Benzene metabolism in the isolated perfused mouse liver

The hematotoxicity of benzene (BZ) requires its hepatic metabolism, the release of metabolites into the circulation, and the access of metabolites to the bone marrow. Although a range of potentially toxic metabolites produced by the liver was identified using subcellular systems and isolated hepatocytes, these models do not allow identification of the metabolites released from the liver with respect to time and flow through the liver. We developed an isolated perfused mouse liver model to evaluate metabolites released following a single-pass of radiolabeled BZ and after recirculation of single-pass metabolites back through the liver. Reversing the path of flow through the liver changes the orientation of hepatic oxidizing and conjugating enzymes with respect to perfusate flow. Comparison of metabolite production following normal (orthograde, portal vein to hepatic vein) perfusion with reversed (retrograde) perfusion permitted an evaluation of the impact of zonal distributions of these enzymes on BZ metabolism. The major metabolites detected by HPLC, irrespective of the direction of perfusion, were free phenol (P), phenylsulfate (PS), and phenylglucuronide (PG), plus lesser amounts of hydroquinone (HQ) and hydroquinone glucuronide (HQG). Recirculation of the products of single pass orthograde perfusion through the liver yielded P conjugates as well as low levels of free and conjugated HQ. No free P was detected after recirculation. Although no qualitative differences between orthograde and retrograde perfusion were observed, the percentage of free P and P conjugates (PS + PG) found as free P was twice as great following orthograde perfusion as compared to retrograde perfusion. These results suggest that regional differences in the zonation of enzymes involved in oxidation and conjugation may play a critical role in hepatic BZ metabolism.