熟地黄多糖提取纯化工艺研究及其结构组成的初探
|
摘要
熟地黄多糖是“药食同源”熟地黄中特有的活性成分之一,药理研究表明,熟地黄多糖有增强免疫、抗肿瘤、补血等生理功能。研究熟地黄多糖的提取、纯化工艺及其结构性质,对熟地黄资源的深加工和熟地黄多糖食品、保健品的开发有重要意义。
本文在研究加热法提取熟地黄多糖的基础上,将超声技术引入熟地黄多糖的提取,用响应面设计法寻找超声提取的最佳工艺;并利用SephadexG-200葡聚糖凝胶与DEAE-Sepharose柱层析纯化熟地黄多糖,在结构分析中,采用了紫外光谱,红外光谱和薄层色谱。
熟地黄多糖加热提取实验表明,加热提取熟地黄多糖的最佳条件:颗粒粒度40目、浸体温度90℃、料液比1:70、浸提时间3h、此时熟地黄多糖的提取率为6.679%。以颗粒粒度40目熟地黄为原料超声提取熟地黄多糖能显著提高生产效率并简化提取工艺,其最佳条件为:料液比1:56,提取温度64.5℃,提取时间3.1min,熟地黄多糖的提取率为6.801%。得出回归方程:y=6.6463+0.304x1+0.1415x2+0.212x3-0.061x1x2-0.2872x1x3-0.0815x2x3-0.2527x12+0.0665x22-0.2727x32
熟地黄多糖分离纯化实验表明:熟地黄多糖提取液浓缩后用sevage法除蛋白,再用蒸馏水透析,经醇沉后的熟地黄多糖,冻干后备用。SephadexG-200葡聚糖凝胶纯化的的熟地黄多糖只有1个洗脱蜂,经DEAE-Sepharose纯化的熟地黄多糖呈现2个洗脱蜂,说明DEAE-Sepharose比SephadexG-200葡聚糖凝胶具有高的分辨率。在纯化过程中SephadexG-200葡聚糖凝胶几乎对色素无吸附作用,而DEAE-Sepharose对色素具有很强的吸附作用,因此,本实验选择DEAE-Sepharose纯化熟地黄多糖。
熟地黄多糖的单糖分析:采用薄层色谱分析法对熟地多糖的单糖组成进行了分析。将熟地黄多糖完全酸解后,采用硅胶薄层板,以正丁醇:乙酸:水(9:4:2)为展开剂,10%硫酸-乙醇溶液为显色剂。分析结果表明:熟地黄多糖主要由葡萄糖、半乳糖、果糖和水苏糖等单糖组成。
Rehmannia glutinosa polyses(RGP) exists particularly in Radix Rehmannia Preparate,which not only is food ,but is chinese tradtional medicine.Pharmacological investigations indicated RGP can resist tumor、adjust immunity,increase blood and so on.To study the extracting technology of and purification and structure are important to deeper the process of Radix Rehmannia Preparate and the development of functional food and medicine of RGP.
This paper based on studying hot extracting of RGP, the supersonic was applied to strengthen the extraction and the supersonic extracting was optimized using response surface methodology. The crude polysaccharides were isolated and purified by chromatographying with SephadexG-200 and DEAE-Sepharose column. During the structure analysis of RGP, purity and component sugars were carried out by UV, IR, TLC.
The extracting test showed that the best hot extracting technology of RGP was that:the particle of 40 mesh size;extraction temperature90℃sample ratio : solvent=1:70(m/V);extraction time 2h , the extracting rate is 6.679%.Supersonic extracting can improve the extracting rate and reduce the extracting time and simplify the extracting process. The better supersonic extracting technology was that: the particle of dried Radix Rehmannia Preparate 40 mesh size;sample ratio : solvent=1:56(m/V); extraction time 3.1min and extraction temperature 64.5℃. The extracting rate is 6.801%. After regression analysis , factors which influenced response variables significantly new selected. Consequently multinomial regression equation was obtained: y=6.6463+0.304x1+0.1415x2+0.212x3-0.061x1x2-0.2872x1x3-0.0815x2x3-0.2527x12+0.0665x22-0.2727x32
The separation and purification technology of RGP test showed: The extracting solution of RGP concentrated by vacuum rotary evaporator, then eliminated protein using sevage reagent and dialysed by distilled water. When ethanol of certain concentration added, many white deposit were formed. Then the deposit dried by vacuum freeze drier. The crude polysaccharides were isolated and purified by chromatographying with SephadexG-200 and DEAE-Sepharose column. DEAE-Sepharose column had a higher partition rate. During the purification process, DEAE-Sepharose column had more adsorption for pigment than SephadexG-200 column. Therefore, DEAE-Sepharose column was chosen for the purification of RGP.
