[关键词]
[摘要]
目的 从粗茎秦艽Gentiana crassicaulis中分离纯化多糖,初步分析其结构特征并研究其体外抗炎活性。方法 采用热水浸提粗茎秦艽粗多糖(Gentiana crassicaulis polysaccharide,GCP),再经Cellulose DE-52纤维素离子交换柱、SephadexG-100柱进行分离纯化,得到GCP-1。采用高效凝胶色谱法(HPGPC)鉴定纯度并测定其相对分子质量;采用高效液相色谱(HPLC)测定其单糖组成;采用红外光谱(IR)、气相色谱-质谱联用(GC-MS)和核磁共振波谱法对该多糖的结构进行初步分析;采用CCK-8法检测其对小鼠单核巨噬细胞RAW 264.7增殖的影响;利用ELISA试剂盒测定其对白细胞介素-1β(interleukin-1β,IL-1β)、一氧化碳(NO)和肿瘤坏死因子(tumor necrosis factor-α,TNF-α)释放量的影响。结果 从粗茎秦艽中分离纯化得到GCP-1多糖,相对分子质量为6.87×104,由甘露糖、鼠李糖、葡萄糖醛酸、半乳糖醛酸、葡萄糖、半乳糖和阿拉伯糖7种单糖组成,其物质的量比为0.07:3.07:0.42:1.98:1.33:5.22:6.76;IR、GC-MS和核磁共振波谱分析表明,GCP-1是一种典型的果胶多糖,含有HG和RGI区域及AGI和AGII侧链;质量浓度在10~200 μg/mL时,GCP-1能明显促进小鼠单核巨噬细胞RAW 264.7的增殖(P<0.05),并能明显抑制小鼠单核巨噬细胞RAW 264.7分泌IL-1β、NO和TNF-α。结论 GCP-1为从粗茎秦艽中分离得到的天然来源的均一多糖,其结构和活性初步研究为秦艽多糖成分的开发提供了一定参考。
[Key word]
[Abstract]
Objective Gentiana crassicaulis polysaccharides (GCP-1) was extracted and purified to study the structural features and anti-inflammatory activity in vitro. Methods The crude polysaccharides of Gentiana crassicaulis (GCP) was extracted by hot water. The Cellulose DE-52 and SephadexG-100 columns were used to separate and purify homogeneous polysaccharides. The relative molecular mass was analyzed by high-performance gel permeation chromatography, and the monosaccharide composition and structure were preliminarily identified by HPLC, IR, GC-MS, and NMR. The effects on proliferation function of RAW 264.7 cells were determined by CCK-8, and the release capacity of IL-1β, NO, and TNF-α were determined by the ELISA kit. Results The molecular weights of the GCP-1 polysaccharides extracted and purified from G. crassicaulis was 6.87×104. GCP-1 was consisted of mannose, rhamnose, glucuronic acid, galacturonic acid, glucose, galactose and arabinose. The molar ratios of these monosaccharide were 0.07:3.07:0.42:1.98:1.33:5.22:6.76. The structure of GCP-1 was characterized by IR, GC-MS and NMR, and GCP-1 was a typical pectin polysaccharide, with HG region, RG-I region and AG-I/AG-II side chains. When the concentration was 10-200 μg/mL, GCP-1 significantly promoted the proliferation of RAW 264.7 cells (P<0.05) and inhibited the release of IL-1β, NO, and TNF-α. Conclusion GCP-1 is a kind of natural homogeneous polysaccharides. Structural characterization and anti-inflammatory activity of GCP-1 provide the significant reference for the further development and elucidation of G. crassicaulis polysaccharides.
[中图分类号]
R284.1
[基金项目]
国家重点研发计划(2017YFC1700705);四川省重点研发项目(2021YFS0043);四川省留学人员科技活动项目择优资助经费计划(2018-68)