TOXICOLOGY AND APPLIED PHARMACOLOGY 93,8 1-88 (1988) Detection of CCI,-Induced Oxidation of Hepatic Tissue in Vivo by Oxygen-l 8 Tracing’ GARY E. HATCH,* JEFFREY SANTROCK,~” RALPH SLADE,* AND J. M. HAYES? *Toxicology Branch, Inhalation Toxicology Division, Health Efects Research Laboratory, Environmental Protection Agency, Research Triangle Park, North Carolina 2771 I, and 7 Department ofchemistry, Indiana University, Bloomington, Indiana 47405 Received September 2, 1987; accepted November 17, 1987 Detection of CC&-Induced Oxidation of Hepatic Tissue in Viva by Oxygen- 18 Tracing. HATCH, G. E., SANTRDCK, J., SLADE, R., AND HAYES, J. M. (1988). Toxicol. Appl. Pharmacol. 93, 8 l-88. Oxidation of rat hepatic tissue was measured as incorporation of oxygen-l 8 (“0) from “02 following exposure to carbon tetrachloride (Ccl.,). Anesthetized rats were injected with CC& and allowed to breathe ‘*02 for 1 hr, and then livers were homogenized, fractionated, and dried. The dried fractions were pyrolyzed to CO by an oxygen elemental analyzer, the CO was oxidized to CO*, and isotope ratio mass spectrometry was used to determine the abundance of ‘*O in the CO*. Rats that breathed “02 (2 1% in Nz) for 1 hr had significant incorporation of “0 into lipids, solutes, and macromolecules of the liver. Injection with CC4 increased incorpo- ration of “0 into all liver fractions, although this increase was significant only in the lipid frac- tion. Rats pretreated with phenobarbital and then given Ccl,, had significantly increased “0 in all liver fractions although it was greatest in the lipid fraction. About 5 pmol of “0 per gram of dry liver was incorporated in phenobarbital/CCL+-treated rats, of which 60% was in the water- soluble fraction, 17% in the lipids, and 16% in the macromolecules. Piperonyl butoxide adminis tration abolished the Ccl,-induced “0 incorporation. Thus, “0 incorporation appeared to pro- vide a measure of oxidation in all tissue fractions following in vivo CCL,-induced liver injury. 0 1988 Academic Press. Inc. Recent reviews of a rapidly expanding litera- ture on the effects of free radicals in tissue in- jury suggest that undesirable reactions of oxy- gen may be involved in diverse pathological processes including chemical toxicity, tumor promotion, atherosclerotic plaque forma- tion, radiation-induced injury, and normal ’ The research described in this article has been re- viewed by the Health Effects Research Laboratory, U. S. Environmental Protection Agency, and approved for publication. Approval does not signify that the con- tents necessarily reflect the views and policies of the agency nor does mention of trade names or commercial products constitute endorsement or recommendation for use. ’ Present address: Biomedical Science Department, General Motors Research Laboratory, Warren, MI 48090-9055. aging (Ames, 1983; Slater, 1984). Much of the evidence which implicates tissue autoxi- dation in disease causation has been derived from studies of chemical reaction mecha- nisms, from detection of oxidized lipids in affected tissues, and from inhibition of patho- logical effects by administration of free-radi- cal scavengers. While the above measure- ments have been useful, an increase in our understanding of tissue oxidation could probably be achieved if it were possible con- veniently to trace the oxygen atom in viva and measure oxidations as oxygen uptake or binding to tissue components. Positron emis- sion tomography has been used to determine the gross disposition of 150-labeled gases in the body (Bigler and Sgouros, 1983); how- 81 0041-008X/88 $3.00 Copyright 0 1988 by Academic Press, Inc. All rights of reproduction in any form reserved.