六种蜂花粉果胶的结构分析及抑制神经氨酸酶的活性研究
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摘要
蜂花粉是蜜蜂采集花粉后，加入少量的分泌物和花蜜，加工成的扁圆形花粉团。蜂花粉具有多种生物活性，例如提高机体免疫力、抗肿瘤、抗病毒、抗氧化和美容养颜等。多糖是蜂花粉的重要活性成分，本论文研究了六种蜂花粉果胶的结构以及抑制流感病毒神经氨酸酶的活性。主要包括；蜂花粉中I型聚鼠李半乳糖醛酸（RG-I）和聚半乳糖醛酸（HG）果胶的精细结构比较研究；蜂花粉果胶结构及其抑制流感病毒神经氨酸酶的活性；以荷花蜂花粉RG-I果胶（WNPP-2-RG）为例分析抗病毒活性的可能机制；对WNPP-2-RG的精细结构进行了分析，探讨了其结构与免疫活性的关系。果胶在自然界中含量丰富，结构复杂，活性多样。本论文的实验结果将为研究果胶结构与活性的关系奠定基础，促进果胶类多糖的进一步应用。
论文选取荷花（Nelumbo nucifera）、虞美人（Papaver rhoeas）、玫瑰（Rosa rugosa）、荞麦（Fagopyrum esculentum）、五味子（Schisandra chinensis）和油菜（Brassicacampestris）六种蜂花粉为原料，通过系统地分级与纯化，得到六种蜂花粉RG-I和六种HG型果胶。结构分析表明，玫瑰蜂花粉RG-I型果胶只含有I型阿拉伯半乳聚糖（AG-I）侧链，其余五中RG-I型果胶都同时含有I型和II型阿拉伯半乳聚糖（AG-II）侧链；鼠李糖（Rha）与半乳糖醛酸（GalA）比例约为1:1，不含或仅含少量HG结构域；Rha和GalA构成的主链结构域含量在16%~50%之间；半乳糖和阿拉伯糖的总含量多数在70%~75%之间；油菜蜂花粉的RG-I型果胶除含有上述结构外，还含有葡聚糖和木聚糖侧链；RG-I型果胶分子量多在3.8×10~5Da左右。六种蜂花粉HG型果胶都含有较多的GalA；少量的RG-I和大量的HG结构域构成核心结构；侧链存在一定的差异，荷花蜂花粉HG果胶同时含有AG-I和AG-II两种类型的侧链，虞美人和玫瑰蜂花粉HG果胶含有1,5-arabinan和1,3-galactan侧链，五味子蜂花粉HG果胶含有1,5-arabinan和1,4-galactan侧链，荞麦蜂花粉HG果胶含有1,4-xylan侧链；虞美人和油菜蜂花粉HG型果胶中含有20%左右的葡聚糖；分子量在8×103~3.6×10~4Da之间。
为了分析果胶的构效关系，对六种RG-I和六种HG果胶抑制流感病毒神经氨酸酶的活性进行了研究。神经氨酸酶是流感病毒发挥活性的关键酶，抑制其活性可达到抗流感病毒的作用，因此该结果可用于探讨果胶结构与抗流感病毒活性的关系。利用本实验室建立的生物发光法和经典的底物荧光法，对果胶级分地抑制流感病毒H1N1和H5N1神经氨酸酶的活性进行了研究。结果表明，富含HG结构域的果胶级分抑制活性明显高于RG-I级分。其中，来源于虞美人和油菜蜂花粉中的HG型果胶对流感病毒神经氨酸酶抑制活性高于其它HG型果胶，对两种酶的IC50均在1mg/mL以下。根据各个级分的结构特征可以推测，HG结构域是具有抑制流感病毒神经氨酸酶活性的主要活性单元，而葡萄糖侧链的存在会大大促进这种抑制作用。
RG-I果胶没有直接抑制流感病毒神经氨酸酶的活性，可能是通过其它途径达到抗流感病毒效果。选择荷花蜂花粉RG-I级分WNPP-2-RG为代表，研究结果表明，它能够显著促进免疫指标，免疫调节作用可能是其发挥抗流感病毒活性的途径之一。对WNPP-2-RG的精细结构进行了鉴定。它以RG-I结构域为核心，AG-I和AG-II为侧链，分子量为3.8×10~5Da。侧链中1,6-Gal构成骨架结构，末端主要为Gal、少量Glc和Ara。为了进一步研究果胶结构与活性的关系，对WNPP-2-RG结构与免疫活性的关系进行了分析。通过部分酸水解除去侧链中的Ara，剩余核心结构为RG-I和AG-II结构域。体外免疫活性结果表明，侧链中的Ara的除去和Gal的暴露，对淋巴细胞增殖作用影响不大，但能够显著促进巨噬细胞的吞噬作用，降低NO释放能力。将WNPP-2-RG与人参RG-I型果胶（WGPA-2-RG）的结构和活性进行了对比研究。结果表明，侧链的类型、糖残基连接方式、甲酯化度、乙酰化度和分子量等不同，对活性的影响也不同。
Bee pollen is the male gametophyte of gymnosperm and angiosperm, which containsmost of the nutrients afford to plant growth and development. It is well known as a naturalnutrition and healthy food especially in improving human’s immunity. Bee pollen had manyactivities, such as antitumor, antivirus and immunobiological activities. Polysaccharides arethe main active components in the bee pollen. In this paper, we analyzed the structure ofpectic polysaccharides from six types of bee pollen and their antivirus activity, including thestructure comparision of type I rhamnogalacturonan (RG-I) and homogalacturonan (HG); therelationship of structure and inhibitory activity on H1N1and H5N1neuraminidase; the fluvirus inhibitory mechanism of RG-I pectin; the fine structure of RG-I pectin from bee pollenof Nelumbo nucifera (WNPP-2-RG) and its structure-activity relationship. The results of thispaper can offer some datas about structure-activity relationship of pectins, improved theapplication of pectin.
In this paper, we choosed six kinds of bee pollens, including Nelumbo nucifera,Papaver rhoeas, Rosa rugosa, Fagopyrum esculentum, Schisandra chinensis and Brassicacampestris, for studying the structure features and structure-activity relationship of pectins.Six RG-I and six HG pectins were obtained by a combination of DEAE-Cellulose andSepharose CL-6B colume. Structure analysis showed six RG-I pectins from bee pollencontained RG-I core region, both type I arabinogalactan (AG-I) and type II arabinogalactan(AG-II) sidechains except RG-I pectin from Rosa Rugosa which only contained AG-Isidechains; the ratio of rhamnose (Rha) and galacturonic acid (GalA) was nearly1:1,contained little HG region; RG-I mainchain contents was about16%~50%; total contents ofgalactose (Gal) and arabinose (Ara) were among70%~75%; the RG-I pectin from Brassicacampestris was contained glucan and xylan sidechains; the molecular weight of RG-Ifractions is about3.8×10~5Da. HG pectins from six kinds of bee pollen contains largeamounts of GalA; HG and RG-I region composed the core structure, ratio of Ara and Galwas nearly1:1; HG pectin from Nelumbo nucifera contained both AG-I and AG-IIsidechains, HG pectin from Papaver rhoeas and Rosa rugosa contained1,5-arabinan and1,3-galactan sidechains, HG pectin from Schisandra chinensis contained1,5-arabinan and1,4-galactan sidechains, HG pectin from Fagopyrum esculentum contained1,4-xylansidechains; HG pectin Papaver rhoeas and Brassica campestris contained glucose (Glc,about20%); the molecular weight was among8×103~3.6×10~4Da.
The inhibitory activity of RG-I and HG fractions on H1N1and H5N1neuraminidaseswas studied by bioluminescent assay and typical fluorescent method. The results indicatedHG fractions had better inhibition activity than RG-I fractions, especially the HG fracionsfrom Papaver rhoeas and Brassica campestris, IC50were below1mg/mL. According to thestructure features of each fraction, we can concluded HG region was the important region oninhibitory activity of H1N1and H5N1neuraminidases, the Glc sidechains can improve thisactivity.
The immunological activity of RG-I pectin (WNPP-2-RG) from bee pollen of Nelumbonucifera was analysed, it can improve the immunological activities, suggested RG-I mighthave antivirus activity through immunological way. Structure analysis showed WNPP-2-RGhas an RG-I core region, both AG-I and AG-II sidechains.1,6-Gal composed of thebackbone of sidechains, branched at O-3or O-4. Most of the Gal residues were determinedto be close to the molecular core, some Gal and Ara on the surface of the molecules. Theimmunological activitiy of WNPP-2-RG was related to its side chains. Ara residues affectedits macrophage phagocytosis and NO production, but no effect on lymphocyte proliferation.The comparation of structure-immunological activity between WNPP-2-RG and ginsengRG-I fraction WGPA-2-RG was performed. Ara residues in different RG-I pectins haddifferent effects on their activities.
