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Liquid chromatography–tandem mass spectrometric determination of tenofovir-diphosphate in human peripheral blood mononuclear cells
Ist Teil von
Journal of chromatography. B, Analytical technologies in the biomedical and life sciences, 2006-11, Vol.843 (2), p.147-156
Ort / Verlag
Amsterdam: Elsevier B.V
Erscheinungsjahr
2006
Link zum Volltext
Quelle
MEDLINE
Beschreibungen/Notizen
To facilitate the evaluation of drug safety, virologic activity, and pharmacokinetics, an anion exchange isolation of tenofovir-diphosphate (TFV-DP) from human peripheral blood mononuclear cells (hPBMCs), coupled with dephosphorylation, desaltation, and detection by LC–MS–MS was validated. hPBMCs were harvested from whole blood, lysed, and a suspension of intracellular tenofovir moieties was produced. TFV-DP was isolated from TFV-monophosphate (TFV-MP) and tenofovir (TFV), dephosphorylated with acid phosphatase to form TFV and then desalted and concentrated, making it possible for tandem mass spectral detection. An LC–MS–MS methodology was developed and validated for the determination of TFV concentrations, which directly correspond with the intra-hPBMC TFV-DP concentration. The assay was linear in the range of 50–10,000
fmol per sample. The lower limit of quantitation (LLOQ) of the method is 10
fmol per million cells with 5 million hPBMCs used. This paper outlines the development and validation of the determination of TFV-DP concentrations in femtomoles per million hPBMCs.