高等学校化学学报 ›› 2016, Vol. 37 ›› Issue (7): 1335-1341.doi: 10.7503/cjcu20150863

• 有机化学 • 上一篇    下一篇

低抗凝肝素寡糖的制备、 结构表征及免疫活性评价

张晓, 单鑫迪, 赵小亮, 李国云, 王小江, 蔡超, 郝杰杰, 于广利   

  1. 山东省糖科学与糖工程重点实验室, 海洋药物教育部重点实验室, 中国海洋大学医药学院, 青岛 266003
  • 收稿日期:2015-11-11 出版日期:2016-07-10 发布日期:2016-06-06
  • 基金资助:
    国家科技支撑计划项目(批准号: 2013BAB01B02)和国家自然科学基金-山东省政府联合基金(批准号: U1406402)资助

Preparation, Characterization and Immunological Activity Evaluation of Low Anticoagulant Heparin Oligosaccharides

ZHANG Xiao, SHAN Xindi, ZHAO Xiaoliang, LI Guoyun, WANG Xiaojiang, CAI Chao, HAO Jiejie, YU Guangli*()   

  1. Shandong Provincial Key Laboratory of Glycoscience and Glycotechnology, Key Laboratory of Marine Drugs, Ministry of Education, School of Medicine and Pharmacy, Ocean University of China, Qingdao 266003, China
  • Received:2015-11-11 Online:2016-07-10 Published:2016-06-06
  • Contact: YU Guangli E-mail:glyu@ouc.edu.cn
  • Supported by:
    † Supported by the National Science and Technology Support Program of China(No.2013BAB01B02) and the National Natural Science Foundation of China-Shandong Joint Fund(No;U1406402)

摘要:

以猪肠黏膜来源低抗凝肝素为原料, 采用苄酯碱水解法制备了寡糖. 通过聚丙烯酰胺凝胶电泳(PAGE)、 高效凝胶渗透色谱(HPGPC)-多角激光光散射法(MALLs)联用、 核磁共振波谱(NMR)和液相色谱-质谱(LC-MS)联用技术对所制备寡糖的分子量分布及化学结构进行了表征. 以该寡糖对活化X因子(FXa)及人凝血酶(FⅡa)活性的抑制作用评价了其抗凝血活性, 并通过测定其对RAW264.7巨噬细胞吞噬能力及NO释放量的影响评价其免疫活性. 结果表明, 所制备寡糖的聚合度分布于2~22之间, 平均分子质量为5300, 无明显的抗凝血活性, 但具有显著的免疫增强活性.

关键词: 低抗凝肝素寡糖, 苄酯碱水解, 液相色谱-质谱联用, 免疫增强活性

Abstract:

Anticoagulant activities limit applications of heparin and enoxaparin sodium in immune-promoting activity. A novel and natural low anticoagulant heparin(LAH) was purified from crude porcine intestinal. In this work, oligosaccharide of LAH(BE-LAH) was prepared by benzyl ester alkaline hydrolysis. The molecular distribution and chemical structure of BE-LAH were determined based on combination technologies including polyacrylamide gel electrophoresis(PAGE), high performance gel permeation chromatography(HPGPC)-multi angel laser light scattering(MALLs), nuclear magnetic resonance(NMR) and liquid chromatography-mass spectrum(LC-MS). Meanwhile, the anticoagulant activities were evaluated by measuring the inhibitory activities on coagulation factor Xa(FXa) and thrombin(FⅡa), while the immunological activities were evaluated by its modulation effects on phagocytic ability and NO release of RAW 264.7 cells. The results demonstrated that BE-LAH was composed of fragments from dp2 to dp22 with an average molecular weight of 5300. Expectedly, BE-LAH showed no anticoagulant activity, but remarkable immune-promoting activity. A low anticoagulant heparin oligosaccharide was prepared and promised as an excellent immunoenhancer.

Key words: Low anticoagulant heparin oligosaccharide, Benzyl ester alkaline hydrolysis, Liquid chromatography-mass spectrum, Immune-promoting activity

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