论文选取荷花（Nelumbo nucifera）、虞美人（Papaver rhoeas）、玫瑰（Rosa rugosa）、荞麦（Fagopyrum esculentum）、五味子（Schisandra chinensis）和油菜（Brassicacampestris）六种蜂花粉为原料，通过系统地分级与纯化，得到六种蜂花粉RG-I和六种HG型果胶。结构分析表明，玫瑰蜂花粉RG-I型果胶只含有I型阿拉伯半乳聚糖（AG-I）侧链，其余五中RG-I型果胶都同时含有I型和II型阿拉伯半乳聚糖（AG-II）侧链；鼠李糖（Rha）与半乳糖醛酸（GalA）比例约为1:1，不含或仅含少量HG结构域；Rha和GalA构成的主链结构域含量在16%~50%之间；半乳糖和阿拉伯糖的总含量多数在70%~75%之间；油菜蜂花粉的RG-I型果胶除含有上述结构外，还含有葡聚糖和木聚糖侧链；RG-I型果胶分子量多在3.8×10~5Da左右。六种蜂花粉HG型果胶都含有较多的GalA；少量的RG-I和大量的HG结构域构成核心结构；侧链存在一定的差异，荷花蜂花粉HG果胶同时含有AG-I和AG-II两种类型的侧链，虞美人和玫瑰蜂花粉HG果胶含有1,5-arabinan和1,3-galactan侧链，五味子蜂花粉HG果胶含有1,5-arabinan和1,4-galactan侧链，荞麦蜂花粉HG果胶含有1,4-xylan侧链；虞美人和油菜蜂花粉HG型果胶中含有20%左右的葡聚糖；分子量在8×103~3.6×10~4Da之间。
为了分析果胶的构效关系，对六种RG-I和六种HG果胶抑制流感病毒神经氨酸酶的活性进行了研究。神经氨酸酶是流感病毒发挥活性的关键酶，抑制其活性可达到抗流感病毒的作用，因此该结果可用于探讨果胶结构与抗流感病毒活性的关系。利用本实验室建立的生物发光法和经典的底物荧光法，对果胶级分地抑制流感病毒H1N1和H5N1神经氨酸酶的活性进行了研究。结果表明，富含HG结构域的果胶级分抑制活性明显高于RG-I级分。其中，来源于虞美人和油菜蜂花粉中的HG型果胶对流感病毒神经氨酸酶抑制活性高于其它HG型果胶，对两种酶的IC50均在1mg/mL以下。根据各个级分的结构特征可以推测，HG结构域是具有抑制流感病毒神经氨酸酶活性的主要活性单元，而葡萄糖侧链的存在会大大促进这种抑制作用。
RG-I果胶没有直接抑制流感病毒神经氨酸酶的活性，可能是通过其它途径达到抗流感病毒效果。选择荷花蜂花粉RG-I级分WNPP-2-RG为代表，研究结果表明，它能够显著促进免疫指标，免疫调节作用可能是其发挥抗流感病毒活性的途径之一。对WNPP-2-RG的精细结构进行了鉴定。它以RG-I结构域为核心，AG-I和AG-II为侧链，分子量为3.8×10~5Da。侧链中1,6-Gal构成骨架结构，末端主要为Gal、少量Glc和Ara。为了进一步研究果胶结构与活性的关系，对WNPP-2-RG结构与免疫活性的关系进行了分析。通过部分酸水解除去侧链中的Ara，剩余核心结构为RG-I和AG-II结构域。体外免疫活性结果表明，侧链中的Ara的除去和Gal的暴露，对淋巴细胞增殖作用影响不大，但能够显著促进巨噬细胞的吞噬作用，降低NO释放能力。将WNPP-2-RG与人参RG-I型果胶（WGPA-2-RG）的结构和活性进行了对比研究。结果表明，侧链的类型、糖残基连接方式、甲酯化度、乙酰化度和分子量等不同，对活性的影响也不同。
Bee pollen is the male gametophyte of gymnosperm and angiosperm, which containsmost of the nutrients afford to plant growth and development. It is well known as a naturalnutrition and healthy food especially in improving human’s immunity. Bee pollen had manyactivities, such as antitumor, antivirus and immunobiological activities. Polysaccharides arethe main active components in the bee pollen. In this paper, we analyzed the structure ofpectic polysaccharides from six types of bee pollen and their antivirus activity, including thestructure comparision of type I rhamnogalacturonan (RG-I) and homogalacturonan (HG); therelationship of structure and inhibitory activity on H1N1and H5N1neuraminidase; the fluvirus inhibitory mechanism of RG-I pectin; the fine structure of RG-I pectin from bee pollenof Nelumbo nucifera (WNPP-2-RG) and its structure-activity relationship. The results of thispaper can offer some datas about structure-activity relationship of pectins, improved theapplication of pectin.
In this paper, we choosed six kinds of bee pollens, including Nelumbo nucifera,Papaver rhoeas, Rosa rugosa, Fagopyrum esculentum, Schisandra chinensis and Brassicacampestris, for studying the structure features and structure-activity relationship of pectins.Six RG-I and six HG pectins were obtained by a combination of DEAE-Cellulose andSepharose CL-6B colume. Structure analysis showed six RG-I pectins from bee pollencontained RG-I core region, both type I arabinogalactan (AG-I) and type II arabinogalactan(AG-II) sidechains except RG-I pectin from Rosa Rugosa which only contained AG-Isidechains; the ratio of rhamnose (Rha) and galacturonic acid (GalA) was nearly1:1,contained little HG region; RG-I mainchain contents was about16%~50%; total contents ofgalactose (Gal) and arabinose (Ara) were among70%~75%; the RG-I pectin from Brassicacampestris was contained glucan and xylan sidechains; the molecular weight of RG-Ifractions is about3.8×10~5Da. HG pectins from six kinds of bee pollen contains largeamounts of GalA; HG and RG-I region composed the core structure, ratio of Ara and Galwas nearly1:1; HG pectin from Nelumbo nucifera contained both AG-I and AG-IIsidechains, HG pectin from Papaver rhoeas and Rosa rugosa contained1,5-arabinan and1,3-galactan sidechains, HG pectin from Schisandra chinensis contained1,5-arabinan and1,4-galactan sidechains, HG pectin from Fagopyrum esculentum contained1,4-xylansidechains; HG pectin Papaver rhoeas and Brassica campestris contained glucose (Glc,about20%); the molecular weight was among8×103~3.6×10~4Da.
The inhibitory activity of RG-I and HG fractions on H1N1and H5N1neuraminidaseswas studied by bioluminescent assay and typical fluorescent method. The results indicatedHG fractions had better inhibition activity than RG-I fractions, especially the HG fracionsfrom Papaver rhoeas and Brassica campestris, IC50were below1mg/mL. According to thestructure features of each fraction, we can concluded HG region was the important region oninhibitory activity of H1N1and H5N1neuraminidases, the Glc sidechains can improve thisactivity.
The immunological activity of RG-I pectin (WNPP-2-RG) from bee pollen of Nelumbonucifera was analysed, it can improve the immunological activities, suggested RG-I mighthave antivirus activity through immunological way. Structure analysis showed WNPP-2-RGhas an RG-I core region, both AG-I and AG-II sidechains.1,6-Gal composed of thebackbone of sidechains, branched at O-3or O-4. Most of the Gal residues were determinedto be close to the molecular core, some Gal and Ara on the surface of the molecules. Theimmunological activitiy of WNPP-2-RG was related to its side chains. Ara residues affectedits macrophage phagocytosis and NO production, but no effect on lymphocyte proliferation.The comparation of structure-immunological activity between WNPP-2-RG and ginsengRG-I fraction WGPA-2-RG was performed. Ara residues in different RG-I pectins haddifferent effects on their activities.
引文
[1] Paulsen B S, Barsett H. Bioactive pectic polysaccharides[J]. Advances in Polymer Science,2005,186:69-101.
[2]郭振楚.糖类化学[M].北京:化学工业出版社,2005,80-87.
[3] Cosgrove D J. Assembly and enlargement of the primary cell wall in plants[J]. Annu Rev Cell DevBiol,1997,13:171-201.
[4] Ridley B L, O'Neill M A, Mohnen D. Pectins: structure, biosynthesis, and oligogalacturonide-relatedsignaling[J]. Phytochemistry,2001,57:929-967.
[5] Schols H A, Voragen A G J. Complex pectins: Structure elucidation using enzymes[J]. Progress inBiotechnology,1996,14:3-19.
[6] Prade R A, Zhan D, Ayoubi P, et al. Pectins, pectinases and plant-microbe interactions[J]. BiotechnolGenet Eng Rev,1999,16:361-391.
[7] McNeil M, Darvill A G, Albersheim P. Structure of Plant Cell Walls: X. Rhamnogalacturonan I, astructurally complex pectic polysaccharide in the walls of suspension-cultured sycamore cells[J].Plant Physiol,1980,66(6):1128-1134.
[8] Sengkhamparn N, Bakx E, Verhoef R, et al. Okra pectin contains an unusual substitution of itsrhamnosyl residues with acetyl and alpha-linked galactosyl groups[J]. Carbohydrate Research,2009,344:1842-1851.
[9] Renard C M G C, Crepeau M J, Thibault J F. Glucuronic acid directly linked to galacturonic acid inthe rhamnogalacturonan backbone of beet pectins[J]. European journal of Biochemistry,1999,266:566-574.
