Study On Antioxidant Capacity Of Peptides Designed By GSH

The antioxidant peptides are nutrient as well as safety, so they are paid attention to formany years. The researches are about their application on food and medicine. Nowadays,researchers concentrated in the separation and purification of antioxidant peptides from thevarious types of protein hydrolysates, but rarely in relationship between the antioxidantpeptide structure and activity. Glutathione(GSH, Glu-Cys-Gly) is a well-knownmultifunctional antioxidant that exists in animals and plants to defend against oxidative stress,It has been demonstrated that extracellular supplementation of GSH in culture mediumsignificantly provided useful radical scavengers to abolished all forms of oxidant-mediatedcell death. So based on the structure of glutathione, we synthetized eleven peptides, at thesame time, partial least squares method was applied to build the quantitative structure-activityrelationship. Additionally, we measured the protection of PC12cells from hydrogen peroxideinduced cytotoxicity by these antioxidant peptides.For the HAT-based assays, it was Tyr-Glu-Gly that showed extraordinary oxygen radicalabsorption capacity (ORAC value=1.20μmol Trolox/μmol sample) and ABTS free radicalscavenging ability (850.18μmol Trolox/mmol sample). For SET-based assay, Glu-Cys-Hisdisplayed the highest DPPH radical scavenging activity (IC500.11mM) and the strongestreducing power (61.23μmol Trolox/mmol sample). The peptides which contain Cys PhePro His, demonstrated strong DPPH radical scavenging activity and reducing power. Takenfive chemical antioxidant capacities as dependent variable and the structure parameters ofpeptides as independent variable, the equations were built. Models were estimated usingpartial least squares regression and validated through full cross-validation and externalvalidation(R2=0.9130, R2ext=0.6074). The results found an important relationship between thephysicochemical properties of the N-terminal regions and antioxidant potency. TheN-terminal amino acid should be a high hydrophobic and low Steric Property (such as Val,Pro and Leu).Cellular antioxidant activity (CAA) assay is a new approach quantify antioxidant activityat the physiological conditions, so we applied a new method to investigate the cellularantioxidant activity of synthetic peptides. The detail of the method was as following: Rat pheochromocytoma cell line (PC12cells) was selected as model, and DCFH-DA (finalconcentration25μM) and AAPH (final concentration600μM) was applied for assays offluorescence intensity. In our study, the order of cellular antioxidant activity wasGlu-Cys-His>GSH>Pro-Cys-Gly>Phe-Lys-Leu>Tyr-Glu-Gly. The IC50values of Glu-Cys-Hiswas0.56mM. The result of correlation demonstrated that CAA activities were significantlycorrelated to DPPH radical scavenging activities and reducing power, and the assays based onsame mechanism were significantly correlated.In our study, it was found that extracellular supplementation of the synthetic peptides inculture media significantly abolished the H2O2-induced cell death. The viability of PC12cellsdecreased to21%as caused by the oxidation of700μM H2O2. Among the peptides,Phe-Lys-Leu demonstrated the best ability to resist H2O2-induced cell injuries at theconcentration of3mM by increasing for50%. As for the protection mechanism, it mayinduce by altering the antioxidant enzymes expression in cells, not for enable of MARK.