RESUMO
A sensitive and rapid LC-MS/MS method was developed and validated for the simultaneous quantitation of four HDAC inhibitors, namely belinostat (BST), panobinostat (PST), rocilinostat (RST) and vorinostat (VST), in mouse plasma as per regulatory guidelines. The analytes and internal standard were extracted from 50 µL mouse plasma by protein precipitation, followed by chromatographic separation using an Atlantis C18 column with an isocratic mobile phase comprising 0.1% formic acid-acetonitrile (25:75, v/v) at a flow rate of 0.5 mL/min within 2.5 min. Detection and quantitation were done by multiple reaction monitoring on a triple quadrupole mass spectrometer following the transitions: m/z 319 â 93, 350 â 158, 434 â 274 and 265 â 232 for BST, PST, RST and VST, respectively, in the positive ionization mode. The calibration curves were linear from 2.92 to 2921 ng/mL for BST and PST and from 1.01 to 1008 ng/mL for RST and VST with r2 ≥ 0.99 for all of the analytes. The intra- and inter-batch accuracy and precision (CV) across quality controls varied from 85.5 to 112% and from 2.30 to 12.5, respectively, for all of the analytes. Analytes were found to be stable under different stability conditions. The method was applied to an i.v. pharmacokinetic study in mice.
Assuntos
Cromatografia Líquida/métodos , Inibidores de Histona Desacetilases/sangue , Espectrometria de Massas em Tandem/métodos , Animais , Calibragem , Inibidores de Histona Desacetilases/farmacocinética , Limite de Detecção , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Padrões de Referência , Reprodutibilidade dos TestesRESUMO
A sensitive and rapid LC-MS/MS method was developed and validated for the simultaneous quantitation of darolutamide and its active metabolite i.e. ORM-15341 in 50µL mice plasma using bicalutamide as an internal standard (I.S.) as per regulatory guidelines. Sample processing was accomplished through liquid-liquid extraction. Chromatographic separation was achieved using an Atlantis C18 column with an isocratic mobile phase comprising 0.2% formic acid:acetonitrile (35:65, v/v) at a flow rate of 0.8mL/min within 2.5min. Detection and quantitation were done by multiple reaction monitoring on a triple quadrupole mass spectrometer following the transitions: m/z 397â202, 395â202 and 429â255 for darolutamide, ORM-15341 and I.S, respectively in the negative ionization mode. The calibration curve was linear from 0.61-1097ng/mL for both darolutamide and ORM-15341. The intra- and inter-day precisions were in the range of 1.34-13.8 and 4.85-12.9 and 3.91-13.7 and 6.54-14.2%, for darolutamide and ORM-15341, respectively. Darolutamide and ORM-15341 were found to be stable under different stability conditions. The validated method was applied to a pharmacokinetic study in mice.