A novel condition for capillary electrophoretic analysis of reductively aminated saccharides without removal of excess reagents
A novel condition for capillary electrophoretic analysis of reductively aminated saccharides without removal of excess reagents
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DOI：
10.1002/elps.201200612
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发表时间：
2013-12
期刊：
影响因子：
2.9
通讯作者：
Masahiro Yodoshi;Natsumi Ikeda;Naoko Yamaguchi;Mana Nagata;N. Nishida;K. Kakehi;T. Hayakawa;Shigeo Suzuki
中科院分区：
文献类型：
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作者：
Masahiro Yodoshi;Natsumi Ikeda;Naoko Yamaguchi;Mana Nagata;N. Nishida;K. Kakehi;T. Hayakawa;Shigeo Suzuki
我们已经确定了从糖蛋白中分离 7-氨基-4-甲基香豆素标记的单糖和寡糖的新毛细管电泳条件。使用中性涂层毛细管和含有羟丙基纤维素和乙腈的碱性硼酸盐缓冲液，糖衍生物形成阴离子硼酸盐复合物，在电场中从阴极移动到阳极，并在阳极端附近检测到。过量的标记试剂和其他荧光产物保留在阴极端。使用 LED 作为光源的荧光检测能够测定至少 0.4 至 400 μM 之间具有良好线性的单糖衍生物，可能对应于 140 amol 至 140 fmol。样品溶液中的较低 LOD (S/N = 5) 仅 80 nM (约 28 amol)。结果与使用荧光检测 LC 报告的值相当。该方法还应用于分析从糖蛋白酶促释放的寡糖。精细的分辨率可以对糖蛋白中的聚糖进行分析。通过将该方法应用于其他用非酸性标签标记的衍生物（如对氨基苯甲酸乙酯和 2-氨基吖啶酮标记的糖类）来检验该方法的适用性。
We have identified novel CE conditions for the separation of 7‐amino‐4‐methylcoumarin‐labeled monosaccharides and oligosaccharides from glycoproteins. Using a neutrally coated capillary and alkaline borate buffer containing hydroxypropylcellulose and ACN, saccharide derivatives form anionic borate complexes, which move from the cathode to the anode in an electric field and are detected near the anodic end. Excess labeling reagents and other fluorescent products remain at the cathodic end. Fluorimetric detection using an LED as a light source enables determination of monosaccharide derivatives with good linearity between at least 0.4 and 400 μM, may correspond to 140 amol to 140 fmol. The lower LOD (S/N = 5) is only 80 nM in the sample solution (ca. 28 amol). The results were comparable to reported values using fluorometric detection LC. The method was also applied to the analysis of oligosaccharides that were enzymatically released from glycoproteins. Fine resolution enables profiling of glycans in glycoproteins. The applicability of the method was examined by applying it to other derivatives labeled with nonacidic tags such as ethyl p‐aminobenzoate‐ and 2‐aminoacridone‐labeled saccharides.