A novel method of measuring hydroxyl radical-scavenging activity of antioxidants using gamma-irradiation

A new method using ESR spin trapping was proposed for measuring the scavenging activity of antioxidants for the hydroxyl (OH) radical. (-)-Epigallocatechin gallate (EGCg) and 5,5-dimethyl-1-pyrroline N-oxide (DMPO) were used as the antioxidant and spin trapping agent, respectively. The conventional method using a Fenton reaction had problems associated with the estimation of activity, because the antioxidant disturbs the system for generating OH radical by coordinating on Fe2+ and by consuming H2O2, besides scavenging the spin adduct (DMPO-OH). Intense gamma-irradiation was therefore used to generate OH radicals, and the intensity decrease in DMPO-OH after irradiation was followed to obtain the rate constant for the scavenging of DMPO-OH by EGCg. The intensities were extrapolated to zero time to estimate the quantity of DMPO-OH formed during gamma-irradiation. By using these values, the reaction rate constant between OH radical and EGCg was calculated as a ratio to that of DMPO. It was shown that this method is useful for comparing the OH radical-scavenging activity of various antioxidants.