Quantification of tamoxifen and three of its phase-I metabolites in human plasma by liquid chromatography/triple-quadrupole mass spectrometry

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• 作者：Lisette Binkhorst^a ; Ron H.J. Mathijssen^b ; Inge M. Ghobadi Moghaddam-Helmantel^b ; Peter de Bruijn^b ; Teun van Gelder^a ; Erik A.C. Wiemer^b ; Walter J. Loos^b ;   ; ^{w.loos@erasmusmc.nl}
• 关键词：Tamoxifen ; N-desmethyl-tamoxifen ; 4-Hydroxy-tamoxifen ; Endoxifen ; LC&ndash ; MS/MS
• 刊名：Journal of Pharmaceutical and Biomedical Analysis
• 出版年：2011
• 出版时间：15 December 2011
• 年：2011
• 卷：56
• 期：5
• 页码：1016-1023
• 全文大小：383 K

文摘

In view of future pharmacokinetic studies, a highly sensitive ultra performance liquid chromatography/tandem mass spectrometry (UPLC¨CMS/MS) method has been developed for the simultaneous quantification of tamoxifen and three of its main phase I metabolites in human lithium heparinized plasma. The analytical method has been thoroughly validated in agreement with FDA recommendations. Plasma samples of 200 ¦Ìl were purified by liquid¨Cliquid extraction with 1 ml n-hexane/isopropanol, after deproteination through addition of 50 ¦Ìl acetone and 50 ¦Ìl deuterated internal standards in acetonitrile. Tamoxifen, N-desmethyl-tamoxifen, 4-hydroxy-tamoxifen and endoxifen were chromatographically separated on an Acquity UPLC^? BEH C18 1.7 ¦Ìm 2.1 mm ¡Á 100 mm column eluted at a flow-rate of 0.300 ml/min on a gradient of 0.2 mM ammonium formate and acetonitrile, both acidified with 0.1 % formic acid. The overall run time of the method was 10 min, with elution times of 2.9, 3.0, 4.1 and 4.2 min for endoxifen, 4-hydroxy-tamoxifen, N-desmethyl-tamoxifen and tamoxifen, respectively. Tamoxifen and its metabolites were quantified by triple-quadrupole mass spectrometry in the positive ion electrospray ionization mode. The multiple reaction monitoring transitions were set at 372 > 72 (m/z) for tamoxifen, 358 > 58 (m/z) for N-desmethyl-tamoxifen, 388 > 72 (m/z) for 4-hydroxy-tamoxifen and 374 > 58 (m/z) for endoxifen. The analytical method was highly sensitive with the lower limit of quantification validated at 5.00 nM for tamoxifen and N-desmethyl-tamoxifen and 0.500 nM for 4-hydroxy-tamoxifen and endoxifen, which is equivalent to 1.86, 1.78, 0.194 and 0.187 ng/ml for tamoxifen, N-desmethyl-tamoxifen, 4-hydroxy-tamoxifen and endoxifen, respectively. The method was also precise and accurate, with within-run and between-run precisions within 12.0 % and accuracy ranging from 89.5 to 105.3 % . The method has been applied to samples from a clinical study and cross-validated with a validated LC¨CMS/MS method in serum.