Development and Validation of RP- HPLC Method for Baricitinib using Quality by Design Approach and its Application to Stability Studies

The present paper reports a simple, sensitive, precise, and robust reverse phase high performance liquid chromatography (RP-HPLC) method using quality by design (QbD) approach and has been developed and validated for analysis of baricitinib in bulk drug. Design of experiment (DoE) using a Box Behnken design approach was employed for method development and optimization where the critical method variables like mobile phase pH, gradient time, flow rate, and the interaction effects on the drug response parameters i.e., retention time, NTP, asymmetry factor were evaluated. Method: The optimal chromatographic separation was carried out by gradient elution mode on a ZORBAX ODS 250x4.6mm, 5 um column using ammonium formate buffer (pH 7): acetonitrile (ACN) as mobile phase at 25°C with a flow rate of 1-mL/min and injection volume of 10ul. Quantitation was achieved using UV detection at 251nm on Waters Alliance 2695 system with a PDA detector. Result: The retention time for baricitinib was found to be 8.14 minutes. The calibration curve was linear over a range of 1-3 ug/mL with limit of detection (LoD) and limit of quantitation (LoQ) values found to be 0.1 ug/mL and 0.5 ug/mL, respectively. The percent recovery was found to be within an acceptable limit of 98-102%. Forced degradation studies were carried out under acid, base, oxidative, photolytic, and thermal conditions indicating the well-resolved peak of drug and degradation products. Conclusion: The optimized chromatographic method was validated as per ICH Q2 (R1) guidelines and proved to be accurate, precise, specific, linear, and robust; also, all the parameters were within acceptance criteria. Forced degradation studies showed that the method developed was specific and can be employed for monitoring the stability of Baricitinib.