Enantiospecific determination of arotinolol in rat plasma by LC-MS/MS: application to a stereoselective pharmacokinetic study.

A highly sensitive and selective liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed and fully validated for quantification of arotinolol enantiomers in rat plasma using haloperidol as the internal standard. After solid phase extraction of 50.0 μL rat plasma in 96 well plate, a baseline resolution of arotinolol enantiomers was achieved on a CHIRALPAK AD-H column using the mobile phase of n-hexane and ethanol in 0.02% diethylamine (20:80, v/v) at a flow rate of 0.550 mL/min within 11.0 min. Acquisition of mass spectrometric data was performed on a triple-quadrupole mass spectrometer in multiple-reaction-monitoring (MRM) mode with an ESI source using the transition m/z 372.1 → 316.1 for (±)-arotinolol and m/z 376.1 → 165.1 for haloperidol. The calibration curves of both enantiomers were linear over the range of 1.00-200.0 ng/mL (r(2)>0.992) and the lower limit of quantification was 1.00 ng/mL. Intra- and inter-day precision ranged from 5.6% to 8.9% for R-(-)-arotinolol and 4.6-7.4% for S-(+)-arotinolol. Accuracy varied from 0.0% to 7.0% for R-(-)-arotinolol and 5.0-10.0% for S-(+)-arotinolol. For R-(-)-arotinolol, the recovery ranged from 87.2% to 99.2% and the matrix factor was 1.03-1.09; for S-(+)-arotinolol, the recovery ranged from 88.0% to 92.4% and the matrix factor was 0.84-0.95, both were not concentration dependent. The method was demonstrated with acceptable accuracy, precision and specificity for the determination of arotinolol enantiomers and has been successfully applied to a stereoselective pharmacokinetic study.