Print Email Facebook Twitter Simultaneous determination of endogenous deoxynucleotides and phosphorylated nucleoside reverse transcriptase inhibitors in peripheral blood mononuclear cells using ion-pair liquid chromatography coupled to mass spectrometry Title Simultaneous determination of endogenous deoxynucleotides and phosphorylated nucleoside reverse transcriptase inhibitors in peripheral blood mononuclear cells using ion-pair liquid chromatography coupled to mass spectrometry Author Coulier, L. van Kampen, J.J.A. de Groot, R. Gerritsen, H.W. Bas, R.C. van Dongen, W.D. Brüll, L.P. Luider, T.M. TNO Kwaliteit van Leven KvL Publication year 2008 Abstract Nucleoside reverse transcriptase inhibitors (NRTIs) are activated intracellularly to their triphosphate (TP) form, which compete with endogenous deoxynucleotide-triphosphates (dNTP) as substrate for HIV reverse transcriptase. The activity of NRTIs is thus described by the NRTI-TP to-dNTP ratio in relevant cell types. Therefore, we developed an ion-pair (IP) LC-MS method for the simultaneous analysis of the mono-, di-, and TP forms of NRTIs and endogenous deoxynucleosides in peripheral blood mononuclear cells (PBMC). The IP-LC method was applied on an ITmass spectrometer using the MS-mode as well as on a triple quadrupole mass spectrometer using the MS/MS mode. The MS/MS approach on the triple quadrupole mass spectrometer demonstrated the best clinical applicability due to its higher sensitivity. The LOD (minimum amount on column) were 25 fmol for the TP forms of zidovudine, lamivudine, and stavudine, as well as for their endogenous dNTP counterparts. The linearity (R2) of the calibration curves were>0.99. The obtained LOD readily allow for clinical applications using just one million PBMC obtained from HIV-infected patients under therapy. © 2008 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim. Chemicals / CAS: RNA directed DNA polymerase, 37213-50-6, 9068-38-6; abacavir, 136470-78-5, 188062-50-2; carbovir, 118353-05-2; lamivudine, 134678-17-4, 134680-32-3; stavudine, 3056-17-5; tenofovir, 147127-19-3, 147127-20-6; zidovudine, 30516-87-1 Subject BiologyAnalytical researchHIVIntracellularIon-pairNucleoside reverse transcriptase inhibitorNucleotideabacavircarbovirstavudinetenofovirzidovudinecell lysatecontrolled studydrug cell levelhuman cellion pair chromatographyisolation procedureperformance measurement systemperipheral blood mononuclear cellprotein phosphorylationquadrupole mass spectrometryseparation technique To reference this document use: http://resolver.tudelft.nl/uuid:3965ea12-baf8-4322-903d-863eec6fab9b DOI https://doi.org/10.1002/prca.200800002 TNO identifier 241169 ISSN 1862-8346 Source Proteomics - Clinical Applications, 2 (10-11), 1557-1562 Document type article Files To receive the publication files, please send an e-mail request to TNO Library.