Simultaneous determination of genotoxic nitrosamine impurities in sitagliptin/metformin combination tablets:...

Genotoxic nitrosamine impurities have increasingly posed substantial safety concerns for pharmaceutical formulations, particularly sitagliptin/metformin combination tablets. In particular, N-nitrosodimethylamine (NDMA) and 7-N-nitroso-3-(trifluoromethyl)-5,6,7,8-tetrahydro[1,2,4]triazolo[4,3-a]pyrazine (NTTP)-a newly identified nitrosamine drug-substance-related impurity (N-nitroso-sitagliptin or N-nitroso-STG-19)-have been detected in these pharmaceutical formulations. However, there are no established analytical methods addressing the simultaneous quantification of these two distinct classes of nitrosamine impurities in a single analysis. Therefore, we aimed to develop and validate a robust analytical approach utilising liquid chromatography-mass spectrometry (LC-MS) coupled with atmospheric-pressure chemical ionisation (APCI). Comparison of the ionisation modes and mass analysers revealed that LC-APCI-MS significantly outperformed electrospray ionisation for NDMA, whereas both ionisation methods were suitable for NTTP. Furthermore, both high-resolution quadrupole time-of-flight and triple quadrupole mass spectrometry platforms met the regulatory validation criteria, with excellent linearity (R² > 0.997), recovery (95-102%), and precision (RSD < 5.7%) observed across both instruments. In contrast, GC-MS analysis, one of the major methods for NDMA or nitrosamine detection, was unsuitable for NTTP quantification because of inadequate volatility and chromatographic resolution. As sitagliptin/metformin remains the primary therapy for type 2 diabetes, the proposed method offers a practical solution for quality control monitoring and ensures a safe and stable supply of this widely used combination product.