Composition analysis of Rehmannia glutinose Polysaccharides was studied. The TLC method was established to analysis composition of Rehmannia glutinose Polysaccharides Silica gel plate, n-butanol : HAc : H2O (9:4:2) as developing solvent, 10%sulfuric acid-ethanol eoloration. Rehmannia glutinose Polysaccharides was composed of galactose, glucose, fructose and stachyose.
本文在研究加热法提取熟地黄多糖的基础上,将超声技术引入熟地黄多糖的提取,用响应面设计法寻找超声提取的最佳工艺;并利用SephadexG-200葡聚糖凝胶与DEAE-Sepharose柱层析纯化熟地黄多糖,在结构分析中,采用了紫外光谱,红外光谱和薄层色谱。
熟地黄多糖加热提取实验表明,加热提取熟地黄多糖的最佳条件:颗粒粒度40目、浸体温度90℃、料液比1:70、浸提时间3h、此时熟地黄多糖的提取率为6.679%。以颗粒粒度40目熟地黄为原料超声提取熟地黄多糖能显著提高生产效率并简化提取工艺,其最佳条件为:料液比1:56,提取温度64.5℃,提取时间3.1min,熟地黄多糖的提取率为6.801%。得出回归方程:y=6.6463+0.304x1+0.1415x2+0.212x3-0.061x1x2-0.2872x1x3-0.0815x2x3-0.2527x12+0.0665x22-0.2727x32
熟地黄多糖分离纯化实验表明:熟地黄多糖提取液浓缩后用sevage法除蛋白,再用蒸馏水透析,经醇沉后的熟地黄多糖,冻干后备用。SephadexG-200葡聚糖凝胶纯化的的熟地黄多糖只有1个洗脱蜂,经DEAE-Sepharose纯化的熟地黄多糖呈现2个洗脱蜂,说明DEAE-Sepharose比SephadexG-200葡聚糖凝胶具有高的分辨率。在纯化过程中SephadexG-200葡聚糖凝胶几乎对色素无吸附作用,而DEAE-Sepharose对色素具有很强的吸附作用,因此,本实验选择DEAE-Sepharose纯化熟地黄多糖。
熟地黄多糖的单糖分析:采用薄层色谱分析法对熟地多糖的单糖组成进行了分析。将熟地黄多糖完全酸解后,采用硅胶薄层板,以正丁醇:乙酸:水(9:4:2)为展开剂,10%硫酸-乙醇溶液为显色剂。分析结果表明:熟地黄多糖主要由葡萄糖、半乳糖、果糖和水苏糖等单糖组成。
Rehmannia glutinosa polyses(RGP) exists particularly in Radix Rehmannia Preparate,which not only is food ,but is chinese tradtional medicine.Pharmacological investigations indicated RGP can resist tumor、adjust immunity,increase blood and so on.To study the extracting technology of and purification and structure are important to deeper the process of Radix Rehmannia Preparate and the development of functional food and medicine of RGP.
This paper based on studying hot extracting of RGP, the supersonic was applied to strengthen the extraction and the supersonic extracting was optimized using response surface methodology. The crude polysaccharides were isolated and purified by chromatographying with SephadexG-200 and DEAE-Sepharose column. During the structure analysis of RGP, purity and component sugars were carried out by UV, IR, TLC.
