Characterization of a synergistic antioxidant synthetic peptide from sea cucumber and pine nut

Abstract

We compared antioxidant activity of the synthetic peptide Val-Leu-Leu-Tyr-Gln-Asp-His-Cys-His (VLLYQDHCH), sea cucumber peptide Val-Leu-Leu-Tyr (VLLY) and pine seed peptide Gln-Asp-His-Cys-His (QDHCH). The structure-activity relationship was analyzed based on radical scavenging ability and Raman, circular dichroism (CD) and nuclear magnetic resonance spectroscopy (NMR). Based on RP-HPLC, the contents of peptides in simulated gastrointestinal tract and digestive juices in rat intestinal sac were determined, and their absorption stability were explored. These results showed that the DPPH clearance rate of VLLYQDHCH was 45.90% higher than the sum of VLLY and QDHCH at 3 mmol/L. The α-helix, β-sheet and random coil of VLLYQDHCH increased, β-turn decreased, and the active hydrogen site shifted. After simulated digestion and absorption, the retention rate of VLLYQDHCH was 80.86 ± 0.88% in simulated stomach and 45.75 ± 0.97% in simulated intestine. There was no significant difference in the absorption rates of the three peptides (P > 0.05). This research provided a new idea for the development of safe and green food-derived animal-plant protein antioxidant peptides.

Keywords: Antioxidant activity; Gastrointestinal digestion; Pine nut peptide; Sea cucumber peptide; Secondary structure; Synthetic peptide.

Fig. 1

In vitro antioxidant activity determination of the VLLY, QDHCH and VLLYQDHCH. A The purity and identification of peptide VLLYQDHCH was confirmed according to the HPLC and MS spectra; B Effects of VLLY, QDHCH and VLLYQDHCH on DPPH radical scavenging capacity at three different concentrations; C Effects of VLLY and QDHCH and VLLYQDHCH on ABTS radical scavenging capacity at three different concentrations; D The EPR spectrum of peptides; E Hydroxyl radical scavenging rate of antioxidant peptides. The different letters (a–e) mean that the variance of two samples is significant (P < 0.05), # means compared to the control group, * means compared to the injury group (P < 0.05), a-d indicate significant differences in hydroxyl radicals among different components (P < 0.05)

Fig. 2

In vivo antioxidant activity determination of the VLLY, QDHCH and VLLYQDHCH. A Effect of antioxidant polypeptides on survival rate of HepG2 Cells; B Comparison of the ability of three peptides to scavenge intracellular oxygen free radicals; C Effects of three peptides on intracellular antioxidant enzyme SOD; D Effects of three peptides on intracellular antioxidant enzyme CAT. The different letters (a–e) mean that the variance of two samples is significant (P < 0.05), # means compared to the control group, * means compared to the injury group (P < 0.05), a-d indicate significant differences in hydroxyl radicals among different components (P < 0.05)

Fig. 3

A Raman spectra of VLLY, QDHCH and VLLYQDHCH between 200 and 3500 cm⁻¹; B CD spectra of VLLY, QDHCH and VLLYQDHCH secondary structures

Fig. 4

The investigation of active hydrogen atoms. A NMR of VLLYQDHCH. B NMR comparison of VLLY, QDHCH and VLLYQDHCH. The green line represents before adding D₂O and the red line means after adding D₂O in A. the orange line represents QDHCH, the cyan line represents VLLY, and the purple line represents VLLYQDHCH after adding D₂O in B

Fig. 5

Simulate the digestive liquid chromatogram and tandard curve. VLLY (A), QDHCH (C), and VLLYQDHCH (E) simulate the digestive liquid chromatogram and tandard curve of them, VLLY (B), QDHCH (D) and VLLYQDHCH (F) end simulation of digestive liquid chromatogram