[10] Cartmell A, McKee L S, Pena M J, et al. The structure and function of an arabinan-specificalpha-1,2-arabinofuranosidase identified from screening the activities of bacterial GH43glycosidehydrolases[J]. J Biol Chem,2011,286:15483-15495.
[11] Zykwinska A, Thibault J F, Ralet M C. Organization of pectic arabinan and galactan side chains inassociation with cellulose microfibrils in primary cell walls and related models envisaged[J]. J ExpBot,2007,58:1795-1802.
[12] Ramon D, Veen P, Visser J. Arabinan degrading enzymes from Aspergillus nidulans: induction andpurification[J]. FEMS Microbiol Lett,1993,113:15-22.
[13] Thakur B R, Singh R K, Handa A K. Chemistry and uses of pectin--a review[J]. Crit Rev Food SciNutr,1997,37:47-73.
[14] Beda M Y. Pectic substances: From simple pectic polysaccharides to complex pectins-A newhypothetical model[J]. Carbohydrate polymers,2011,86:373-385.
[15]高小荣,刘培勋.多糖构效关系研究进展[J].中草药,2004,35:229-231.
[16] Koyanagi S, Tanigawa N, Nakagawa H. Oversulfation of fucoidan enhances its anti-angiogenis andanti-tumor activities[J]. Biochemical Pharmacology,2003,65:173-179.
[17] Shi B J, Nie X H, Xu H Y, et al. Preparation of Grifola frondosa polysaccharide sulfate and itsantitumor activity[J]. Chinese Journal of Pharmaceuticals,2003,34(8):383-385.
[18] Kolender A A, Matulewicz M C. Sulfated polysaccharides from the red sea-weed Geogiellaconfluens [J]. Carbohydrate Research,2002,337(1):57-68.
[19]田庚元.天然多糖的研究和应用（上）[J].上海化工,2000,10:29-31.
[20] Huang F, Meng Y W. Studies on polysaccharies with biological activity[J]. Nat Prod Res Dev,1999,11(5):90-98.
[21] Liang Z Y, Miao C Y, Zhang Y S. Influence of chemical modified structures on antitumor activity ofpolysaccharides from Clitopilus caespitosus[J]. Chin P harm J,1996,31(10):613-615.
[22] Huang C H, Li Y, Wang X F. Studies on extraction and analysis of pachyman and its structure[J].Guangzhou Food Sci Tech,2001,17(3):13-15.
[23] Fang J N, Wang S C. Review on advancement of polysaccharide from lentinula edodes [J]. ChinPharm J,1997,32(6):431-433.
[24] Schepetkin I A, Quinn M T. Botanical polysaccharides: Macrophage immunomodulation andtherapeutic potential [J]. Int Immunopharmacol,2006,6(3):317-333.
[25] Li S, Wu Q H, Chen C, et al. Recent research progress on anti-tumor activity of polysaccharides [J].Chinese Journal of Biochemical Pharmaceutics,2007,28(3):213-216.
[26]张旭.人参多糖的系统分析及其免疫活性研究[D]:[博士学位论文].长春:东北师范大学,2009.
[27]倪维华.人参多糖免疫活性及抗肿瘤作用[D]:[博士学位论文].长春:东北师范大学,2010.
[28]范玉莹.人参果胶对细胞迁移的影响及其机理研究[D]:[博士学位论文].长春:东北师范大学,2010.
[29] Kulicake W M. Correlation between immunological activity, molar mass, and molecular structure ofdifferent (1→3)-β-D-glucans [J]. Carbohydr Res,1997,297:135-143.
[30] Matsuoka H. Lentinan potentiates immunity and prolongs the survival time of some patients [J].Anticancer Res,1997,17(4A):2751-2755.
[31] Zhang P, Zhang L, Cheng S. Effect of urea and sodium hydroxide on the molecular weight andconformation of β-(1,3)-D-glucan from letinus edodes in aqueous solution [J]. Carbohydr Res,2000,327(2):431-438.
[32] Young S H, Jacobs R R. Sodiun hydroxide-induced conformational change in schizophylan detectedby the fluorescencedye, aniline blue [J]. Carbohydr Res,1998,310(1):91-99.
[33] Kazuo, Yumi, Yideaki, et al. Anti-tumor activity of an enzymatically synthesized α-1,6branched-1,4-Glucan glycogen [J]. Biosci,2004,68(11):2332-2340.
[34] Lever R, Lo W T, Faraidoun M, et al. Size-fractionated heparins have differential effects on humanneutrophil function in vitro [J]. British Journal of Pharmacology,2007,151:837-843.
[35] Walker A, Turnbull J E, Gallapher J T, et al. Specific heparan sulfate saccharides mediate the activityof basic fibroblast growth factor [J]. The Journal of Biological Chemistry,1994,269(2):931-935.
[36]毕洪涛.胶陀螺多糖的系统分析及生物学活性研究[D]:[博士学位论文].长春:东北师范大学,2010.
[37] Zhang Y F, Wu F L. Study on the anti-tumor and anti-oxidative of astragalus in mouse [J]. ChineseJournal of Public Health.1997,16(4):229-230.
[38] Zhu J W, Xie P. Study on effects of saccharum germanium of glossy ganoderma on immunity andantitumor [J]. Chinese Journal of Biochemical Pharmaceutics,2000,21(4):189-190.
[39] Wang S C, Bligh S W A, Shi S S, et al. Structural features and anti-HIV-1activity of novelpolysaccharides from red algae Grateloupia longifolia and Grateloupia filicina [J]. InternationalJournal of Biological Macromelecules,2007,41:369-375.
[40] Tang F X, Chen F, Li F. Preparation and potential in vivo anti-influenza virus activity of lowmolecular-weight κ-carrageenans and their derivatives [J]. Journal of Applied Polymer Science.2013,127(3):2110-2115.
[41] Chen C Y, Huang X H, Zhou J Y, et al. Sulfation of polysaccharides isolated from Indocalamustesselatus and their anticytopathic effect on human immunodeficiency virus type I [J]. Yao Xue XueBao.1998,33(4):264-268.
[42] Yoshida O, Nakashima H, Yoshida T, et al. Sulfation of the immunomodulating polysaccharidelentinan: a novel strategy for antivirals to human immunodeficiency virus (HIV)[J]. Biochem Pharm,1998,37(7):2887-2889.
[43] Wang C Y, Guan H S. Advances of researches on antiviral activities of polysaccharides Ⅱ. Antiviralactivities of sulfated polysaccharides [J]. China Biotechnology,2000,20(2):3-8.
[44]杨威.流感病毒神经氨酸酶的检测及人参多糖对其抑制作用的研究[D]:[博士学位论文].长春:东北师范大学,2012.
[45] Chen X M, Pan J Q, Zhang Z N. The structures and biology activities of the low-molecular heparin[J]. Chinese Pharmacological Bulletin,1992,8(2):86-90.
[46] Mulloy B, Mourao P A S, Gray E. Structure/function studies of anticoagulant sulphatedpolysaccharides using NMR [J]. J Biotechnol,2000,77(1):123-135.
[47] Du M, Zhang S. Mechanism of edible fungal polysaccharide on reducing blood sugar [J]. Journal ofmicrobiology,2007,27(2):83-87.
[48]王元凤.茶多糖的分离纯化、结构及构效关系研究[D]:[博士学位论文].无锡:江南大学,2005.
[49]陈艳.人参水提物降血糖作用的研究[D]:[博士学位论文].长春:东北师范大学,2010.
[50] Ou-yang J M, Hu P. Chemical bases and research progress on inhibition of urinary stone by sulfatedpolysaccharide from seaweeds [J]. China Pharmaceutica,2005,14(10):21-23.
[51]邱琳,辛现良,耿美玉.多糖构效关系研究进展[J].现代生物医学进展,2009,9(9):1764-1768.
[52]王佳.人参多糖抗疲劳和抗抑郁作用及其机制的研究[D]:[博士学位论文].长春:东北师范大学,2010.
[53] Mizuno M, Minato K, Ito H, et al. Anti-tumor polysaccharide from the mycelium of liquid-culturedAgaricus blazei mill[J]. Biochem Mol Biol Int,1999,47:707-714.
[54] Sun Y X. Immunological adjuvant effect of a water-soluble polysaccharide, CPP, from the roots ofCodonopsis pilosula on the immune responses to ovalbumin in mice[J]. Chem Biodivers,2009,6:890-896.
[55] Zhao Y, Son Y O, Kim S S, et al. Antioxidant and anti-hyperglycemic activity of polysaccharideisolated from Dendrobium chrysotoxum Lindl[J]. J Biochem Mol Biol,2007,40:670-677.
[56] Wang J, Li S, Fan Y, et al. Anti-fatigue activity of the water-soluble polysaccharides isolated fromPanax ginseng C. A. Meyer[J]. J Ethnopharmacol,2010,130:421-423.
[57] Brecker L, Wicklein D, Moll H, et al. Structural and immunological properties of arabinogalactanpolysaccharides from pollen of timothy grass (Phleum pratense L.)[J]. Carbohydr Res,2005,340:657-663.