The extracting test showed that the best hot extracting technology of RGP was that:the particle of 40 mesh size;extraction temperature90℃sample ratio : solvent=1:70(m/V);extraction time 2h , the extracting rate is 6.679%.Supersonic extracting can improve the extracting rate and reduce the extracting time and simplify the extracting process. The better supersonic extracting technology was that: the particle of dried Radix Rehmannia Preparate 40 mesh size;sample ratio : solvent=1:56(m/V); extraction time 3.1min and extraction temperature 64.5℃. The extracting rate is 6.801%. After regression analysis , factors which influenced response variables significantly new selected. Consequently multinomial regression equation was obtained: y=6.6463+0.304x1+0.1415x2+0.212x3-0.061x1x2-0.2872x1x3-0.0815x2x3-0.2527x12+0.0665x22-0.2727x32
The separation and purification technology of RGP test showed: The extracting solution of RGP concentrated by vacuum rotary evaporator, then eliminated protein using sevage reagent and dialysed by distilled water. When ethanol of certain concentration added, many white deposit were formed. Then the deposit dried by vacuum freeze drier. The crude polysaccharides were isolated and purified by chromatographying with SephadexG-200 and DEAE-Sepharose column. DEAE-Sepharose column had a higher partition rate. During the purification process, DEAE-Sepharose column had more adsorption for pigment than SephadexG-200 column. Therefore, DEAE-Sepharose column was chosen for the purification of RGP.
Composition analysis of Rehmannia glutinose Polysaccharides was studied. The TLC method was established to analysis composition of Rehmannia glutinose Polysaccharides Silica gel plate, n-butanol : HAc : H2O (9:4:2) as developing solvent, 10%sulfuric acid-ethanol eoloration. Rehmannia glutinose Polysaccharides was composed of galactose, glucose, fructose and stachyose.
引文
[1]王培根,苯酚-硫酸法测定小柴胡汤口服液多糖的含量[J].中成药,1993,15(3):12-13
[2]王旭,胖大海中多糖的成分分析和含量测定[J].中国实验方剂学杂志,1997,3(5):39-40
[3]陈健民,紫外分光光度法测定灵芝菌丝体发酵液中多糖的含量[J].中药材,1994,17(8):32-33
[4]苗三明,李振国主编.现代实用中药质量技术[M].北京,人民卫生出版社,2000,381-382
[5]倪慕云,边宝林,王宏生,干地黄化学成分的研究[J].中国中药杂志,1992,17(6):297-298
[6]倪慕云,边宝林,王宏杰,地黄及其炮制品石油提取物中酸性成分的分离、鉴定和比较[J].中国中药杂志,1991,16(6):339-340
[7]扬云,苗三明,王浴铭,怀地黄多糖化学研究[J].时珍国医国药,1999,10(8):564-565
[8]张玲,徐新刚,时延增,地黄提取工艺的研究[J].中草药,1998,29(5):308-310
[9]杜灿屏,刘鲁生,张恒,21世纪有机化学发展战略[M].北京:化学工业出版社,2002:298-316
[10]宋建德,袁丽萍,梁勇,药用植物多糖的免役作用研究进展[J].中兽医医药杂志[J],2004,2:22-24
[11]韩果萍,段玉峰,我国天然活性多糖药理研究进展[J].江苏临床医学杂志,2000,4(4):227
[12]胡文祥,王来曦,多糖及其衍生物的医药学研究[J].科学(中文版),1994,187(3):4-8
[13]姚新生主编,天然药物化学[M] .第三版,北京:人民卫生出版社,2001:101
[14]谭周进,谢达平,多糖的研究进展[J].食品科技,2002,3:10-12
[15]王卫国,赵永亮,韩山宝,香菇多糖分离纯化技术的研究,中国食用菌,2001,21(2):30-32
[16]张晓静,刘会东,植物多糖提取分离及药物作用的研究进展,时珍国医国药,2003,14(8):495—497
[17]池玉梅,李伟,文红梅,白术多糖的分离纯化和化学结构研究[J].中药材,2001,24(9):647-648
