Mechanistic study of phenolic acid inhibition of α-amylase: combining enzyme assays, spectroscopy, and molecular docking

Type 2 diabetes (T2D) is a chronic metabolic disorder marked by persistent hyperglycemia and insulin resistance. Inhibiting α-amylase activity is a key strategy to mitigate postprandial hyperglycemia. This study investigated the inhibitory effects of structurally distinct phenolic acids—benzoic acid-type and cinnamic acid-type on α-amylase. Among them, p-hydroxybenzoic acid (PHBA) and ferulic acid (FA) showed superior inhibitory activity. PHBA (IC₅₀ = 3.08 μg/mL) demonstrated stronger inhibition than FA (76.73 μg/mL) and acarbose (16.7 μg/mL). Kinetic analysis revealed mixed-type inhibition dominated by competitive binding. Spectroscopic analyses (UV, fluorescence, CD, FT-IR) indicated that PHBA more effectively altered enzyme conformation through hydrogen bonding. Molecular docking and dynamics simulations confirmed PHBA's stronger binding affinity (−4.94 kcal/mol > −4.34 kcal/mol for FA) and greater interaction stability at the α-amylase active site. These findings provide mechanistic insight into structure–activity relationships of phenolic acids and support their potential in the development of functional foods for glycemic control. • The inhibition mechanism of phenolic acids on α-amylase was systematically investigated. • Both benzoic acid- and cinnamic acid-type phenolics inhibit α-amylase; benzoic acid-type is stronger. • Spectroscopy showed PHBA enhances α-amylase binding and conformation. • Docking and dynamics confirmed PHBA forms more stable interactions than FA. • The findings support the application of phenolic acids in functional food development.