Application of a UPLC–MS/MS method to the protein binding study of TM-2 in rat, human and beagle dog plasma

Abstract

TM-2 known as a potential antitumor drug is a novel semi-synthetic taxane derivative. As drug–protein interactions contribute to insights into pharmacokinetic and pharmacodynamic properties, we elucidated the binding of TM-2 to plasma protein. In this study, a simple, rapid and reliable method was developed and validated employing equilibrium dialysis for the separation of bound and unbound drugs and ultra-performance liquid chromatography–tandem mass spectrometry (UPLC–MS/MS) for the quantitation. Protein binding reached equilibrium within 24h of incubation at 37°C. After liquid–liquid extraction with methyl tert-butyl ether, the samples were separated on Thermo Syncronis UPLC® C18 (2.1mm×50mm, 1.7µm), and acquisition of mass spectrometric data was performed in multiple reaction monitoring (MRM) mode via positive electrospray ionization. The assay was linear over the concentration rang of 5–2000ng/mL. The intra- and inter-day precisions were 0.1%–14.8%, and the accuracy was from −6.4% to 7.0%. This assay has been successfully applied to a protein binding study of TM-2 in rat, human and beagle dog plasma. TM-2 showed high protein binding of 81.4%±6.5% (rat), 87.9%±3.6% (human) and 79.4%±4.0% (beagle dog). The results revealed that there was an insignificant difference among the three species.