Determination of Gemcitabine Triphosphate in Peripheral Blood Mononuclear Cells by Reverse-Phase HPLC

This paper describes a high-performance liquid chromatographic assay coupled with UV detection at 272 nm, to determine the levels of the nucleotide analogue gemcitabine triphosphate, one of the active metabolites of the antitumoral drug gemcitabine, in peripheral blood mononuclear cells. Isocratic ion-pair chromatography on a C18 column was used. Samples were treated with trichloroacetic acid 40%, cleaned and neutralized with freon:trioctylamine (4:1). The method was linear on the concentration range tested, and the evaluated precision was found satisfactory (RSD < 6.8). The detection limit was 31 pmol, and the quantitation limit 102 pmol. The recovery of gemcitabine triphosphate ranged between 95 and 114%. The procedure was used to provide pharmacokinetic data from cancer patients treated with intravenous gemcitabine.