Zidovudine (3′-azido-2′,3′-dideoxythymidine [AZT]) inhibits human immunodeficiency virus replication and delays progression of acquired immune deficiency syndrome. We have recently found that, in muscle, AZT causes oxidative damage to mitochondrial DNA (mtDNA) and other signs of mitochondrial oxidative damage. The aim of this work was to test if AZT causes oxidative damage to liver mtDNA. In our study, an experimental mouse model was used in which mice were administered AZT (10 mg/kg body weight/d) in drinking water. Liver mtDNA of mice treated with AZT had 40% more of the oxidized, mutagenic nucleoside, 8-oxo-7,8-dihydroxy-2′deoxyguanosine (8-oxo-dG) than untreated controls. This oxidative damage to mtDNA is caused by a significant increase (of over 240%) in peroxide production by liver mitochondria from AZT-treated mice, which was prevented by dietary administration with vitamins C and E.