High-performance liquid chromatographic assay of biphenyl metabolism by hepatocytes cultured in an embryo/hepatocyte co-culture medium

• Published in: Analytical biochemistry Vol. 175; no. 1; p. 74
• Main Authors: Mole, M L, Sanders, L, Oglesby, L A
• Format: Journal Article
• Language: English
• Published: United States 15-11-1988
• Subjects: Animals; Biphenyl Compounds - metabolism; Biphenyl Compounds - standards; Cells, Cultured; Chromatography, High Pressure Liquid - methods; Embryo, Mammalian; Liver - metabolism; Mixed Function Oxygenases - metabolism; Rabbits; Rats; Reference Standards
• ISSN: 0003-2697
• DOI: 10.1016/0003-2697(88)90363-6

A high-performance liquid chromatographic method has been modified for the evaluation of both Phase I and II metabolism of biphenyl by hepatocytes maintained in an embryo/hepatocyte co-culture medium. Extracts of the media, before and after hydrolysis of conjugates, are directly injected onto the HPLC and the major hydroxylated metabolites plus unmetabolized biphenyl are detected by fluorescence after separation under gradient or isocratic conditions. The method is almost free of interferences and is relatively simple and rapid. In the case of the monohydroxylated derivatives, the minimum media concentrations which can be measured are 7 to 20 nM (0.07 to 0.2 pmol on-column). Recoveries from culture medium to which known amounts of biphenyl and metabolites had been added were quantitative (90-103%) and the reproducibility good (interassay CV less than 5%). The assay was applied to cultures of hepatocytes derived from rabbit and from phenobarbital induced and noninduced rat.