LAUSR

This study aimed at isolating and identifying antioxidative peptides from Agaricus bisporus protein hydrolysate ultrafiltration fraction. Pancreatin was used to hydrolyze A. bisporus protein isolate and ultrafiltration was used to obtain 1–3 kDa fraction. Through consecutive chromatographic purification, two antioxidative fractions (C and D) were isolated. Pretreatment of HepG2 and Caco-2 cells with the peptide fractions prior to H2O2-induced stress exposure exhibited excellent intracellular ROS scavenging ability (178.3 ± 6.90% and 186.81 ± 14.32% at 20 µg/mL of C and D, respectively, compared to 480.02 ± 7.69% and 378.05 ± 6.77% of H2O2-stressed HepG2 and Caco-2 cells, respectively). The peptide fractions also preserved cell membrane integrity by decreasing lactate dehydrogenase (LDH) leakage and suppressing malondialdehyde (MDA) increase. Superoxide dismutase (SOD) and catalase (CAT) activities and reduced glutathione (GSH) levels were increased in the protected cells compared to the H2O2-damaged cells. Nano-LC-ESI-MS/MS and de novo sequencing identified eight peptide sequences rich in hydrophobic amino acids. These cytoprotective effects against H2O2-induced oxidative stress suggested that the peptide fractions may be considered as potential natural antioxidants for use in functional foods formulation as a strategy for delaying onset and progression of chronic diseases associated with oxidative stress.