Chromatographic Separation and Quantification of Nine Nitrosamine Genotoxic Impurities in a Single Method in Nebivolol Tablet by Using Validated Ultra-Sensitive Liquid Chromatography: Mass Spectrometry Analytical Method.

Abstract

N-nitrosamine impurities have been detected in a vast variety of drug substances and drug products, showing concern for regulatory aspects. To meet the regulatory requirement for the concerned impurity, a sensitive analytical method capable of quantifying these impurities at a lower level with accuracy and precision is required. This article focuses on the development and validation of an analytical method for the simultaneous detection of nine nitrosamine impurities in a single method for nebivolol drug product using liquid chromatography-mass spectrometry/mass spectrometry-atmospheric pressure chemical ionization (LC-MS/MS-APCI). The chromatographic separation was performed using the LC-MS column Allure BiPh C18 (250 × 4.6 mm), 5 μm employed a gradient mode elution program using 0.002 M Ammonium acetate buffer pH 4.5 as mobile phase A and methanol as mobile phase B. The method was challenged for accuracy, precision and linearity in accordance with International Council for Harmonization guidelines to ensure its suitability for the intended usage. The developed method was specific, accurate and linear with square of correlation coefficient (r2) found to be greater than 0.99 (0.9970-0.9992). The LOQ obtained in the range of 9.85-19.62 ppb for nine nitrosamines showed good sensitivity. The results demonstrated that method can be applied to quantify the nitrosamines in nebivolol drug products.