Kang J W, Van Schepdael A, Roets E, Hoogmartens J
Laboratorium voor Farmaceutische Chemie en Analyse van Geneesmiddelen, Faculteit Farmaceutische Wetenschappen, Katholieke Universiteit Leuven, Belgium.
Electrophoresis. 2001 Aug;22(12):2588-92. doi: 10.1002/1522-2683(200107)22:12<2588::AID-ELPS2588>3.0.CO;2-X.
A fast and highly selective micellar electrokinetic capillary chromatography (MEKC) method for quantitative analysis of vancomycin and related impurities is described. Among the tested surfactants, cetyltrimethylammonium chloride (CTAC) offered the best selectivity. Another important parameter, which strongly influenced the selectivity, was buffer pH. It was found that the selectivity increased with buffer pH decreasing from 9 to 5. Using Tris-phosphate buffer containing CTAC, satisfactory separation could be obtained in the pH range from 5.0 to 5.5. Excellent repeatability in terms of migration time and peak area could be obtained when the capillary was carefully washed between two runs. In order to obtain optimal conditions and to evaluate the method robustness, a central composite experimental design was carried out. The optimal conditions were: 44 cm length of fused-silica capillary with 50 microm ID, 120 mM Tris-phosphate buffer (pH 5.2) containing 50 mM CTAC, -15 kV applied voltage, UV detection at 210 nm, and a column temperature of 25 degrees C. Under the optimal conditions, more than 20 peaks could be separated within 8 min. The method has a linearity range from 0.004 to 1.2 mg/ml (concentration of vancomycin B, active component). The limit of detection (LOD) and limit of quantitation (LOQ) were 0.4 microg/mL vancomycin, equivalent to 0.3 microg/mL vancomycin B (0.04%) and 1.1 microg/mL vancomycin, equivalent to 0.9 microg/mL vancomycin B (0.1%), respectively.
本文描述了一种用于定量分析万古霉素及其相关杂质的快速且高选择性的胶束电动毛细管色谱法（MEKC）。在测试的表面活性剂中，十六烷基三甲基氯化铵（CTAC）具有最佳的选择性。另一个对选择性有强烈影响的重要参数是缓冲液的pH值。研究发现，随着缓冲液pH值从9降至5，选择性增加。使用含有CTAC的磷酸三乙胺缓冲液，在pH值为5.0至5.5的范围内可获得令人满意的分离效果。当两次进样之间仔细冲洗毛细管时，迁移时间和峰面积方面可获得出色的重复性。为了获得最佳条件并评估该方法的稳健性，进行了中心复合实验设计。最佳条件为：内径50微米的熔融石英毛细管长度为44厘米，含50 mM CTAC的120 mM磷酸三乙胺缓冲液（pH 5.2），施加电压-15 kV，210 nm处紫外检测，柱温25℃。在最佳条件下，8分钟内可分离出20多个峰。该方法的线性范围为0.004至1.2 mg/ml（万古霉素B，活性成分的浓度）。检测限（LOD）和定量限（LOQ）分别为0.4微克/毫升万古霉素，相当于0.3微克/毫升万古霉素B（0.04%）和1.1微克/毫升万古霉素，相当于0.9微克/毫升万古霉素B（0.1%）。
