Preparation Of Dendrobium Officinale Polysaccharide And Its Protective Effect On Acute Alcoholic Liver Injury

Objective To optimizate of extraction process of polysaccharides of Dendrobium by single factor and orthogonal expriment.Dendrobium polysaccharides purified by DEAE-cellulose and Sephacryls-300 chromatography,and the separated components for structural analysis.Investigation Dendrobium polysaccharides on prevention and treatment of acute alcoholic liver injury in mice,providing experimental data for the comprehensive development and utilization of DendrobiumMethods?Firstly,To investigate factors and levels determined by single factor experiment,On the basis of the design L9(34)orthogonal,Optinization of extraction process of polysaccharides of Dendrobium,In the factor of extraction temperature,extraction time,solid-liquid ratio and extraction times,as an indicator to sugar content of Dendrobium extraction?After Washed?homogenized and crushed of fresh Dendrobium,extracted by orthogonal process,then centrifuged supernatant?oncentrated?removal of protein?ethanol precipitatio?the precipitate which dried under vacuum to obtain Dendrobium polysaccharides(DOP).Polysaccharide content determined by method of sulfuric acid phenol,uronic acid content eterminated of sulfuric acid carbazole,protein content was measured with Coomassie Brilliant Blue.Polysaccharide first eluted through the DEAE-cellulose column with buffer Tris-cl,with a solution of NaCl gradient elution,the peak of fractions component eluted and concentrated,and then through Sephacryls-300 for further separation and purification,Then,the components to give high-performance liquid,infrared analysis?Male ICR mice were randomly divided into control group,model group,DDB positive group,different doses of DOP(10,50,250 mg·kg-1)group.To create acute alcoholic liver injury model mice and to measure ALT,AST,TG,SOD,MDA content of liver homogenates,ALT,AST,SOD,MDA content in serum.Detecting the expression of liver ATGL and HO-1 mRNA by RT-PCRResults?Water as a solvent,to acquire the best extraction process of Dendrobium Polysaccharide by study was:extraction temperature 70?,extraction time 4 h,solid-liquid ratio 1:40,extraction 1 times;The order of the factors of influence:extraction temperature>extraction time>extraction times>solid-liquid ratio.?Polysaccharide of laboratory extracted was a dark brown solid,polysaccharide content of 88.83%,uronic acid content was 7.44%,soluble protein content was 0.41%.Dendrobium polysaccharides isolated three component peak over DEAE-cellulose column,named DOP1,DOP2,DOP3;The component peak concentrated over Sephacryls-300,were isolated singlet,doublet and quartet respectively,named DOP1-1,DOP2-1,DOP2-2,DOP3-1,DOP3-2,DOP3-3,DOP3-4.After HPLC analysis of DOP1-1,DOP2-1 showed that two peaks were mannose and glucose,mannose which accounted for the major part;peaks of DOP3-1,DOP3-2,DOP3-3 corresponding was glucose.Component DOP2-2,DOP3-4 wered not detected peak.IR analysis shows that polysaccharide of DOP1-1,DOP2-1,DOP3-1 with the general characteristics of polysaccharides?Compared with the control group,ALT,AST content were significantly increased(P<0.05),show that the model of alcoholic liver injury was established;Compared with the model group,DOP each dose group can reduce ALT,AST,TG and MDA levels,increase SOD content;The expression of each dose group of ATGL and HO-1 mRNA have changesConclusion To optimize the extraction of the polysaccharides by single factor and orthogonal experiment,Polysaccharides were isolated and purified with DEAE-52 cellulose and Sephacryls-300 Dendrobium.The results show that the process is simple,feasible and reliable method.DOP can reduce ALT,AST,TG and MDA levels increased due to the one-time heavy drinking,increased SOD levels,thus acute alcoholic liver injury in mice play a protective role in the prevention,which may be related to increase in liver ATGL and HO-1 mRNA expression,thereby accelerate the decomposition of TG and increase antioxidant capacity.