小蓟多糖的分离纯化及生物学作用研究
|
摘要
小蓟Cephalanoplos segetum(Bge.) Kitam俗称刺儿菜、野红花、青青菜,为菊科植物,在我国分布广泛,资源丰富。它具有凉血止血,祛瘀消肿的功能,为传统的中药。小蓟同时也可以食用,作为药食两用的野生植物资源含有丰富的营养物质,为人们喜爱的民间野菜。多糖具有丰富的药理活性,且无毒副作用,已广泛应用于临床及保健品市场。为了进一步开发利用小蓟资源,本试验对小蓟多糖的提取分离、组成结构及其生物活性进行了研究。
本实验以陕西周至的小蓟干料为原料,进行了小蓟营养成分的测定;小蓟多糖的最佳提取工艺、分离纯化及对所得组分进行了纯度鉴定、分子量测定和组成分析;采用H_2O_2/Fe~(2+)体系和Fe~(2+)诱发卵黄脂蛋白PUFA过氧化体系研究小蓟多糖半纯品的体外抗氧化活性;通过动物实验进行了小蓟粗多糖生物学作用研究。通过以上研究,为综合开发利用小蓟资源提供参考和依据。
主要实验结果如下:
1.小蓟中灰分含量为8.760%、粗蛋白质含量为4.470%粗脂肪含量为0.430%。在已测定的17种氨基酸中,氨基酸总含量为10.302%,其中人体必需的氨基酸有7种,占氨基酸总含量的42.526%。在已测定的15种矿物元素中,含有9种人体必需元素,其中Ca、Mg的含量较高;有害元素Hg、As、Pb及Cd的含量较低。
2.热水浸提小蓟多糖的最佳工艺为:提取温度90℃,提取时间120min,料液比为1∶10。
3.超声波辅助提取小蓟多糖的最佳工艺参数为:功率250W,提取20min,料液比为1∶15。
4.木瓜蛋白酶+Sevag法为最佳脱蛋白方法,即可较完全的脱除粗多糖中的蛋白成分,又能大大降低多糖的损失率。蛋白脱除率达96.45%。
5.经过DEAE-52和Sephadex G-200柱层析分离到CSK-A'、CSK-B'、CSK-C'、CSK-D'四种多糖组分。
6.四种精制级分纯度鉴定:紫外光谱显示在260nm、280nm处均未发现核酸和蛋白质的吸收峰;醋酸薄膜电泳都得到单一色带;SephadexG-200柱洗脱后,四种多糖的洗脱峰基本均匀对称。
7.组成和结构分析:Sephadex G-200柱层析测定它们的分子量分别为20.7kD、28.6kD、148.0kD、501.2kD;气相色谱分析结果表明CSK-A'主要有木糖、半乳糖、葡萄糖组成,质量比为6.31∶8.52∶1.03,CSK-B'主要有鼠李糖、山梨糖和葡萄糖
Cephalanoplos segetum(Bge.) Kitam called thorn vegetable, wild red flower, green vegetable, is one kind of feverfew and is extensively grow in China and rich in resource. As the traditional Chinese medicine, it can cool the blood and stop bleeding, and other pharmacological activities. Besides, this wild plant is fonded by people because of its abundant nutritional ingredient and can be consumed as food and medical materials as well. Cephalanoplos segetum(Bge.) Kitam is very active in pharmacological aspect while poses no poisonous effect to the body, so it is widely applied to clinical service and sold in health product market. This experiment studies its extraction, separation, components, and active functions.This experiment adopts dry Cephalanoplos segetum(Bge.) Kitam which grows in Zhouzhi within Shaanxi province as the raw material researchs its nutrition ingredients and studies the best way of extraction, separation, and also researches its purity, molecular weight, component. What's more, this paper also focuses on its antioxidative activities by the means of H_2O_2/Fe~(2+) system and polyunsaturated fatty acid(PUFA) of the lipoprotein induced by Fe~(2+) system. This experiment is carried out by putting the polysaccharides of Cephalanoplos segetum(Bge.) Kitam in animal's body to research its biology action. This study provides reference and gives evidence to exploit and utilize Cephalanoplos segetum(Bge.) Kitam.The main experiment result is as following:1. The content of ash in Cephalanoplos segetum(Bge.) Kitam was 8.760% , crude protein 4.470%crude fat 0.430%. The content of 17 determined total amino acids added up to 10.302%, which included 7 essential amino acids accounting for 42.526%of total amino acids. 15 mineral elements had been determined, including 9 essential elements among which the contents of Ca and Mg were higher. The contents of harmful elements Hg, As, Pb and Cd were lower.2. The best extracting technology is in hot water is: at 90℃, extracting 120min, and the ratio of material to liquid is 1:10.3. With the help of ultrasonic, the best technology parameter of extracting is: at the power of 250W, extracting 20 mins, and the ratio of material to liquid is 1:15.4. The best way of extracting protein is using enzyme combined with Sevag method, which can nearly extract the total protein in raw polysaccharide, while degrade the loss rate of polysaccharide. The protein extraction rate is 96.45%.5. Four kinds of polysaccharide as CSK-A'、 CSK-B'、 CSK-C'、 CSK-D' are
