Ozone Degrades into Hydroxyl Radical under Physiological Conditions : A Spin Trapping Study

Defining the reactants is a critical step towards elucidating the mechanism of ozone toxicity to biomembranes. To document ozone-induced HO.radicals, the spin trap 5,5-dimethyl-1-pyrroline-N-oxide was used and the resulting spin adduct was monitored with electron spin resonance spectroscopy. Chelexed potassium phosphate buffer (10 millimolar and 0.2 molar) at pH 7.2 and 7.8 was exposed to ozone (1-40 microliters per liter) by directing a stream of ozone over the surface for 60 seconds. Under these conditions, no HO. was detected. Using 0.5 x 10(-4) molar caffeic acid in phosphate buffer, strong DMPO.OH electron spin resonance signals were obtained, indicating HO. production. Air controls yielded no signal. High pH (7.8) enhanced signal strength. Furthermore, with sorbitol (0.4 osmolal final concentration), a net HO. signal loss of 28% was observed, while a carbon-centered sorbitol radical adduct appeared. Although HO. radicals were produced, no breakage of Daucus carota protoplast plasma membranes was observed nor were differences in membrane fluidity observed as determined by 5-doxyl stearic acid.