Inhibitory Effects of Grape Seed Extract and Epicatechin on Malondialdehyde-Mediated Protein Lipoxidation

Malondialdehyde (MDA) is one of the principal and most studied reactive carbonyl species derived from polyunsaturated fatty acid peroxidation. This aldehyde is a chemically reactive molecule and should not be considered merely as a marker of lipid peroxidation. Its interaction with proteins, commonly termed protein lipoxidation, modifies protein structure and functions, thus extending the hazards of lipid peroxidation. In this study, a simplified model mimicking MDA-mediated protein oxidation in bovine serum albumin (BSA) was established to simulate protein lipoxidation in food systems. Grape seed extract (GSE, 50-200 mg/L) and epicatechin (EC, 145-580 mg/L) significantly inhibited MDA-mediated protein lipoxidation in BSA, as evidenced by a marked decrease in lipofuscin-like fluorescence and confirmed via Western blot analysis of MDA-protein adduct formation. Nevertheless, concentration-dependent dual effects were observed by protein conformational changes (reduced sulfhydryl, surface hydrophobicity, and altered microstructure) at high concentrations of GSE and EC, although they exhibited potent inhibitory effects in MDA-mediated protein lipoxidation. HPLC quantification demonstrated that GSE scavenged 10-60% of MDA while EC scavenged 70-95%, indicating their direct contribution to MDA removal. Molecular docking simulations further implied a possible binding mode in which EC associates with BSA more favorably (lower free energy) than MDA, which may competitively block the sites accessible to MDA. Collectively, these findings support a mitigation strategy in which phenolic nucleophiles can be employed to counteract MDA-mediated protein lipoxidation.