HPLC法检测醋氯芬酸有关物质及主要杂质鉴别

目的：建立HPLC法检测醋氯芬酸有关物质，并采用NMR、MS法对主要杂质进行结构鉴定。方法：采用Shim-pack VP-ODS（250 mm×4.6 mm，5 μm）色谱柱，以1.12 mg·mL^-1磷酸溶液（用1.0 mol·L^-1氢氧化钠溶液调节pH至7.0）为流动相A，乙腈为流动相B，梯度洗脱，流速1.0 mL·min^-1，检测波长276 nm，柱温25℃。用半制备液相色谱分离纯化主要杂质，采用SunFire C₁₈色谱柱，0.50 mg·mL^-1磷酸溶液-甲醇为流动相梯度洗脱，通过NMR、ESI-MS确定结构。结果：主要杂质为双氯芬酸乙酯、醋氯芬酸乙酯、醋氯芬酸叔丁酯。建立的HPLC有关物质分析方法专属性强，各杂质在1.6~17.6 μg·mL^-1范围内呈良好的线性关系，回收率在92%~105%之间，准确度高。对5批醋氯芬酸样品进行检测，结果符合规定。结论：建立的方法可作为醋氯芬酸原料药的质控方法。

Objective:To establish an HPLC method for determination of related substances in aceclofenac and to identify the structure of the main impurity by NMR and MS.Methods:A Shim-pack VP-ODS(250 mm×4.6 mm,5 μm)was adopted for the study.The mobile phase A was 1.12 mg·mL^-1 phosphoric acid solution(adjusted to pH 7.0 using 1.0 mol·L^-1 sodium hydroxide solution),and the mobile phase B was acetonitrile with gradient elution at the flow rate of 1.0 mL·min^-1,the detection wavelength was 276 nm,and the column temperature was 25℃.The main impurity was separated and purified by semi-preparative HPLC.The determination was performed on a SunFire C₁₈ column and using 0.50 mg·mL^-1 phosphoric acid solution-methanol as the mobile phase with gradient elution.The structure was confirmed by NMR and ESI-MS.Results:The main impurity was ethyl ester of diclofenac,ethyl ester of aceclofenac and benzeneacetic acid,respectively.The established method for the determination of related substances in aceclofenac was specific,and the impurities showed a good linearity in the range of 1.6-17.6 μg·mL^-1.The recovery rates were 92%-105%.The results of 5 batches of aceclofenac sample met with the requirement.Conclusion:The established HPLC method can be applied in the quality control of the aceclofenac bulk drug.