separated by DE-52 and Sephadex G-200 chromatography.6. Four kinds of purification identification for the refined products: no nucleic acid and protein absorption peaks appear in the UV spectrum at the places of 260nm and 280nm. After the electrophoresis, there emerges a sole ribbon in the acetic acid membrane. After extracting by SephadexG-200 then, peaks of the four polysaccharides are basically and evenly symmetrical.7. Composing and Structure analysis: their molecular weights are 20.7kD, 28.6kD^ 1481dX 501.2kD respectively measured by Sephadex G-200 chromatography. The result in the spectrum indicates that CSK-A' is mainly composed of xylose, galactose and glucose, with the mass ratio of 6.31:8.52:1.03;CSK-B' is mainly made up of rhamnose, sorbose and glucose, with the mass ratio of 6.97:1.52:0.98;CSK-C is chiefly composed of xylose, fructose, sorbose and glucose, with the mass ratio of 7.01:8.93:3.04:1.03;CSK-D' is formed mainly by xylose, fructose, sorbose and glucose, with the mass ratio of 2.10:9.85:1.05. Infrared spectrum reveals that the Infrared spectrums of these four polysaccharides are classical absorption spectrums, that is, at the place of 1400 cm"'-1200 cm"1 there is angle change vibration, at the place of 2930 cm" there appears C-H stretching out and drawing back vibration area, at the place of 3400 cm"1 there emerges absorption wide peak caused by O-H stretching out and drawing back vibration, at the place of 1200cm"'-1030 cm"1 there is absorption peak that is formed by C-0 stretching out and drawing back vibration. In addition, the absorption peak around the area of 1110 cm"1 shows that the monosaccharide in the polysaccharide is hexahydric heterocycle, with the point of 1640 cm"'-1600 cm"1 as the characteristical peak. At the place of 1075 cm"1-1000 in the Infrared spectrum there appears an absorption condition. So we can deduce that it is one type of B-pyrane polyoses that contains acetamide group.8. The test of antioxidative activities indicates:(1) CSK-B, CSK-C, CSK-D can remove OH that is produced by Fenton reaction through H2O2/Fe2+ system. With the increase of amount, the rate of reduction goes with the same trend, so there is a mutual relationship between reduction rate and quantity of polysaccharides, in which CSK-D poses the highest restraining effect, 1.8ml usage reaching 67.29%, with CSK-B coming as a close second. Although CSK-C has some reduction effect, it is comparatively weaker. While CSK-A almost has no such effect.(2) CSK-A, CSK-B, CSK-C has antioxidative effect to the polyunsaturated fatty acid (PUFA) of the lipoprotein induced by Fe2+ system. With the increase of amount, the rate of reduction goes with the same trend, so there is a mutual relationship between reduction rate and quantity of polysaccharides, in which CSK-C poses the strongest restraining effect, 2.5mg/ml usage reaching 53.64%, followed by CSK-A, CSK-B.
While CSK-D almost has no such effect. 9. Animal experiment indicates:(1) The polysaccharides of Cephalanoplos segetum(Bge.) Kitam can help decrease the content of triglyceride(TG) and total cholesterol(TC), increase the ratio of high-density lipoprotein (HDL-C) in the blood serum of mouse, and obviously regulate its metabolization of lipid.(2) The polysaccharides of Cephalanoplos segetum(Bge.) Kitam can stop mouse from getting old that is caused by D-galactose: it could increase the activities of blood superoxide dismutas(SOD), glutathione peroxidase(GSH-PX), catalase (CAT) of mice, liver and brain organization, decreased the content of the malondialdehyde(MDA)and the activity of monoamine oxidase(MAO). Which indicate that it can reduce the speed of getting old from D-galactose.(3) The polysaccharides of Cephalanoplos segetum(Bge.) Kitam can improve the motion capability of mouse, lower the content of blood urea nitrogen(BUN) of post-exercise mice, decrease the consumption of liver glycogen and increase the activity of lactic dehydrogenase(LDH). Which prove that it can effectively ease the hurt to mouse that is caused by over-motion.
