二维超高效液相色谱-QTof质谱联用技术在来氟米特片杂质谱研究中的应用

目的：建立二维超高效液相色谱-QTof质谱联用研究来氟米特片杂质谱的方法。方法：通过二维液相色谱在线脱盐技术，在不改变一维液相色谱条件的基础上，将目标化合物通过阀切换转换至二维液相色谱柱，在二维液相流动相的带动下进质谱，进行结构鉴定。一维色谱柱为Inertsil ODS-3（250 mm×4.6 mm 5 μm），流动相为0.025 mol·L^-1的磷酸二氢钾溶液（用磷酸调节pH至3.0）-乙腈（57：43），检测波长为210 nm；二维液相色谱柱为Waters BEH C₁₈（50 mm×2.1 mm 1.7 μm），流动相A为0.1%甲酸溶液，流动相B为0.1%甲酸乙腈溶液，梯度洗脱；采用ESI负离子模式进行质谱扫描，离子源温度120℃，雾化气温度500℃，雾化气流速900 L·h^-1，采集模式为MS^E。结果：对样品中存在的杂质进行分析，解析出可能杂质结构4个，部分结构片段3个；对杂质的毒性预测显示，杂质7具高毒性风险，所有杂质遗传毒性均属5级，没有警示结构。结论：本法建立了二维超高效液相色谱-QTof质谱联用技术测定、分析来氟米特片杂质的方法，可快速实现杂质定性，结合软件解析结构更便捷；国内产品与参比制剂Sanofi产品相比，杂质谱更复杂，建议生产企业优化处方、生产工艺，保证药品的质量稳定性。

Objective: To establish a two-dimensional UPLC-QTof MS method for the study of impurity profile of leflunomide tablets.Methods: By two-dimensional liquid chromatography online desalination technology,there was no need to change the onedimensional liquid chromatography conditions,and converse the target compound to two-dimensional liquid chromatography column by valve switching,which led it into the mass spectrometer under the second-dimensional mobile phase elution,and the compound structure was identified.First-dimensional column was Inertsil ODS-3(250 mm×4.6 mm 5 μm),the mobile phase was 0.025 mol·L^-1 potassium dihydrogen phosphate solution(adjust pH to 3.0 using phosphoric acid)-acetonitrile(57:43),detection wavelength:210 nm;second-dimensional column was Waters BEH C₁₈(50 mm×2.1 mm 1.7 μm),mobile phase A was 0.1% formic acid solution,mobile phase B was 0.1% formic acid acetonitrile solution,gradient elution. Electrospray ionization(ESI) source was used as negative ion detection model,the temperature of ESI source was 120℃,nebulizing temperature was 500℃,nebulizing flow rate was 900 L·h^-1,and the acquisition mode was MS^E.Results: The existing impurities in the sample were determined,and four possible impurities structure and three structure segment were obtained;toxicity prediction of impurities showed that impurity 7 had high risk of toxicity,all impurity's genetic toxicity is level 5 with no warning structure.Conclusion: This article was focused on the method establishment to determine leflunomide impurities by two-dimensional UPLC-QTof MS technology.The qualitative of impurity was quickly obtained by using 2-dimensional HPLC-MS,and it is convenient to identify the structure of impurities combining the software.The domestic products' impurity profile was more complex compared with Sanofi products,which can suggest production enterprises to optimize the prescription and production process to ensure the quality stability of the drug.