Bioanalytical Method Development and Validation of Ritlecitinib Using Crizotinib as an Internal Standard in Rat Plasma and its Application to Pharmacokinetics Study

liIn this work, the amount of Ritlecitinib in rat plasma was determined using a liquid chromatography- tandem mass spectrometry approach. Crizotinib is an internal standard(IS). With a 3.5µm particle size and dimensions of 150 x 4.6 mm, the Waters X-Terra RP-18 column was used for the chromatographic separation. Acetonitrile: TEA pH- 2.5/Formic acid in a 60:40% v/v ratio was used as the mobile phase at a flow rate of 1.0 mL/min. The drug and IS were extracted using the liquid-liquid extraction technique. Proton adducts of Ritlecitinib and Crizotinib were detected at m/z 286.159-105.7458 and 451.3378-282.4251, respectively, in the positive mode of multiple reaction monitoring (MRM). The validation was carried out within the linear concentration range of 12.50–100.0 ng/mL. The coefficient of variation (%CV) for Ritlecitinib was determined to be 2.80% and 96.47%. Further, the precision and accuracy results for HQC, MQC, LQC, and LLQC were determined to be 98.02%, 98.42%, 98.62%, and 96.13%. With this method the average recovery rate was found to be 98.78%.