[18]杨洪彩,张月明,邹红云,植物多糖药效研究进展,地方病通报,2004,19(2):88-90
[19]吴亚林,黄静,潘远江,无花果多糖的分离、纯化和鉴定[J].浙江大学学报,2004,31(2):178-179,187
[20]鞠海,张建民,魏峰,天然多糖的分离、纯化和结构鉴定,国外医药.植物药分册,2000,15(3):107-113
[21]田庚元,冯宇澄,林颖,植物多糖的研究进展[J].中国中药杂志,1995,20(7):441-444
[22]王长云,多糖抗病毒作用研究进展[J].生物工程进展,2000,20(1),17
[23]王洪斌,郑钦岳,钱定,商陆多糖对小鼠免疫功能的影响[J].中国药理学报,1993,14(3):24-26
[24]王莉,刘志勇,鲁建江,黄芪多糖的微波提取及含量测定[J].中医药学报,2001,29(6):35-36
[25]魏敏,江雪华,正交设计法优选黄芪提取工艺研究[J].基层中药杂志,2000,14(6):23-25
[26]吴光枫,汤坚,芦荟多粉的纯化与体外抗氧化活性的研究[J].食品科学,2002,23(9):129-132
[27]张安强,张劲松,潘迎捷.食药用菌多糖的提取、分离纯化与结构分析[J].食用菌学报,2005,12(2):62-68
[28]聂凌鸿,宁正祥,活性多糖的构效关系,林产化学与工业,2003,23(4):89-94
[29]黄霞,庆慧,王慧森,熟地水煎液及其提取物对小鼠外周血象的比较的研究.中成药,2002,24(2):111
[30]元阿萍,刘超,黄霞,熟地多糖联合康力龙治疗慢性在生障碍性贫血34例.中国中西医结合杂志,1998,18(6):351
[31]宋建德,袁丽萍,梁勇,药用植物多糖的免疫作用研究进展,中兽医医药杂志,2004,2:22-24
[32]熊子仙,植物多糖与保健,云南师范大学学报,2004,(3):41-43
[33]陈怡,天然多糖的研究概况[J].世界科学技术—中药现代化,2000,2(6):52-55
[34]徐娟华,马武翔,石莼多糖的提取分离纯化及其降血脂作用的初步研究,中国中医药科技,2002,9(3):167-168
[35]梁英,扬宏志,夏远亮,黑木耳硒多糖对小鼠及过氧化物酶的影响,营养学报,2000,22(3):250-252
[36]汪东风,茶叶多糖抗辐射能力的研究,茶叶科学,1996,16(1):1-8
[37]张以芳,段刚,螺旋藻及其多糖、多糖蛋白提取物对体外癌细胞的抑制作用,海洋科学,2000,3:16-18
[38]曲显俊,崔淑香,螺旋藻多糖抗癌作用的实验研究,中国海洋药物,2000,(4):10-14
[39]衣艳君,枸杞降血脂作用的实验研究,首都师范大学学报,2000,21(4):68-70
[40]王蓉,吴剑波,多糖生物活性的研究进展.国外医药抗声素分册,2001,22(5):97-100
[41]赵骏,钟蓉,王洪章,桑叶多糖提取工艺的优选,中草药,2000,31(5):347-348
[42]黄民权,蔡体育,黄步汉,铁皮石斛多糖的提取分离和分析,中草药,1994,25(3):128-129
[43]汪程远,张浩,孟莉,大孔吸附树枝分离纯化生地黄中苷类与糖类,中药材,2003,26(3):202-203
[44]肖建军,蒋依辉,食药用真菌多糖研究进展,生命的化学,2002,22(2):148-151
[45]胡庭俊,生物活性多糖与机体免疫系统的信号转导,中兽医医药杂志,2000,2:17-18
[46]郑敏,王亚军,中药多糖抗肿瘤的药理学研究进展.国外医学(中医中药分册),2000,22(5):259-263
[47]许爱华,陈华圣,银杏外种皮多糖对人癌细胞的抑制作用及阿霉素的协同效应,中国新药杂志,2000,9(11):753-755
[48]Gao H著,方唯硕译,具有抗HIV活性的天然产物,国外医药植物分册,1993,8(2):65
[49]王钦茂,洪浩,赵帜平,丹皮多糖-2b对2型糖尿病大鼠模型的作用及其降解作用机制,中国药理学同报,2002,18(4):456-459
[50]廖建民,沈子龙,张瑾,海带多糖中不同组分降血脂及抗肿瘤作用的研究,中国药科大学学报,2002,33(1):55-57
[51]宫司进之,大花紫薇和桑叶提取物对大鼠血糖值的影响,国外医学,中医中药分册,2000,22(4):232
[52]张维杰.糖复合物生化研究技术[M].杭州:浙江人民出版社,1999,56
[53]中国药典(Ch P)[S].VolII.2000
[54]Zhang P,Zhang L,Cheng S.Effect of urea and sodium hydroxide on the molecular weight and comformation ofβ-(1,3)-D-glucan from Letinus edodes in aqueous solution[J] .Carbohydr Rs,2000,327(2):431-438
[55]Bing Lu;Douglas Westerlund;Indirect UV detection of carbohydrates in cappillary zone electrophoresis by using tryptophan as a maker [J].Electroporesis.1996,17,325-332
[56]Y-H. Lee,T-I.Lin.Determination of carbohydrates by high-performance capillary electrophoresis with indirect absorbance detection[J].J.Chro-matogrB,1996,681:87-97
[57]A Possible role for protein kinase C.J Biol Chem,1999,274(20):13790-13799
[58]Ana P Bartolome,Pilar Ruperez.Polysacchrides from the walls of pineapple fruit.Journal of agriculture and food chemistry,1995,43,608-612
[59] Ryoko Conda,Masashi Tomoda. Structural features of Vkonan C,a reticuloendotheliar system-activaing polysaccharide from the Rhizome of curcuma longa.Pharm.Bull,1991,39(2):441
