title: Development and validation of a simple LC method for the determination of phenacetin, coumarin, tolbutamide, chlorzoxazone, testosterone and their metabolites as markers of cytochromes 1A2, 2A6, 2C11, 2E1 and 3A2 in rat microsomal medium.
abstract: Cytochrome P450 enzymes are responsible for the oxidative metabolism of most pharmaceutical compounds. A "cocktail" approach which employs simultaneous administration of a mixture of substrates of CYP enzymes was often used to assess the metabolic activity of multiple P450 forms in one experiment. Phenacetin, coumarin, tolbutamide, chlorzoxazone and testosterone are commonly used as probe substrates to evaluate cytochrome P450 function. An analytical strategy to simultaneously extract and analyze the five probe substrates and their major metabolites by HPLC-DAD was developed. The incubation was done with all the substrates in one step. The ten analytes were extracted simultaneously by solid-phase extraction (SPE) from rat liver microsomes. A C18 analytical column and mobile phase composed of acetonitrile and 0.02% aqueous phosphoric acid were used for the chromatographic separation with DAD detection. Limits of quantification varied between 0.02378 and 0.2361 microg/mL which contributed to quantify all these drugs and metabolites with UV detection. The method is applicable for the modeling and description of pharmacological interactions on rat cytochromes P450 or can be used for in vitro evaluation of cytochromes 1A2, 2A6, 2C11, 2E1 and 3A2.
authors: Department of Pharmacy, Union Hospital, Huazhong University of Science and Technology, Wuhan, China.
authors: Xuejia
authors: X
authors: Zhai
authors: Yongning
authors: Y
authors: Lu
mesh_terms: D018712:Analgesics, Non-Narcotic / Q000032:analysis*; D000818:Animals; D000925:Anticoagulants / Q000032:analysis*; D002138:Calibration; D002753:Chlorzoxazone / Q000032:analysis*; D002851:Chromatography, High Pressure Liquid; D003374:Coumarins / Q000032:analysis*; D003470:Culture Media; D003577:Cytochrome P-450 Enzyme System / Q000032:analysis*; D007004:Hypoglycemic Agents / Q000032:analysis*; D066298:In Vitro Techniques; D059625:Liquid-Liquid Extraction; D008862:Microsomes, Liver / Q000201:enzymology*; D009125:Muscle Relaxants, Central / Q000032:analysis*; D010615:Phenacetin / Q000032:analysis*; D011786:Quality Control; D051381:Rats; D012015:Reference Standards; D015203:Reproducibility of Results; D013056:Spectrophotometry, Ultraviolet; D013739:Testosterone / Q000032:analysis*; D014044:Tolbutamide / Q000032:analysis*
chemical_list: D018712:Analgesics, Non-Narcotic; D000925:Anticoagulants; D003374:Coumarins; D003470:Culture Media; D007004:Hypoglycemic Agents; D009125:Muscle Relaxants, Central; D013739:Testosterone; D003577:Cytochrome P-450 Enzyme System; D014044:Tolbutamide; C030123:coumarin; D010615:Phenacetin; D002753:Chlorzoxazone