Novel anti-oxidative peptides from equine hemoplasma protein hydrolysates: Purification, identification and protective effects on Caco-2 cells

Abstract

In this study, we purified and identified antioxidant peptides from equine plasma protein hydrolysates and assessed their protective effects against H₂O₂-induced oxidative stress in Caco-2 cells. Four antioxidant peptides were identified by liquid chromatography-tandem mass spectrometry (LC-MS/MS) in equine plasma protein hydrolysate, namely: GTMVGC (567.69 Da), FGMTST (662.88 Da), VGYHSHF (847.01 Da) and ALSPFFKE (939.18 Da). Among them, ALSPFFKE showed the strongest antidigestive properties after modelled digestion studies. Moreover, ALSPFFKE enhanced intracellular superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), and catalase (CAT) activities while significantly reducing reactive oxygen species accumulation and malondialdehyde formation in Caco-2 cells. The molecular docking analysis suggested that ALSPFFKE achieves regulation of the Keap1-Nrf2 pathway mainly by forming multiple hydrogen bonds and hydrophobic interactions with key amino acids (Arg380, Ser555, Gln530, Tyr334) in Keap1. These findings suggested that equine plasma peptides hold significant promise for the development of novel, potent, and stable antioxidant functional foods.