| ObjectivesAs people's living standard improve gradually,people pay more and more attention to the body health,such as Chinese caterpillar fungus that can increase the body's immune ability gradually,but the Chinese caterpillar fungus is very scare,north cordyceps and Chinese caterpillar fungus had similar substances(such as cordycepin,etc).The price is relatively cheap and gradually entered into people's sight.Traditional north cordyceps cordycepin extraction efficiency is not very high,cordycepin and shell oligosaccharides can enhance immune function,but the combination of these two substances improving the immune function is less.The purpose of this experiment research is to study eutectic solvent on the extraction effect of cordycepin and explore the best extraction conditions about the eutectic solvent extracting cordycepin.And another purpose of this experiment research is to study the combination cordycepin and shell oligosaccharides in kunming mice on immune function.Methods1 The extraction of cordycepin1.1 The shattering of the north Chinese caterpillar fungusPut Chinese caterpillar fungus in 60 ? oven,dry for 8 hours north and use crusher crushing.Then use 60 mesh screens to filtrate and place in a dry place.1.2 The preparation of solventThe choline chloride and urea,choline chloride and glycerin,choline chloride and ethylene glycol in accordance with the ratio of 1:2 to join in the conical flask,heated and stirred at 60 ? temperature,until a colorless transparent solution is formed.The ethanol and water according to the 3:1 proportion is joined in the conical flask,stirred to form mixed solution.1.3 The single factor experimentChange of solvent,temperature,time,moisture content,such as the ratio of material liquid,the condition of fixed other conditions,using the ultrasonic extraction method to extract of cordycepin,and observe the influence of these factors on the extraction efficiency of cordycepin.1.4 The orthogonal experimentSelect temperature,liquid ratio,extraction time and moisture content for the four factors,each factor to select three levels,study four factors influence on the extraction rate of Chinese caterpillar fungus,choose table L9 orthogonal analysis to study the best extraction conditions of the extraction rate of Chinese caterpillar fungus.2 The immune function of cordycepin,shell oligosaccharide and cordycepin joint shell oligosaccharide.2.1 The experimental animal grouping and dosageBlank control group,low concentration cordycepin group(40 mg/kg),high concentration cordycepin group(80 mg/kg),low concentration shell oligosaccharide group(200 mg/kg),high concentration shell oligosaccharide group(400 mg/kg),low cordycepin concentration(40 mg/kg)and low shell oligosaccharide concentration(200 mg/kg)mixed group,high cordycepin concentration(80 mg/kg)and high shell oligosaccharide concentration group(400 mg/kg).2.2 The immune function experiment of experimental animalsThe immune function experiments include ConA induced mouse spleen lymphocyte transformation test,sheep red blood cells(SRBC)induced mice delayed-type allergy DTH(foot plantar thickening method),the determination of serum hemolysin method(blood clot),indexes of the clearance rate of carbon,NK cell activity determination,and weigh weight,thymus weight and spleen weight of mice.Results1 The extraction of cordycepin1.1 Choline chloride and urea 1:2 as solvent extract the highest efficiency.1.2 With the extension of time,the extraction yield of cordycepin rises.But after 30 minutes,with the extension of time,the extraction yield of cordycepin drops.The highest extraction yield of cordycepin is in 30 minutes.1.3 With the increase of solid-liquid ratio,material liquid extraction yield of cordycepin increases,the extraction yield of cordycepin in material liquid extraction yield is similar between 1:300 and 1:200.1.4 With the increasing of temperature,higher extraction yield of cordycepin reaches the highest at 60 ?,extraction rate reaches 80.8%,when the temperature rises further,extraction yield of cordycepin reduces instead.1.5 With the increase of water content,extraction yield of cordycepin becomes higher,when water content decreases,further extraction yield of cordycepin reduces instead,reaches the highest when the water content of 40%.1.6 The optimal extraction conditions is material liquid ratio of 1:300,extraction temperature of 60 ?,the extracting time for 50 minutes,the water content is 20%,and the extraction yield reaches 84.2%.2 The immune function of cordycepin,shell oligosaccharide and cordycepin joint shell oligosaccharide.2.1 The experimental group and control group in mice spleen weight/body weight ratio,serum hemolysin,thymus weight/body weight ratio,NK cell activity,creatinine and urea nitrogen are not statistically significant(P>0.05).2.2 Low dose cordycepin group and normal control group lymphocyte proliferation ability are statistically difference(P<0.05),the other group and the control group are not statistically difference(P>0.05).2.3 Low dose cordycepin group,high mixed dose group and the control group in late onset of allergy are statistically difference(P<0.05),the other group and the control group are not statistically difference(P>0.05).2.4 Low dose cordycepin group and normal group in mononuclear-macrophages carbon clearance ability statistically significant,other group and normal group are not statistically different.2.5 The experimental group and control group in glutamic oxalacetic transaminase and glutamic-pyruvic transaminase has no statistical difference,and has the difference in total cholesterol and bilirubin.In total cholesterol,low shell,mix and normal group are statistically difference(P<0.05).In the index of bilirubin,mixed low group and the control group are statistically difference(P<0.05),the other group and normal group are not statistically difference(P>0.05).Conclusions1.This experiment adopts the eutectic solvent as extraction solvent,and adopts single factor change as law of temperature,liquid ratio,extraction time and moisture content,using the orthogonal experiment method to get the optimal extraction conditions.The optimal extraction conditions is material liquid ratio of 1:300,extraction temperature of 60 ?,the extracting time for 50 minutes,the water content is 20%,and the extraction yield reaches 84.2%.2.Cordycepin and shell oligosaccharides can enhance the ability of delayed-type allergy,but can't increase lymphocyte proliferation ability,enhance lymphocyte proliferation ability,the ability of macrophage cell,the activity of NK cells and hemolysin level.3.Low concentration cordycepin group can enhance the ability of lymphocyte proliferation and macrophage phagocytosis,but cannot improve the capacity of late onset of allergy,NK cell activity and hemolysin index.4.Shell oligosaccharide cannot enhance the ability of lymphocyte proliferation,macrophage phagocytosis and late onset allergy,NK cell activity and hemolysin index. |
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