Carbon tetrachloride as been shown previously to be metabolized to the trichloromethyl radical and to a novel oxygen-containing carbon dioxide anion radical in the perfused rat liver. The carbon dioxide anion radical adduct also was observed in urine following the intragastric administration of carbon tetrachloride or carbon bromo-trichloride and spin trap. The rate of formation of carbon dioxide anion radical adduct was decreased 2 - 3 fold following inhibition of cytochrome P-450-dependent mono-oxygenases by metyrapone (0.5 mM) and was increased about two-fold by induction of cytochrome P-450 by phenobarbital pretreatment. Toxicity of halocarbons in the perfused liver was assessed by measuring the release of lactate dehydrogenase (LDH) into the effluent perfusate in livers from phenobarbital-treated rats under conditions identical to those employed to detect radical adducts. Metabolism of halocarbons to the carbon dioxide anion radical adduct was 6-8 fold faster during perfusion with nitrogen-saturated rather than with oxygen-saturated perfusate. Concomitantly, liver damage detected from LDH release occurred much sooner during halocarbon infusion in the presence of nitrogen-saturated perfusate. A good correlation (r = -0.8) between the rate of formation of PBN/carbon dioxide anion radical and the time to onset of LDH release following halocarbon infusion was observed. Therefore, it is concluded that PBN/carbon dioxide anion is a useful marker for the free radical intermediates that are casually related to halocarbon-induced hepatotoxicity. The trichloromethyl radical and carbon dioxide anion - radical adducts also have been detected in the bile from anesthetized rats. Hypoxia or pretreatment with phenobarbital has been reported to enhance the hepatotoxicity of carbon tetrachloride in vivo; these treatments also produced an increase in the biliary concentration of the PBN/trichloromethyl radical adduct and in the trichloromethyl-derived PBN/carbon dioxide anion radical adduct as well. ESR analysis of bile from animals treated with free radical traps and other xenobiotics, such as ethanol, have proved useful in monitoring hepatic free radical-adduct formation in vivo.
