Antioxidant Activity Of Oat Peptides In Hepatic HepG2 cells

The health benefits of oats have long been established, with their protective activities against cardiovascular diseases and blood sugar being the most prevalent, although the protective effect against cancer has been demonstrated as well.In all these conditions, antioxidants and fibres present in oats are believed to be the most beneficial molecules.Recently however, it has been demonstrated that hydrolyzed proteins and their fractions have biological properties beyond their basic nutritional function.Many studies have been performed to demonstrate the antioxidant activities of hydrolyzed proteins and peptides from cereals such as corn, wheat, and barley using in vitro chemical assays.Two studies have also demonstrated the antioxidant properties of hydrolyzed oat proteins in chemical-based assays.Oat protein hydrolysates produced using different enzymes were analyzed.From the oat protein hydrolysate fraction with highest radical scavenging, which was produced using viscozyme and alcalase, four peptides were identified.These peptides include GQTVFNDRLRQGQLL (P4), YHNAPGLVYIL (P6), and DVNNNANQLEPR (P7), as well as their fragments GQTV (P3), FNDRLRQGQLL (P1), YHNAP (P5), and GLVYIL (P2).The purpose of the current study was to establish the antioxidant activities of these peptides using chemical and cell culture based assays.The current research found that all seven peptides exhibited radical scavenging activity in ORAC assay, with P2 (0.67 µM Trolox equivalent (TE)/µM peptide), P5 (0.61 µM TE/µM peptide), and P3 (0.52 µM TE/µM peptide) having the highest activities, followed by P6, P4, P1, and P7 (0.14-0.37 µM TE/µM peptide.The cytoprotective effect of the peptides was determined using the human hepatoma HepG2 cell model.Oxidative stress was i du ed i HepG ells , ′-azobis (2-methylpropionamidine) dihydrochloride (AAPH), and ii protective activities of peptides against AAPH including cell death, alteration of the antioxidant enzymes glutathione peroxidase (GPx), superoxide dismutase (SOD), and catalase (CAT), change in total glutathione and levels of reactive oxygen species (ROS).Two different concentrations, 50 and 100 µM of each peptide were used to establish whether the effects were dosedependent.None of the peptides showed cytotoxicity.P2 increased cell viability by approximately 5-fold at both concentrations.However, P1 and P7 at 50 µM as well as P3, P4, and P5 at both 50 and 100 µM did not protect cells from AAPH-induced death.On the other hand, P1 and P7 at 100 µM, as well as P2 and P6 at both concentrations enhanced cell survival to between 65 -143%, with P2 increasing cell survival passed that of untreated cells.Treatment with AAPH decreased activities of GPx, SOD, and total glutathione by 18%, 29%, and 48%, respectively.Meanwhile AAPH treatment increased the CAT activity by almost 2-fold and ROS by 36.9%.For both GPx and glutathione, treatments with P1, P2, and P6 at 50 and 100 µM as well as P7 at 50 µM only significantly inhibited the effect of AAPH.In the SOD assay, only P2 (50 and 100 µM), and P1 (100 µM) attenuated the decrease of SOD activity by AAPH.P2 and P7 brought the activity of SOD back to that of untreated cells (negative control, NEG).The activity catalase, which increased after AAPH treatment, was further increased in the presence of P1, P2, P6, and P7 at both concentrations.The decrease SOD and GPx activities of HepG2 cells after treatment with AAPH were associated to the increase of intracellular ROS.The protection provided by peptides (P1, P2, P6, and P7) can then be attributed to their radical scavenging activity and to their potential to increase GSH levels.In most cases, the activity of the peptides correlated with their hydrophobicity.iii DEDICATION This thesis is dedicated to my father, mother, and my sister Sylvia.For the completion of this work would not have been possible without their unconditional love, support, and encouragement.providing expertise, valuable guidance, patience and encouragement to me.

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