Research Article | Volume: 8, Issue: 9, September, 2018

Simultaneous Determination of Pyrazinamide, Rifampicin, Ethambutol, Isoniazid and Acetyl Isoniazid in Human Plasma by LC-MS/MS Method

Le Thi Luyen Ta Manh Hung Le Thu Huyen Le Anh Tuan Duong Thi Ly Huong Hoang Van Duc Bui Thanh Tung   

Open Access   

Published:  Sep 30, 2018

DOI: 10.7324/JAPS.2018.8910

Monitoring of anti-tuberculosis drug concentrations in human plasma is very important in the treatment of TB disease. In this study, we described a method that can rapidly and simultaneously measure the plasma concentrations of four anti-tuberculosis drugs (isoniazid, rifampicin, pyrazinamide, and ethambutol) and one major metabolite (acetylisoniazid) using high-performance liquid chromatography/tandem mass spectrometry (HPLC/MS/MS). To adjust for degradation and losses during sample preparation, Diltiazem hydrochloride was used as an internal standard. Samples were prepared by using protein precipitation with methanol. Four drugs and one metabolite were simultaneously separated by using the high-performance liquid chromatography system with Gemini C18 column and a mobile phase consisting of MeOH and Ammonium acetate 5mM, pH 3.5. The method was found to have high selectivity. Precision estimated by the coefficient of variation was <15% for all drugs. The linear range of the calibration curve for PZA 1.0–100 μg/mL; RIF 0.2–20 μg/mL; INH 0.1–10 μg/mL; AcINH 0.1–10 μg/mL; and EMB 20–5000 ng/mL. The lower limit of quantification was 1 μg/mL; 0.2 μg/mL; 0.1 μg/mL; 0.1 μg/mL and 20 ng/mL, respectively. The results of validation for specificity/selectivity, precision, accuracy, linearity, limit of quantification, recovery, and stability show that the method meets the requirements of a bioanalytical method. Our method may serve well for routine therapeutic monitoring of the first-line anti-TB drugs in patient plasma.

Keyword:     Pyrazinamide Rifampicin Isoniazid Acetyl Isoniazid Ethambutol LC-MS/MS Tuberculosis.


Luyen LT, Hung TM, Huyen LT, Tuan LA, Huong DTL, Duc HV, Tung BT. Simultaneous determination of Pyrazinamide, Rifampicin, Ethambutol, Isoniazid and Acetyl Isoniazid in human plasma by LC-MS/MS method. J App Pharm Sci, 2018; 8(09): 061-073.

Copyright: © The Author(s). This is an open-access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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