Molecular Basis for the Divergent Inhibition of α-Amylase and α-Glucosidase by Phenolic Acids: The Critical Role of Hydroxyl Substitution

The global rise in metabolic disorders demands novel interventions targeting starch digestion. This study investigated two dietary phenolic acids (caffeic acid (CA) and p-hydroxycinnamic acid (p-HA)) as inhibitors of α-amylase and α-glucosidase through integrated experimental and computational approaches. Molecular docking showed distinct binding modes, and CA formed stable hydrogen bonds with catalytic residues of α-glucosidase, while p-HA interacted mainly with α-amylase via hydrophobic contacts. Enzyme kinetics revealed concentration-dependent mixed-type inhibition, with CA being more potent against α-glucosidase and p-HA against α-amylase. Spectroscopic analysis indicated both acids induced structural changes in the enzymes, with CA causing greater α-helix reduction (Δ7.03% vs. Δ2.10%) by altering the tryptophan microenvironment. Moreover, both compounds significantly suppress glucose absorption in the proximal small intestine in an ex vivo everted gut sac model, with p-HA exhibiting exceptional efficacy in the duodenum. These findings clarify structure–activity relationships and support the potential use of CA and p-HA as local intestinal agents for modulating carbohydrate absorption.