AQBD Assisted Development and Validation of a Stability-Indicating UPLC Method for Assay of Pirtobrutinib in Bulk and Pharmaceutical Formulation

Abstract

Objective: Utilizing the principles of Analytical Quality by Design (AQbD), a stability-indicating UPLC method for estimating Pirtobrutinib in bulk and formulation was developed. Critical method variables were identified by risk assessment using an Ishikawa diagram, and multivariate optimization of the experimental conditions for the Ultra-Performance Liquid Chromatography (UPLC) technique was accomplished by central composite design using the design of experiments (DoE) software. Methods: Elution was achieved by using an Acquity UPLC BEH Shield RP-18 column (50 mm × 1.0 mm, 1.7 µm), mobile phase composed of Acetonitrile and Buffer in the ratio of 27:73 v/v, 0.3 ml/min flow rate and 5 µl** injection volume with PDA detection at 229 nm. In the developed method the retention time (RT) of Pirtobrutinib was 1.174 min, but from AQBD the RT of Pirtobrutinib was selected as 0.853 min. The concentrations of LOD (Limit of Detection) and LOQ (Limit of Qantification) were found to be 0.4 µg/ml and 1.2 µg/ml, respectively. Results: The developed method was validated as per ICH Q2 (R1) guidelines and demonstrated linearity in the range of 20-120 µg/ml (R2 = 0.99966) with good accuracy (99.8-101.8 % recovery) and precision (%RSD 0.72). Forced degradation studies, such as acid, alkali, oxidative stress etc., were under limits. The results were found to be within the acceptable limits of USFDA (United States Food and Drug Administration). Conclusion: Thus, the proposed method was sensitive, simple, precise, and suitable for stability testing of Pirtobrutinib, which can be employed for routine analysis.