Purification and identification of novel antioxidant peptides from watermelon seed protein hydrolysates and their cytoprotective effects on H 2 O 2 -induced oxidative stress.

The aim of this study was to purify and identify antioxidant peptides from watermelon seed protein hydrolysates (WSPHs-I: Mw < 1 kDa) and further evaluate their cytoprotective effects against H ₂ O ₂ -induced oxidative stress in HepG2 cells. After purification by Sephadex G-15 and semi-preparative reversed-phase high performance liquid chromatography (RP-HPLC), five peptides, RDPEER (P1), KELEEK (P2), DAAGRLQE (P3), LDDDGRL (P4), and GFAGDDAPRA (P5) were sequenced by LC-MS/MS and synthesized with solid-phase synthesis method. These peptides showed desirable 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging capacity (IC ₅₀ : 0.216 ± 0.01-0.435 ± 0.03), 2,2'-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) radical scavenging capacity (IC ₅₀ : 0.54 ± 0.02-1.23 ± 0.03), and oxygen radical absorbance capacity (ORAC) (82.36 ± 1.2-130.67 ± 2.2 μM TE/mg). Among them, peptide P1 exhibited the strongest antioxidant capacity. Moreover, the results suggested that peptide P1 may protect HepG2 cells from H ₂ O ₂ -induced oxidative damage by significantly inhibiting reactive oxygen species (ROS), [Ca ²⁺ ]i, malondialdehyde (MDA) levels and increasing antioxidative enzyme activities.
Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
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