Separation and Identification of a Novel Antioxidant and Calcium-Binding Peptide Derived from Phosvitin Hydrolysates: In Vitro and In Silico Prediction

This study aimed to identify a novel peptide from phosvitin hydrolysates with enhanced antioxidant and calcium absorption-promoting properties. Simultaneous enzymatic hydrolysates (trypsin + alkaline protease) exhibited better bifunctional activity than sequential enzymatic hydrolysates. Three components of phosvitin hydrolysates (PPP0, PPP1, and PPP2) were isolated by anion-exchange chromatography, and PPP2 displayed the highest activity. Molecular docking results revealed that AEFGTEPDAK, AEFGTEPDAKT, and KILDDTDNQ exhibited lower binding energies to Keap1 or TRPV6 compared with other PPP2-derived peptides, primarily via hydrogen bonding and hydrophobic interactions. Synthesized KILDDTDNQ demonstrated exceptional ABTS radical scavenging activity (80.21 ± 0.30%), lipid oxidation inhibition rate (79.41 ± 0.59%), 2-fold higher cellular antioxidant activity than glutathione, and calcium-binding capacity (140.87 ± 0.04 mg calcium/g peptide). Additionally, the results of the calcium content in Caco-2 cells revealed that KILDDTDNQ could greatly improve calcium absorption under oxidative stress conditions. These findings will aid in the development of calcium dietary calcium supplements.