A Sensitive HPLC-MS/MS Method for the Quantification of Selegiline in Beagle Dog Plasma: Application to a Pharmacokinetic Study

<P>Objective: To develop a reliable and sensitive high-performance liquid chromatographytandem mass spectrometry (HPLC-MS/MS) method for the quantification of selegiline in Beagle dog plasma and apply the validated method to study the pharmacokinetics and bioavailability of oral selegiline lyophilizate in Beagle dogs. Methods: Following alkalization with 1 M sodium hydroxide solution, selegiline and the Internal Standard (IS) zolmitriptan were extracted using tert-butyl methyl ether and separated on a CAPCELL PAK C18 column under isocratic conditions. They were detected by MS/MS using electrospray ionization (ESI) in the positive mode. Quantification was performed using multiple reaction monitoring (MRM) with transitions of m/z 188.05→90.9 for selegiline and m/z 288.05→57.95 for IS. Results: Calibration curves were constructed in the concentration range of 0.2–200 ng/mL with a lower limit of quantification (LLOQ) of 0.21 ng/mL. The matrix effect of dog plasma on the selegiline signal ranged from 98.8 to 105.6%, and the mean extraction recovery ranged from 79.0% to 81.4% at concentrations of 1.04, 20.8, and 166 ng/mL. The intra-day precision was lower than 6.86% and the inter-day precisions were lower than 4.63%. Conclusion: The validation results demonstrated the reliability of this bioanalytical method, which was successfully applied to study the pharmacokinetics and bioavailability of 1.25 mg of orally administered selegiline lyophilizate in Beagle dogs. The pharmacokinetic results were also compared with those obtained following intragastric (i.g.) and intravenous (i.v.) administration. Buccal delivery of selegiline was found to significantly increase its bioavailability.