[58] Liu H G, Liang Q Y, Meng H L, et al. Hypoglycemic effect of extracts of cactus pear fruitpolysaccharide in rats[J]. Zhong Yao Cai,2010,33:240-242.
[59] Sun C, Rosendahl A H, Wang X D, et al. Polysaccharide-K (PSK) in Cancer-Old Story, NewPossibilities?[J]. Curr Med Chem,2012,19:757-762.
[60] Layton R C, Petrovsky N, Gigliotti A P, et al. Delta inulin polysaccharide adjuvant enhances theability of split-virion H5N1vaccine to protect against lethal challenge in ferrets[J]. Vaccine,2011,29:6242-6251.
[61] Han X Q, Chai X Y, Jia Y M, et al. Structure elucidation and immunological activity of a novelpolysaccharide from the fruit bodies of an edible mushroom, Sarcodon aspratus (Berk.) S. Ito[J]. IntJ Biol Macromol,2010,47:420-424.
[62]李晓东,李娟,杨丽霞,等.中药植物多糖降血糖作用的研究进展[J].甘肃中医,2010,11:77-80.
[63]曾先知.浓缩的营养库――蜂花粉[J].食品与生活,2011,1:33.
[64]张秋蓉,周浓,张筱岚.花粉的营养及保健功能[J].亚太传统医药,2010,07:148-150
[65]蒋枫,朱威,张颖,等.莲花花粉营养成分分析[J].中国蜂业,2007,9:5-7.
[66] Xie Y, Wan B, Li W. Effect of bee pollen on maternal nutrition and fetal growth[J]. Hua Xi Yi KeDa Xue Xue Bao,1994,25:434-437.
[67]傅作申,李树凯,黄士领,等.玉米花粉营养成分的分析[J].临床军医杂志.2000.28:2.
[68] Human H, Nicolson S W. Nutritional content of fresh, bee-collected and stored pollen of Aloegreatheadii var. davyana (Asphodelaceae)[J]. Phytochemistry,2006,67:1486-1492.
[69] Yang X, Guo D, Zhang J, et al. Characterization and antitumor activity of pollen polysaccharide[J].Int Immunopharmacol,2007,7:427-434.
[70] Li F, Yuan Q, Rashid F. Isolation, purification and immunobiological activity of a newwater-soluble bee pollen polysaccharide from Crataegus pinnatifida Bge[J]. Carbohydrate Polymers,2009,78:80-88.
[71] Wang B, Diao Q Y, Zhang Z Y, et al. Antitumor activity of bee pollen polysaccharides from RosaRugosa. Molecular Medicine Reports,2013,7:1555-1558.
[72]耿越,王开发,张玉兰.玉米花粉多糖PMA-I的结构解析[J].天然产物研究与开发,2006,2:263-265.
[73]杨晓萍,罗祖友,吴谋成.油菜花粉多糖的制备及其对荷瘤小鼠的影响[J].食品科学,2005,12:202-204.
[74]杨新跃,刘志勇,汪礼国,等.蜂花粉多糖液抑制肿瘤作用的实验研究[J].江西农业大学学报,2006,2:293-294.
[75]王开发,邹朝中,陆明.玉米花粉多糖抑制肿瘤的作用效应研究[J].蜜蜂杂志,2001,2:3-4.
[76]刘伟,付中民,杨文超,等.山茶蜂花粉多糖对四氧嘧啶致小鼠糖尿病的降血糖作用研究[J].蜜蜂杂志,2009,2:4-6.
[77]曾志将,汪礼国,饶波,等.蜂花粉多糖对大鼠降血脂效果研究[J].江西农业大学学报,2004,3:406-408.
[78]屈洪岩,郭玉璞,陆钢,等.玉米花粉多糖对鸡新城疫弱毒疫苗免疫应答的影响[J].中国兽医杂志,1998,1:37-39.
[79]胡积清,程良新,方凯,等.蜂花粉的营养特性及在畜禽业中应用前景[J].安徽农学通报,2010,9:196-197.
[80]张国锋,刁其玉,屠焰,等.蜂花粉及其多糖对犊牛生长性能及血液生化指标的影响[J].中国畜牧杂志,2010,13:52-56.
[1]蒋枫,朱威,张颖,等.莲花花粉营养成分分析[J].中国蜂业,2007,9:5-7.
[2] Paulsen B S, Barsett H. Bioactive pectic polysaccharides[J]. Advances in Polymer Science,2005,186:69-101.
[3]方积年,丁侃.天然药物—多糖的主要生物活性及分离纯化方法[J].中国天然药物,2007,5:338-347.
[4]张旭.人参多糖的系统分析及其免疫活性研究[D]:[博士学位论文].长春:东北师范大学,2009.
[5] Zhang X, Yu L, Bi H T, et al. Total fractionation and characterization of the water-solublepolysaccharides isolated from Panax ginseng C. A. Meyer[J]. Carbohydrate Polymers,2009,77:544-552.
[6] Yu L, Zhang X, Li S S, et al. Rhamnogalacturonan I domains from ginseng pectin[J]. CarbohydratePolymers,2010,79:811-817.
[7] Chambers R E, Clamp J R. An assessment of methanolysis and other factors used in the analysis ofcarbohydrate-containing materials[J]. Biochem J,1971,125:1009-1018.
[8] Guttman A. Analysis of monosaccharide composition by capillary electrophoresis[J]. J ChromatogrA,1997,763:271-277.
[9] Fu D, O'Neill R A. Monosaccharide composition analysis of oligosaccharides and glycoproteins byhigh-performance liquid chromatography[J]. Anal Biochem,1995,227:377-384.
[10] Dubois M, Gilles K, Hamilton J K, et al. A colorimetric method for the determination of sugars[J].Nature,1951,168:167.
[11] Blumenkrantz N, Asboe-Hansen G. New method for quantitative determination of uronic acids[J].Anal Biochem,1973,54:484-489.
[12] Bennett J, Scott K J. Quantitative staining of fraction I protein in polyacrylamide gels usingCoomassie brillant blue[J]. Anal Biochem,1971,43:173-182.
[13] Paulsen B S, Barsett H. Bioactive pectic polysaccharides[J]. Advances in Polymer Science,2005,186:69-101.
[14] Ridley B L, O'Neill M A, Mohnen D. Pectins: structure, biosynthesis, andoligogalacturonide-related signaling[J]. Phytochemistry,2001,57:929-967.
[15] Gronhaug T E, Ghildyal P, Barsett H, et al. Bioactive arabinogalactans from the leaves of Opiliaceltidifolia Endl. ex Walp.(Opiliaceae)[J]. Glycobiology,2010,20:1654-1664.
[16] Alves A, Caridade S G, Mano J F, et al. Extraction and physico-chemical characterization of aversatile biodegradable polysaccharide obtained from green algae[J]. Carbohydr Res,2010,345:2194-2200.
[1] Prade R A, Zhan D, Ayoubi P, et al. Pectins, pectinases and plant-microbe interactions[J]. BiotechnolGenet Eng Rev,1999,16:361-391.
[2] Paulsen B S, Barsett H. Bioactive pectic polysaccharides[J]. Advances in Polymer Science,2005,186:69-101.
[3]张旭.人参多糖的系统分析及其免疫活性研究[D]:[博士学位论文].长春:东北师范大学,2009.
[4] Needs P W, Selvendran R R. Avoiding oxidative degradation during sodium hydroxide/methyliodide-mediated carbohydrate methylation in dimethyl sulfoxide[J]. Carbohydrate Research,1993,245:1-10.
[5] McNeil M, Darvill A G, Albersheim P. Structure of Plant Cell Walls: X. Rhamnogalacturonan I, astructurally complex pectic polysaccharide in the walls of suspension-cultured sycamore cells[J].Plant Physiol,1980,66(6):1128-1134.
[6] Ridley B L, O'Neill M A, Mohnen D. Pectins: structure, biosynthesis, and oligogalacturonide-relatedsignaling[J]. Phytochemistry,2001,57:929-967.
[7] Raju T S, Davidson E A. Structural features of water-soluble novel polysaccharide components fromthe leaves of Tridax procumbens Linn[J]. Carbohydr Res,1994,258:243-254.
[8] Schols H A, Voragen A G J. Complex pectins: Structure elucidation using enzymes[J]. Progress inBiotechnology,1996,14:3-19.
[9] Dong Q, Liu X, Yao J, et al. Structural characterization of a pectic polysaccharide from Neriumindicum flowers[J]. Phytochemistry,2010,71:1430-1437.
[10] Duan J, Wang X, Dong Q, et al. Structural features of a pectic arabinogalactan with immunologicalactivity from the leaves of Diospyros kaki[J]. Carbohydr Res,2003,338:1291-1297.
[11] Yu L, Zhang X, Li S S, et al. Rhamnogalacturonan I domains from ginseng pectin. CarbohydratePolymers,2010,79(4):811-817.
[12] Zhang X, Li S S, Sun L, et al. Further analysis of the structure and immunological activity of an RG-Itype pectin from Panax ginseng. Carbohydrate Polymers,2012,89(2):519-525.
[13] Zhang X, Yu L, Bi H T, et al. Total fractionation and characterization of the water-solublepolysaccharides isolated from Panax ginseng C. A. Meyer. Carbohydrate Polymers,2009,77:544-552.
