黑灵芝葡聚糖高活性硫酸化产物的制备及分离纯化

Preparation and Isolation of Bioactive Sulfated Derivatives from Water-insoluble Glucan in Ganoderma arum

目的:通过化学修饰手段改善黑灵芝葡聚糖的水溶解性,探讨修饰产物的抗肿瘤活性及溶液性质。方法:采用氯磺酸-吡啶法制备不同酯化度的硫酸化修饰产物(AGAP),通过凝胶渗透色谱和红外光谱进行表征,MTT法检测对S-180肉瘤细胞毒性,筛选具有高活性的硫酸化多糖;对高活性硫酸化多糖进行分离纯化,并研究其溶液性质。结果:制备了9种不同酯化度的修饰产物,筛选出酯化度(DS)为1.14的硫酸化多糖AGAPS5,其糖含量为66.3%,对S-180肉瘤细胞生长的抑制率达到67.74%。凝胶柱分离得到高纯度的AGAPS5-1组分,其在0.1mol/L NaNO3溶液中的重均分子质量(M w)和数均分子质量(M n)分别为5.19×104u和2.63×104u,呈多分散性,以无规线团状形式存在。结论:硫酸化修饰能改善黑灵芝葡聚糖的水溶解性及溶液性质,增强抗肿瘤活性。

Objective： To improve the water solubility of a water-insoluble glucan from Ganoderma atrum by sulfated modification technique and to investigate the in vitro antitumor activity and properties of sulfated derivatives. Methods： The water-insoluble glucan was modified using chlorosulfonic acid-pyridine （CSA-Pyr） method to obtain sulfated products with different degrees of sulfation （DS）. The sulfated polysaccharides were characterized by gel filtration chromatography and FT-IR spectroscopy. The cytotoxicity on S-180 sarcoma cells was measured to screen a fraction with significant antitumor activity. The separated fraction with high antitumor activity was purified and its solution was subjected to molecular characterization. Results： Nine sulfated products with different DS were prepared. AGAPS5 （DS 1.14） had significant antitumor activity. The sugar content of AGAPS5 was 66.3%, and the inhibitory rate of AGAPS5 on the growth of S-180 sarcoma cells was 67.74%. The purified AGAPS5-1 showed a random coil chain in 0.1 mol/L NaNO3 solution. The weight-average molecular weight （Mw） of AGAPS5-1 was 5.19 × 10^4 u and the number-average molecular weight （M,） was 2.63 × 10^4 u. Conclusion： Sulfated modification can improve the water-solubility and solution properties, and enhance the in vitro antitumor activity of glucan from Ganoderma atrum.