荧光标记电泳对透明质酸寡糖片段鉴定的方法建立

近年来，透明质酸寡糖片段（hyaluronan oligosaccharides, o-HA）的生物学活性引起国外学者的重视，因为o-HA具有一定的生物学活性，如参与免疫调节、刺激新生血管形成等.本研究建立一种经济、简便的ANTS（8-氨基奈-1,3,6-三磺酸）荧光标记电泳对透明质酸寡糖片段大小鉴定的新实验方法.实验原理为，ANTS能与糖分子发生还原反应，在反应时提供3个电子和1个荧光基团，通过高浓度PAGE分离，在特定波长下呈现颜色反应.采用酶消化法得到不同分子量大小的o-HA片段，测得不同片段大小的o-HA聚合度，分别与高效液相色谱（high-performance liquid chromatography, HPLC）和静电喷雾电离质谱（electrospray ionization mass spectrometry, ESI-MS）进行比较，结果吻合.研究提示，用荧光标记电泳法分析寡糖分子量，操作简单、设备低廉、灵敏度较高且检测速度快，是一种检测鉴定寡糖分子的较好方法.

Application of a Novel Fluorophore-labeled Electrophoresisto Analysis of Hyaluronan Oligosaccharides

To establish a novel and simple electrophoresis for characterize hyaluronan oligosaccharides, native high molecular weight hyaluronan was digested into small fragments, and the oligosaccharides were labeled with ANTS (8-amino naphthalene-1, 3, 6-trissulfonic acid) that provided three negative charges and a fluorophore group to saccharides. The oligosaccharides were then separated by a high voltage polyacrylamide gel and electrophoresis under 4-8℃ condition. The bands ongel were determined on 306 nm and compared with electrospray ionization mass spectrometry (ESI-MS) and HPLC, respectively. The ANTS labeled electrophoresis could determine hyaluronan oligosaccharide molecules as small as 2 disaccharides. All the molecular weights of hyaluronan oligosaccharides on PAGE were the same as they were on ESI-MS and HPLC. Our results showed that the fluorophore-labeled electrophoresis might be a simple, sensitive, and rapid method to characterize oligosaccharides molecules.