The Polysaccharide Structure-activity Correlation Study Based On Sulfate Oyster Glycogen Structure

Natural polysaccharide has many bioactivities, including immunomudulation, antioxidation, antitumor, antivirus, anticoagulant, etc. Research showed that many natural polysaccharides with specific physiological activity have specific molecular structures. These specific molecular structures determine the bioactivity of the polysaccharides. But up to now, research on the relationship between polysaccharides structure and bioactivity progressed slowly. One of the main obstacles is the lack of suitable mode compounds. Such polymeric compounds must have single monosaccharide composition, clear and definite glycosidic bond type and substituent at same substitution positions, consistent anomer configuration, and their molecular weight can be effectively controlled. It’s difficult to obtain such polysaccharides as discribed above and this difficulty cannot be easily overcome in the structure-activity relationship studies.Natural polysaccharides are of complicated in structure, complex monosaccharide composition, varied connecting type, substituent types and replace position and so on. In addition, polysaccharide with different molecular weight and same structure are hardly to obtain. These problems made the structure-activity relationship research a hard task. In this study, polysaccharide was extracted and purified from oyster Ostrea talienwhanensis Crosse. After structure identification, the polysaccharide was proved to be a kind of well purified glycogen from oyster. The oyster glycogen was then sulfated, and the substitution patterns were precisely determined to be C-6in the sulfated oyster glycogen. This kind of glycogen, with simple monosaccharide composition and anomer configuration, consistent glycoside type, ascertain molecular length, consistent and determined position of the sulfuric base, thus can be used as an excellent model compound in the study of structure-activity relationship.In order to obtain purified oyster glycogen, the polysaccharide content was used as an indix to optimize the extraction method to remove the conjugated pretein. The optimum extraction conditions were determined:the optimize extraction enzyme is pepsin, the ratio of solid material to liquid is1:45, enzyme percentage is1.5%(m/m, substrate), pH2.0, temperature as37℃, enzymolysis time is2h. Ethanol (v/v) was then added into the enzymatic hydrolysate to obtain the crude oyster polysaccharide. The crude oyster polysaccharide was then purified by a series of column chromatography, including dimethylaminoethyl negion exchange column (DEAE-cellulose-52,2.0cm×40cm) and Sephadex G-100column (1.6cm×70cm). The major fraction was collected, concentrated, desalted by dialysis and then vacuum freeze-dried to obtain the homogeneous polysaccharide.After derivatization, the structure of oyster polysaccharide polymer was analyzed by gas chromatography and high performance liquid chromatography (HPLC). Results showed that the oyster polysaccharide consisted solely of glucose. IR scan results proved the obtained polymer was polysaccharide. The fully methylated products were analyzed by GC-MS and were affirmed to be a kind of glycogen because of its typical glycogen linkage,1-4Glc,1-4,6Glc and1-Glc. Meanwhile, there exist trace other glycogen linkage type. The ratio of1,4-linkage to1,4,6-linkage was6.6, which indicated that the oyster glycogen chain branched for6.6glucose residues averagely. And it was further confirmed by NMR analysis that the oyster polysaccharide was a kind of glycogen and the oyster glycogen chain branched for6.6glucose residues-averagely.The oyster glycogen was chemically sulfated and further purified. Three fractions of sulfated oyster glycogen were collected, named as SOG, SOG1, and SOG2. SOG was sulfate substituted at C-6position. SOG1was sulfate substituted at C-2and C-3positions. SOG showed better activity in promoting spleen lymphocyte proliferation than SOG1. SOG and SOG1shared similar structure properties, except the sulfate position. It’s proved that the position of sulfate substitution played an important role in promoting lymphocyte proliferation ability. The polysaccharide with C-6substitution demonstrated the most significant activity.After periodate oxidation, SOGF1was obtained by gel chromatography of the degradation products of SOG. The content of sulfate group in SOGF1was40.6%. Molecular weight of SOGF1was determined as6.3×106. Thus SOGF1, SOG, SOG1and SOG2, with different sulfate group content, different molecular weight, but similar structure, could be used as model polysaccharides to research the structure-activity relationship. Activity studies showed that all the above four kinds of glycogens could promote lymphocyte proliferation with an ascending series of SOG>SOGF1>SOG1>SOG2.Analysis showed that different structural properties of the polysaccharides played different role in promoting lymphocyte proliferation, with an ascending series of sulfate positon>sulfate content> molecular weight.Results showed that there was a proportional relation betrelationship between sulfate content and lymphocyte proliferation activity. But molecular weight did not demonstrate any effect on the bioactivity.Results showed that C-6position sulfate substitution has decisive influence on the bioactivity of polysaccharide if other structural properties were similar. The content of sulfuric acid played the second role, more sulfate content, stronger bioactivity. Finally, molecular weight of sulfate polysaccharides also could influence the bioactivity, but there is no clear correlation between them. Perhaps the molecular weight might play some role when reaches a certain value.