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Simultaneous quantification of four first line antitubercular drugs and metabolites in human plasma by hydrophilic interaction chromatography and tandem mass spectrometry

Artikel i vetenskaplig tidskrift
Författare Jesper Sundell
E. Bienvenu
Sofia Birgersson
Angela Äbelö
Michael Ashton
Kurt-Jürgen Hoffmann
Publicerad i Journal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences
Volym 1105
Sidor 129-135
ISSN 1570-0232
Publiceringsår 2019
Publicerad vid Institutionen för neurovetenskap och fysiologi, sektionen för farmakologi
Sidor 129-135
Språk en
Länkar dx.doi.org/10.1016/j.jchromb.2018.1...
Ämnesord Biomolecules, Diseases, Extraction, Formic acid, Hydrazine, Hydrophilicity, Liquid chromatography, Liquids, Mass spectrometry, Metabolites, Plasma (human), Chromatographic separations, Hydrophilic interaction chromatography, Liquid chromatography-tandem mass spectrometry, Lower limit of quantifications, Mass spectrometric detection, Multiple-reaction monitoring modes, Tandem mass spectrometry, US Food and Drug Administration, Drug interactions
Ämneskategorier Farmakologi, Analytisk kemi


Co-infection of tuberculosis in HIV-patients is a major health concern worldwide and especially so in Sub-Saharan Africa. To enhance the study of potential drug-drug interactions when simultaneously treating the two infections, a liquid chromatography tandem mass spectrometry method was developed for the quantitation of the four first line anti-tuberculosis drugs isoniazid, rifampicin, pyrazinamide, ethambutol and four of their major metabolites in human plasma. Analytes were extracted from 200 μL of plasma using a sequential liquid-liquid extraction with ethyl acetate at neutral and acidic pH. The combined extracts were analyzed by liquid chromatography with mass spectrometric detection in a multiple reaction monitoring mode. The chromatographic separation was performed on a hydrophilic interaction column using a stepwise gradient with two mobile phases consisting of water with 0.3% formic acid and methanol with 0.3% formic acid, respectively. The total run time of each analysis was 4 min. The lower limit of quantification applied was 40 ng/mL for ethambutol, acetylisoniazid and 25-desacetylrifampicin, 60 ng/mL for 5-hydroxypyrazinamide, 80 ng/mL for isoniazid and isonicotinic acid, 200 ng/mL for rifampicin and 320 ng/mL for pyrazinamide. The method was validated according to US Food and Drug Administration guidance. The method exhibited adequate accuracy (87.1–114.9%), precision (CV < 12.8%) and specificity. Recovery and matrix effect were consistent (CV < 11.9%). The extracted samples were stable in the autosampler at 8 °C for up to 24 h as well as after three freeze-thaw cycles (recovery > 86.3%). The method has been shown to be robust for the analysis of the stated drugs and metabolites in human plasma obtained from 73 patients receiving these four first line anti-tuberculosis drugs. © 2018 Elsevier B.V.

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