Continuous monitoring of hydroperoxide-induced peroxidation in human erythrocytes by low-level chemiluminescence

Abstract

We studied the effect of cumene hydroperoxide, t-butyl hydroperoxide, and hydrogen peroxide on intact healthy human erytyrocytes (15 g hemoglobin/dl) using chemiluminescence to monitor peroxidation. We measured the chemiluminescence spectrum, the process of hemolysis, the pH shift, and absorbance spectrum during the incubation with chemicals producing oxidative stress. Maximum chemiluminescence was reached with cumene hydroperoxide at about 50 min, but with t-butyl hydroperoxide at 100 min. The effect of organic hydroperoxide was concentration dependent, whereas the effect of hydrogen peroxide was independent of concentration. Peroxides induced hemolysis after 30 min. The pH shift to alkaline was observed in the first 20-min period. Incubation with organic hydroperoxides induced a decrease in absorption at 580, 545, and 345 nm. Hydrogen peroxide induced a decrease in the same period of time but this returned to the normal range by 120 min. There was no change in absorption at 420 nm with any of the peroxidative agents. Our results suggest that low-level chemiluminescence is a useful model for studying hydroperoxide-induced peroxidation in human erythrocytes.