Solínová Veronika, Kasicka Václav, Barth Tomislav, Hauzerová Linda, Fanali Salvatore
Institute of Organic Chemistry and Biochemistry, Academy of Sciences of the Czech Republic, Flemingovo 2, 16610 Prague 6, Czech Republic.
J Chromatogr A. 2005 Jul 15;1081(1):9-18. doi: 10.1016/j.chroma.2005.01.046.
Capillary zone electrophoresis (CZE) has been applied to qualitative and quantitative analysis and separation of synthetic analogues and fragments of enkephalins ([Leu5]enkephalin, H-Tyr-Gly-Gly-Phe-Leu-OH, [Met5]enkephalin, H-Tyr-Gly-Gly-Phe-Met-OH), and dalargin (H-Tyr-D-Ala-Gly-Phe-Leu-Arg-OH), biologically active peptides with morphin-like effects acting as ligands for the opiate receptors in the brain. These oligopeptides (dipeptides to hexapeptides) were analyzed as cations in two acidic background electrolytes (BGEs), BGE I (100mM H3PO4, 50mM Tris, pH 2.25), BGE II (100mM iminodiacetic acid, pH 2.30), and both as cations and anions in alkaline BGE IV (40 mM Tris, 40 mM Tricine, pH 8.10). Purity degrees of peptides, expressed in three different ways (relative peak height, relative peak area and relative corrected peak area), were determined by their CZE analyses in the above BGEs, and their values were compared with respect to the peak shapes and migration times of the main synthetic products and their admixtures. Selected analogues and fragments of enkephalins and dalargin were successfully separated by CZE in acidic isoelectric buffers, 100 and 200 mM iminodiacetic acid, pH 2.30 and 2.32, respectively. The effective electrophoretic mobilities at standard temperature 25 degrees C, and effective and specific charges of all analyzed peptides in the above three BGEs were determined. Correlation between effective electrophoretic mobility of the analyzed peptides and their charge and size (relative molecular mass) was investigated, which revealed different molecular shape of analyzed peptides in acidic and alkaline BGEs. In addition, the selected characteristics of the UV-absorption detector (noise, signal to noise ratio, sensitivity, and limits of detection and quantification) were determined.
毛细管区带电泳（CZE）已应用于脑啡肽（[亮氨酸5]脑啡肽，H-Tyr-Gly-Gly-Phe-Leu-OH，[甲硫氨酸5]脑啡肽，H-Tyr-Gly-Gly-Phe-Met-OH）以及达勒argin（H-Tyr-D-Ala-Gly-Phe-Leu-Arg-OH）的合成类似物和片段的定性、定量分析与分离，这些具有吗啡样作用的生物活性肽可作为大脑中阿片受体的配体。这些寡肽（二肽至六肽）在两种酸性背景电解质（BGE）中作为阳离子进行分析，即BGE I（100mM磷酸，50mM Tris，pH 2.25）、BGE II（100mM亚氨基二乙酸，pH 2.30），并且在碱性BGE IV（40mM Tris，40mM三（羟甲基）甲基甘氨酸，pH 8.10）中作为阳离子和阴离子进行分析。通过在上述BGE中进行CZE分析，以三种不同方式（相对峰高、相对峰面积和相对校正峰面积）表示的肽纯度得以确定，并将其值与主要合成产物及其混合物的峰形和迁移时间进行比较。脑啡肽和达勒argin的选定类似物和片段在酸性等电缓冲液（分别为100和200mM亚氨基二乙酸，pH 2.30和2.32）中通过CZE成功分离。测定了在标准温度25℃下所有分析肽的有效电泳迁移率，以及在上述三种BGE中的有效电荷和比电荷。研究了分析肽的有效电泳迁移率与其电荷和大小（相对分子质量）之间的相关性，结果表明在酸性和碱性BGE中分析肽的分子形状不同。此外，还测定了紫外吸收检测器的选定特性（噪声、信噪比、灵敏度以及检测限和定量限）。
