| Abstract: | A highly sensitive, specific and enantioselective assay has been validated for the quantitation of OTX015 enantiomers (+)‐OTX015 and (?)‐OTX015] in mice plasma on LC–MS/MS‐electrospray ionization as per regulatory guidelines. Protein precipitation was used to extract (±)‐OTX015 enantiomers and internal standard (IS) from mice plasma. The active (?)‐OTX015] and inactive (+)‐OTX015] enantiomers were resolved on a Chiralpak‐IA column using an isocratic mobile phase (0.2% ammonia/acetonitrile 20 : 80, v /v) at a flow rate of 1.2 mL/min. The total run time was 6.0 min. (+)‐OTX015, (?)‐OTX015 and IS eluted at 3.34, 4.08 and 4.77 min, respectively. The MS/MS ion transitions monitored were m/z 492 → 383 for OTX015 and m/z 457 → 401 for IS. The standard curves for OTX015 enantiomers were linear (r 2 > 0.998) in the concentration range 1.03–1030 ng/mL. The inter‐ and intraday precisions were in the range 2.20–13.3 and 8.03–12.1% and 3.80–14.4 and 8.97–13.6% for (+)‐OTX015 and (?)‐OTX015, respectively. Both the enantiomers were found to be stable in a battery of stability studies. This novel method has been applied to the study of stereoselective oral pharmacokinetics of (?)‐OTX015 and unequivocally demonstrated that (?)‐OTX015 does not undergo chiral inversion to its antipode in vivo in mice. |
