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Quantification of tamoxifen and three of its phase-I metabolites in human plasma by liquid chromatography/triple-quadrupole mass spectrometry
Ist Teil von
Journal of pharmaceutical and biomedical analysis, 2011-12, Vol.56 (5), p.1016-1023
Ort / Verlag
Amsterdam: Elsevier B.V
Erscheinungsjahr
2011
Quelle
Elsevier ScienceDirect Journals
Beschreibungen/Notizen
In view of future pharmacokinetic studies, a highly sensitive ultra performance liquid chromatography/tandem mass spectrometry (UPLC–MS/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
μl were purified by liquid–liquid extraction with 1
ml
n-hexane/isopropanol, after deproteination through addition of 50
μl acetone and 50
μl 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
μm 2.1
mm
×
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–MS/MS method in serum.