Validation of Methods for in Vitro - in Vivo Extrapolation Using Hepatic Clearance Measurements in Isolated Perfused Fish Livers.

In vitro biotransformation assays using hepatocytes or liver subcellular fractions, combined with in vitro-in vivo extrapolation (IVIVE) models, have been proposed as an alternative to live fish bioconcentration studies. The uncertainty associated with IVIVE approaches to date has been attributed to assay protocols, model assumptions, or variability of in vivo data. An isolated perfused trout liver model that measures hepatic clearance has been proposed for validating IVIVE predictions in the absence of other confounding factors. Here, we investigated the hepatic clearances of five chemicals (pyrene, phenanthrene, 4- n -nonlyphenol, deltamethrin, and methoxychlor) in this model and compared measured rates to values predicted from published in vitro intrinsic clearances for validation of IVIVE models. Additionally, we varied protein concentrations in perfusates to test binding assumptions of these models. We found that measured and predicted hepatic clearances were in very good agreement (root mean squared error 16.8 mL h -1 g -1 ) across three levels of protein binding and across a more diverse chemical space than previously studied within this system. Our results show that current IVIVE methods can reliably predict in vivo clearance rates and indicate that discrepancies from measured bioconcentration factors might be driven by other processes, such as extrahepatic biotransformation, etc., and help streamline optimization efforts to the processes that truly matter.