本实验以陕西周至的小蓟干料为原料,进行了小蓟营养成分的测定;小蓟多糖的最佳提取工艺、分离纯化及对所得组分进行了纯度鉴定、分子量测定和组成分析;采用H_2O_2/Fe~(2+)体系和Fe~(2+)诱发卵黄脂蛋白PUFA过氧化体系研究小蓟多糖半纯品的体外抗氧化活性;通过动物实验进行了小蓟粗多糖生物学作用研究。通过以上研究,为综合开发利用小蓟资源提供参考和依据。
主要实验结果如下:
1.小蓟中灰分含量为8.760%、粗蛋白质含量为4.470%粗脂肪含量为0.430%。在已测定的17种氨基酸中,氨基酸总含量为10.302%,其中人体必需的氨基酸有7种,占氨基酸总含量的42.526%。在已测定的15种矿物元素中,含有9种人体必需元素,其中Ca、Mg的含量较高;有害元素Hg、As、Pb及Cd的含量较低。
2.热水浸提小蓟多糖的最佳工艺为:提取温度90℃,提取时间120min,料液比为1∶10。
3.超声波辅助提取小蓟多糖的最佳工艺参数为:功率250W,提取20min,料液比为1∶15。
4.木瓜蛋白酶+Sevag法为最佳脱蛋白方法,即可较完全的脱除粗多糖中的蛋白成分,又能大大降低多糖的损失率。蛋白脱除率达96.45%。
5.经过DEAE-52和Sephadex G-200柱层析分离到CSK-A'、CSK-B'、CSK-C'、CSK-D'四种多糖组分。
6.四种精制级分纯度鉴定:紫外光谱显示在260nm、280nm处均未发现核酸和蛋白质的吸收峰;醋酸薄膜电泳都得到单一色带;SephadexG-200柱洗脱后,四种多糖的洗脱峰基本均匀对称。
7.组成和结构分析:Sephadex G-200柱层析测定它们的分子量分别为20.7kD、28.6kD、148.0kD、501.2kD;气相色谱分析结果表明CSK-A'主要有木糖、半乳糖、葡萄糖组成,质量比为6.31∶8.52∶1.03,CSK-B'主要有鼠李糖、山梨糖和葡萄糖
Cephalanoplos segetum(Bge.) Kitam called thorn vegetable, wild red flower, green vegetable, is one kind of feverfew and is extensively grow in China and rich in resource. As the traditional Chinese medicine, it can cool the blood and stop bleeding, and other pharmacological activities. Besides, this wild plant is fonded by people because of its abundant nutritional ingredient and can be consumed as food and medical materials as well. Cephalanoplos segetum(Bge.) Kitam is very active in pharmacological aspect while poses no poisonous effect to the body, so it is widely applied to clinical service and sold in health product market. This experiment studies its extraction, separation, components, and active functions.This experiment adopts dry Cephalanoplos segetum(Bge.) Kitam which grows in Zhouzhi within Shaanxi province as the raw material researchs its nutrition ingredients and studies the best way of extraction, separation, and also researches its purity, molecular weight, component. What's more, this paper also focuses on its antioxidative activities by the means of H_2O_2/Fe~(2+) system and polyunsaturated fatty acid(PUFA) of the lipoprotein induced by Fe~(2+) system. This experiment is carried out by putting the polysaccharides of Cephalanoplos segetum(Bge.) Kitam in animal's body to research its biology action. This study provides reference and gives evidence to exploit and utilize Cephalanoplos segetum(Bge.) Kitam.The main experiment result is as following:1. The content of ash in Cephalanoplos segetum(Bge.) Kitam was 8.760% , crude protein 4.470%crude fat 0.430%. The content of 17 determined total amino acids added up to 10.302%, which included 7 essential amino acids accounting for 42.526%of total amino acids. 15 mineral elements had been determined, including 9 essential elements among which the contents of Ca and Mg were higher. The contents of harmful elements Hg, As, Pb and Cd were lower.2. The best extracting technology is in hot water is: at 90℃, extracting 120min, and the ratio of material to liquid is 1:10.3. With the help of ultrasonic, the best technology parameter of extracting is: at the power of 250W, extracting 20 mins, and the ratio of material to liquid is 1:15.4. The best way of extracting protein is using enzyme combined with Sevag method, which can nearly extract the total protein in raw polysaccharide, while degrade the loss rate of polysaccharide. The protein extraction rate is 96.45%.5. Four kinds of polysaccharide as CSK-A'、 CSK-B'、 CSK-C'、 CSK-D' are
separated by DE-52 and Sephadex G-200 chromatography.6. Four kinds of purification identification for the refined products: no nucleic acid and protein absorption peaks appear in the UV spectrum at the places of 260nm and 280nm. After the electrophoresis, there emerges a sole ribbon in the acetic acid membrane. After extracting by SephadexG-200 then, peaks of the four polysaccharides are basically and evenly symmetrical.7. Composing and Structure analysis: their molecular weights are 20.7kD, 28.6kD^ 1481dX 501.2kD respectively measured by Sephadex G-200 chromatography. The result in the spectrum indicates that CSK-A' is mainly composed of xylose, galactose and glucose, with the mass ratio of 6.31:8.52:1.03;CSK-B' is mainly made up of rhamnose, sorbose