Simultaneous determination of four probe drugs of CYP450 in rat plasma by LC-MS/MS

Abstract

Abstract:

AIM:To establish a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for determination of four probe drugs of phenacetin (1A2), tolbutamide (2C9), dextromethorphan (2D6) and midazolam (3A4) in rat plasma. METHODS:Chromatographic separation was achieved on a 2.1 mm×50 mm, 3.5 μm particle, symmetry C18 column at 35 ℃. The mobile phase was constituted of methanol and 0.1% formic acid in water (67∶33,V/V), running at a flow rate of 0.2 mL/min. Mass spectrometry was performed under the multiple reaction monitoring (MRM) mode with an ESI source. With the internal standard carbamazepin, the specific probe substrates of phenacetin, tolbutamide, dextromethorphan and midazolam could be detected in rat plasma. This method was applied into the analysis of plasma samples in rat and was evaluated in terms of recovery, stability, precision and accuracy, and. RESULTS:A good linear relationship was found in phenacetin (10-5 000 ng/mL), tolbutamide (20-10 000 ng/mL), dextromethorphan (4-1 000 ng/mL) and midazolam (4-1 000 ng/mL). The correlation coefficiency (R²) was greater than 0.99. The accuracy of data ranged from 95.96% to 103.10%. RSD(%) of intra-day and inter-day precision was less than 10%. The extraction recoveries of probe drugs were greater than 77% and RSD(%) less than 7%. The range of matrix effect was 90.7% to 100.3% and RSD(%) less than 5.9%. The data exhibited excellent stability. CONCLUSION: The established method is accurate, sensitive, and suitable for determining plasma concentrations of the four probe drugs (phenacetin, tolbutamide, dextromethorphan and midazolam).The plasma samples demonstrate excellent stability and it could be used for evaluating the CYP1A2, CYP2C9, CYP2D6 and CYP3A4 activities in the drug-drug interaction studies.

Key words: LC-MS/MS, CYP450, cocktail

CLC Number:

R965.2

LIU Jingjing, LIANG Zhi, LUO Xiangchong, HE Gonghao, FENG Enfu, XU Guili. Simultaneous determination of four probe drugs of CYP450 in rat plasma by LC-MS/MS[J]. Chinese Journal of Clinical Pharmacology and Therapeutics, 2017, 22(2): 144-150.

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