HPLC-MS Analysis of Four Potential Genotoxic Impurities in Alogliptin Pharmaceutical Materials.

BACKGROUND: Pyridine, 3-aminopyridine, 4-dimethylaminopyridine, and N, N-dimethylamine are reactive bases that may be used in the preparation of the pharmaceutical ingredient alogliptin (ALO). They are considered as potentially genotoxic impurities (PGIs) since they contain electrophilic functional groups. Therefore, they should be monitored at the allowed limits in ALO. OBJECTIVE: The aim of this study was to develop a novel liquid chromatography-mass spectrometry (LC-MS) method to estimate quantities of pyridine, 3-aminopyridine, 4-dimethylaminopyridine, and N, N-dimethylaniline impurities in ALO drug material. METHODS: The separation was performed on Kromasil CN (250 mm × 3.9 mm, 3.5 µm) column in reversed phase mode. The mobile phase was a mixture of water-methanol (55:45, v/v) containing 2.5 mM ammonium acetate and 0.1% formic acid. Impurities were detected by MS in selected ion monitoring mode at m/z = 80, 95, 122, and 123 for pyridine, 3-aminopyridine, N, N-dimethylaniline, and 4-dimethylaminopyridine, respectively. The flow rate of the method was 0.5 mL/min. RESULTS: The sensitivity of the method was excellent at levels much less than the allowed limits. The method had excellent linearity in the concentration ranges of Quantitation Limit (QL)-150% of allowed limits and coefficients of determination were above 0.9990. The recovery ratios were in the range of 93.56-110.28%. CONCLUSION: Results showed good linearity, precision, accuracy, sensitivity, selectivity, robustness, and solution stability. The studied method was applied to test two samples of raw materials and one sample of tablets. HIGHLIGHTS: The method discussed here could be very useful for controlling potentially genotoxic impurities (PGIs) levels in ALO during its synthesis, and for QC testing of ALO raw materials before using them in the preparation of pharmaceutical products.

Development and validation of an analytical method for quantitative determination of three potentially genotoxic impurities in vildagliptin drug material using HPLC-MS.

A novel high-performance liquid chromatography-mass spectrometry method was developed to determine the quantities of pyridine, 4-dimethylaminopyridine, and N, N-dimethylaniline impurities in vildagliptin drug material. These impurities are reactive bases that may be used in synthesis of vildagliptin pharmaceutical ingredients. They are considered as potentially genotoxic impurities since they contain electrophilic functional groups. Therefore, these impurities should be monitored at the allowed limits in vildagliptin. Hence a high-performance liquid chromatography-mass spectrometry method was developed to quantify the amounts of these impurities in vildagliptin. The column was KROMASIL CN (250 mm × 3.9 mm, 3.5 µm) in reversed-phase mode. The mobile phase was a mixture of water-methanol (55:45) containing 2.5 mM ammonium acetate and 0.1% formic acid. The mass spectrometer was used to detect the amounts of impurities using selected ionization monitoring mode at m/z = 80, 122, and 123 for pyridine, N, N-dimethylaniline, and 4-dimethylaminopyridine, respectively. The flow rate was 0.5 mL/min. The sensitivity of the method was excellent at levels very less than the allowed limits. The method had excellent linearity in the concentration ranges of limit of quantification-150% of the permitted level with coefficients of determination above 0.9990. The recovery ratios were in the range of 93.70-108.63%. Results showed good linearity, precision, accuracy, sensitivity, selectivity, robustness, and solution stability.

Quantitative determination of potential genotoxic impurity 3-aminopyridine in linagliptin active pharmaceutical ingredient using HILIC-UV.

This study aimed to develop an analytical method to determine the quantity of the impurity 3-aminopyridine (3AP). 3-Aminopyridine is a reactive reagent in the synthesis of linagliptin. The method was sensitive at level of 30.0 ppm of 3AP relative to linagliptin. The analysis was carried out using hydrophilic interaction liquid chromatography. The analytical column was Tracer Extrasil Silica (150 × 4.0 mm, 3 µm). A mobile phase of water-acetonitrile (10:90, v/v) containing 10.0 mM ammonium acetate was prepared and adjusted to pH 6.0. A UV detector was used to detect the amount of 3AP at a wavelength of 298 nm. Validation of the method was performed as per the International Council for Harmonization of Technical Requirements for Pharmaceuticals for Human Use in terms of detection limit, quantitation limit, linearity, accuracy, precision, specificity and robustness. The calibration curve was linear (r2 = 0.999) for 3AP concentration in the range of 30.0-450.0 ppm. This method showed a good sensitivity with a detection limit and a quantitation limit of 7.5 and 25.0 ppm, respectively.