and glucose, with the mass ratio of 6.97:1.52:0.98;CSK-C is chiefly composed of xylose, fructose, sorbose and glucose, with the mass ratio of 7.01:8.93:3.04:1.03;CSK-D' is formed mainly by xylose, fructose, sorbose and glucose, with the mass ratio of 2.10:9.85:1.05. Infrared spectrum reveals that the Infrared spectrums of these four polysaccharides are classical absorption spectrums, that is, at the place of 1400 cm"'-1200 cm"1 there is angle change vibration, at the place of 2930 cm" there appears C-H stretching out and drawing back vibration area, at the place of 3400 cm"1 there emerges absorption wide peak caused by O-H stretching out and drawing back vibration, at the place of 1200cm"'-1030 cm"1 there is absorption peak that is formed by C-0 stretching out and drawing back vibration. In addition, the absorption peak around the area of 1110 cm"1 shows that the monosaccharide in the polysaccharide is hexahydric heterocycle, with the point of 1640 cm"'-1600 cm"1 as the characteristical peak. At the place of 1075 cm"1-1000 in the Infrared spectrum there appears an absorption condition. So we can deduce that it is one type of B-pyrane polyoses that contains acetamide group.8. The test of antioxidative activities indicates:(1) CSK-B, CSK-C, CSK-D can remove OH that is produced by Fenton reaction through H2O2/Fe2+ system. With the increase of amount, the rate of reduction goes with the same trend, so there is a mutual relationship between reduction rate and quantity of polysaccharides, in which CSK-D poses the highest restraining effect, 1.8ml usage reaching 67.29%, with CSK-B coming as a close second. Although CSK-C has some reduction effect, it is comparatively weaker. While CSK-A almost has no such effect.(2) CSK-A, CSK-B, CSK-C has antioxidative effect to the polyunsaturated fatty acid (PUFA) of the lipoprotein induced by Fe2+ system. With the increase of amount, the rate of reduction goes with the same trend, so there is a mutual relationship between reduction rate and quantity of polysaccharides, in which CSK-C poses the strongest restraining effect, 2.5mg/ml usage reaching 53.64%, followed by CSK-A, CSK-B.
While CSK-D almost has no such effect. 9. Animal experiment indicates:(1) The polysaccharides of Cephalanoplos segetum(Bge.) Kitam can help decrease the content of triglyceride(TG) and total cholesterol(TC), increase the ratio of high-density lipoprotein (HDL-C) in the blood serum of mouse, and obviously regulate its metabolization of lipid.(2) The polysaccharides of Cephalanoplos segetum(Bge.) Kitam can stop mouse from getting old that is caused by D-galactose: it could increase the activities of blood superoxide dismutas(SOD), glutathione peroxidase(GSH-PX), catalase (CAT) of mice, liver and brain organization, decreased the content of the malondialdehyde(MDA)and the activity of monoamine oxidase(MAO). Which indicate that it can reduce the speed of getting old from D-galactose.(3) The polysaccharides of Cephalanoplos segetum(Bge.) Kitam can improve the motion capability of mouse, lower the content of blood urea nitrogen(BUN) of post-exercise mice, decrease the consumption of liver glycogen and increase the activity of lactic dehydrogenase(LDH). Which prove that it can effectively ease the hurt to mouse that is caused by over-motion.
引文
[1] ranz G et al, Polysaccharides in Pharmacy: Current Applications and Future Concepts [J]. Plant Medical, 1989, 55: 493-497.
[2] 周建华.国外对多糖的抗肿瘤作用研究简况[J].国外医学中医中药分册,1991,6:1-4.
[3] 闵三弟.真菌的药用价值[J].食用菌学报,1996,4:55-59.
[4] Imberty A, Boume Y Cambillau C et al,Oligosaccharide Conformation in rotei/Carbohydrat Computers[M]. Adv.Biophys.Chem, 1993, 3: 71-I17.
[5] Bourne Y Tibeurgh H Y, Cambillau C. Protein Carbonhydrate Iteraction [M]. Curr. Opin.Stru. Biol. 1993, 3: 681-684.
[6] 张艺,杨明等.日本研究多糖的新进展.国外医学中医中药册,1997,3:46-48.
[7] Rademacher TN, et al. Current Advance in Glycobiology [M]. Ann Rev Biochem, 1988, 57: 785.
[8] 张树政.糖生物学:生命科学种的新前沿[J].生命的化学,1999,3:103-106.
[9] 张翼伸,李治平等.长白云芝多糖的毒性和活性研究及口服制剂对200慢性肝炎临床使用的近期疗效观察[J].吉林师大学报 1997,2:96-106.
[10] 王一飞,吕玉民主编.中药与肿瘤免疫研究[M].北京中国医药科学出版社,1996:278-290.
