Kinetic Model Of Extraction And Structural Characterization Of Water-soluble Polysaccharide From Polygonatum Odoratum

Many bioactive studys have shown that polysaccharide can be improving immunoregulation, anticancer, antivirus, anti-aging, anti-infection and radio-resistant. Thus polysaccharide has become one of the important projects for both domestic and foreign pharmaceutical companies in the research and development of supplements and carbohydrate drugs. Polygonatum odoratum,(Mill.) Druce(PO) is stipulated by Ministry of Health of China to be used both for food and medicine. Its main component, polysaccharide, is effective in preventing and curing hyperglycaemia and hyperlipemia, as well as in anti-oxidation and immunity enhancement. The research on PO has so far focused on the pharmacological activity of polysaccharide, the improvement of traditional extraction process and studying other chemical components, while research on the extraction under the kinetic theory of PO as well as a more systematic structural identification remain unseen. In order to further elaborate on the kinetic of the extraction process of PO and its structural identification, this thesis consists of three parts: studying PO respectively from the kinetic model of extraction, parameter analysis, preparation and purification of separation, primary structure identification, thus providing theoretical support to the production and technical development of polysaccharide. The main research content is as follows: 1.Extraction process kinetic model and parameter research of PO polysaccharideThis study based on the Fick’s second law with the spherical model assumptions. The extraction process kinetic model is derived by the mathematical equations.Through the PO polysaccharide extract test conditions which included that the temperature range was form 323 K to 373 K, the liquid-solid ratio was 50,100,200 m L·g-1, grain size was 177.5μm, at the extract time from 30 to 210 min. The results of mass concentration and 4 kinds of kinetic parameters were obtained.And the kinetic equation of PO polysaccharide extraction process was established. The forecast results of the extraction process has been demonstrated by t-Verify method(P>0.05).The predictive ability of this extraction process kinetic model equation has been verified experimentally. 2.Isolation, purification process of PO polysaccharide researchThe dried and defatted pretreated PO was extracted with hot water and then was purified by AB-8 macroporous resin decolorization, deproteinization.The precipitate was centrifuged when the supernatant was mixed with absolute ethanol to a final ethanol concentration of 60% and 80%,and named by POP60 and POP80. The purified polysaccharides, NPOP60-I and POAP60-I, were purified by DEAE-Cellulose52 chromatography and Sephadex G-100 chromatography from crude POP60. The purity identification of NPOP60-I and POAP60-I were through UV spectroscopy and GPC chromatography. UV analysis result showed that the polysaccharides has no obvious absorption peak under 260-280 nm UV scanning,indicated that NPOP60-I and POAP60-I contained no nucleic acids and proteins.GPC analysis showed that GPC chromatography elution curves was a single symmetrical peak, indicated that they were homogeneous polysaccharide components.Anthrone-sulfuric method was used to determine content of the polysaccharide by a conversion factor(f =1.48) of refined NPOP60-I to glucose as standard sample. The polysaccharide content determination of crude polysaccharide and extract of PO were 89.41% and 7.24%. 3.physical and chemical properties and structure characterization of POP60 researchThe primary structure of PO polysaccharide about physico-chemical characteristics, monosaccharide composition, glycosidic bond configuration, and the position and connection mode of the main and side chains were determined by HPLC,FT-IR,NMR,GC-MS,SEM, combined with completely acid hydrolysis, PMP derivatization and methylation.The NPOP60-I and POAP60-I were no reducing sugar, protein and starch, that confirmed by the methods of Sulphuric acid- phenol chromogenic reaction, Molish reaction, Film reaction, Biuret reaction and Iodostarch reaction.Sulfuric acid-Phenylphenol reaction showed that POAP60-I had uronic acid.The monosaccharide composition and the mole ratio determined by HPLC of NPOP60-I was: Mannose:Glucose:Galactose=1.00:12.07:9.55. And the result of POAP60-I was: Mannose: Rhamnose:Galacturonic acid:Glucose:Galactose=1.00:0.82:0.22:12.61:4.87:2.23.Infrared spectroscopy showed that NPOP60-I and POAP60-I were β- form pyranose ring. There was asymmetric scale symmetric absorption peak of C=O of POAP60-I at 1741cm-1, it showed that POAP60-I had uronic acid.GC-MS and NMR results showed that NPOP60-I and POAP60-I were pyranose rings. NPOP60-I were linked mainly by β-configuration glycosidic bond, and contained backbone chains of(1,6)-linked D-glucose and(1,2)-linked D-glucose and(1,6)-linked D-galactose, and non-reducing terminal mannose, the mole ratio of them were 2.25:0.76:0.80:1.00.POAP60-I were linked mainly by β-configuration glycosidic bond, and contained backbone chains of(2,6)-linked D-mannose,(1,6)-linked D-glucose,(1,4)-linked D-glucose,(1,6)-linked D-galactos, and non-reducing terminal glucose and the branch chains was with arabinose, the mole ratio of them were 1.00:9.87:2.13:5.84:1.45:3.24:1.77.The SEM space structure showed that: NPOP60-I were regular, agglomerative, folded schistose substances that monitored under 50 to 200 times of Scanning Electron Microscope.It maybe directional during the carbohydrate chains assembled. POAP60-I were fibriform substances, it maybe related to stronger interaction between polysaccharide molecules, or had branch structure of polysaccharide molecule.