A Comprehensive LC-MS/MS Method for Detecting Genotoxic Nitrosamine Impurities in Favipiravir API
Indian Journal of Pharmaceutical Education and Research
Abstract
Background: Favipiravir, a purine nucleic acid analogue (T-705), was used to treat SARS-CoV-2 patients. Existing methods often target specific nitrosamines in various matrices but not necessarily Favipiravir. These findings highlight the lack of a comprehensive method for detecting all seven potential nitrosamine impurities in Favipiravir API. Materials and Methods: The current approach discusses the trace-level quantification of nitrosamine impurities (NDEA, NDIPA, NEIPA, NMBA, NMPA, NDMA and NDBA) in Favipiravir API. The set of nitrosamine impurities were separated on gradient elution mode (1.0%v/v formic acid and 100.0% methanol) throughout a 20 min run period using Symmetry C18 (150X4.6 mm, 5 µm) with a flow rate of 0.8 mL/min. The Column oven was saturated to attain a temperature of 40º±1.0ºC, where the auto sampler was maintained at 5 . A rinse volume of 1200 µL was employed before and after aspiration, with a dip time of 5 seconds. All the nitrosamine impurities were quantified and ionized in positive polarity mode of Electron Spray Ionization (ESI) using Multiple Reaction Monitoring (MRM). Results: The retention times of the impurities NDEA, NDIPA, NEIPA, NMBA, NMPA, NDMA, NDBA and Favipiravir were found to be 8.51, 10.54, 9.60, 6.45, 11.06, 5.41, 12.67 and 6.96 min respectively. % Individual impurity in un spiked test solution has not been detected with any of the impurities. 1.29-1.98 was the %RSD for precision with 6-time repetitions Linearity was tested for specific impurities at six levels (5-100 ng/mL), with correlation coefficients (r2 ) ranging from 0.995-0.999. The percentage recovery of each impurity at LOQ level was observed to be 83.7-107.2, whereas 91.1-101.8 at three levels of accuracy (50%, 100%, 200%) injected in triplicate. Conclusion: This novel LC-MS/ MS method effectively separates and quantifies seven nitrosamine impurities in Favipiravir API, offering a reliable tool for quality control. This study validates an LC-MS/MS method for detecting nitrosamine impurities in Favipiravir API according to International Conference on Harmonization (ICH) guidelines
Keywords
- NEIPA
- NDIPA
- NMPA
- Favipiravir API
- NDBA
- Genotoxic impurities
- ESI(+ve).