CYP2C19-mediated (S)-mephenytoin 4'-hydroxylation assayed by high-performance liquid chromatography with radiometric detection.
A reverse-phase, high-performance liquid chromatography method is described for the quantification of 4'-hydroxymephenytoin formed enzymatically from 14C-labeled (S)-mephenytoin following incubation with cDNA-expressed CYP2C19 or human liver microsomes. Analytical separation is achieved using a C18 column developed with a gradient from 10 to 100% methanol, with detection using a scintillation detector. This method is applicable to enzymatic studies for determination of CYP2C19-catalyzed (S)-mephenytoin 4'-hydroxylation activity.
Crespi
Charles L
CL
Discovery Labware, BD Biosciences, Woburn, MA, USA.
Chang
Thomas K H
TK
Waxman
David J
DJ
D001189:Aryl Hydrocarbon Hydroxylases / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D065731:Cytochrome P-450 CYP2C19; D006801:Humans; D008617:Mephenytoin / Q000378:metabolism*; D008862:Microsomes, Liver / Q000201:enzymology; D006899:Mixed Function Oxygenases / Q000378:metabolism*; D011874:Radiometry / Q000379:methods*
D006899:Mixed Function Oxygenases; D001189:Aryl Hydrocarbon Hydroxylases; C045793:CYP2C19 protein, human; D065731:Cytochrome P-450 CYP2C19; D008617:Mephenytoin