[14] Prade R A, Zhan D, Ayoubi P, et al. Pectins, pectinases and plant-microbe interactions[J].Biotechnol Genet Eng Rev,1999,16:361-391.
[1] Paulsen B S, Barsett H. Bioactive pectic polysaccharides[J]. Advances in Polymer Science,2005,186:69-101.
[2] Hoshino T, Suzuki Y, Takeichi M, et al. Concomitant intraperitoneal therapy with the antitumorpolysaccharide Sizofiran and rG-CSF for ovarian cancer[J]. Gan To Kagaku Ryoho,1993,20:1638-1641.
[3] Ohwada S, Kawate S, Ikeya T, et al. Adjuvant therapy with protein-bound polysaccharide K andtegafur uracil in patients with stage II or III colorectal cancer: randomized, controlled trial[J]. DisColon Rectum,2003,46:1060-1068.
[4] Hwang I, Ahn G, Park E, et al. An acidic polysaccharide of Panax ginseng ameliorates experimentalautoimmune encephalomyelitis and induces regulatory T cells[J]. Immunol Lett,2011,138:169-178.
[5] Subramanian M, Chintalwar G J, Chattopadhyay S. Antioxidant and radioprotective properties of anOcimum sanctum polysaccharide[J]. Redox Rep,2005,10:257-264.
[6] Qi H, Zhang Q, Zhao T, et al. Antioxidant activity of different sulfate content derivatives ofpolysaccharide extracted from Ulva pertusa (Chlorophyta) in vitro[J]. Int J Biol Macromol,2005,37:195-199.
[7] Xu W, Zhang F, Luo Y, et al. Antioxidant activity of a water-soluble polysaccharide purified fromPteridium aquilinum[J]. Carbohydr Res,2009,344:217-222.
[8]方积年,丁侃.天然药物—多糖的主要生物活性及分离纯化方法[J].中国天然药物,2007,5:338-347.
[9]张旭.人参多糖的系统分析及其免疫活性研究[D]:[博士学位论文].长春:东北师范大学,2009.
[10] Clark C C. Collaborative study of an on-column periodate reaction method for the analysis ofphenylpropanolamine hydrochloride in elixirs[J]. J Assoc Off Anal Chem,1973,56:100-104.
[11] Politis D J, Goodman R N. Fine structure of extracellular polysaccharide of erwinia amylovora[J].Appl Environ Microbiol,1980,40:596-607.
[12] Shingel K I. Current knowledge on biosynthesis, biological activity, and chemical modification ofthe exopolysaccharide, pullulan[J]. Carbohydr Res,2004,339:447-460.
[13] Zhang X, Li S S, Sun L, et al. Further analysis of the structure and immunological activity of anRG-I type pectin from Panax ginseng. Carbohydrate Polymers,2012,89:519-525.
[14] Blumenkrantz N, Asboe-Hansen G. New method for quantitative determination of uronic acids[J].Anal Biochem,1973,54:484-489.
[15] Ni W H, Zhang X, Wang B, et al. Antitumor activities and immunomodulatory effects of ginsengneutral polysaccharides in combination with5-Fluorouracil. Journal of Medicinal Food,2010,13(2):1–8.
[16] Nair P K, Melnick S J, Ramachandran R, et al. Mechanism of macrophage activation by(1,4)-alpha-D-glucan isolated from Tinospora cordifolia[J]. Int Immunopharmacol,2006,6:1815-1824.
[17] Wakshull E, Brunke-Reese D, Lindermuth J, et al. PGG-glucan, a soluble beta-(1,3)-glucan,enhances the oxidative burst response, microbicidal activity, and activates an NF-kappa B-like factorin human PMN: evidence for a glycosphingolipid beta-(1,3)-glucan receptor[J].Immunopharmacology,1999,41:89-107.
[18] Ni W H, Zhang X, Bi H T, et al. Preparation of a glucan from the roots of Rubus crataegifolius Bge.and its immunological activity. Carbohydr Res,2009,344:2512-2518.
[19] Sykes B W, Furr M, Giguere S. In vivo pretreatment with PGG-glucan fails to alter cytokine mRNAexpression of equine peripheral blood mononuclear cells exposed to endotoxin ex vivo[J]. Vet Ther,2005,6:67-76.
[20] Bellon T, Martinez V, Lucendo B, et al. Alternative activation of macrophages in human peritoneum:implications for peritoneal fibrosis[J]. Nephrol Dial Transplant,2011,26:2995-3005.
[21] Bredt D S, Snyder S H. Nitric oxide, a novel neuronal messenger[J]. Neuron,1992,8:3-11.
[22] Gan L, Zhang H S, Yang L X, et al. Immunomodulation and antitumor activity by apolysaccharide-protein complex from Lycium barbarum[J]. Int Immunopharmacol,2004,4:563-569.
[23] Prade R A, Zhan D, Ayoubi P, et al. Pectins, pectinases and plant-microbe interactions[J].Biotechnol Genet Eng Rev.1999.16:361-391.
[24]毕洪涛.胶陀螺多糖的系统分析及生物学活性研究[D]:[博士学位论文].长春:东北师范大学,2010.
[25] Schols H A, Voragen A G J. Complex pectins: Structure elucidation using enzymes[J]. Progress inBiotechnology,1996,14:3-19.
[26] Dong Q, Liu X, Yao J, et al. Structural characterization of a pectic polysaccharide from Neriumindicum flowers[J]. Phytochemistry,2010,71:1430-1437.
[27] Brecker L, Wicklein D, Moll H, et al. Structural and immunological properties of arabinogalactanpolysaccharides from pollen of timothy grass (Phleum pratense L.)[J]. Carbohydr Res,2005,340:657-663.
[28] Tanaka T. Mechanism of antitumor action of polysaccharide fraction prepared from bagasse. II.Immunological reactivity of mice and properties of their sera[J]. Gann,1967,58:451-457.
[29] Gordon S. Alternative activation of macrophages[J]. Nat Rev Immunol,2003,3:23-35.
[30] Gordon S, Martinez F O. Alternative activation of macrophages: mechanism and functions[J].
Immunity,2010,32:593-604.
[1] Salom D, Hill B R, Lear J D, et al. pH–Dependent tetrmerization and amantadine binding of thetransmembrane helix of M from the influenza a virus[J]. Biochemistry,2000,39(46):14160-14170.
[2] Bright R A, Medina M J, Xu X, et al. Incidence of adamantine resistance among influenza A (H3N2)viruses isolated worldwide from1994to2005: a cause for concern[J]. Lancet,2005,366(9492):1175-81.
[3] Moscona A. Medical management of influenza infection[J]. Annu Rev Med,2008,59:397-413.
[4] Hayden F G. Perspectives on antiviral use during pandemic influenza[J]. Philos Traps R Soc Lond BBiol Sci,2001,356(1416):1877-1884.
[5] White M R, Crouch E, Hartshorn M, et al. Cooperative anti-influenza activities of respiratory innateimmune proteins and neuraminidase inhibitor[J]. Am J Physiol-Lung C,2005,288(5):831-840.
[6] Govorkova E A, Fang H B, Tan M, et al. Neuraminidase inhibitor-rimantadine combinations exertadditive and synergistic anti-influenza virus effects in MDCK cells[J]. Antimicrob Agents CH,2004,48(12):4855-4863.
[7] Lindegardh N, Hien T T, Farrar J, et al. A simple and rapid liquid hromatographic assay forevaluation of potentially counterfeit Tamiflu[J]. J Pharmaceut Biomed,2006,42(4):430-433.
[8]杨威.流感病毒神经氨酸酶的检测及人参多糖对其抑制作用的研究[D]:[博士学位论文].长春:东北师范大学,2012.
[9] Potier M, Mameli L, Belisle M, et al. Fluorometric assay of neuraminidase with a sodium(4-methylumbelliferyl-(-D-N-acetylneuraminate)substrate[J]. Anal Biochem,1979,94:287-296.
[10] Ortiz A R, Pisabarro M T, Gago F, et al. Prediction of drug binding affinities by comparativebinding energy analysis[J]. J Med Chem,1995,38:2681-2691.
[11] Escuret V, Frobert E, Bouscambert-Duchamp M, et al. Detection of human influenza A(H1N1) andB strains with reduced sensitivity to neuraminidase inhibitors[J]. J Clin Virol,2008,41:25-28.
[2]郭振楚.糖类化学[M].北京:化学工业出版社,2005,80-87.
[3] Cosgrove D J. Assembly and enlargement of the primary cell wall in plants[J]. Annu Rev Cell DevBiol,1997,13:171-201.
[4] Ridley B L, O'Neill M A, Mohnen D. Pectins: structure, biosynthesis, and oligogalacturonide-relatedsignaling[J]. Phytochemistry,2001,57:929-967.
[5] Schols H A, Voragen A G J. Complex pectins: Structure elucidation using enzymes[J]. Progress inBiotechnology,1996,14:3-19.
[6] Prade R A, Zhan D, Ayoubi P, et al. Pectins, pectinases and plant-microbe interactions[J]. BiotechnolGenet Eng Rev,1999,16:361-391.
