| In this paper,natural selenium-enriched and artificial selenium-enriched green tea were used as raw materials.The basic components and selenium contents were determined firstly.Then,the tea polysaccharides were extracted,separated and purified.Finally,the antitumor activity and structure were studied.The main contents of this paper are as follows:(1)Firstly,the basic components and selenium contents of the two kinds of tea were determined,the results showed that the content of tea polyphenols in natural selenium-enriched green tea was higher than that in artificial selenium-enriched green tea,and the other components and selenium contents were lower than those of artificial selenium-enriched green tea.Next,the extraction yield and the content of the two selenium-enriched polysaccharides were measured,the results showed that the yield,total soluble sugar,soluble protein and selenium content of artificial selenium-enriched crude polysaccharides were higher than those of natural selenium-enriched crude polysaccharides,they are respectively,4.29%,24.78%,1.27%,18.42mg/kg and the content of uronic acid in natural selenium-enriched crude polysaccharides was 40.96%,which was significantly higher than that of artificial selenium-enriched polysaccharides.Finally,two kinds of selenium-enriched crude polysaccharides were separated by DEAE Sepharose FF to obtain four different components.Firstly,the homogeneity and molecular weight of the two tea polysaccharides were found to have four peaks,the maximum molecular weight is more than one million,presumably for glycoprotein,purified polysaccharides ASe-TPS2 and NSe-TPS2 are Single symmetric peak,indicating that it is pure polysaccharide without impurities,which can be used for subsequent detection of the structure.The yield and composition of the purified polysaccharides were analyzed.The results showed that NSe-TPS2 had the highest yield,the yield of NSe-NTPS was the lowest in the natural purified polysaccharides,while the artificial purified polysaccharides were the opposite.The results of uronic acid determination showed that all the four components in the natural purified polysaccharides were acid sugar,and in addition to ASe-TPS3 the rest are acidic sugar and a small amount of protein was also detected in the purified polysaccharides,indicating that after a series of purification steps,the polysaccharides still bound to a small amount of protein.(2)The antitumor effect in vivo and in vitro of tea polysaccharides and seleniumwas studied by using the crude and refined products of the two kinds of tea polysaccharides extracted and purified in the previous step as raw materials.The results showed that tea polysaccharides and selenium all had certain antitumor effect.The antitumor effect of inorganic selenium was better than that of organic selenium.The antitumor effect of common tea polysaccharides was bigger than they are mixed with yeast selenium.Antitumor effect of natural selenium-enriched tea polysaccharides > Artificial selenium-enriched tea polysaccharides > common tea polysaccharides mixed with yeast selenium group,and they all showed dose-dependent characteristics.The results also showed that selenium polysaccharides had stronger antitumor effect than polysaccharides simply mixed with selenium,and selenium could enhance the antitumor activity of tea polysaccharides.(3)Finally,the structures of the two selenium-enriched polysaccharides were observed and analyzed.The results of the determination of monosaccharide composition showed that the crude polysaccharides NSe-TPS and the fine polysaccharides of the natural selenium-enriched polysaccharides were acidic sugars containing higher amount of uronic acid,the artificial selenium-enriched crude polysaccharides ASe-TPS and the fine polysaccharides were also acidic sugars containing higher amount of uronic acid.The results of UV scanning showed that the two kinds of crude polysaccharides had obvious protein absorption peaks,in which the natural purified polysaccharides NSe-TPS3 and the artificial purified polysaccharides ASe-TPS1 also showed obvious protein absorption peaks,indicating that these components were Glycoprotein,the other components have a weak protein absorption peak,indicating that crude polysaccharides after purified still contain a small amount of protein.The results of infrared scanning showed that the two kinds of selenium-enriched polysaccharides had similar infrared spectra.The difference was that the characteristic absorption peaks of uronic acid were found in the four natural selenium-enriched tea polysaccharides,and that was only found in crude polysaccharides and purified polysaccharides ASe-TPS1 in the artificial selenium-enriched tea polysaccharides,indicating that the natural selenium-rich polysaccharides are acidic sugar.