Abstract: Objective To establish an analytical method based on UPLC-MS/MS for the determination of genotoxic impurities in clozapine. Methods The impurities were separated using an Inert Sil-ODS-3 (75 mm×?.? mm×? mm) with a mobile phase composed of 0.1% formic acid aqueous solution (mobile phase A) and acetonitrile (mobile phase B) by gradient elution at a flow rate of 0.7 mL·min-1. Multiple reaction monitoring (MRM) was performed on a mass spectrometer equipped with an ESI source in the negative mode. Results The linearity of anthranilic acid, cyclized and condensed compounds, and cyclized acid precipitates was desirable within ranges of 20 to 100 ng·mL-1, 0.16 to 3.2 ng·mL-1 and 0.48 to 3.2 ng·mL-1. The recoveries of spiked samples at low（80%）, medium（100%） and high（120%）concentrations ranged from 90.8% to 117.2%, and RSD was between 3.9% and 12.8%. The limit of detection ranged from 0.08 to 10 ng·mL-1 and the limits of quantification of anthranilic acid, cyclized and condensed compounds, and cyclized acid precipitates were 20 ng·mL-1, 0.16 ng·mL-1 and 0.48 ng·mL-1, separately. The detected concentration was below the limit. Conclusion The established method is simple and reliable, which is applicable to quantifications of three genotoxic impurities in clozapine.

Key words: quantification of anthranilic acid, cyclized and condensed compounds, cyclized acid precipitates, UPLC-MS/MS, clozapine, genotoxic impurity, analytical method