[11] 方积年.多糖研究的现状[J].药学学报,1986,21:944-947.
[12] Let LS, Lin ZHB. Effect of Ganoderma Polysaccharides on T Cell Subpopulatons and Production of Inteleukin 2 in Mixed Lymphocyte Response [J]. Acta Pharmaceutica Sinica, 1992, 5: 331-334.
[13] Let LS, Lin ZHB. Effect of Ganoderma Polyaccharides on the Activity of DNA Polymerasea in Spleen Cells Stimulated by Alloantigens in Mice in Vtro [J]. Beijing Med.Unv., 1991, 4: 329-332.
[14] 周爱如等.银耳多糖抗肿瘤作用的研究[J].北京医科大学学报,1987,3:39-41.
[15] 吴波等.茯苓多糖抗肿瘤作用与机理的试验研究[J].中国药理学通报,1994,4:300-303.
[16] 徐朝辉.金针菇子实体多糖提取物对人肝癌SMMC-7721细胞的抑增殖作用[J].解剖学杂志,1998,4:187-190.
[17] 陈春英,黄雪华.硫酸酯化箬叶多糖的结构修饰及其抗艾滋病病毒养性[J].药学学报,1998,4:264-268.
[18] 沈自尹.中医药对免疫功能影响的综述与评价[J].中西医结合杂志,1992,7:443-446.
[19] 谭竹钧,韩雅莉.动物药物提取制备实用技术[M].北京:中国农业出版社,2000,132-134.
[20] G.Franz D., Pauper S. Alban. Pharmacological activities of sulfated carbohydr carbohydrate polymers [J]. Proceedings of the phytochemical society of Europe, 2000, 44: 47-52.
[21] Fujihara. M., et al. Sugar Constituents of fucoidans from Sorgassum Ringgoldianum and theivbiological wtivity [J]. CarbohydrRes, 1984, 125: 97-99.
[22] 丁源等.我国药用海藻名录及其应用[J].海洋药物,1982,2(2):44-45.
[23] Marsin JR, Topalova H. Radioprotection by polysaccharide [J]. Pharmacol. Ther., 1987, 39: 255-266.
[24] 吴剑波,王蓉.多糖生物活性的研究进展[J].国外医药抗生素册,2001,22(3):97-100.
[25] 王长云,管华诗.多糖抗病毒作用研究进展.Ⅰ.多糖抗病毒作用[J].生物工程进展,2000,20(1):17-20.
[26] 左绍远.螺旋藻多糖对D-半乳糖所致衰老小鼠的作用[J].中国生化药物杂志,1998,19(1):15-18.
[27] 钱新华,王俏先,唐晓玲等.绞股兰多糖对免疫功能的影响[J].中国药科大学学报,1998,30(1):51-53.
[28] 龚晓健,季晖,卢顺高等.人工虫草多糖对小鼠免疫功能的影响[J].中国药科大学学报,2000,31(1):53-55.
[29] 齐春会,张永祥,赵修南等.拘杞粗多糖的免疫活性[J].Chinese Journal of Pharmmacology and Toxicology,2001,15(3):180-184.
[30] 庞战军,陈缓,周玫.云芝多糖对巨噬细胞Gpx基因表达的影响[J].生物化学与生物物理学报.1999,31(3):284-288.
[31] 张庆,雷林生,林勤保等.大枣多糖体外抗补体活性及促进小鼠脾细胞的增殖作用[J].中药药理与临床,1998,14(5):19-21.
[32] 田庚元,冯宇澄.多糖类免疫调节剂的研究与应用[J].化学进展,1994,6(2):113-115.
[33] Liu F, Fung M C, Ooi V E C, et al. Induction in the mouse of gen expression f immunomodulating cytokine by Mushroom polysaccharide-protein complexes [J]. Life Science, 1996, 58(21): 1795-1803.
[34] 孔庆胜,袁慧,昊福国等.玉米穗多糖对四氧嘧啶糖尿病小鼠高血糖的防 治作用[J].济宁医学院学报,1996,19(2):4-5.
[35] 王靖,葛盛芳,陈琦等.知母多糖降血糖活性研究[J].中草药,1996,27(10):605-606.
[36] Tadashi K, Hotaka M, Shigeyuki U, et al. Antidiabetic activity of an acidic polysaccharide from the fruiting bodies of Tremella aurantia [J]. Front Biomed Biotechnol, 1996, 3, 243-248.
[37] 陈琼华.《生物化学》[M].人民卫生出版社,1987,6.
[38] 周慧萍,蒋巡天,王淑如等.浒苔多糖的降血脂及其对SOD活力和LPO含量的影响[J].生物化学杂志,1995,11(2):161-165.
[39] 孔庆胜,王彦英,蒋滢.南瓜多糖的分离、纯化及其降血脂作用[J].中国生化药物杂志,2000,21(3):130-132.
