Identification and molecular mechanism of novel antioxidant peptides from okara fermentation broth with Bacillus amyloliquefaciens YP2: In silico and in vitro analysis

This study aimed to identify novel antioxidant peptides extracted from okara fermentation broth, and to investigate the molecular mechanisms using in silico and in vitro approaches. The results demonstrated that three novel peptides—CKLLL, FMAV and SVCC—were identified, among which CKLLL exhibited the strongest DPPH radical scavenging activity (IC 50  = 30.21 ± 4.27 μM). Simulated gastrointestinal digestion showed CKLLL had stable DPPH scavenging activity and improved ABTS and hydroxyl radical scavenging capacities. Furthermore, all three antioxidant peptides effectively protected HepG2 cells against H 2 O 2 -induced damage, with CKLLL>SVCC>FMAV ranking highest to lowest in terms of protective effect. Peptide and Keap1 protein docking results indicate that the -CDOCKER energy of CKLLL (89.13 kcal/mol) is higher than that of FMAV and SVCC (55.35 and 48.01 kcal/mol). These findings suggest that these three peptides, especially CKLLL, have the potential to serve as antioxidant peptides, and provide a more reliable screening method for developing natural plant-derived antioxidant peptides.