Extraction, Separation, Structure Characterization And Antioxidant Activities Of Polysaccharide From Leaves Of Eucalyptus Grandis X E.Urophylla

The extraction, separation, structure characterization and antioxidant activities of polysaccharides from leaves of Eucalyptus Grandis x E. Urophylla were investigated, which is the abundant resource in Guangxi provience. The main research contents are:(1) The quantitative analysis method of total sugar, reducing sugar and polysaccharides in Eucalyptus Grandis x E.Urophylla by using 3, 5-dinitrosalicylic acid absorptiometry was established. The RSD results were 2.7% and 1.7% in precision experiment and repeatability, respectively, and the recoveries were 99.3%～101.3%. The content of total sugar of Eucalyptus Grandis x E. Urophylla was 15.26%, and that reducing sugar was 1.15%.(2) The quantitative analysis method of polysaccharides in Eucalyptus Grandis x E. Urophylla probed by phonel-sulfate absorptiometry was established. Standard curve is:y=0.0145x+0.0008, R2=0.9996, where the linear range is 10.0～60.0 μg·mL-1. The RSD<3% in precision experiment, whereas the RSD≤1% in repeatable experiment, and the recoveries were in the range between 98%～107%.(3) The extraction process of polysaccharides in Eucalyptus Grandis x E. Urophylla was optimized. The optimum conditions were:microwave power of 560 W, liquid/material ratio of 45:1 and extraction time of 7 minute, where the yield of polysaccharides was 8.94%. The yield of polysaccharides was increased 8.63% comparing with the common extraction method (8.23%). The effect degree of these factors were:liquid/material ratio> extraction time> microwave power.(4) The decontamination of polysaccharides in Eucalyptus Grandis x E. Urophylla was investigated. The clearance effect of protein in polysaccharides was probed by analysis method of Coomassie Brilliant Blue G-250, and the result demonstrated that the protein was strongly absorpted by polyamide, whereas the adsorption of polysaccharides was slighter. The Protein removal rate, bleaching rate and polysaccharide retention rates were 75.2%,79.0% and 78.0%.(5) The separation of polysaccharides in Eucalyptus Grandis x E. Urophylla was investigated. The polysaccharides was purified by polyamide firstly. Then it was classified chromatographic with the cellulose column of DEAE-52. The components were merged and concentrated afterwards the elution of water and 0.1 mol·L-1 NaCl respectively. Finally, polysaccharides was separated by chromatogram column of Sephadex G-100, and two components, EP1 and EP2, were obtained.(6) The structure characterization of polysaccharides was conducted by HPGPC, and result indicated that the component EP2 was a single component with the molecular weight of 9430.269. The UV spectrum was used to detect the EP2, which reflected none protein and nucleic acid in polysaccharides after purified. The result of IR spectroscopy showed a typical absorption peak of polysaccharides.The EP2 was analyzed by GC-MS more precisely, and the monosaccharide rate is arabinose:mannose=1:44.5.(7) The antioxidant activities of polysaccharides was investigated. The antioxidant to DPPH· and hydrogen peroxide were conducted by polysaccharides, purified polysaccharides (ESP) and EP2 with the reference substance of ascorbic acid. The results indicated that the maximum scavenging rates of polysaccharides, purified polysaccharides (ESP) and EP2 to DPPH· were 70.05%,62.73% and 45.00% respectively, where the maximum scavenging concentration is 0.10 mg·mL-1. whereas the maximum scavenging rates of polysaccharides, purified polysaccharides (ESP) and EP2 to hydrogen peroxide were 84.70%,70.90% and 64.00% respectively, with the maximum scavenging concentration of 0.11 mg·mL-1. The antioxidant ability of these components were:ascorbic acid>polysaccharides>purified polysaccharides (ESP)>EP2.