PREDICTION OF IN-VIVO DISPOSITION FROM IN-VITRO SYSTEMS - CLEARANCE OF PHENYTOIN AND TOLBUTAMIDE USING RAT HEPATIC-MICROSOMAL AND HEPATOCYTE DATA

The kinetics of oxidation of phenytoin and tolbutamide were determined in freshly isolated hepatocytes and hepatic microsomes from male Spra gue-Dawley rats. Similar enzyme kinetic models are applicable to the d ata from both in vitro systems; a two-site model for phenytoin with a high affinity (K-m = 1-5 mu M, based on unbound drug concentration), l ow capacity site and a low affinity, high capacity site, and a one-sit e model for tolbutamide. Steady-state infusion studies were performed to characterize the Michaelis-Menten parameters for phenytoin disposit ion in vivo, these data could also be described by a two-site metaboli sm model (K-m 1.3 mu M, intrinsic clearance 62 ml/min for unbound drug for the high affinity site). Comparison of in vivo and in vitro param eters (after scaling the latter parameters for either hepatocyte yield or microsomal recovery) showed excellent prediction of in vivo cleara nce of unbound drug from hepatocyte data (55 ml/min) but underpredicti on from microsomal data (17 ml/min). In contrast to phenytoin, the in vivo clearance of tolbutamide (1.5 ml/min for unbound drug) was equall y well predicted by both hepatocyte (2.4 ml/min) and microsomal (3.1 m l/min) studies. The difference between the utility of in vitro systems to predict the in vivo clearance of these two drugs, which show simil ar pharmacokinetic properties (low clearance restricted to unbound dru g concentration in blood), may be a consequence of the particular term inal metabolite formed in each in vitro system. Whereas for tolbutamid e a similar terminal metabolite is formed in microsomes and hepatocyte s as occurs in vivo, for phenytoin the lack of an active conjugation s ystem in microsomes precludes the clearance of the primary hydroxy met abolite, a known end-product inhibitor. If this phenomenon can be gene ralized to other drugs that undergo sequential phase 2 metabolism, the n the use of microsomes for predicting the in vivo clearance would app ear to be limited.