[40] 宋丽艳,马文霞,于荣敏等.银杏细胞培养物多糖和银杏叶多糖生物活性的研究[J].中国生化药物杂志,1999,20(6):278-280.
[41] 陶海南,刘辉,薛喜文等.紫萁多糖抗菌活性初步研究[J].南开大学学报,1996,20(4):305-308.
[42] 秦华珍,夏新华,李钟文.黄花倒水莲多糖的抗应激作用[J].广西中医药,1996,14(3):52-54.
[43] 田庚元.植物多糖的研究进展[J].中国中药杂志,1998,20(7):441.
[44] 黄泰康.党参水煎液及多糖的药理研究[J].中国中药杂志,1997,16(7):3.
[45] Czop J K, Austen K F. Properties of glycans that activate the human alternative complement pathway and interact with human monocyte fl-glucan receptor[J]. Immunol, 1985, 135(1): 3388-3393.
[46] Willian L, Polysaccharides from lichems[J]. Characteristics and biological activity, 2001, 208(3): 199.
[47] Demleitner S, Kraus J, Franz C.Synthesis and anti-tumor activity of derivatives of curdlan and Iichenan branched[J]. Carbohydrate Research, 1992, 226(2): 26-29.
[48] Matsuzaki K, Yamamoto I, Sato T, etal. C-NMR investigation of synthetic branched polysaccharides[J]. Carbohydrate Polymers, 1986, 6(2): 155-163.
[49] Perret J, Bruneteau M, Michel G, et al. Effect of growth conditions on the structure of β-D-glucans from Phytophthora parasitica Dastur[J]. Carbohydrate Polymer, 1991, 17(2): 231-236.
[50] 胡文祥,王文曦.多糖及其衍生物的医药学研究[J].多糖药理学研究,1994,187(3):4-8.
[51] Gao Y, Fukuda A, Katsuraya K, et al. Synthesis of regioselective substituted curdlan sulfates with medium molecular weights and their specific activi- ties[M]. Macrom, 1997, 30(11): 3224-3228.
[52] Groth T, Wagenknect W. Anticoagulant potential of regioselective derivatized cellulose[M]. Biomaterials, 2001, 22(8): 2719-2729.
[53] Misaki A, Kawagnchi K. Structure of pestalotan-a highly branched (1,3)-β-D-glucan elaborated by pestlotia, and the enhancement of its antitumor activity by polyol modification of the side chains[J]. Carbohydrate Researchs, 1984, 129(1): 209-227.
[54] Mischnick P.Determination of the substitution pattern in the polymer chain of cellulose sulfates[J]. Carbohydr Res, 1998, 309(1): 109-110.
[55] Godes M, Mischnick P. Determination of the substitution pattern in the polymer chain of cellulose sulfates[J]. Carbohydr Res, 1998, 309(1): 109-115.
[56] 张丽萍,汉丽萍,王月秋等.硫酸化高山红景天多糖(RSASL)的置备及鉴定[J].分子科学学报,1999,15(3):205-210.
[57] 陈惠黎,王克夷.糖复合物的结构和功能[M].上海医科大学出版社,1998,353-354.
[58] 田庚元.天然多糖的研究和应用(下)[M].上海化工,2000,11:23-25.
[59] 陈毓,丁安伟,杨星昊,张丽.小蓟化学成分药理作用及临床应用研究述要[J].中医药学刊,2005,4:614-615.
[60] 曾再新.漫话刺儿菜[J].上海蔬菜,1998,2:52.
[61] 李清华.小蓟止血成分的研究[J].中草药,1982,13(9):393.
[62] 姚乾元,赵渤年,吴保杰等.小蓟升压成分的化学研究[J].中草药,1992,23(10):517.
[63] 顾玉诚,屠呦呦.小蓟化学成分研究[J].中国中药杂志,1992,17(9):547.
[64] 周文序,田珍.中药大、小蓟中柳穿鱼甙和芦丁的薄层扫描法定量分析[J].药物分析杂志,1994,6:35.
[65] 张来新.小蓟中提取分离小蓟类化合物的实验研究[J].中成药,2004,26(6):503-504.
[66] 张来新.从小蓟中提取止血药成分咖啡酸[J].中成药,2002,24(10):807-808.
[67] 顾玉诚.小蓟化学成分研究[J].中国中药杂志,1992,(9):547-548.
[68] 李德华.小蓟临床研究[J].山东医学院学报,1959,(7):42.
[69] 魏彦,邱乃英,欧阳青.大蓟、小蓟的鉴别与临床应用[J].北京中医杂志,2002,21(5):296-297.
[70] 中国医学科学院药物研究所抗菌工作组[J].中药通报,1960,(2):59.
[71] 赵学建.小蓟药理研究进展[J].中国医药学刊,2005,23(4):614-615.
