COX-2 Inhibition by Bioactive Peptides from Peanut Worm (Siphonosoma australe) Collagen Through In Vitro Digestion Simulation

Research background. Chronic, unregulated inflammation is a crucial factor in developing numerous diseases and is closely linked to the increased expression of cyclooxygenase-2 (COX-2). While various bioactive peptides from marine organisms have exhibited COX-2 inhibitory effects, peptides derived from the collagen of the peanut worm (Siphonosoma australe) have not yet been demonstrated. This study aimed to evaluate peanut worm collagen's potential COX-2 inhibitory activity through in vitro simulated digestion using pepsin-pancreatin followed by molecular docking. Experimental approach. During simulated in vitro digestion, commercial pepsin (at pH 3) and pancreatin (at pH 7.5) were applied for 240 min at 37 °C to evaluate the degree of hydrolysis, peptide concentration, and COX-2 inhibitory activity. Samples showing the most significant COX-2 inhibitory activity were subsequently separated into fractions and identified. Results and conclusions. The 210 min in vitro simulated digestion showed the highest COX-2 inhibitory activity (64.31 %). This finding was confirmed by the elevated degree of hydrolysis (DH) and peptide concentrations observed during the in vitro simulated digestion. The peptide fraction of <1 kDa exhibited the highest inhibitory activity (89.05 %), followed by peptide sequencing. Three novel peptides, ADIAGQAAQVLR, LNNEITTLR, and VGTVEK, were identified and contain crucial amino acids, confirming them as COX-2 inhibitors. VGTVEK has the most potent interaction, as shown by the lowest binding energy (−4.41 kcal/mol). The molecular docking revealed that VGTVEK (631.35 Da) binds to the active side of COX-2, forming hydrogen bonds with Gln178, Leu338, Ser339, Tyr371, Ile503, Phe504, Val509, and Ser516 and hydrophobic interactions with Met99, Val102, Val330, Ile331, Tyr334, Val335, Leu345, Trp373, Leu517, and Leu520. Other biological activities of the produced peptides included ACE inhibitors, DPP-IV inhibitors, and α-glucosidase inhibitors. According to the toxicity prediction, peptides have been classified as non-toxic. Novelty and scientific contribution. The study found that peptides generated from peanut worm collagen exhibit potential as novel, natural agents for anti-inflammatory therapy. Their broader application in functional foods, nutraceuticals, and pharmaceuticals could offer new options for individual sufferers of inflammation, supporting both treatment and overall health maintenance.