A simple LC-MS/MS method for the simultaneous quantification of drug metabolic enzymes

• Published in: Journal of chromatography. B, Analytical technologies in the biomedical and life sciences Vol. 1214; p. 123536
• Main Authors: Guo, Xuan, Zhang, Lei, Lei, Zihan, Hou, Zhe, Li, Hui, Li, Xiaodong, Dong, Jing, Song, Ling, Chen, Dingding, Liu, Dongyang
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01.01.2023
• Subjects: Chromatography, Liquid - methods; CYP450 enzymes; Cytochrome P-450 CYP3A - metabolism; Cytochrome P-450 Enzyme System - metabolism; Humans; LC-MS/MS; Microsomes, Liver - metabolism; Proteomics; Tandem Mass Spectrometry - methods; LC-MS/MS; CYP450 enzymes; Proteomics
• ISSN: 1570-0232; 1873-376X
• DOI: 10.1016/j.jchromb.2022.123536

•A targeted proteomics method to quantify the abundance of drug-related metabolizing enzymes by UPLC-MS/MS.•Ethylene glycol is used to enhance the mass spectral response of surrogate peptides.•Needle washing method and needle washing solution are optimized to avoid serious crossover. The aim of this study is to develop a LC-MS/MS method for the quantitation of seven cytochrome P450 (CYP450) enzymes. A high-performance liquid chromatography-tandem mass spectrometry method was developed using multiple reaction monitoring mode with positive electrospray ionization. The method was validated with selectivity, linearity, stability, accuracy and precious. In addition, the abundance of seven CYP450 enzymes in human liver microsomes and CYP3A4 in placenta were determined using the current method. The linear range for CYP1A2, CYP2B6 and CYP2C8 was 0.036–3.6 nM and for CYP2C9, CYP2C19, CYP2D6 and CYP3A4 was 0.090–9.0 nM. No interference was found between the blank matrix and each specific peptides. The accuracy and precious results were in accord with the requirement of analytical methods for biological samples in Chinese Pharmacopoeia. In addition, the peptides were stable under current stability conditions. The content of CYP3A4 in placenta and the seven CYP450 enzymes in human liver microsomes were accurately quantified. The developed method is sensitive and specific and can be applied to the quantification of enzymes abundance in different human derived samples like placenta and liver microsomes.