[72] 余冰宾(主编).生物化学实验指导[M].清华大学出版社,2004,136-137.
[73] 陈业高主编.《植物化学成分》[M].化学工业出版社,2004,4:41-44.
[74] 北京医学院编.《中草药化学成分》[M].人民卫生出版社,1983,5:32-35
[75] 陈业高主编.《植物化学成分》[M].化学工业出版社,2004,4:49-50.
[76] 李俊,韩向晖,李钟洪等.茯苓多糖的提取及含量测定[J].中国现代应用药学杂志,2000,17(2):49-50.
[77] 马如复主编.《生物化学试验》[M].武汉大学出版社,1998,1:45-46.
[78] 方积年.多糖的分离纯化及其纯度鉴别与分子量测定[J].药学通报,1984,19(10):46-49.
[79] 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.
[80] 欧阳平凯主编.《生物分离原理及技术》[M].化学工业出版社,1999,2.
[81] 郭玫,余晓晖.多糖的研究概况[J].甘肃中医学院学报,1994,11(2):55-56.
[82] 赵永芳主编.《生物化学技术原理及其应用》[M].武汉大学出版社,1991,2.
[83] 任金山,吴梧桐,李颖等.花粉多糖的分离纯化及其组成单糖分析[J].中国药科大学学报,1990,21(3):173-175.
[84] 张林维.当归水溶性多糖级分As-Ⅲa和As-Ⅲb的纯瘾定与结构研究[J].激光生物学报,1999,8(2):123-126.
[85] 陶乐平,丁在富.气相色谱在多糖结构测定中的应用[J].色谱,1994,12(5):351-354.
[86] 孟令芝,何永炳.有机波谱分析[M].武汉大学出版社,1997,10:71-73.
[87] 凌关庭.抗氧化食品与健康[M].北京:化学工业出版社,2004,38-39.
[88] 方允中等.自由基生物学地理论与应用[M].科学出版社,2002,42-43.
[89] 金鸣,蔡亚欣,李金荣等.邻二氮菲-Fe~(2+)氧化法检测H_2O_2/Fe~(2+)产生的羟自由基[J].生物化学与生物物理进展,1996,23(6):553-555.
[90] 张尔贤,俞丽君,周意琳.Fe~(2+)诱发脂蛋白PUFA过氧化体系及对若干天然产物抗氧化作用的评价[J].生物化学与生物物理学报,1996,28(2):218-222.
[91] 郑健仙编著.《功能性食品》(第三卷)[M].中国轻工业出版社,1999,181-435.
[92] Criqui, M.H. and Ringel, B.L. Does diet or alcohol explain the French paradaox[J]. Lancet 1994, 344: 1719-1723.
[93] 黄益丽,廖鑫凯等.香菇多糖的生物活性[J].生命化学,2001,21(5):371-373.
[94] 施新酋犬.医学动物实验方法[M].北京:人民卫生出版社,1980,103-104.
[95] 陈钧辉著.生物化学实验[M].科学出版社,2003(3):63-64.
[96] 何来英,严卫星,楼密密等.保健食品抗疲劳作用试验方法研究[J].中国食品卫生杂志,1997,9(4):1-6.
[97] 卫生部卫生监督司.保健食品检验与评价技术规范[M].北京:中科多媒体电子出版社,2003,87-93
[2] 周建华.国外对多糖的抗肿瘤作用研究简况[J].国外医学中医中药分册,1991,6:1-4.
[3] 闵三弟.真菌的药用价值[J].食用菌学报,1996,4:55-59.
[4] Imberty A, Boume Y Cambillau C et al,Oligosaccharide Conformation in rotei/Carbohydrat Computers[M]. Adv.Biophys.Chem, 1993, 3: 71-I17.
[5] Bourne Y Tibeurgh H Y, Cambillau C. Protein Carbonhydrate Iteraction [M]. Curr. Opin.Stru. Biol. 1993, 3: 681-684.
[6] 张艺,杨明等.日本研究多糖的新进展.国外医学中医中药册,1997,3:46-48.
[7] Rademacher TN, et al. Current Advance in Glycobiology [M]. Ann Rev Biochem, 1988, 57: 785.
[8] 张树政.糖生物学:生命科学种的新前沿[J].生命的化学,1999,3:103-106.
[9] 张翼伸,李治平等.长白云芝多糖的毒性和活性研究及口服制剂对200慢性肝炎临床使用的近期疗效观察[J].吉林师大学报 1997,2:96-106.
[10] 王一飞,吕玉民主编.中药与肿瘤免疫研究[M].北京中国医药科学出版社,1996:278-290.
[11] 方积年.多糖研究的现状[J].药学学报,1986,21:944-947.
[12] Let LS, Lin ZHB. Effect of Ganoderma Polysaccharides on T Cell Subpopulatons and Production of Inteleukin 2 in Mixed Lymphocyte Response [J]. Acta Pharmaceutica Sinica, 1992, 5: 331-334.
