RESUMEN
A liquid chromatography-tandem mass spectrometric (LC-MS-MS) method for the highly sensitive determination of a new bone-anabolic agent, TAK-778 in human serum was developed. The internal standard (I.S.) used was deuterated TAK-778. TAK-778 and I.S. were extracted from serum samples with diethyl ether at neutral pH. A turbo ion spray interface was used as the ion source of LC-MS-MS, and the analysis was performed in the selected reaction monitoring mode. The lower limit of quantification was 0.02 ng/ml when 0.4 ml of serum was used, and the standard curve was linear in the range of 0.02-10 ng/ml. The method was precise; the intra- and inter-day precision of the method was not more than 17.9%. The accuracy of the method was good with the deviations between added and calculated concentration of TAK-778 being typically within 9.0%.
Asunto(s)
Anabolizantes/sangre , Benzotiepinas/sangre , Anabolizantes/química , Benzotiepinas/química , Cromatografía Líquida de Alta Presión/métodos , Cromatografía Líquida de Alta Presión/estadística & datos numéricos , Humanos , Espectrometría de Masas/métodos , Espectrometría de Masas/estadística & datos numéricosRESUMEN
We investigated the application of alkylamines, as additives to the mobile phase, to a quantification method for the metabolites, M-III and M-IV, of TAK-778, which is a new bone anabolic agent, in human serum using liquid chromatography/tandem mass spectrometry (LC/MS/MS). Prior to setting up the analytical method, we found that 1-alkylamines co-existing with M-III and M-IV in the turbo ionsprayed solution formed 1-alkylammonium adduct molecules of these metabolites during the ionization process, and the abundance of the adduct ions was considerably higher than that of protonated molecules ([M + H](+)s) of these metabolites. Based on these findings, we investigated a variety of 1-alkylamines and their spiked concentrations in the mobile phase for LC/MS/MS analysis to obtain higher sensitivities for the quantification of these metabolites. After these examinations, we found that 1-hexylamine at a final concentration of 0.05 mmol l(-1) was the most suitable additive for the mobile phase, and set the selected reaction monitoring (SRM) ions for the 1-hexylammonium adduct molecule and [M + H](+), allowing about a fivefold gain in the SRM chromatographic peak compared with that without 1-hexylamine. The adduct ion was considered to be formed by interaction between the amino group of 1-hexylamine and the phosphoryl group of M-III and M-IV. The internal standard (I.S.) used was deuterated M-III for each metabolite. The analytes and I.S. were extracted with diethyl ether from serum samples at neutral pH and injected into the LC/MS/MS system with a turbo ionspray interface. The limit of quantification for both analytes was 0.5 ng ml(-1) when 0.1 ml of serum was used, and the calibration curves were linear in the range 0.5-100 ng ml(-1). The method was precise; the intra- and inter-day precisions of the method were not more than 5.6%. The accuracy of the method was good, with deviations between added and calculated concentrations of M-III and M-IV being typically within 16.6%. This method provided reliable pharmacokinetic data for M-III and M-IV after the intramuscular administration of TAK-778 sustained-release formulation in humans.