[60] Yokota S, Amano K,Hayashi S,et al.Low antigenicity of the polysaccharide region of Helicobacter pylori lipopolysaccharides derived from tumors of patients with gastric cancer . Infection-and-Immunity,1997,65(9):3509-3512
[61]Maisin J R.Radio prontection by polysaccharides,Journal of Clinical Pharmacy and Therapeutics,1998,339,225
[62]Zhao J F,Hiroaki K,Haruki Y, Heterogeneity and characterisation of mitogenic and anti-complementary pectic polysaccharides from the roots of Glycyrrhiza uralensis Fisch et D.C[J] Carbohydr.Res,1991:219:149-172
[63]Hiroaki K, Norito T,Zhao J F, et al. Pectic polysaccharides from roots of glycyrrhiza uralensis:possible contribution of neutral oligosaccharides in the galacturonase-resistant region to anti-complementary and mitogenic activities[J].PlantMed,1996,62:14-19
[64]Xiaoming Qin et al. Structural features of arabinogalactan-proteins from the fruit of Lycium Chinese Mill [J]. Cabohohydrate Res,2001,333:79-85
[65]A.Q. Arifkhodzhave.Galactans and Galactan-containing polysaccides of Higher plants. Chemistry of Natural Compouds,2000,36(3):229-242
[66]Wyss D F,Choi J S. et al.Conformation and Function of the N-linked Glycan in the Adbesion Domain of Human CD2.1995,269(5228):1276-1278
[67]Maria I B, Alexey A G, Nadezhda E U, et al.A highly regula fraction of a fucoidan from the brown seaweed Fucus distichusL. Carbohydrate Res,2003, 339;500-517
[68]T.Yoshida, Synthesis of polysaccharides having specific biological activities. Prog, Polvm. Sci,2001,26:379-441
[69]Dubois M Gilles, K A Hamilton, J K Rebers, et al.Colorimetric method for determination of sugars and related substance [J]. Anal Chem ,1956,28:350-356
[70]Michael A. Mc Donough et al. Rhamnogalacturonan lyase reveals a unique three-domain modular structure for polysaccharide lyase family. FEBS Letters,2004,565:188-194
[71]Yusuke Edashige et al.Rhamonogalacturonan I from xylem differentiating zones of Cruptomeria japonica.Carbohydrate Res, 1997,304:357-365
[72]Valerie Rion et al. Aggregation of grape seed tannins in model wine-effect of wine polysaccharides. Food Hydrocollids,2002,16:17-23
[73]P. Pellerin et al. Characterization of five type II arabiongalactan-protein fraction from red wine of increasing uronic acid content.Carbohydrate Res,1995,277:135-143
[74]Christophe Rihhouey, et al.Structural features of CDTA-solubie pectins from flax hypocotyls. Carbohydrate Res,1995,28:159-166
[75]Ian J . Colquhoun et al.Identification by n.m.r.spectroscopy of oligosaccharides obtaened by treatment of the hairy regions of apple pectin with rhamnogalacturonase. Carbohydrate Res,1990,201:131-144
[76]Michael Mcneil et al.Structure of plant Cel Walls. Plant Physical,1980,66:1128-1134
