Metabolism of [14C]phenol in the isolated perfused mouse liver.

ABSTRACT

A previous report from this laboratory focused on the metabolism of [14C]benzene (BZ) in the isolated, perfused, mouse liver (C. C. Hedli, et al., 1997, Toxicol. Appl. Pharmacol. 146, 60-68). Whereas administration of BZ to mice results in bone marrow depression (R. Snyder et al., 1993, Res. Commun. Chem. Pathol. Pharmacol. 20, 191-194), administration of phenol (P), the major metabolite of BZ, does not. It was, therefore, of interest to determine whether the metabolic fate of P produced during BZ metabolism differed from that of P metabolized in the absence of BZ. Mouse livers were perfused with a solution of [14C]P in both the orthograde (portal vein to central vein) and retrograde (central vein to portal vein) direction to investigate the metabolic zonation of enzymes involved in P hydroxylation and conjugation. Perfusate samples were collected, separated by HPLC, and tested for radioactivity. Unconjugated metabolites were identified by comparing their retention times with nonradiolabeled standards, which were detected by UV absorption. Conjugated metabolites were identified and collected on the basis of radiochromatogram results, hydrolyzed enzymatically, and identified by co-chromatography with unlabeled BZ metabolites. The objective was to compare and quantify the metabolites formed during the perfusion of P in the orthograde and retrograde directions and to compare the orthograde P-perfusion results with the orthograde BZ results reported previously. Regardless of the direction of P perfusion, the major compounds released from the liver were P. phenylgucuronide, phenylsulfate, hydroquinone (HQ), and HQ glucuronide. A comparison of the results of perfusing P in the orthograde versus the retrograde direction showed that more P was recovered unchanged and more HQ was formed during retrograde perfusion. The results suggest that enzymes involved in P hydroxylation are generally closer to the central vein than those involved in conjugation, and that during retrograde perfusion, P metabolism may be limited by the sub-optimal conditions of perfusion. Comparison of the orthograde perfusion studies of P and BZ revealed that a larger percentage of the radioactivity released from the liver was identified as unconjugated HQ after BZ perfusion than after P perfusion. In addition, the amount of radioactivity covalently bound to liver macromolecules was measured after each perfusion and determined to be proportional to the amount of HQ and HQG detected in the perfusate samples.