Synergistic Orchestration of Chlorogenic Acid Analogs in Sugarcane Molasses Polyphenols for Sucrase Inhibition: Insights into a Multilevel Inhibitory Mechanism.

This study systematically explored the inhibitory mechanisms against sucrase exerted by sugarcane molasses polyphenol extract (SMPE) and its three principal chlorogenic acid analogs: neochlorogenic acid (NA), chlorogenic acid (CA), and methyl chlorogenate (MC). By integrating in vitro assays, molecular simulations, and spectroscopic analyses, we demonstrated synergistic inhibition among the components. SMPE exhibited the greatest inhibitory potency (IC50 = 3.58 ± 0.16 mg/mL), with an IC50 significantly lower than those of the individual compounds. Synergy analysis indicated marked synergistic effects for both the CA-NA pair and the ternary mixture. Simultaneous multi-ligand docking further suggested complementary co-binding configurations and interactions with residues surrounding the catalytic region, providing a possible structural interpretation of the experimentally observed synergistic effects. Mechanistically, CA and NA exhibited competitive inhibition, whereas MC and SMPE showed mixed-type inhibition; fluorescence, CD, and thermal stability analyses revealed compound-dependent changes in aromatic residue microenvironments, secondary structure, and protein stability, while molecular modeling predicted peripheral binding of CA/NA and catalytic-region interactions of MC, suggesting distinct but complementary routes for perturbing substrate recognition and catalysis. This study provides mechanistic insight into the synergistic inhibition of sucrase by SMPE and supports its potential development as a natural agent for modulating postprandial glycemia.