Combination of on‐line CE assay with MS detection for the study of drug metabolism by cytochromes P450. Issue 11 (6th May 2015)

• Record Type: Journal Article
• Authors: Langmajerová, Monika; Řemínek, Roman; Pelcová, Marta; Foret, František; Glatz, Zdeněk
• Other Names: Guttman Andras guestEditor.; Kilar Ferenc guestEditor.
• Is Part Of: Electrophoresis. Volume 36:Issue 11/12(2015)
• Page Start: 1365
• Page End: 1373
• Publication Date: 2015-05-06
• Subjects: CE -- Cytochrome P450 -- Drug metabolism -- In‐capillary reaction -- MS; Electrophoresis -- Periodicals; Electrophoresis -- Periodicals; 541.372
• DOI: 10.1002/elps.201400394
• Languages: English
• ISSNs: 0173-0835

Abstract

Abstract : A new CE‐MS method with enzymatic reaction inside the capillary was developed for the study of drug metabolism by cytochromes P450. This automated method, based on the transverse diffusion of laminar flow profiles methodology, is comprised of the injection of substrates and enzyme, their mixing, incubation, and separation of the reaction products, all performed by CE, and their detection, identification, and quantification by MS. The developed and validated method was finally used to conduct a kinetic study of cytochrome P450 isoform 2C9 or human liver microsomes with diclofenac in order to demonstrate its practical functionality. All the estimated kinetic values—apparent Michaelis constants and apparent maximum reaction velocities were in agreement with literature data obtained using other techniques. In addition, the consumption of reactants was in the tens of nL per analysis. The method's usability was further demonstrated on tolbutamide, the other probe substrate of cytochrome P450 isoform 2C9. As a result, the method is conceptually applicable for the screening of any other cytochrome P450 isoform and its substrates and inhibitors after adapting the incubation and separation conditions.