Rapid solid-phase extraction method to quantify [11C]-verapamil, and its [11C]-metabolites, in human and macaque plasma

IntroductionP-glycoprotein (P-gp), an efflux transporter, is a significant barrier to drug entry into the brain and the fetus. The positron emission tomography (PET) ligand, [11C]-verapamil, has been used to measure in vivo P-gp activity at various tissue–blood barriers of humans and animals. Since verapamil is extensively metabolized in vivo, it is important to quantify the extent of verapamil metabolism in order to interpret such P-gp activity. Therefore, we developed a rapid solid-phase extraction (SPE) method to separate, and then quantify, verapamil and its radiolabeled metabolites in plasma.MethodsUsing high-performance liquid chromatography (HPLC), we established that the major identifiable circulating radioactive metabolite of [11C]-verapamil in plasma of humans and the nonhuman primate, Macaca nemestrina, was [11C]-D-617/717. Using sequential and differential pH elution on C8 SPE cartridges, we developed a rapid method to separate [11C]-verapamil and [11C]-D-617/717. Recovery was measured by spiking the samples with the corresponding nonradioactive compounds and assaying these compounds by HPLC.ResultsVerapamil and D-617/717 recovery with the SPE method was >85%. When the method was applied to PET studies in humans and nonhuman primates, significant plasma concentration of D-617/717 and unknown polar metabolite(s) were observed. The SPE and the HPLC methods were not significantly different in the quantification of verapamil and D-617/717.ConclusionsThe SPE method simultaneously processes multiple samples in less than 5 min. Given the short half-life of [11C], this method provides a valuable tool to rapidly determine the concentration of [11C]-verapamil and its [11C]-metabolites in human and nonhuman primate plasma.