胶束电动毛细管色谱法分析头孢吡肟与其E异构体等杂质

Research on the separation of cefepime and cefepime E isomer and its related substances by micellar electrokinetic capillary chromatography

目的以十二烷基硫酸钠(SDS)胶束为准固定相,考察头孢吡肟、头孢吡肟E异构体及其他未知杂质在胶束电动毛细管色谱(MECC)分离模式下的分离行为。方法研究了运行缓冲液的pH值、磷酸盐浓度、SDS浓度、甲醇体积分数、分离电压、分离温度等因素对头孢吡肟、E异构体及其他杂质的迁移时间、分离度以及可分离出的杂质个数的影响。结果发现上述因素对头孢吡肟与诸杂质间的分离及检测有显著的影响,尤以pH值为最。它不仅影响它们的迁移时间和分离效率,还直接影响头孢吡肟及其杂质峰的检测。优化后的分离条件:运行缓冲液为70mmol/L磷酸盐/100mmol/LSDS-5%甲醇(pH7.0),分离电压为15kV,分离温度为25℃。用非涂渍石英毛细管51cm×75μm(有效长度42.5cm),压力进样5s,检测波长254nm。结论本法可分离出34个杂质,诸杂质彼此间及与头孢吡肟间可得到有效分离,可用于测定盐酸头孢吡肟的含量和有关物质。

Objective The separation of cefepime, cefepime E isomer and its unknown related substances by micellar electrokinetic capillary chromatography （MECC） using sodium dodecyl sulphate （SDS） as the micellar phase was investigated. Methods The effects ofpH, concentration of phosphate buffer solution, SDS micelle concentration ,methanol volume fraction, applied voltage and temperature on the separation were studied. Results It was found that the migration of these compounds was affected by these factors, especially by pH of the solution. The elution, as well as the migration time and separation efficiency of cefepime, cefepime E isomer and others related substances changes with the acidity of the solution. The optimized separation conditions consisted of a running buffer of 70mmol/ L sodium phosphate buffer, at pH7.0, containing 100mmol/L SDS and 5% of methanol ,with an applied voltage of 15 kV and a temperature of 25℃. The uncoated fused-silica capillary of 51cm×75μm （42.5cm of effective length） was used. The sample was injected to the column by pressure （50mbar） for 5s. The detection wavelength was set at 254 nm with cefalexin as internal standard. Conclusion 34 impurities in cefepime could be detected .Cefepime and the degradations could be separated well. The method was applied to separate and determine cefepime and its related substances successfully.