[7] McNeil M, Darvill A G, Albersheim P. Structure of Plant Cell Walls: X. Rhamnogalacturonan I, astructurally complex pectic polysaccharide in the walls of suspension-cultured sycamore cells[J].Plant Physiol,1980,66(6):1128-1134.
[8] Sengkhamparn N, Bakx E, Verhoef R, et al. Okra pectin contains an unusual substitution of itsrhamnosyl residues with acetyl and alpha-linked galactosyl groups[J]. Carbohydrate Research,2009,344:1842-1851.
[9] Renard C M G C, Crepeau M J, Thibault J F. Glucuronic acid directly linked to galacturonic acid inthe rhamnogalacturonan backbone of beet pectins[J]. European journal of Biochemistry,1999,266:566-574.
[10] Cartmell A, McKee L S, Pena M J, et al. The structure and function of an arabinan-specificalpha-1,2-arabinofuranosidase identified from screening the activities of bacterial GH43glycosidehydrolases[J]. J Biol Chem,2011,286:15483-15495.
[11] Zykwinska A, Thibault J F, Ralet M C. Organization of pectic arabinan and galactan side chains inassociation with cellulose microfibrils in primary cell walls and related models envisaged[J]. J ExpBot,2007,58:1795-1802.
[12] Ramon D, Veen P, Visser J. Arabinan degrading enzymes from Aspergillus nidulans: induction andpurification[J]. FEMS Microbiol Lett,1993,113:15-22.
[13] Thakur B R, Singh R K, Handa A K. Chemistry and uses of pectin--a review[J]. Crit Rev Food SciNutr,1997,37:47-73.
[14] Beda M Y. Pectic substances: From simple pectic polysaccharides to complex pectins-A newhypothetical model[J]. Carbohydrate polymers,2011,86:373-385.
[15]高小荣,刘培勋.多糖构效关系研究进展[J].中草药,2004,35:229-231.
[16] Koyanagi S, Tanigawa N, Nakagawa H. Oversulfation of fucoidan enhances its anti-angiogenis andanti-tumor activities[J]. Biochemical Pharmacology,2003,65:173-179.
[17] Shi B J, Nie X H, Xu H Y, et al. Preparation of Grifola frondosa polysaccharide sulfate and itsantitumor activity[J]. Chinese Journal of Pharmaceuticals,2003,34(8):383-385.
[18] Kolender A A, Matulewicz M C. Sulfated polysaccharides from the red sea-weed Geogiellaconfluens [J]. Carbohydrate Research,2002,337(1):57-68.
[19]田庚元.天然多糖的研究和应用（上）[J].上海化工,2000,10:29-31.
[20] Huang F, Meng Y W. Studies on polysaccharies with biological activity[J]. Nat Prod Res Dev,1999,11(5):90-98.
[21] Liang Z Y, Miao C Y, Zhang Y S. Influence of chemical modified structures on antitumor activity ofpolysaccharides from Clitopilus caespitosus[J]. Chin P harm J,1996,31(10):613-615.
[22] Huang C H, Li Y, Wang X F. Studies on extraction and analysis of pachyman and its structure[J].Guangzhou Food Sci Tech,2001,17(3):13-15.
[23] Fang J N, Wang S C. Review on advancement of polysaccharide from lentinula edodes [J]. ChinPharm J,1997,32(6):431-433.
[24] Schepetkin I A, Quinn M T. Botanical polysaccharides: Macrophage immunomodulation andtherapeutic potential [J]. Int Immunopharmacol,2006,6(3):317-333.
[25] Li S, Wu Q H, Chen C, et al. Recent research progress on anti-tumor activity of polysaccharides [J].Chinese Journal of Biochemical Pharmaceutics,2007,28(3):213-216.
[26]张旭.人参多糖的系统分析及其免疫活性研究[D]:[博士学位论文].长春:东北师范大学,2009.
[27]倪维华.人参多糖免疫活性及抗肿瘤作用[D]:[博士学位论文].长春:东北师范大学,2010.
[28]范玉莹.人参果胶对细胞迁移的影响及其机理研究[D]:[博士学位论文].长春:东北师范大学,2010.
[29] Kulicake W M. Correlation between immunological activity, molar mass, and molecular structure ofdifferent (1→3)-β-D-glucans [J]. Carbohydr Res,1997,297:135-143.
[30] Matsuoka H. Lentinan potentiates immunity and prolongs the survival time of some patients [J].Anticancer Res,1997,17(4A):2751-2755.
[31] Zhang P, Zhang L, Cheng S. Effect of urea and sodium hydroxide on the molecular weight andconformation of β-(1,3)-D-glucan from letinus edodes in aqueous solution [J]. Carbohydr Res,2000,327(2):431-438.
[32] Young S H, Jacobs R R. Sodiun hydroxide-induced conformational change in schizophylan detectedby the fluorescencedye, aniline blue [J]. Carbohydr Res,1998,310(1):91-99.
[33] Kazuo, Yumi, Yideaki, et al. Anti-tumor activity of an enzymatically synthesized α-1,6branched-1,4-Glucan glycogen [J]. Biosci,2004,68(11):2332-2340.
[34] Lever R, Lo W T, Faraidoun M, et al. Size-fractionated heparins have differential effects on humanneutrophil function in vitro [J]. British Journal of Pharmacology,2007,151:837-843.
[35] Walker A, Turnbull J E, Gallapher J T, et al. Specific heparan sulfate saccharides mediate the activityof basic fibroblast growth factor [J]. The Journal of Biological Chemistry,1994,269(2):931-935.
[36]毕洪涛.胶陀螺多糖的系统分析及生物学活性研究[D]:[博士学位论文].长春:东北师范大学,2010.
[37] Zhang Y F, Wu F L. Study on the anti-tumor and anti-oxidative of astragalus in mouse [J]. ChineseJournal of Public Health.1997,16(4):229-230.
[38] Zhu J W, Xie P. Study on effects of saccharum germanium of glossy ganoderma on immunity andantitumor [J]. Chinese Journal of Biochemical Pharmaceutics,2000,21(4):189-190.
[39] Wang S C, Bligh S W A, Shi S S, et al. Structural features and anti-HIV-1activity of novelpolysaccharides from red algae Grateloupia longifolia and Grateloupia filicina [J]. InternationalJournal of Biological Macromelecules,2007,41:369-375.
[40] Tang F X, Chen F, Li F. Preparation and potential in vivo anti-influenza virus activity of lowmolecular-weight κ-carrageenans and their derivatives [J]. Journal of Applied Polymer Science.2013,127(3):2110-2115.
[41] Chen C Y, Huang X H, Zhou J Y, et al. Sulfation of polysaccharides isolated from Indocalamustesselatus and their anticytopathic effect on human immunodeficiency virus type I [J]. Yao Xue XueBao.1998,33(4):264-268.
[42] Yoshida O, Nakashima H, Yoshida T, et al. Sulfation of the immunomodulating polysaccharidelentinan: a novel strategy for antivirals to human immunodeficiency virus (HIV)[J]. Biochem Pharm,1998,37(7):2887-2889.
[43] Wang C Y, Guan H S. Advances of researches on antiviral activities of polysaccharides Ⅱ. Antiviralactivities of sulfated polysaccharides [J]. China Biotechnology,2000,20(2):3-8.
[44]杨威.流感病毒神经氨酸酶的检测及人参多糖对其抑制作用的研究[D]:[博士学位论文].长春:东北师范大学,2012.
[45] Chen X M, Pan J Q, Zhang Z N. The structures and biology activities of the low-molecular heparin[J]. Chinese Pharmacological Bulletin,1992,8(2):86-90.
[46] Mulloy B, Mourao P A S, Gray E. Structure/function studies of anticoagulant sulphatedpolysaccharides using NMR [J]. J Biotechnol,2000,77(1):123-135.
[47] Du M, Zhang S. Mechanism of edible fungal polysaccharide on reducing blood sugar [J]. Journal ofmicrobiology,2007,27(2):83-87.
[48]王元凤.茶多糖的分离纯化、结构及构效关系研究[D]:[博士学位论文].无锡:江南大学,2005.
[49]陈艳.人参水提物降血糖作用的研究[D]:[博士学位论文].长春:东北师范大学,2010.
[50] Ou-yang J M, Hu P. Chemical bases and research progress on inhibition of urinary stone by sulfatedpolysaccharide from seaweeds [J]. China Pharmaceutica,2005,14(10):21-23.
[51]邱琳,辛现良,耿美玉.多糖构效关系研究进展[J].现代生物医学进展,2009,9(9):1764-1768.
[52]王佳.人参多糖抗疲劳和抗抑郁作用及其机制的研究[D]:[博士学位论文].长春:东北师范大学,2010.
[53] Mizuno M, Minato K, Ito H, et al. Anti-tumor polysaccharide from the mycelium of liquid-culturedAgaricus blazei mill[J]. Biochem Mol Biol Int,1999,47:707-714.
[54] Sun Y X. Immunological adjuvant effect of a water-soluble polysaccharide, CPP, from the roots ofCodonopsis pilosula on the immune responses to ovalbumin in mice[J]. Chem Biodivers,2009,6:890-896.
