Development and Validation of a Sensitive and Specific LC-MS/MS Cocktail Assay for CYP450 Enzymes; Application to Study the Effect of Catechin on Rat Hepatic CYP Activity.

Development and Validation of a Sensitive and Specific LC-MS/MS Cocktail Assay for CYP450 Enzymes; Application to Study the Effect of Catechin on Rat Hepatic CYP Activity. Biomed Chromatogr. 2019 Dec 29;:e4789 Authors: Nguyen V, Espiritu M, Elbarbry F Abstract A sensitive and specific liquid chromatography tandem mass spectrometric (LC-MS/MS) method was developed and validated that enables the simultaneous quantification of Cytochrome P450 (CYP) enzymes probe substrates and their metabolites. These substrates (metabolites) were coumarin (7-hydroxycoumarin), Tolbutamide (4-hydroxytolbutamide), S-mephenytoin (4-hydroxymephenytoin), Dextromethorphan (Dextrophan), and Testosterone (6β-hydroxytestosterone) were utilized as markers for the activities of the major human CYP enzymes CYP2A6, 2C9, 2C19, 2D6, and 3A4, respectively. Analytes were separated on Kinetex C18 column (2.1 × 50 mm, 5 μm using a binary gradient mobile phase of 0.1% formic acid in water and 0.1% formic acid in acetonitrile. Metabolites were detected and quantified by the mass spectrometry using multiple reaction monitoring at m/z 163 →107.2 for 7-hydroxycoumarin, m/z 235 → 150.1 for 4-hydroxymephenytoin, m/z 287 →171 for 4-hydroxytolbutamide, m/z 258 →157.1 for dextrophan, m/z 305 → 269 for 6β-hydroxytestosterone, and m/z 237 → 194 for the internal standard. The assay shows good linearity over a range of 10-500 ng/mL with acceptable accuracy and precision...
Source: Biomedical Chromatography : BMC - Category: Biomedical Science Authors: Tags: Biomed Chromatogr Source Type: research