The role of 8-hydroxy-2'-deoxyguanosine in rifamycin-induced DNA damage.

The effect of rifamycin SV on the formation of 8-hydroxy-2'-deoxyguanosine (8-0HdG) has been investigated in vitro and in vivo. Oxidative modification of 2'-deoxyguanosine has been measured as an indication of DNA damage using high-performance liquid chromatography with electrochemical detection. Rifamycin SV in the presence of copper(II) ions induces the formation of 8-0HdG in calf thymus DNA. The effect is enhanced by increasing the antibiotic concentration and inhibited by catalase and hydroxyl radical (.0H) scavengers, such as thiourea and ethanol, in a rifamycin SV concentration-dependent manner. The reduced glutathione (GSH) inhibits DNA damage, and this effect is proportional to the final concentration of the tripeptide in the incubation medium. A significant increase in the formation of 8-0HdG and of malondialdehyde (MDA) in rat liver DNA was observed only in GSH-depleted animals after 5 days of rifamycin SV treatment. These results support the involvement of hydrogen peroxide (H2(0)2) and .0H in the mechanism of the oxidative modification of DNA achieved by rifamycin SV. The role of other reactive species and the antioxidant properties of GSH against oxidative damage is also discussed.

The role of glutathione in protection against DNA damage induced by rifamycin SV and copper(II) ions.

Incubation of calf thymus DNA in the presence of rifamycin SV induces a decrease in the absorbance of DNA at 260 nm. The effect, was found to be proportional to the antibiotic concentration and enhanced by copper(II) ions. In the presence of rifamycin SV and copper(II), a significant increase in thiobarbituric acid-reactive (TBA-reactive) material is also observed. This effect is inhibited to different degrees by the following antioxidants: catalase 77%; thiourea 72%; glutathione (GSH) 62%; ethanol 52%; and DMSO 34%, suggesting that both hydrogen peroxide (H2O2) and hydroxyl radicals (OH.) are involved in DNA damage. Rifamycin SV-copper(II) mixtures were also found to induce the production of peroxidation material from deoxyribose and, in this case, glutathione and ethanol were the most effective antioxidant substrates with inhibition rates of 91% and 88% respectively. Electrophoretic studies show that calf thymus DNA becomes damaged after 20 min. incubation in the presence of both agents together and that the damaged fragments run with migration rates similar to those obtained by the metal chelating agent 1,10-phenanthroline. Normal DNA electrophoretic pattern was found to be preserved by catalase, and GSH at physiological concentrations and by thiourea. No protection is observed in the presence of ethanol or DMSO. The results obtained indicate the involvement of different reactive species in the degradation process of DNA due to rifamycin SV-copper(II) complex and emphasize the role of reduced glutathione as an oxygen free radical scavenger.