Drug metabolism

High-performance liquid chromatography

Chromatography

Metabolism

Sample preparation

Biochemistry

In vitro

Hepatocyte

Mass spectrometry

Chemistry

Liquid chromatography–mass spectrometry

Combinatorial Chemistry & High Throughput Screening

We have assessed the metabolic activity of freshly isolated and commercially preserved rat, monkey, and human primary hepatocytes in a 96 well plate format utilizing eight beta-adrenolytic drugs as model compounds. Sample introduction from 96 well plates, HPLC solvent delivery, mass spectrometric (MS) detection, and/or UV detection were fully integrated and operated unattended. After drugs were incubated with hepatocytes for three or six hr, LC-MS analyses were carried out to determine the amount of drug which was not metabolized. Two LC-MS methods were used which had a sample throughput of 4 samples/hr and 12 samples/hr. Under optimal conditions, this hepatic assay could screen 300 samples/week or 1200 samples/month. Although freshly isolated hepatocytes were more active, commercially available rat, monkey, and human primary isolated hepatocytes metabolized drug substrates in similar relative rank orders. This drug-hepatocyte assay provides useful information for prioritizing pharmaceutical leads in relative rank orders or in a high/low manner according to their resistance toward liver metabolism.

High Throughput Liquid Chromatography-Mass Spectrometry Assessment of the Metabolic Activity of Commercially Available Hepatocytes from 96-Well Plates