Abstract: OBJECTIVE To establish an UPLC method for determination of related substances of Cetrorelix Acetate injection. METHODS The separation was performed on ACQUITY UPLC CSHTMC₁₈ column(2.1 mm×150 mm, 1.7 μm), 0.05 mol·L^-1 sodium perchlorate solution(adjust the pH with phosphoric acid to 2.0) was used as mobile phase A, and 0.05 mol·L^-1 sodium perchlorate solution-acetonitrile(30︰70)(adjust the pH with phosphoric acid to 2.0) was used as mobile phase B, with a gradient elution. Flow rate was 0.2 mL·min^-1, the detection wavelength was 226 nm and sample volume was 1 μL. RESULTS The linear ranges of impurity A, B, C, D, E and F were 0.127 8-6.390 4, 0.124 4-6.221 3, 0.126 8-6.340 8, 0.120 5-6.025 6, 0.120 9-6.047 3, 0.127 7-6.389 4 μg·mL^-1, respectively. The limits of detection of each impurities was about 0.04 ng. The limits of quantitation of impurity B, D, E were about 0.12 ng, that of impurity A, C, F were about 0.13 ng. The average recoveries were 97.4%, 95.2%, 101.5%, 105.3%, 100.7%, 97.4%, respectively. RSDs were 1.7%, 3.0%, 3.8%, 2.4%, 0.8%, 2.8%(n=9), respectively. CONCLUSION The method is rapid, simple, accurate and specific and can be used for the quantitative study on related substances in Cetrorelix Acetate for injection.