[77]Masumi H.Sakuri et al. Characterization of antigenic epitopes in anti-ulcer pectic polysaccharides from Bupleurum falcatum L. using several carbohvdrates.Carbohvdrate Res,1998,311:219-229
[78]Alexander Osterceld et al. Characterization of hop pectins shows the presence of an arabinogalactan-protein. Carbohydrate Polymers,2002,49:407-413
[79]Thierry Doco et al. Rhamnogalacturonan II, a dominant polysaccharide in juices produces by enzymic liquefaction of fruits and vegetables. Carbohydrate Res,1997,297:181-186
[80]Siobhain Reid et al.Characterisation of extracellular ploysaccharides from suspension cultures of apple(Malus domestica).Carbohydrate polymer,1999,39:369-376
[81]Stefan Karacsonyi et al . Structure study on arabinogalactan-protein from Picea abies L. Karst.Carbohydrate Res,1998,307:271-279
[82]Drissa Diallo et al . Polysaccharides from the roots of Entada africana Guill. Et Perr, Mimosaceae,with complement fixing activity. Journal of Ethnopharmacology.2001,74:159-171
[2]王旭,胖大海中多糖的成分分析和含量测定[J].中国实验方剂学杂志,1997,3(5):39-40
[3]陈健民,紫外分光光度法测定灵芝菌丝体发酵液中多糖的含量[J].中药材,1994,17(8):32-33
[4]苗三明,李振国主编.现代实用中药质量技术[M].北京,人民卫生出版社,2000,381-382
[5]倪慕云,边宝林,王宏生,干地黄化学成分的研究[J].中国中药杂志,1992,17(6):297-298
[6]倪慕云,边宝林,王宏杰,地黄及其炮制品石油提取物中酸性成分的分离、鉴定和比较[J].中国中药杂志,1991,16(6):339-340
[7]扬云,苗三明,王浴铭,怀地黄多糖化学研究[J].时珍国医国药,1999,10(8):564-565
[8]张玲,徐新刚,时延增,地黄提取工艺的研究[J].中草药,1998,29(5):308-310
[9]杜灿屏,刘鲁生,张恒,21世纪有机化学发展战略[M].北京:化学工业出版社,2002:298-316
[10]宋建德,袁丽萍,梁勇,药用植物多糖的免役作用研究进展[J].中兽医医药杂志[J],2004,2:22-24
[11]韩果萍,段玉峰,我国天然活性多糖药理研究进展[J].江苏临床医学杂志,2000,4(4):227
[12]胡文祥,王来曦,多糖及其衍生物的医药学研究[J].科学(中文版),1994,187(3):4-8
[13]姚新生主编,天然药物化学[M] .第三版,北京:人民卫生出版社,2001:101
[14]谭周进,谢达平,多糖的研究进展[J].食品科技,2002,3:10-12
[15]王卫国,赵永亮,韩山宝,香菇多糖分离纯化技术的研究,中国食用菌,2001,21(2):30-32
[16]张晓静,刘会东,植物多糖提取分离及药物作用的研究进展,时珍国医国药,2003,14(8):495—497
[17]池玉梅,李伟,文红梅,白术多糖的分离纯化和化学结构研究[J].中药材,2001,24(9):647-648
[18]杨洪彩,张月明,邹红云,植物多糖药效研究进展,地方病通报,2004,19(2):88-90
[19]吴亚林,黄静,潘远江,无花果多糖的分离、纯化和鉴定[J].浙江大学学报,2004,31(2):178-179,187
[20]鞠海,张建民,魏峰,天然多糖的分离、纯化和结构鉴定,国外医药.植物药分册,2000,15(3):107-113
[21]田庚元,冯宇澄,林颖,植物多糖的研究进展[J].中国中药杂志,1995,20(7):441-444
[22]王长云,多糖抗病毒作用研究进展[J].生物工程进展,2000,20(1),17
[23]王洪斌,郑钦岳,钱定,商陆多糖对小鼠免疫功能的影响[J].中国药理学报,1993,14(3):24-26
[24]王莉,刘志勇,鲁建江,黄芪多糖的微波提取及含量测定[J].中医药学报,2001,29(6):35-36
[25]魏敏,江雪华,正交设计法优选黄芪提取工艺研究[J].基层中药杂志,2000,14(6):23-25
[26]吴光枫,汤坚,芦荟多粉的纯化与体外抗氧化活性的研究[J].食品科学,2002,23(9):129-132
[27]张安强,张劲松,潘迎捷.食药用菌多糖的提取、分离纯化与结构分析[J].食用菌学报,2005,12(2):62-68
[28]聂凌鸿,宁正祥,活性多糖的构效关系,林产化学与工业,2003,23(4):89-94
[29]黄霞,庆慧,王慧森,熟地水煎液及其提取物对小鼠外周血象的比较的研究.中成药,2002,24(2):111
[30]元阿萍,刘超,黄霞,熟地多糖联合康力龙治疗慢性在生障碍性贫血34例.中国中西医结合杂志,1998,18(6):351
[31]宋建德,袁丽萍,梁勇,药用植物多糖的免疫作用研究进展,中兽医医药杂志,2004,2:22-24
[32]熊子仙,植物多糖与保健,云南师范大学学报,2004,(3):41-43
[33]陈怡,天然多糖的研究概况[J].世界科学技术—中药现代化,2000,2(6):52-55
[34]徐娟华,马武翔,石莼多糖的提取分离纯化及其降血脂作用的初步研究,中国中医药科技,2002,9(3):167-168
[35]梁英,扬宏志,夏远亮,黑木耳硒多糖对小鼠及过氧化物酶的影响,营养学报,2000,22(3):250-252
