Isolation,Purification,Structural Elucidation And Bioactivity In Vitro Of Polysaccharides From Edwardsia Sipunculoides

The polysaccharides of Edwardsia Sipunculoides were used as the research object in the study.The extraction,isolation,purification,physical-chemical properties,structural elucidation and biological activities in vitro were researched systematically and deeply in this paper.Biological activities mainly included antioxidant,anti-tumor and immunomodulatory.The results were as follows:Based on single factor tests,the response surface methodology(RSM)was used for optimizing homogenate extraction processing of polysaccharide from E.sipunculoides(SAP).Results showed that the optimum extraction condition of polysaccharide from Edwardsia sipunculoides was as follows: 7 min homogenate extraction time,extraction temperature 40℃,and ratio of material to liquid 1:10.Under the conditions,the extracting yield reached 8.29%. Three polysaccharides,sea anemone polysaccharide 30(SAP30),sea anemone polysaccharide 60(SAP60)and sea anemone polysaccharide 80(SAP80)were separated from the body of E.sipunculoides by tissue homogenate and papain hydrolysis under the optimal condition.The antioxidant activities of three fractions of crude polysaccharides from E.sipunculoides were evaluated by in vitro assay of reducing power,and ABTS radical.The antioxidant power of crude polysaccharides decreased in the order of SAP80 > SAP30 > SAP60 in the two assays.The results shown that the inhibitory effects of SAP against different tumor cells decreased in order of SAP30 > SAP80 > SAP60.The same as immune activity in vitro.SAP30 was selected for further study and purified with ion exchange chromatography on DEAE-Sepharose fast flow.And two major elution peaks: SAP30 n and SAP30 a were detected.Then PP80 W was further purified by preparative liquid phase GS620 gel-permeation chromatography,and two major elution peaks: SAP30n-1 and SAP30n-2 were detected.SAP30n-1 appeared as a single symmetrical peak in the HPLC chromatograph,indicating that SAP30n-1 was a homogeneous polysaccharide,with an average molecular weight of 1800 kDa.The results of UV scanning indicated that SAP30n-1 didn’t contain protein or nucleic acid.Compared between SAP30n-1,SAP30n-2,SAP30 a and SAP30,the inhibitory effects of SAP30 against different tumor cells was higher than the other three polysaccharides.The structure characterization of SAP30n-1 was elucidated by IR,GC-MS analysis and NMR spectrum analysis.Gas chromatography of its alditol acetates has shown that SAP30n-1 was composed of an unkown mount,D-mannose,D-glucose and D-galactose as the molar ratio of 0.93:1.00:34.69:1.03.The methylation analysis and NMR analysis showed that the primary structure of SAP30n-1 mainly consisted of 1,3-linked 6-deoxy-Galp,1,4-disubstituted-D-Glcp,1,2,3,4-tetrasubstituted-D-Glcp and 1-substituted-D-Glcp.1,6-disubstituted-D-Glcp and 1,6-disubstituted-D-Galp were secondary contents.In addition,it also contained trace amount of galactosamine.