase, and intestine α-glucosidase. Molecular docking experiments confirmed the enzyme-binding affinity of these drugs, demonstrating interactions stronger than the conventional medication acarbose. In vivo testing on healthy and diabetic rats demonstrated the extracts' ability to lower blood glucose levels without harm, even at high doses (up to 3000 mg/kg). These findings indicate that O. dillenii pulp and peel extracts contain bioactive chemicals with promise as natural antidiabetic drugs, necessitating additional research for therapeutic applications.