Isolation, Purification And Biological Effects Of Polysaccharides From Colocasla Antiquorum

Colocasla Antiquorum is commonly known as taro. Taro is nutrient-rich, high content of crude fiber and low in fat content. It is also rich in vitamins and mineral elements, which is the homology of medicine and food. Especially taro is rich in mucilage, the main component of mucilage is viscous polysaccharide, which can improve the immune system. A large number of studies have shown that plant polysaccharides have pharmacologically activities of anti-tumor, enhancing immunity, anti-oxidation, anti-aging, lowering blood sugar, lowering blood pressure, anti-virus and so on. Due to their wide range of sources and low toxicity, many plant polysaccharides have been applied to the field of health food, animal breeding and medicine. Nearly a hundred species of plant polysaccharides had extracted and made some structural identification at present. But there was a little research on taro polysaccharides. In this paper, the study aimed to isolate and purify the non-starch polysaccharides from taro. The chemical characteristics and structure were explored of each homogeneous polysaccharides component, comparing the immune cells regulation functions in order to find a good immune activity polysaccharide fraction. Then the animal experiments were carried out to explore the immune regulation of the best biological activity component on the normal mice.(1) The first part study aimed to develope an optimal procedure for the microwave-assisted extraction of polysaccharides from taro. Based on the single factor experimental results, Box-Benhnken experimental design was used to select the most important ones affecting extraction of taro polysaccharides, the optimal values of selected parameters, namely water extraction time, water extraction temperature, microwave power and microwave treatment time, response surface experiments were used to optimize the extraction process, the mathematic modle of the factors and indications was established and analyzed. Results showed that the optimal extraction parameters were water extraction time of3h, water extraction temperature of82℃, microwave power of395W, microwave treatment time of77s and extracted two times, the actual polysaccharides yield was5.57%under this condition, compared with the theoretical value the relative error was less than0.5%.(2) The crude taro polysaccharides were extracted with hot water, then protein was removed by means of enzymatic-assisted Seveg methods. TPS was obtained after dialysis and freeze-drying. After further purification by DEAE-52celluse and Sephadex G-75column chromatography, three major homogeneous polysaccharides components were obtained, which were named as TPS1p, TPS2p1and TPS3P2respectively. The UV spectroscopy and HPLC assays indicated that TPS1p and TPS2p1were two kinds of homogeneous polysaccharides, however, TPS3p2may be a polysaccharide protein complexes. The molecular weight of TPS1p, TPS2P1and TPS3p2were7.9×104Da,1.1×105Da,1.3×105Da, respectively. These three homogeneous polysaccharides components were both composed of arabinose, mannose, glucose and galactose by GC assays. The molar ratio was2.76:2.127.61:3.30,2.44:4.07:1.87:12.1,1.27:1.89:1.13:5.97respectively. The IR scanning results showed that these three components both have obvious polysaccharide characteristic absorption peaks, and they are all have pyran ring structure which were connected together by the (3glycosidic bond.(3) The immune cells regulation functions were explored of four kinds of polysaccharides TPS, TPS1p, TPS2P1, TPS3P2, and investigated the effects of three different concentrations25,50,100μg/mL. The results showed that any doses of TPS1p could assist ConA in promoting the proliferation of spleen lymphocytes of normal mouse (P<0.05). At the high doses of TPS2P1, the high and middle doses of TPS and TPS3P2also had significant impact on assisting ConA(P<0.05). Meanwhile, the different doses of TPS1p can markedly improve phagocytosis of macrophages of mouse. At the high doses of TPS2P1, the high and middle doses of TPS and TPS3P2also both showed significant impact on phagocytosis of macrophages(P<0.05). At the high doses of TPS, the high and middle doses of TPS1p and TPS2P1could assist LPS in promoting the proliferation of peritoneal macrophages(P<0.05), however, any doses of TPS3p2showed no remarkable effects on assisting LPS. Compared with the other three polysaccharides, the neutral polysaccharide TPS1p had the best immune cells modulate activities.(4) The neutral polysaccharide TPS1p on the immune system in normal mice was investigated. The mice were treated p.o. with TPS1p (75,150and225mg/kg·d) for21days. The normal control group was administered with the same volume of saline. After the administration for21days, their humoral, cellular and nonspecific immune functions were determined by relative assays. The results showed that any doses of TPS1p could significantly improve the spleen indexes and NK cells activities (P<0.05) when compared with the normal control group. The high and middle doses TPS1p could significantly improve phagocytic function of peritoneal macrophage, meanwhile spleen lymphocyte induced by ConA proliferation and thymus index also increased, and the formulation of haemolytic plaque and serum haemolysin were also significantly promoted at these doses. However, different doses of TPS1P had no remarkable effects on the DTH assay. In conclusion, TPS1p could improve the immune function of normal mice. It laid the foundation for the wide application of the TPS1p in such fields as food industry and health care.