Flavonoids inhibit α-glucosidase and α-amylase, enzymes involved in carbohydrate digestion. This study compared the inhibitory activity of six flavonoid groups against yeast α-glucosidase, rat small intestinal α-glucosidase, and porcine pancreatic α-amylase. The structures of these flavonoids were evaluated to determine their inhibitory activity. Anthocyanidin, isoflavone, and flavonol groups were potent inhibitors of yeast α-glucosidase, with IC50 values less than 15 μM. Structures such as an unsaturated C ring, 3-OH, 4-CO, and hydroxyl substitution on the B ring enhanced inhibitory activity. In contrast, 3-OH reduced inhibitory activity. For rat small intestinal α-glucosidase, many flavonoids showed weak inhibition, with the anthocyanidin and isoflavone groups showing slightly higher activity. Hydroxylation at the 3 position and hydroxyl substitution on the B ring increased inhibitory activity. In porcine pancreatic α-amylase, luteolin, myricetin, and quercetin were potent inhibitors with IC50 values less than 500 μM. Structures such as the 2,3-double bond, 5-OH, and hydroxyl substitution on the B ring enhanced inhibitory activity, while 3-OH reduced it. The inhibitory activity of flavonoids was influenced by the structures of the A, B, and C rings. Hydroxylation at the 3 and 5 positions of flavone enhanced inhibitory activity. The linkage of the B ring at the 3 position enhanced inhibitory activity, while saturation of the 2,3-double bond in the C ring decreased it. The galloyl group at the 3 position of epigallocatechin increased inhibitory activity. The study also compared the inhibitory activity of flavonoids against different enzymes and found that the inhibitory activity varied depending on the structure of the flavonoid. The results suggest that the structures of the A, B, and C rings are closely related to the inhibitory activity of flavonoids against these enzymes. The findings have implications for the development of natural inhibitors for carbohydrate digestion and glucose absorption.Flavonoids inhibit α-glucosidase and α-amylase, enzymes involved in carbohydrate digestion. This study compared the inhibitory activity of six flavonoid groups against yeast α-glucosidase, rat small intestinal α-glucosidase, and porcine pancreatic α-amylase. The structures of these flavonoids were evaluated to determine their inhibitory activity. Anthocyanidin, isoflavone, and flavonol groups were potent inhibitors of yeast α-glucosidase, with IC50 values less than 15 μM. Structures such as an unsaturated C ring, 3-OH, 4-CO, and hydroxyl substitution on the B ring enhanced inhibitory activity. In contrast, 3-OH reduced inhibitory activity. For rat small intestinal α-glucosidase, many flavonoids showed weak inhibition, with the anthocyanidin and isoflavone groups showing slightly higher activity. Hydroxylation at the 3 position and hydroxyl substitution on the B ring increased inhibitory activity. In porcine pancreatic α-amylase, luteolin, myricetin, and quercetin were potent inhibitors with IC50 values less than 500 μM. Structures such as the 2,3-double bond, 5-OH, and hydroxyl substitution on the B ring enhanced inhibitory activity, while 3-OH reduced it. The inhibitory activity of flavonoids was influenced by the structures of the A, B, and C rings. Hydroxylation at the 3 and 5 positions of flavone enhanced inhibitory activity. The linkage of the B ring at the 3 position enhanced inhibitory activity, while saturation of the 2,3-double bond in the C ring decreased it. The galloyl group at the 3 position of epigallocatechin increased inhibitory activity. The study also compared the inhibitory activity of flavonoids against different enzymes and found that the inhibitory activity varied depending on the structure of the flavonoid. The results suggest that the structures of the A, B, and C rings are closely related to the inhibitory activity of flavonoids against these enzymes. The findings have implications for the development of natural inhibitors for carbohydrate digestion and glucose absorption.