Extraction,Purification And Bioactivity Of Eucalyptus Citri Resin Polysaccharide

Eucalyptus citriodora resin is a secretion flowing out from the wound of tree trunk.The resin contains a large number of natural effective components.The resin contains a large number of effective natural components and polysaccharide is one of them.Polysaccharides have become a research hotspot due to their wide range of biological activities.Through the study on the extraction,purification,structural characterization and biological activity of eucalyptus citriodora resin polysaccharide,the following main results were obtained:1.The phenol-sulphuric acid method was used to determining the polysaccharide content of eucalyptus citriodora resin as raw material.And the effects of extraction material-to-liquid ratio,extraction temperature and extraction frequency on the extraction rate of lemon eucalyptus resin polysaccharides were investigated by single-factor experiments.Based on the single factor experimental results,the appropriate material-liquid ratio,extraction temperature and extraction times were selected as the reference factors,and the orthogonal experimental design of three factors and three levels was carried out.The optimum extraction conditions were extraction temperature 95℃,extraction material-liquid ratio 1:20,extraction times 3 times,alcohol precipitation time 6 h and alcohol precipitation ratio 75%.Under these conditions,the extraction rate of Eucalyptus citri resin polysaccharide was 7.79%.2.The extract of eucalyptus citriodora resin polysaccharide was separated and purified.The eucalyptus citriodora resin polysaccharide was deproteinized by Sevage method and trichloroacetic acid method（TCA）.The results showed that the effect of TCA method on the deproteinization of eucalyptus citriodora resin polysaccharide was better.When the concentration of TCA reached 6%,the protein clearance rate was 98.90%and the retention rate of polysaccharide was 82.99%.The decolorization test of eucalyptus citriodora resin polysaccharide was carried out by polyamide chromatography column decolorization and activated carbon decolorization.It was found that decolorization effect of polyamide column chromatography is better than that of activated carbon.The decolorization rate was 77.95%and the retention rate of polysaccharide was 71.94%.The decolorized and deproteinized polysaccharides were further separated and purified,and the polysaccharides of RP1-2 and RP2-1 were obtained by fractional purification with DEAE-52 and Bio gel p-10.The physical and chemical experiments,infrared spectrum analysis,molecular weight and purity determination and monosaccharide composition analysis of the two polysaccharides were carried out.The results showed that RP1-2 was a neutral polysaccharide with a molecular weight of 11461 Da.It was mainly composed of glucose and mannose in the ratio of 1:0.2.RP2-1 is an acidic polysaccharide with a molecular weight of 8915 Da.It is mainly composed of mannose,rhamnose,glucuronic acid,glucose and galactose in the ratio of 0.3:0.5:0.3:0.8:1.0.3.The scavenging DPPH·free radical,scavenging·OH free radical,reducing ability and total antioxidant activity of eucalyptus citriodora resin polysaccharide were determined.The results showed that eucalyptus citriodora resin polysaccharide had good free radical scavenging ability.Through linear fitting calculation,the IC₅₀ of ERPC,ERP1,ERP2,RP1-2 and RP2-1for scavenging DPPH·free radical were 9.82,13.6,11.3,18.05 and 24.09 mg/L respectively,and the IC₅₀ for scavenging·OH free radical were 23.6,26.9,53.7,55.02 and 74.06 mg/m L respectively;eucalyptus citriodora resin polysaccharide has good total antioxidant capacity and reducing capacity.4.The antioxidant activity of eucalyptus citriodora resin polysaccharide in vitro was investigated.MTT colorimetry was used to determining the inhibitory effect of Polysaccharide on the proliferation of colon cancer cell HT,liver cancer cell Hep G2 and lung cancer cell A549in vitro.The results showed that in the concentration range of 1-400 ug/m L,RP1-2 and RP2-1had relatively little inhibitory effect on HT in colorectal cancer cells,and the inhibitory rates were only 6.12%and 6.43%;The inhibition rate of Hep G2 was 35.27%and 28.58%respectively,and the IC₅₀ value was 7.310 and 14.49 mg/m L respectively;The inhibition rate of RP1-2 was better than that of RP2-1,and the IC₅₀ value of RP1-2 was 13.46mg/m L.The research on the resin polysaccharide of eucalyptus citriodora resin shows that the resin polysaccharide of eucalyptus citriodora resin has good biological activity,and that conducting research on their polysaccharides is an important guide to the exploitation of Eucalyptus citriodora.