Quantification of HIV protease inhibitors and non-nucleoside reverse transcriptase inhibitors in peripheral blood mononuclear cell lysate using liquid chromatography coupled with tandem mass spectrometry. Journal of chromatography. B, Analytical technologies in the biomedical and life sciences [J Chromatogr B Analyt Technol Biomed Life Sci] Journal article | | Title | Quantification of HIV protease inhibitors and non-nucleoside reverse transcriptase inhibitors in peripheral blood mononuclear cell lysate using liquid chromatography coupled with tandem mass spectrometry. | | Author(s) | Ter Heine R, Davids M, Rosing H, van Gorp EC, Mulder JW, van der Heide YT, Beijnen JH, Huitema AD | | Institution | Slotervaart Hospital, Department of Pharmacy & Pharmacology, Louwesweg 6, 1066EC Amsterdam, The Netherlands. | | Source | J Chromatogr B Analyt Technol Biomed Life Sci 2009 Jan 14. | | Abstract | For pharmacokinetic monitoring, measurement of antiretroviral agents in plasma is the gold standard. However, human immunodeficiency virus protease inhibitors (PIs) or non-nucleoside reverse transcriptase inhibitors (NNRTIs) exert their action within the infected cell. Cell-associated concentrations may therefore more adequately reflect therapy outcome. Therefore, for the quantification of nine PIs (amprenavir, atazanavir, darunavir, indinavir, lopinavir, nelfinavir, ritonavir, saquinavir and tipranavir), 1 active PI metabolite (nelfinavir M8) and 2 NNRTIs (efavirenz and nevirapine) in lysate of peripheral blood mononuclear cells (PBMCs) an assay was developed and validated, using liquid chromatography coupled with tandem mass spectrometry. Analytes were extracted from a PBMC pellet by means of a one-step extraction with 50% methanol containing the internal standards D6-indinavir, D5-saquinavir, 13C6-efavirenz and dibenzepine. Chromatographic separation was performed on a reversed phase C18 column (150mmx2.0mm, particle size 5mum) with a quick stepwise gradient using an acetate buffer (pH 5) and methanol, at a flow rate of 0.25mL/min. The analytical run time was 10min. The triple quadrupole mass spectrometer was operated in the positive ion-mode and multiple reaction monitoring was used for drug quantification. The method was validated over a range of 1-500ng/mL in PBMC lysate for all analytes. The method was proven to be specific, accurate, precise and robust. The mean precision and accuracy was less than +/-12% at all concentration levels. Using the developed assay and a previously developed assay for these analytes in plasma, the relationship between plasma and intracellular pharmacokinetics and their relationship with therapy outcome can now be determined. | | Language | ENG | | Pub Type(s) | JOURNAL ARTICLE
| | PubMed ID | 19168402 |
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