Extraction,Isolation,Purification And Antioxidant Activity Of Polysaccharides From Elderberry Fruit

Elderberry,also known as elderberry,elderberry,iron bone,etc.,has biological activities such as preventing osteoporosis,antiviral,anti-inflammatory,antioxidant,antitumor,anti-AIDS,hypoglycemic,hypolipidemic,etc.,and is widely used in food,pharmaceutical,greening and toiletries as a functional ingredient worldwide.Studies have shown that polysaccharides are one of the important active substances of elderberry and have antioxidant,anti-inflammatory,anti-cancer and other functions.However,his research is mainly based on basic research,and there is still a lack of in-depth and systematic research on its structure identification and pharmacodynamic mechanism.In this paper,elderberry fruit was used as raw material to study the extraction process,separation and purification,structural characterization and antioxidant activity of polysaccharides in vivo.The main research structure is as follows:（1）Extraction process of elderberry fruit polysaccharides:the ultrasound-assisted e xtraction process of elderberry fruit polysaccharides was optimized by response surfa ce method,and the optimal extraction process parameters were:ultrasonic power 500W,extraction temperature 75°C,ultrasonic time 45 min,material-liquid ratio 1:20（g/m L）,repeated 3 tests,and the yield of elderberry fruit polysaccharides was（3.17±0.48）%.（2）Separation and purification of elderberry fruit polysaccharides:crude polysaccharides（EFP）were separated and purified by defatted and deproteinized,DEAE-52cellulose column and Sephadex G-100 glucan gel column to obtain 5 components EFP-0,EFP-1,EFP-2,EFP-3 and EFP-4,with polysaccharide purity of 77.86%,81.67%,90.65%,81.77%and 82.25%;the protein content was 0.34%,0.32%,0.13%,0.28%and 0.24%,respectively.（3）Structural characterization of elderberry fruit polysaccharides:the monosacchar ide composition and molecular weight size of elderberry fruit polysaccharide compon ents were determined,and the results showed that EFP-0 was mainly composed of a rabinose and fucose,with a ratio of 47.70:17.60 and an average molecular weight of 1.8289×10⁴ Da;The average molecular weights of EFP-1 and EFP-2 were 1.2336×10⁴ Da,1.7536×10⁴ Da,respectively,which were heteropolysaccharides dominated b y arabinose,galactose and glucose.EFP-2,EFP-3 and EFP-4 all contained galacturo nic acid,the ratios was 6.90:22.70:30.60,and the average molecular weights of EFP-3 and EFP-4 were 2.2593×10⁴ Da,5.0097×10⁴ Da,respectively.FT-IR analysis show ed that EFP-0,EFP-2,EFP-3 and EFP-4 all contained glucopyrane rings,αglycosidi c bonds existed in EFP-1,andαandβglycosidic bonds existed in EFP-2,EFP-3 a nd EFP-4.Based on the results of monosaccharide composition,methylation and NM R experiments,the main chain of EFP-2 contains two units:→4)-Manp（1→4）-β-D-G lcp（1→and→4）-β-D-Glcp（1→5）-α-L-Araf（1→）.（4）Study on antioxidant activity of elderberry fruit polysaccharides:the antioxidant activity of EFP-2 was analyzed by in vivo cell experiments and in vivo zebrafish model experiments.The results of cell experiments showed that EFP-2 had a strong protective effect on RAW264.7 cells with H₂O₂-induced damage,could also inhibit the morphological changes of RAW264.7 cells,and protect them from H₂O₂-induced oxidative stress,participate in the process of programmed suicide,remove functionless,unwanted,abnormal and harmful cells.Thereby reducing the death of RAW264.7 cells in the zebrafish oxidative damage model test,zebrafish after adding EFP-2 significantly reduced fluorescence intensity,reduced ROS level in vivo,and also showed obvious antioxidant capacity.The above results show that elderberry fruit polysaccharides have antioxidant activity,which can reduce oxidative damage,and provide theoretical support for the research and development of elderberry polysaccharides in functional foods and natural medicines.