Uptake and intracellular binding of lipophilic amine drugs by isolated rat hepatocytes and implications for prediction of in vivo metabolic clearance.

The hepatic uptake of imipramine and propranolol was investigated after incubation with isolated rat hepatocytes over a wide concentration range (0.04-400 microM). The cell-to-medium concentration ratio (K(p)) was concentration-dependent and could be described using a two-site binding model incorporating a high affinity/low capacity site and a linear component for a site which was apparently not saturated. Maximum (at 0.04 microM) and minimum K(p) values (at 400 microM) were 360 and 280, and 110 and 70 for imipramine and propranolol, respectively. During these incubations, metabolism was inhibited using aminobenzotriazole (an irreversible inhibitor of cytochrome P450). Pretreatment of cells either by freeze-thawing or with saponin (which permeabilizes the plasma membrane) eliminated the saturable process. The saturable uptake process of imipramine was also inhibited by 18 other lipophilic amine drugs (including propranolol). This uptake component may involve membrane transporter(s), whereas the nonsaturable component probably represents partition into the phospholipid component of membranes. Propranolol was further investigated to determine the impact of high K(p) values on hepatocellular clearance. The area under the curve for propranolol concentrations in the total incubate (medium including the cells) from the depletiontime profile was substantially greater than the corresponding area under the curve for the drug concentration in the extracellular medium, and this difference approximated the nonsaturable uptake component. It is concluded that the clearance of propranolol in isolated hepatocytes is not rate-limited by hepatic uptake and is directly proportional to unbound drug concentration, being independent of the higher K(p) value.