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目的对来源于沙蚕(Perinereis aibuhitensis)的硫酸多糖进行结构表征和抗凝血活性研究。方法采用两步酶解法从沙蚕中提取多糖;利用强阴离子交换色谱和凝胶渗透色谱对粗多糖进行分离纯化;通过离子色谱法、高效液相色谱法(HPLC)、高效凝胶渗透色谱-多角度激光光散射仪(HPGPC-MALLS)联用技术、硫酸软骨素酶ABC酶法分析、核磁共振氢谱(1 H-NMR)等方法,研究纯化多糖的化学组成和结构特征;通过测定APTT、PT和TT评价其体外抗凝血活性。结果从沙蚕中纯化得到了2种硫酸多糖组分PAE1和PAE2,PAE1主要由Gal、Glc、GalN、GlcA及少量Fuc组成;PAE2主链为硫酸软骨素C[→4GlcAβ1→3GalNAc6Sβ1→],支链主要由Fuc、Gal和Glc构成。PAE1和PAE2均可明显延长APTT和PT。结论首次从沙蚕中提取、分离得到2种硫酸多糖,其中PAE2是1种含有Fuc、Gal与Glc支链并具有明显的体外抗凝血活性的结构新颖的类硫酸软骨素,该发现为沙蚕硫酸多糖的开发提供了依据。
Objective To study the structural characterization and anticoagulant activity of sulfated polysaccharides derived from Perinereis aibuhitensis. Methods Polysaccharide was extracted from Nereis through a two-step enzymatic method. The crude polysaccharides were separated and purified by strong anion exchange chromatography and gel permeation chromatography. Their structures were characterized by ion chromatography, high performance liquid chromatography (HPLC), high performance gel permeation chromatography The chemical composition and structural characteristics of purified polysaccharides were studied by HPGPC-MALLS, enzymatic chondroitinase ABC, 1H-NMR and other methods. The APTT , PT and TT evaluated their in vitro anticoagulant activity. Results Two kinds of sulfated polysaccharides, PAE1 and PAE2, were purified from Nereis. PAE1 mainly consisted of Gal, Glc, GalN, GlcA and a small amount of Fuc. The main chain of PAE2 was chondroitin sulfate C [→ 4GlcAβ1 → 3GalNAc6Sβ1 →] The chain is mainly composed of Fuc, Gal and Glc. Both PAE1 and PAE2 significantly prolonged APTT and PT. Conclusions Two kinds of sulfated polysaccharides were isolated and purified from the silkworm for the first time. Among them, PAE2 is a novel chondroitin sulfate-like substance containing Fuc, Gal and Glc branches and showing obvious anti-clotting activity in vitro. Silkworm sulfate polysaccharide provided the basis for the development.