[36]汪东风,茶叶多糖抗辐射能力的研究,茶叶科学,1996,16(1):1-8
[37]张以芳,段刚,螺旋藻及其多糖、多糖蛋白提取物对体外癌细胞的抑制作用,海洋科学,2000,3:16-18
[38]曲显俊,崔淑香,螺旋藻多糖抗癌作用的实验研究,中国海洋药物,2000,(4):10-14
[39]衣艳君,枸杞降血脂作用的实验研究,首都师范大学学报,2000,21(4):68-70
[40]王蓉,吴剑波,多糖生物活性的研究进展.国外医药抗声素分册,2001,22(5):97-100
[41]赵骏,钟蓉,王洪章,桑叶多糖提取工艺的优选,中草药,2000,31(5):347-348
[42]黄民权,蔡体育,黄步汉,铁皮石斛多糖的提取分离和分析,中草药,1994,25(3):128-129
[43]汪程远,张浩,孟莉,大孔吸附树枝分离纯化生地黄中苷类与糖类,中药材,2003,26(3):202-203
[44]肖建军,蒋依辉,食药用真菌多糖研究进展,生命的化学,2002,22(2):148-151
[45]胡庭俊,生物活性多糖与机体免疫系统的信号转导,中兽医医药杂志,2000,2:17-18
[46]郑敏,王亚军,中药多糖抗肿瘤的药理学研究进展.国外医学(中医中药分册),2000,22(5):259-263
[47]许爱华,陈华圣,银杏外种皮多糖对人癌细胞的抑制作用及阿霉素的协同效应,中国新药杂志,2000,9(11):753-755
[48]Gao H著,方唯硕译,具有抗HIV活性的天然产物,国外医药植物分册,1993,8(2):65
[49]王钦茂,洪浩,赵帜平,丹皮多糖-2b对2型糖尿病大鼠模型的作用及其降解作用机制,中国药理学同报,2002,18(4):456-459
[50]廖建民,沈子龙,张瑾,海带多糖中不同组分降血脂及抗肿瘤作用的研究,中国药科大学学报,2002,33(1):55-57
[51]宫司进之,大花紫薇和桑叶提取物对大鼠血糖值的影响,国外医学,中医中药分册,2000,22(4):232
[52]张维杰.糖复合物生化研究技术[M].杭州:浙江人民出版社,1999,56
[53]中国药典(Ch P)[S].VolII.2000
[54]Zhang P,Zhang L,Cheng S.Effect of urea and sodium hydroxide on the molecular weight and comformation ofβ-(1,3)-D-glucan from Letinus edodes in aqueous solution[J] .Carbohydr Rs,2000,327(2):431-438
[55]Bing Lu;Douglas Westerlund;Indirect UV detection of carbohydrates in cappillary zone electrophoresis by using tryptophan as a maker [J].Electroporesis.1996,17,325-332
[56]Y-H. Lee,T-I.Lin.Determination of carbohydrates by high-performance capillary electrophoresis with indirect absorbance detection[J].J.Chro-matogrB,1996,681:87-97
[57]A Possible role for protein kinase C.J Biol Chem,1999,274(20):13790-13799
[58]Ana P Bartolome,Pilar Ruperez.Polysacchrides from the walls of pineapple fruit.Journal of agriculture and food chemistry,1995,43,608-612
[59] Ryoko Conda,Masashi Tomoda. Structural features of Vkonan C,a reticuloendotheliar system-activaing polysaccharide from the Rhizome of curcuma longa.Pharm.Bull,1991,39(2):441
[60] Yokota S, Amano K,Hayashi S,et al.Low antigenicity of the polysaccharide region of Helicobacter pylori lipopolysaccharides derived from tumors of patients with gastric cancer . Infection-and-Immunity,1997,65(9):3509-3512
[61]Maisin J R.Radio prontection by polysaccharides,Journal of Clinical Pharmacy and Therapeutics,1998,339,225
[62]Zhao J F,Hiroaki K,Haruki Y, Heterogeneity and characterisation of mitogenic and anti-complementary pectic polysaccharides from the roots of Glycyrrhiza uralensis Fisch et D.C[J] Carbohydr.Res,1991:219:149-172
[63]Hiroaki K, Norito T,Zhao J F, et al. Pectic polysaccharides from roots of glycyrrhiza uralensis:possible contribution of neutral oligosaccharides in the galacturonase-resistant region to anti-complementary and mitogenic activities[J].PlantMed,1996,62:14-19
[64]Xiaoming Qin et al. Structural features of arabinogalactan-proteins from the fruit of Lycium Chinese Mill [J]. Cabohohydrate Res,2001,333:79-85
