Abstract: To develop a sensitive analytical method for the determination of the genotoxic impurity mono ethyl ester of ecabet(Imp-I), an HPLC-MS/MS technique was employed. Imp-I was synthesized according to the previous literatures. MS/MS and NMR were used to confirm the structure of Imp-I. A Thermo C₁₈ column was used for chromatographic separations. The mobile phase consisting of A: 5 mmol/L ammonium acetate(pH adjusted to 3. 0 with formic acid)and B: acetonitrile, with a gradient program: 0 min 50%B, 4 min 50%B, 12 min 80%B, 16 min 80%B, 16. 1 min 50%B and 20 min 50%B. The column was maintained at 40 °C throughout the analysis. All measurements were carried out with the mass spectrometer operated under the negative ESI mode. The selective reaction monitor(SRM)transition was used. Good linearity was obtained for Imp-I over the concentration range of 4- 150 ng/mL with the coefficient of determination(r)of 0. 999. And the LOQ was 4 ng/mL. A rapid and sensitive HPLC-ESI-MS/MS method was developed for quantitative analysis of Imp-I in ecabet sodium APIs. This method can be of used for quality assurance of ecabet sodium in bulk commercial drugs.