Bioinformatic Identification and Experimental Validation of Novel Antioxidant Peptides Derived from Salmon Collagen

The objective of this study was to identify and experimentally validate novel antioxidant peptides derived from salmon collagen through an integrated bioinformatic and experimental workflow. Collagen sequences from Salmo salar were subjected to in silico enzymatic hydrolysis, and the resulting peptides were characterized using physicochemical descriptors. Principal component analysis and k-means clustering were then applied as exploratory tools to prioritize candidates displaying physicochemical proximity to previously reported antioxidant peptides. The selected candidates were chemically synthesized, purified, and evaluated at concentrations of 0.2 and 0.4 mM using the 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical-scavenging assay, the 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) radical-scavenging assay, and a cellular antioxidant activity assay, with vitamin C as a positive control. Cytotoxicity was assessed using the WST-1 assay. The peptide AS-5390 (DGCERCF) demonstrated superior radical scavenging capacity, with activity comparable to that of vitamin C in the DPPH assay at 0.2 mM and maintaining antioxidant activity in the ABTS assay at both tested concentrations. The peptide AS-5394 (PGDIG) exhibited the strongest cellular antioxidant response, reaching approximately 40% activity at 0.4 mM. Both peptides maintained high cell viability. Overall, this integrated strategy enabled the identification of salmon collagen-derived peptides with assay-dependent antioxidant activity and potential applications as functional food or nutraceutical ingredients.