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An automated method for the simultaneous measurement of azole antifungal drugs in human plasma or serum using turbulent flow liquid chromatography-tandem mass spectrometry

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Abstract

Azole antifungal drugs are important in the prophylaxis and treatment of invasive aspergillosis. Therapeutic drug monitoring may be indicated to (1) monitor adherence, (2) guide dosage and (3) minimise the risk of drug–drug interactions and dose-related toxicity. TurboFlowTM technology offers online, automated sample preparation. An Aria TranscendTM TLX-II coupled with a TSQ VantageTM MS was used. Centrifuged samples (25 μL) were mixed with internal standard solution (975 μL) and 30 μL injected directly onto a C18-P-XL TurboFlow column. Analytes were focussed onto a Phenomenex Gemini Phenyl analytical column and eluted using a methanol/water gradient (flow-rate, 0.8 mL/min). Analytes were monitored in selected reaction monitoring mode (two transitions per analyte, positive mode APCI). Calibration ranges were as follows: itraconazole, hydroxyitraconazole, and posaconazole 0.05–5.0 mg/L; voriconazole and fluconazole 0.1–10 mg/L. Total analysis time was 12 min. TurboFlow column recovery was >77% for all analytes. Calibration was linear (R 2 > 0.99) for all analytes. Inter- and intra-assay imprecision (% RSD) was <8% and accuracy (nominal internal quality control values) 90–105% for all analytes. The limit of detection was 0.01 mg/L for all analytes. No matrix effects were observed. This method is simple, robust and suitable for measuring these compounds at concentrations attained during therapy.

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Acknowledgments

Thanks to Tom Whitehouse and Jeff Zonderman (ThermoFisher Scientific) for analytical support, to Mr. A. Noel and Dr. M. Darville from the UKNEQAS Antifungal Scheme and to Prof. D. Uges from the KKGT EQA Scheme for permission to cite data from their schemes. Thanks also to Janssen-Cilag and to Pfizer for the gift of reference materials and to Pfizer and Gilead for sponsoring the Aspergillosis study.

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Authors and Affiliations

  1. Toxicology Unit, Department of Clinical Biochemistry, King’s College Hospital NHS Foundation Trust, Third Floor, Bessemer Wing, Denmark Hill, London, SE5 9RS, UK

    L. Couchman, S. L. Buckner, P. E. Morgan & R. J. Flanagan

  2. Department of Haematological Medicine, King’s College Hospital NHS Foundation Trust, London, SE5 9RS, UK

    M. M. Ceesay & A. Pagliuca

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  1. L. Couchman
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  2. S. L. Buckner
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  3. P. E. Morgan
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  4. M. M. Ceesay
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  5. A. Pagliuca
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  6. R. J. Flanagan
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Correspondence to L. Couchman.

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Couchman, L., Buckner, S.L., Morgan, P.E. et al. An automated method for the simultaneous measurement of azole antifungal drugs in human plasma or serum using turbulent flow liquid chromatography-tandem mass spectrometry. Anal Bioanal Chem 404, 513–523 (2012). https://doi.org/10.1007/s00216-012-6176-3

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  • DOI: https://doi.org/10.1007/s00216-012-6176-3

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