[55] Zhao Y, Son Y O, Kim S S, et al. Antioxidant and anti-hyperglycemic activity of polysaccharideisolated from Dendrobium chrysotoxum Lindl[J]. J Biochem Mol Biol,2007,40:670-677.
[56] Wang J, Li S, Fan Y, et al. Anti-fatigue activity of the water-soluble polysaccharides isolated fromPanax ginseng C. A. Meyer[J]. J Ethnopharmacol,2010,130:421-423.
[57] Brecker L, Wicklein D, Moll H, et al. Structural and immunological properties of arabinogalactanpolysaccharides from pollen of timothy grass (Phleum pratense L.)[J]. Carbohydr Res,2005,340:657-663.
[58] Liu H G, Liang Q Y, Meng H L, et al. Hypoglycemic effect of extracts of cactus pear fruitpolysaccharide in rats[J]. Zhong Yao Cai,2010,33:240-242.
[59] Sun C, Rosendahl A H, Wang X D, et al. Polysaccharide-K (PSK) in Cancer-Old Story, NewPossibilities?[J]. Curr Med Chem,2012,19:757-762.
[60] Layton R C, Petrovsky N, Gigliotti A P, et al. Delta inulin polysaccharide adjuvant enhances theability of split-virion H5N1vaccine to protect against lethal challenge in ferrets[J]. Vaccine,2011,29:6242-6251.
[61] Han X Q, Chai X Y, Jia Y M, et al. Structure elucidation and immunological activity of a novelpolysaccharide from the fruit bodies of an edible mushroom, Sarcodon aspratus (Berk.) S. Ito[J]. IntJ Biol Macromol,2010,47:420-424.
[62]李晓东,李娟,杨丽霞,等.中药植物多糖降血糖作用的研究进展[J].甘肃中医,2010,11:77-80.
[63]曾先知.浓缩的营养库――蜂花粉[J].食品与生活,2011,1:33.
[64]张秋蓉,周浓,张筱岚.花粉的营养及保健功能[J].亚太传统医药,2010,07:148-150
[65]蒋枫,朱威,张颖,等.莲花花粉营养成分分析[J].中国蜂业,2007,9:5-7.
[66] Xie Y, Wan B, Li W. Effect of bee pollen on maternal nutrition and fetal growth[J]. Hua Xi Yi KeDa Xue Xue Bao,1994,25:434-437.
[67]傅作申,李树凯,黄士领,等.玉米花粉营养成分的分析[J].临床军医杂志.2000.28:2.
[68] Human H, Nicolson S W. Nutritional content of fresh, bee-collected and stored pollen of Aloegreatheadii var. davyana (Asphodelaceae)[J]. Phytochemistry,2006,67:1486-1492.
[69] Yang X, Guo D, Zhang J, et al. Characterization and antitumor activity of pollen polysaccharide[J].Int Immunopharmacol,2007,7:427-434.
[70] Li F, Yuan Q, Rashid F. Isolation, purification and immunobiological activity of a newwater-soluble bee pollen polysaccharide from Crataegus pinnatifida Bge[J]. Carbohydrate Polymers,2009,78:80-88.
[71] Wang B, Diao Q Y, Zhang Z Y, et al. Antitumor activity of bee pollen polysaccharides from RosaRugosa. Molecular Medicine Reports,2013,7:1555-1558.
[72]耿越,王开发,张玉兰.玉米花粉多糖PMA-I的结构解析[J].天然产物研究与开发,2006,2:263-265.
[73]杨晓萍,罗祖友,吴谋成.油菜花粉多糖的制备及其对荷瘤小鼠的影响[J].食品科学,2005,12:202-204.
[74]杨新跃,刘志勇,汪礼国,等.蜂花粉多糖液抑制肿瘤作用的实验研究[J].江西农业大学学报,2006,2:293-294.
[75]王开发,邹朝中,陆明.玉米花粉多糖抑制肿瘤的作用效应研究[J].蜜蜂杂志,2001,2:3-4.
[76]刘伟,付中民,杨文超,等.山茶蜂花粉多糖对四氧嘧啶致小鼠糖尿病的降血糖作用研究[J].蜜蜂杂志,2009,2:4-6.
[77]曾志将,汪礼国,饶波,等.蜂花粉多糖对大鼠降血脂效果研究[J].江西农业大学学报,2004,3:406-408.
[78]屈洪岩,郭玉璞,陆钢,等.玉米花粉多糖对鸡新城疫弱毒疫苗免疫应答的影响[J].中国兽医杂志,1998,1:37-39.
[79]胡积清,程良新,方凯,等.蜂花粉的营养特性及在畜禽业中应用前景[J].安徽农学通报,2010,9:196-197.
[80]张国锋,刁其玉,屠焰,等.蜂花粉及其多糖对犊牛生长性能及血液生化指标的影响[J].中国畜牧杂志,2010,13:52-56.
[1]蒋枫,朱威,张颖,等.莲花花粉营养成分分析[J].中国蜂业,2007,9:5-7.
[2] Paulsen B S, Barsett H. Bioactive pectic polysaccharides[J]. Advances in Polymer Science,2005,186:69-101.
[3]方积年,丁侃.天然药物—多糖的主要生物活性及分离纯化方法[J].中国天然药物,2007,5:338-347.
[4]张旭.人参多糖的系统分析及其免疫活性研究[D]:[博士学位论文].长春:东北师范大学,2009.
[5] Zhang X, Yu L, Bi H T, et al. Total fractionation and characterization of the water-solublepolysaccharides isolated from Panax ginseng C. A. Meyer[J]. Carbohydrate Polymers,2009,77:544-552.
[6] Yu L, Zhang X, Li S S, et al. Rhamnogalacturonan I domains from ginseng pectin[J]. CarbohydratePolymers,2010,79:811-817.
[7] Chambers R E, Clamp J R. An assessment of methanolysis and other factors used in the analysis ofcarbohydrate-containing materials[J]. Biochem J,1971,125:1009-1018.
[8] Guttman A. Analysis of monosaccharide composition by capillary electrophoresis[J]. J ChromatogrA,1997,763:271-277.
[9] Fu D, O'Neill R A. Monosaccharide composition analysis of oligosaccharides and glycoproteins byhigh-performance liquid chromatography[J]. Anal Biochem,1995,227:377-384.
[10] Dubois M, Gilles K, Hamilton J K, et al. A colorimetric method for the determination of sugars[J].Nature,1951,168:167.
[11] Blumenkrantz N, Asboe-Hansen G. New method for quantitative determination of uronic acids[J].Anal Biochem,1973,54:484-489.
[12] Bennett J, Scott K J. Quantitative staining of fraction I protein in polyacrylamide gels usingCoomassie brillant blue[J]. Anal Biochem,1971,43:173-182.
[13] Paulsen B S, Barsett H. Bioactive pectic polysaccharides[J]. Advances in Polymer Science,2005,186:69-101.
[14] Ridley B L, O'Neill M A, Mohnen D. Pectins: structure, biosynthesis, andoligogalacturonide-related signaling[J]. Phytochemistry,2001,57:929-967.
[15] Gronhaug T E, Ghildyal P, Barsett H, et al. Bioactive arabinogalactans from the leaves of Opiliaceltidifolia Endl. ex Walp.(Opiliaceae)[J]. Glycobiology,2010,20:1654-1664.
[16] Alves A, Caridade S G, Mano J F, et al. Extraction and physico-chemical characterization of aversatile biodegradable polysaccharide obtained from green algae[J]. Carbohydr Res,2010,345:2194-2200.
[1] Prade R A, Zhan D, Ayoubi P, et al. Pectins, pectinases and plant-microbe interactions[J]. BiotechnolGenet Eng Rev,1999,16:361-391.
[2] Paulsen B S, Barsett H. Bioactive pectic polysaccharides[J]. Advances in Polymer Science,2005,186:69-101.
[3]张旭.人参多糖的系统分析及其免疫活性研究[D]:[博士学位论文].长春:东北师范大学,2009.
[4] Needs P W, Selvendran R R. Avoiding oxidative degradation during sodium hydroxide/methyliodide-mediated carbohydrate methylation in dimethyl sulfoxide[J]. Carbohydrate Research,1993,245:1-10.
[5] McNeil M, Darvill A G, Albersheim P. Structure of Plant Cell Walls: X. Rhamnogalacturonan I, astructurally complex pectic polysaccharide in the walls of suspension-cultured sycamore cells[J].Plant Physiol,1980,66(6):1128-1134.
[6] Ridley B L, O'Neill M A, Mohnen D. Pectins: structure, biosynthesis, and oligogalacturonide-relatedsignaling[J]. Phytochemistry,2001,57:929-967.
[7] Raju T S, Davidson E A. Structural features of water-soluble novel polysaccharide components fromthe leaves of Tridax procumbens Linn[J]. Carbohydr Res,1994,258:243-254.
[8] Schols H A, Voragen A G J. Complex pectins: Structure elucidation using enzymes[J]. Progress inBiotechnology,1996,14:3-19.
[9] Dong Q, Liu X, Yao J, et al. Structural characterization of a pectic polysaccharide from Neriumindicum flowers[J]. Phytochemistry,2010,71:1430-1437.
