A simplified LC-MS/MS method for determination of isoniazid and pyrazinamide in human plasma using protein precipitation technique and its application to a pharmacokinetic study

[Speaker] Phongpan Mokmued:1
[Co-author] Archawin Rojanawiwat:1, Surakameth Mahasirimongkol:1, Ingorn Prasanchaimontri:2, Katsushi Tokunaka:3, Taisei Mushiroda:4
1:Department of Medical Sciences, Medical Life Science Institute, Thailand, 2:Department of Medical Sciences, Bureau of Drug and Narcotic, Thailand, 3:Department of Human Genetics, Graduate School of Medicine, The University of Tokyo, Japan, 4:Laboratory for Pharmacogenomics, RIKEN Center for Integrative Medical Sciences, Yokohama, Japan

A sensitive and rapid liquid chromatography tandem mass spectrometry (LC-MS/MS) method was developed to simultaneous plasma drugs level measurement of Isoniazid (INH) and Pyrazinamide (PZA) first line anti-tuberculosis. Samples with the spiked drug standards were extracted using protein precipitation and then separated on ACQUITY UPLC BEH amide column (2.1 x 150 mm 1.7 um) under isocratic elution of 10 mM ammonium acetate in water and acetonitrile (60:40 v/v) at flow rate of 0.400 ml/min. Detection of all analytes was performed under positive ionization mode by multiple reaction monitoring (MRM). The analytical method was achieved within a run time of 10 minutes. The inter-assay calibrated variability data were obtained through concentrations of 1-100 µg/ml for INH and PZA. The calibration curves were tested a linear 100-fold concentration range (r2 ≥ 0.995). Within-run and between-run precision (%CV) at concentration corresponding to LLOQ medium and high levels for two drugs were 1.45-17.83 % and 2.03-12.37 % respectively at concentration representing low medium and high levels for two drugs. The performance of our MS/MS detection technique was generally acceptable. The method was applied to a pharmacokinetic study in 30 tuberculosis Thai patients.
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