[65]A.Q. Arifkhodzhave.Galactans and Galactan-containing polysaccides of Higher plants. Chemistry of Natural Compouds,2000,36(3):229-242
[66]Wyss D F,Choi J S. et al.Conformation and Function of the N-linked Glycan in the Adbesion Domain of Human CD2.1995,269(5228):1276-1278
[67]Maria I B, Alexey A G, Nadezhda E U, et al.A highly regula fraction of a fucoidan from the brown seaweed Fucus distichusL. Carbohydrate Res,2003, 339;500-517
[68]T.Yoshida, Synthesis of polysaccharides having specific biological activities. Prog, Polvm. Sci,2001,26:379-441
[69]Dubois M Gilles, K A Hamilton, J K Rebers, et al.Colorimetric method for determination of sugars and related substance [J]. Anal Chem ,1956,28:350-356
[70]Michael A. Mc Donough et al. Rhamnogalacturonan lyase reveals a unique three-domain modular structure for polysaccharide lyase family. FEBS Letters,2004,565:188-194
[71]Yusuke Edashige et al.Rhamonogalacturonan I from xylem differentiating zones of Cruptomeria japonica.Carbohydrate Res, 1997,304:357-365
[72]Valerie Rion et al. Aggregation of grape seed tannins in model wine-effect of wine polysaccharides. Food Hydrocollids,2002,16:17-23
[73]P. Pellerin et al. Characterization of five type II arabiongalactan-protein fraction from red wine of increasing uronic acid content.Carbohydrate Res,1995,277:135-143
[74]Christophe Rihhouey, et al.Structural features of CDTA-solubie pectins from flax hypocotyls. Carbohydrate Res,1995,28:159-166
[75]Ian J . Colquhoun et al.Identification by n.m.r.spectroscopy of oligosaccharides obtaened by treatment of the hairy regions of apple pectin with rhamnogalacturonase. Carbohydrate Res,1990,201:131-144
[76]Michael Mcneil et al.Structure of plant Cel Walls. Plant Physical,1980,66:1128-1134
[77]Masumi H.Sakuri et al. Characterization of antigenic epitopes in anti-ulcer pectic polysaccharides from Bupleurum falcatum L. using several carbohvdrates.Carbohvdrate Res,1998,311:219-229
[78]Alexander Osterceld et al. Characterization of hop pectins shows the presence of an arabinogalactan-protein. Carbohydrate Polymers,2002,49:407-413
[79]Thierry Doco et al. Rhamnogalacturonan II, a dominant polysaccharide in juices produces by enzymic liquefaction of fruits and vegetables. Carbohydrate Res,1997,297:181-186
[80]Siobhain Reid et al.Characterisation of extracellular ploysaccharides from suspension cultures of apple(Malus domestica).Carbohydrate polymer,1999,39:369-376
[81]Stefan Karacsonyi et al . Structure study on arabinogalactan-protein from Picea abies L. Karst.Carbohydrate Res,1998,307:271-279
[82]Drissa Diallo et al . Polysaccharides from the roots of Entada africana Guill. Et Perr, Mimosaceae,with complement fixing activity. Journal of Ethnopharmacology.2001,74:159-171