(4)The results of partial acid hydrolysis showed that fucose existed only in the branched chain of two selenium-enriched tea polysaccharides,arabinose,galactose and mannose were mainly in the branched chain of two selenium-enriched tea polysaccharides,glucose and Gal A was mainly in the main chain of twoselenium-enriched tea polysaccharides,the artificial selenium-enriched tea polysaccharides did not contain fructose,when the acid concentration was 0.05 M,the branched chains of the two selenium-enriched tea polysaccharides had been hydrolyzed completely.(5)The results of Periodate oxidation and Smith degradation showed that: From the results of the oxidation of periodate,it can be concluded that the glycosidic linkage of 1→,1→2,1→3,1→4,1→6,1→2,6,1→4,6 may exist in the structure of ASe-TPS2 and 1→、1→2、1→4、1→6、1→2,6、1→4,6 may exist in the structure of NSe-TPS2(Measured in our lab before).From the GC of Smith degradation products,it is presumed that there may be 1→,1→2,1→3,1→5,1→6,1→2,6 linked glycosidic bonds in the ASe-TPS2 structure,in which the 1→3 linkage may account for the main part and there may be 1→、1→2、1→4、1→5、1→6 linked glycosidic bonds in the NSe-TPS2 structure(Measured in our lab before).(6)The results of NMR showed that both selenium-enriched polysaccharides contained α and β glucose,but they are mainly contained by β-D-Glup;Their acidic sugar are all α-D-GalpA and the content of NSe-TPS2 is significantly higher than that of ASe-TPS2;They all contain α and β ramnose;They all contain arabinose,but it’sα-L-Araf in the ASe-TPS2 and it’s β-L-Araf in the NSe-TPS2;They all contain-COOCH3,CH3CO-and a small amount of protein,so their chemical nature is acid glycoprotein,the result is the same as the infrared result.(7)The results of methylation analysis showed that the structures of artificial selenium-enriched polysaccharides were mainly based on the β-D-(1→3)-Glup andα-D-(1→4)-GalpA,and its’ branches are α-L-(1→2,3)-Araf,β-D-(1→4)-Glup,α-L-(1→2)-Rhap and a small amount of α-D-(1→4)-GalpA,and its’ non-reducing end is mainly composed of Araf and Xylp;The structures of natural selenium-enriched polysaccharides were mainly based on the β-D-(1→4)-Glup and α-D-(1→4)-GalpA,and its’ branch is β-L-(1→2)-Araf,α-D-(1→3)-Galp,β-L-(1→2)-Rhap and its’ non-reducing end is mainly composed of Glup and Galp.(8)The results of the Scanning electron microscopy(SEM)showed that rude polysaccharides in two kinds of selenium-enriched green tea compared with the purified polysaccharides,the space conformation changed from the dispersion of sheets or leaves to more scattered filaments,strips and granular,such as irregular state.The results of thermogravimetric analysis showed that the natural selenium-enriched tea polysaccharides were more stable than the artificial selenium-enriched teapolysaccharides.From the above results,it was found that the structure difference of the two polysaccharides was that the chain skeleton of artificial polysaccharide consists ofβ-D-(1→3)-Glup,while the natural polysaccharide consists of β-D-(1→4)-Glup;GalpA exists in the main and side chain in artificial polysaccharide,while it only exists in the main chain in natural polysaccharide;The Araf in the side chain is αstructure in artificial polysaccharide,while it is β structure in the natural polysaccharide;The non-reducing end is mainly composed of Araf and Xylp in artificial polysaccharide,while it is mainly composed of Glup and Galp in natural polysaccharide and the natural polysaccharides were more stable than the artificial polysaccharides.Both have in common is that the main chain of the polysaccharide all consists of β-Glup and α-GalpA,the side chain all consists of Araf and Rhap,and the space conformations are all flakes in the crude polysaccharide,while they are all granulars in the fine polysaccharide. |
PDF Full Text Request