[10] Duan J, Wang X, Dong Q, et al. Structural features of a pectic arabinogalactan with immunologicalactivity from the leaves of Diospyros kaki[J]. Carbohydr Res,2003,338:1291-1297.
[11] Yu L, Zhang X, Li S S, et al. Rhamnogalacturonan I domains from ginseng pectin. CarbohydratePolymers,2010,79(4):811-817.
[12] Zhang X, Li S S, Sun L, et al. Further analysis of the structure and immunological activity of an RG-Itype pectin from Panax ginseng. Carbohydrate Polymers,2012,89(2):519-525.
[13] Zhang X, Yu L, Bi H T, et al. Total fractionation and characterization of the water-solublepolysaccharides isolated from Panax ginseng C. A. Meyer. Carbohydrate Polymers,2009,77:544-552.
[14] Prade R A, Zhan D, Ayoubi P, et al. Pectins, pectinases and plant-microbe interactions[J].Biotechnol Genet Eng Rev,1999,16:361-391.
[1] Paulsen B S, Barsett H. Bioactive pectic polysaccharides[J]. Advances in Polymer Science,2005,186:69-101.
[2] Hoshino T, Suzuki Y, Takeichi M, et al. Concomitant intraperitoneal therapy with the antitumorpolysaccharide Sizofiran and rG-CSF for ovarian cancer[J]. Gan To Kagaku Ryoho,1993,20:1638-1641.
[3] Ohwada S, Kawate S, Ikeya T, et al. Adjuvant therapy with protein-bound polysaccharide K andtegafur uracil in patients with stage II or III colorectal cancer: randomized, controlled trial[J]. DisColon Rectum,2003,46:1060-1068.
[4] Hwang I, Ahn G, Park E, et al. An acidic polysaccharide of Panax ginseng ameliorates experimentalautoimmune encephalomyelitis and induces regulatory T cells[J]. Immunol Lett,2011,138:169-178.
[5] Subramanian M, Chintalwar G J, Chattopadhyay S. Antioxidant and radioprotective properties of anOcimum sanctum polysaccharide[J]. Redox Rep,2005,10:257-264.
[6] Qi H, Zhang Q, Zhao T, et al. Antioxidant activity of different sulfate content derivatives ofpolysaccharide extracted from Ulva pertusa (Chlorophyta) in vitro[J]. Int J Biol Macromol,2005,37:195-199.
[7] Xu W, Zhang F, Luo Y, et al. Antioxidant activity of a water-soluble polysaccharide purified fromPteridium aquilinum[J]. Carbohydr Res,2009,344:217-222.
[8]方积年,丁侃.天然药物—多糖的主要生物活性及分离纯化方法[J].中国天然药物,2007,5:338-347.
[9]张旭.人参多糖的系统分析及其免疫活性研究[D]:[博士学位论文].长春:东北师范大学,2009.
[10] Clark C C. Collaborative study of an on-column periodate reaction method for the analysis ofphenylpropanolamine hydrochloride in elixirs[J]. J Assoc Off Anal Chem,1973,56:100-104.
[11] Politis D J, Goodman R N. Fine structure of extracellular polysaccharide of erwinia amylovora[J].Appl Environ Microbiol,1980,40:596-607.
[12] Shingel K I. Current knowledge on biosynthesis, biological activity, and chemical modification ofthe exopolysaccharide, pullulan[J]. Carbohydr Res,2004,339:447-460.
[13] Zhang X, Li S S, Sun L, et al. Further analysis of the structure and immunological activity of anRG-I type pectin from Panax ginseng. Carbohydrate Polymers,2012,89:519-525.
[14] Blumenkrantz N, Asboe-Hansen G. New method for quantitative determination of uronic acids[J].Anal Biochem,1973,54:484-489.
[15] Ni W H, Zhang X, Wang B, et al. Antitumor activities and immunomodulatory effects of ginsengneutral polysaccharides in combination with5-Fluorouracil. Journal of Medicinal Food,2010,13(2):1–8.
[16] Nair P K, Melnick S J, Ramachandran R, et al. Mechanism of macrophage activation by(1,4)-alpha-D-glucan isolated from Tinospora cordifolia[J]. Int Immunopharmacol,2006,6:1815-1824.
[17] Wakshull E, Brunke-Reese D, Lindermuth J, et al. PGG-glucan, a soluble beta-(1,3)-glucan,enhances the oxidative burst response, microbicidal activity, and activates an NF-kappa B-like factorin human PMN: evidence for a glycosphingolipid beta-(1,3)-glucan receptor[J].Immunopharmacology,1999,41:89-107.
[18] Ni W H, Zhang X, Bi H T, et al. Preparation of a glucan from the roots of Rubus crataegifolius Bge.and its immunological activity. Carbohydr Res,2009,344:2512-2518.
[19] Sykes B W, Furr M, Giguere S. In vivo pretreatment with PGG-glucan fails to alter cytokine mRNAexpression of equine peripheral blood mononuclear cells exposed to endotoxin ex vivo[J]. Vet Ther,2005,6:67-76.
[20] Bellon T, Martinez V, Lucendo B, et al. Alternative activation of macrophages in human peritoneum:implications for peritoneal fibrosis[J]. Nephrol Dial Transplant,2011,26:2995-3005.
[21] Bredt D S, Snyder S H. Nitric oxide, a novel neuronal messenger[J]. Neuron,1992,8:3-11.
[22] Gan L, Zhang H S, Yang L X, et al. Immunomodulation and antitumor activity by apolysaccharide-protein complex from Lycium barbarum[J]. Int Immunopharmacol,2004,4:563-569.
[23] Prade R A, Zhan D, Ayoubi P, et al. Pectins, pectinases and plant-microbe interactions[J].Biotechnol Genet Eng Rev.1999.16:361-391.
[24]毕洪涛.胶陀螺多糖的系统分析及生物学活性研究[D]:[博士学位论文].长春:东北师范大学,2010.
[25] Schols H A, Voragen A G J. Complex pectins: Structure elucidation using enzymes[J]. Progress inBiotechnology,1996,14:3-19.
[26] Dong Q, Liu X, Yao J, et al. Structural characterization of a pectic polysaccharide from Neriumindicum flowers[J]. Phytochemistry,2010,71:1430-1437.
[27] Brecker L, Wicklein D, Moll H, et al. Structural and immunological properties of arabinogalactanpolysaccharides from pollen of timothy grass (Phleum pratense L.)[J]. Carbohydr Res,2005,340:657-663.
[28] Tanaka T. Mechanism of antitumor action of polysaccharide fraction prepared from bagasse. II.Immunological reactivity of mice and properties of their sera[J]. Gann,1967,58:451-457.
[29] Gordon S. Alternative activation of macrophages[J]. Nat Rev Immunol,2003,3:23-35.
[30] Gordon S, Martinez F O. Alternative activation of macrophages: mechanism and functions[J].
Immunity,2010,32:593-604.
[1] Salom D, Hill B R, Lear J D, et al. pH–Dependent tetrmerization and amantadine binding of thetransmembrane helix of M from the influenza a virus[J]. Biochemistry,2000,39(46):14160-14170.
[2] Bright R A, Medina M J, Xu X, et al. Incidence of adamantine resistance among influenza A (H3N2)viruses isolated worldwide from1994to2005: a cause for concern[J]. Lancet,2005,366(9492):1175-81.
[3] Moscona A. Medical management of influenza infection[J]. Annu Rev Med,2008,59:397-413.
[4] Hayden F G. Perspectives on antiviral use during pandemic influenza[J]. Philos Traps R Soc Lond BBiol Sci,2001,356(1416):1877-1884.
[5] White M R, Crouch E, Hartshorn M, et al. Cooperative anti-influenza activities of respiratory innateimmune proteins and neuraminidase inhibitor[J]. Am J Physiol-Lung C,2005,288(5):831-840.
[6] Govorkova E A, Fang H B, Tan M, et al. Neuraminidase inhibitor-rimantadine combinations exertadditive and synergistic anti-influenza virus effects in MDCK cells[J]. Antimicrob Agents CH,2004,48(12):4855-4863.
[7] Lindegardh N, Hien T T, Farrar J, et al. A simple and rapid liquid hromatographic assay forevaluation of potentially counterfeit Tamiflu[J]. J Pharmaceut Biomed,2006,42(4):430-433.
[8]杨威.流感病毒神经氨酸酶的检测及人参多糖对其抑制作用的研究[D]:[博士学位论文].长春:东北师范大学,2012.
[9] Potier M, Mameli L, Belisle M, et al. Fluorometric assay of neuraminidase with a sodium(4-methylumbelliferyl-(-D-N-acetylneuraminate)substrate[J]. Anal Biochem,1979,94:287-296.
[10] Ortiz A R, Pisabarro M T, Gago F, et al. Prediction of drug binding affinities by comparativebinding energy analysis[J]. J Med Chem,1995,38:2681-2691.
[11] Escuret V, Frobert E, Bouscambert-Duchamp M, et al. Detection of human influenza A(H1N1) andB strains with reduced sensitivity to neuraminidase inhibitors[J]. J Clin Virol,2008,41:25-28.