为探究芋头水溶性多糖的免疫活性功能,本文用DEAE-52纤维素阴离子交换柱层析对由水提醇沉的方法得到芋头水溶性粗多糖进行分离纯化,并利用Sephadex G-200凝胶柱层析、SDS-聚丙烯酰胺凝胶电泳和傅里叶红外光谱分析其组成结构。再通过CCK-8试剂法、中性红吞噬实验、Griess法和酶联免疫吸附法测定其对巨噬细胞RAW264.7的增殖活性、吞噬能力以及分泌NO和细胞因子能力的影响。结果表明,得到的两种芋头水溶性多糖TPS1和TPS2纯度分别为87.4%和81.5%。TPS1主要成分为含有甘露糖的α型中性多糖,TPS2主要成分为含有硫酸基团的β型吡喃酸性多糖。细胞实验表明TPS1和TPS2在25~400 μg/mL浓度范围内对RAW264.7无毒害作用。其中,TPS1能够显著提高该细胞的中性红吞噬能力(P<0.05),吞噬指数最大可达到空白对照组的2.07倍;TPS2则具有更强的促进NO、细胞因子分泌的作用。TPS1和TPS2作用下的NO分泌量分别为空白对照组的3.07倍和4.57倍,TNF-α的分泌量为空白对照组的3.04倍和3.76倍,IL-1β的分泌量为空白对照组的1.37和1.54倍。以上结果表明,TPS1和TPS2均具有免疫调节的功能,有望进一步开发作为免疫调节剂添加到食品中。
In order to explore the immune activity of water-soluble polysaccharide of taro,DEAE-52 cellulose anion exchange column chromatography was used to isolate and purify the water-soluble crude polysaccharide of taro,which extracted by water-extracted and alcohol-immersed method. Then,Sephadex G-200 gel column chromatography,SDS-polyacrylamide gel electrophoresis and Fourier infrared spectroscopy were used to analyze its composition. Next,CCK-8 reagent method,neutral red phagocytosis experiment,Griess method and enzyme-linked immunosorption method were used to determine its effect on the proliferation activity,phagocytosis ability and secretion of NO and cytokines in macrophages RAW264.7. Results showed that two kinds of taro water-soluble polysaccharides(TPS1 and TPS2)with purity of 87.4% and 81.5% were obtained. The main components of TPS1 were neutral polysaccharide containing mannose and TPS2 were β-pyranic acid polysaccharide containing sulfate group. Cell experiments showed that TPS1 and TPS2 had no toxic effect on RAW264.7 in the concentration range of 25~400 μg/mL.Among them,TPS1 could significantly improve the phagocytosis of the cells(P<0.05),and the phagocytosis index was up to 2.07 times that of the blank control group. TPS2,on the other hand,played a stronger role in promoting the secretion of NO and cytokines. The secretion of NO under the action of TPS1 and TPS2 was 3.07 and 4.57 times that of control group.The secretion of TNF-α was 3.04 and 3.76 times that of control group.The secretion of IL-1β was 1.37 and 1.54 times that of control group.The above results indicate that both TPS1 and TPS2 have immunomodulatory functions and are expected to be further developed and added to food as immunomodulators.