首页 | 本学科首页   官方微博 | 高级检索  
     


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
Authors:R. ter Heine  M. Davids  H. Rosing  E.C.M. van Gorp  J.W. Mulder  Y.T. van der Heide  J.H. Beijnen  A.D.R. Huitema
Affiliation:Slotervaart Hospital, Department of Pharmacy & Pharmacology, Louwesweg 6, 1066EC Amsterdam, The Netherlands
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 (150 mm × 2.0 mm, particle size 5 μm) with a quick stepwise gradient using an acetate buffer (pH 5) and methanol, at a flow rate of 0.25 mL/min. The analytical run time was 10 min. 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 1–500 ng/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.
Keywords:
本文献已被 ScienceDirect 等数据库收录!
设为首页 | 免责声明 | 关于勤云 | 加入收藏

Copyright©北京勤云科技发展有限公司  京ICP备09084417号