Abstract: In this work, a method for the determination of the amounts of the four genotoxic impurities in gefitinib has been developed. A simple and sensitive high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) approach has been developed, optimized, and validated. The genotoxic impurities were 3-chloro-4-fluoroaniline, 3,4-difluoroaniline, 3-fluoro-4-chloroaniline, and 3,4-dichloroaniline. Separation was achieved on an Inertsil ODS-3 column (100 mm×3.0 mm, 3 μm). The column temperature was 40 ℃, and the running time was 12 min. A triple quadrupole mass detector with positive electrospray ionization in the multiple reaction monitoring (MRM) mode was applied. The method was validated in terms of its specifi-city, linearity, precision, accuracy, stability, and robustness. The correlation coefficient of each impurity was higher than 0.999 in the range of 0.6-96.0 μg/L. The limits of detection (LODs) and limits of quantity (LOQs) were in the ranges of 0.2-2.0 μg/L and 0.6-6.0 μg/L, respectively. The recoveries of all impurities at 6, 30, and 60 μg/L were within 91.0%-98.5%. All impurities were stable within 2 h. After detection, only 3-chloro-4-fluoroaniline was detected in the batch number 16052301 and R16052501-1 gefitinib samples, but its concentration was below the impurity limit (6 mg/L). This method is simple, reliable, and suitable for the determination of four genotoxic impurities in gefitinib. It can be further applied as a reference for quality control.

Key words: high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS), genotoxic impurity, gefitinib