Development and Validation for the Quantification of Genotoxic Impurity in Mirabegron Drug Substance by LCMS

The mirabegron drug substance was shown to have potential genotoxic impurity (PGI); hence, it needs to develop the method for the determination of the 2-(2-Nitrophenyl) ethylamine hydrochloride (4-NPE RC-2) PGI at a 30-ppm limit considering the maximum daily dose of 50 mg/day and a 1.5 mg per day TTC limit as per ICH M7 guideline. A simple high-performance liquid chromatography (HPLC) method can be used due to the chromophore nature of the impurity, but due to sample matrix interference on the HPLC method, a selective and specific method was developed using liquid chromatography-mass spectrometry (LC-MS/MS) method. Using an Inertsil ODS 3-V column, gradient elution mode, 0.8 mL per min flow rate, 0.2% formic acid aqueous solution as mobile phase A, and acetonitrile solvent as mobile phase B was used for better chromatographic separation. Triple quadrupole mass detection with electrospray ionization (ESI) in multiple reaction monitoring (MRM) mode was used for determination and quantification. For this PGI, the procedure was thoroughly verified with good linearity across the mirabegron test concentration range of 9-45 ppm. Linearity was observed with an exceeding 0.990 correlation coefficient and found accuracy in terms of recovery between 100.0% and 103.0% with below 5.0% relative standard deviation.