GENERATION OF REACTIVE OXYGEN SPECIES BY CO(II) FROM H2O2 IN THE PRESENCE OF CHELATORS IN RELATION TO DNA-DAMAGE AND 2'-DEOXYGUANOSINE HYDROXYLATION

The generation of reactive oxygen species by Co(II) from H2O2 in the p resence of chelators and related DNA damage was investigated by electr on spin resonance (ESR), electrophoretic assays, and high-performance liquid chromatography (HPLC). Incubation of Co(II) with beta-alanyl-3- methyl-L-histidine (anserine) and H2O2 generated (OH)-O-. radicals. Om ission of any one component sharply reduced the amount of (OH)-O-. rad icals generated, indicating that anserine modulated the oxidation pote ntial of Co(II) to enhance its capability to generate (OH)-O-. radical s from H2O2. Formate only moderately decreased the (OH)-O-. radical ge neration, while ethanol had no observable effect, indicating that the generation of (OH)-O-. radical is site specific. The metal ion chelato r I, 10-phenanthroline reduced the (OH)-O-. radical generation, and de feroxamine suppressed it with the formation of deferoxamine nitroxide radical. Electrophoretic assays using both lambda Hind III linear DNA and PM2 supercoiled DNA showed that (OH)-O-. radicals generated from a mixture of Co(II), H2O2, and anserine caused DNA strand breaks. A mix ture of Co(II), H2O2, and I,10-phenanthroline also caused DNA strand b reaks, which were inhibited by sodium azide, indicating that O-1(2) wa s involved in DNA damage. HPLC measurements showed that (OH)-O-. radic als and O-1(2) generated by Co(II) reactions caused 2'-deoxyguanosine hydroxylation to form 8-hydroxy-2'-deoxyguanosine. ESR spin trapping m easurements provided evidence for O-1(2), generation by Co(II) from H2 O2 in the presence of I, 10-phenanthroline. The results indicate that the oxidation potential of Co(II) can be modulated by chelators to fac ilitate its generation of reactive oxygen species from H2O2. These spe cies may be involved in Co(II)-induced cellular damage.