Zhou Lei, Wang Weiping, Wang Shumin, Hui Yang, Luo Zhi, Hu Zhide
Department of Chemistry, Lanzhou University, Lanzhou 730000, China.
Anal Chim Acta. 2008 Mar 24;611(2):212-9. doi: 10.1016/j.aca.2008.01.084. Epub 2008 Feb 13.
A novel method based on separation by nonaqueous capillary electrophoresis (NACE) combined with laser-induced fluorescence (LIF) detection was developed and compared with classic aqueous modes of electrophoresis in terms of resolution of solutes of interest and sensitivity of the fluorescence detection. Catecholamines derivatized with 4-chloro-7-nitro-2,1,3-benzoxadiazole (NBD-Cl) were chosen as test analytes for their subtle fluorescence properties. In aqueous systems, capillary zone electrophoresis (CZE) was not suitable for the analysis of test analytes due to complete fluorescence quenching of NBD-labeled catecholamines in neat aqueous buffer. The addition of micelles or microemulsion droplets into aqueous running buffer can dramatically improve the fluorescence response, and the enhancement seems to be comparable for micellar electrokinetic chromatography (MEKC) and microemulsion electrokinetic chromatography (MEEKC). As another alternative, NACE separation was advantageous when performing the analysis under the optimum separation condition of 20mM sodium tetraborate, 20mM sodium dodecyl sulfate (SDS), 0.1% (v/v) glacial acetic acid, 20% (v/v) acetonitrile (ACN) in methanol medium after derivatization in ACN/dimethyl sulfoxide (DMSO) (3:2, v/v) mixed aprotic solvents containing 20mM ammonium acetate. Compared with derivatization and separation in aqueous media, NACE-LIF procedure was proved to be superior, providing high sensitivity and short migration time. Under respective optimum conditions, the NACE procedure offered the best fluorescence response with 5-24 folds enhancement for catecholamines compared to aqueous procedures. In addition, the mechanisms of derivatization and separation in nonaqueous media were elucidated in detail.
开发了一种基于非水毛细管电泳（NACE）分离并结合激光诱导荧光（LIF）检测的新方法，并在目标溶质的分离度和荧光检测灵敏度方面与经典的水性电泳模式进行了比较。选择用4-氯-7-硝基-2,1,3-苯并恶二唑（NBD-Cl）衍生化的儿茶酚胺作为测试分析物，因为它们具有微弱的荧光特性。在水性体系中，毛细管区带电泳（CZE）不适用于测试分析物的分析，因为在纯水性缓冲液中NBD标记的儿茶酚胺会完全荧光猝灭。向水性运行缓冲液中添加胶束或微乳液滴可显著改善荧光响应，并且对于胶束电动色谱（MEKC）和微乳液电动色谱（MEEKC），这种增强效果似乎相当。作为另一种选择，在乙腈/二甲基亚砜（DMSO）（3:2，v/v）混合非质子溶剂中用20mM乙酸铵衍生化后，在甲醇介质中20mM硼酸钠、20mM十二烷基硫酸钠（SDS）、0.1%（v/v）冰醋酸、20%（v/v）乙腈（ACN）的最佳分离条件下进行NACE分离是有利的。与在水性介质中的衍生化和分离相比，NACE-LIF方法被证明更优越，具有高灵敏度和短迁移时间。在各自的最佳条件下，与水性方法相比，NACE方法对儿茶酚胺的荧光响应最佳，增强了5-24倍。此外，还详细阐明了非水介质中衍生化和分离的机制。
