Isolation, Purification, Structural Characterization And Biological Activity Of Polysaccharides From Polygonatum Cyrtonema Hua

Background:Polygonatum cyrtonema Hua which is a common medicinal and edible plant from Liliaceae family.Polygonatum sibiricum Red.and Polygonatum kingianum Coll.et Hemsl were collected as the basic plants of Polygonatum in the Chinese pharmacopoeia 2015 edition.Aims:In this paper,the extraction process,isolation,purification,structure analysis,antioxidant activity and immunomodulatory activity of polysaccharide from Polygonatum cyrtonema Hua were studied to offer reference for utilization and exploration of Polygonatum cyrtonema Hua.Methods and Results:The article is composed of four parts:First,optimization of ultrasonic assisted extraction of polysaccharide from Polygonatum cyrtonema Hua.Second,isolation and purification of polysaccharide from Polygonatum cyrtonema Hua.Third,the antioxidant activity of polysaccharide from Polygonatum cyrtonema Hua was analyzed.Forth,the immunological activity of polysaccharide from Polygonatum cyrtonema Hua was studied.The experiment results are as follows:1.Research on optimal extraction condition of PCP:Box-Behnken design（BBD）was applied to optimize the ultrasonic-assisted extraction（UAE）condition of polysaccharides from Polygonatum cyrtonema Hua（PCP）.The effect of ratio of liquid to solid,ultrasonic power,extraction temperature and extraction time on the extraction yield of PCP were studied by single factor experiment.Based on single-factor design experiments,by using Design Expert 10.0 software,the data was analyzed and the optimized extraction condition was as follows,extraction temperature of 63.8℃,extraction time of 36.9 min,ratio of liquid to solid of 47.6 mL/g and ultrasonic power of 225 W.Under the optimal conditions,the extraction yield was 24.61%.2.Isolation,purification and structure analysis of purified polysaccharide PCP-1:Crude PCP was preliminary isolated by Cellulose DEAE-52,revealing three elution peaks subsequently designated PCP-1,PCP-2 and PCP-3 with the yields of 55.3%,15.7%and 8.1%,respectively.Among them,PCP-1 showed the highest yield,Therefore,PCP-1 was further purified by Sephadex G-100 chromatography.UV spectrum showed that PCP-1 did not contain protein,nucleic acid and other impurities.The results of infrared spectrum scanning showed that PCP-1 had the characteristic absorption peak of polysaccharide.Structural analysis revealed that PCP-1 was relatively homogeneous with the average molecular weight of 2966,composed of mannose,glucose,galactose,xylose and arabinose in a molar ratio of1:17.53:7.02:0.27:0.59,The scanning results of electron microscopy showed that PCP-1 was mainly composed of irregularly distributed sheets.¹ H NMR and ¹³C NMR indicated that PCP-1 mainly containedβ-type glycosidic bonds,a small amount ofα-type glycosidic bonds,and 6 kinds of sugar residues.3.The antioxidant activity of polysaccharide PCP-1:The vitro antioxidant experiment results showed that the PCP-1 has scavenging free radicals and antioxidant activity,compared with Vc.PCP-1 had different effect on different systems,which has the strong scavenging effect on superoxide anion and DPPH free radical,also had strong ability in the fangs of Fe²⁺.In a certain concentration range,the scavenging ability was enhanced with the increasing of PCP-1 concentration.4.The immunomodulatory activity of polysaccharide PCP-1:Methyl thiazolyl tetrazolium（MTT）assay showed that PCP-1 could promote the growth of macrophage RAW264.7 in the range of test concentration,with no cytotoxicity.The results of neutral red experiment showed that PCP-1 could significantly improve the phagocytic capacity of cells,and the content analysis of NO showed that PCP-1 could promote the macrophage RAW264.7 to produce NO,which was dose-dependent.Conclusion:By the research of this study,it was found that the PCP-1 obtained from the crude polysaccharide from Polygonatum cyrtonema Hua had good antioxidant activity and immunomodulatory activity,which provided new ideas and clues for the development of natural antioxidants and immunomodulatory agent.