XOD inhibition peptides obtained via fermentation of Salmon by Bacillus amyloliquefaciens and evaluation of their anti-hyperuricemia mechanism in HK-2 cells

Hyperuricemia (HUA) is a metabolic disorder that can trigger gout and cause renal dysfunction. In this study, microbial fermentation with Bacillus amyloliquefaciens Y11 was employed to ferment salmon dark meat. Subsequently, novel peptides with high xanthine oxidase (XOD) inhibitory activity were derived from the fermented samples, purified, and identified. The hypouricemic effect of these peptides was evaluated using an in vitro HK-2 cell HUA model. Five fractions (G1–G5) were isolated via ultrafiltration and Sephadex G15 gel chromatography, and the fractions with higher XOD inhibition rates (G1 and G5) were combined to prepare a mixed fraction. Subsequently, 4122 peptides were identified from the mixed fraction using liquid chromatography with tandem mass spectrometry. Based on predictive bioactivity, toxicity, allergenicity, and solubility scores, five peptides and seven corresponding truncated peptides were selected for synthesis. Their XOD inhibitory activity was assessed, and molecular docking analysis was performed. Ultimately, GEADFMDYGR and its truncated peptide GEADF, which showed high XOD inhibitory activity in vitro (inhibition rates of 90.72 ± 2.85% and 94.06 ± 1.30%, respectively), were screened out. In the HK-2 HUA model, both peptides significantly reduced intracellular uric acid levels. Specifically, 0.5 mg/mL GEADFMDYGR and GEADF decreased the uric acid levels in these cells by 39.70 ± 8.75 and 27.43 ± 3.31 μmol/L, respectively. Further analyses revealed that GEADFMDYGR and GEADF are reversible mixed-type XOD inhibitors. These findings provide new insights and experimental evidence guiding the development of novel therapeutic peptides for HUA. • Novel XOD inhibitory peptides were obtained from salmon using Bacillus amyloliquefaciens fermentation. • GEADFMDYGR and its truncated peptide GEADF, which showed high XOD inhibitory activity in vitro. • 0.5 mg/mL GEADFMDYGR and GEADF decreased the uric acid levels in HUA cells by 39.70 ± 8.75 and 27.43 ± 3.31 μmol/L, respectively.