[13] Let LS, Lin ZHB. Effect of Ganoderma Polyaccharides on the Activity of DNA Polymerasea in Spleen Cells Stimulated by Alloantigens in Mice in Vtro [J]. Beijing Med.Unv., 1991, 4: 329-332.
[14] 周爱如等.银耳多糖抗肿瘤作用的研究[J].北京医科大学学报,1987,3:39-41.
[15] 吴波等.茯苓多糖抗肿瘤作用与机理的试验研究[J].中国药理学通报,1994,4:300-303.
[16] 徐朝辉.金针菇子实体多糖提取物对人肝癌SMMC-7721细胞的抑增殖作用[J].解剖学杂志,1998,4:187-190.
[17] 陈春英,黄雪华.硫酸酯化箬叶多糖的结构修饰及其抗艾滋病病毒养性[J].药学学报,1998,4:264-268.
[18] 沈自尹.中医药对免疫功能影响的综述与评价[J].中西医结合杂志,1992,7:443-446.
[19] 谭竹钧,韩雅莉.动物药物提取制备实用技术[M].北京:中国农业出版社,2000,132-134.
[20] G.Franz D., Pauper S. Alban. Pharmacological activities of sulfated carbohydr carbohydrate polymers [J]. Proceedings of the phytochemical society of Europe, 2000, 44: 47-52.
[21] Fujihara. M., et al. Sugar Constituents of fucoidans from Sorgassum Ringgoldianum and theivbiological wtivity [J]. CarbohydrRes, 1984, 125: 97-99.
[22] 丁源等.我国药用海藻名录及其应用[J].海洋药物,1982,2(2):44-45.
[23] Marsin JR, Topalova H. Radioprotection by polysaccharide [J]. Pharmacol. Ther., 1987, 39: 255-266.
[24] 吴剑波,王蓉.多糖生物活性的研究进展[J].国外医药抗生素册,2001,22(3):97-100.
[25] 王长云,管华诗.多糖抗病毒作用研究进展.Ⅰ.多糖抗病毒作用[J].生物工程进展,2000,20(1):17-20.
[26] 左绍远.螺旋藻多糖对D-半乳糖所致衰老小鼠的作用[J].中国生化药物杂志,1998,19(1):15-18.
[27] 钱新华,王俏先,唐晓玲等.绞股兰多糖对免疫功能的影响[J].中国药科大学学报,1998,30(1):51-53.
[28] 龚晓健,季晖,卢顺高等.人工虫草多糖对小鼠免疫功能的影响[J].中国药科大学学报,2000,31(1):53-55.
[29] 齐春会,张永祥,赵修南等.拘杞粗多糖的免疫活性[J].Chinese Journal of Pharmmacology and Toxicology,2001,15(3):180-184.
[30] 庞战军,陈缓,周玫.云芝多糖对巨噬细胞Gpx基因表达的影响[J].生物化学与生物物理学报.1999,31(3):284-288.
[31] 张庆,雷林生,林勤保等.大枣多糖体外抗补体活性及促进小鼠脾细胞的增殖作用[J].中药药理与临床,1998,14(5):19-21.
[32] 田庚元,冯宇澄.多糖类免疫调节剂的研究与应用[J].化学进展,1994,6(2):113-115.
[33] Liu F, Fung M C, Ooi V E C, et al. Induction in the mouse of gen expression f immunomodulating cytokine by Mushroom polysaccharide-protein complexes [J]. Life Science, 1996, 58(21): 1795-1803.
[34] 孔庆胜,袁慧,昊福国等.玉米穗多糖对四氧嘧啶糖尿病小鼠高血糖的防 治作用[J].济宁医学院学报,1996,19(2):4-5.
[35] 王靖,葛盛芳,陈琦等.知母多糖降血糖活性研究[J].中草药,1996,27(10):605-606.
[36] Tadashi K, Hotaka M, Shigeyuki U, et al. Antidiabetic activity of an acidic polysaccharide from the fruiting bodies of Tremella aurantia [J]. Front Biomed Biotechnol, 1996, 3, 243-248.
[37] 陈琼华.《生物化学》[M].人民卫生出版社,1987,6.
[38] 周慧萍,蒋巡天,王淑如等.浒苔多糖的降血脂及其对SOD活力和LPO含量的影响[J].生物化学杂志,1995,11(2):161-165.
[39] 孔庆胜,王彦英,蒋滢.南瓜多糖的分离、纯化及其降血脂作用[J].中国生化药物杂志,2000,21(3):130-132.
[40] 宋丽艳,马文霞,于荣敏等.银杏细胞培养物多糖和银杏叶多糖生物活性的研究[J].中国生化药物杂志,1999,20(6):278-280.
[41] 陶海南,刘辉,薛喜文等.紫萁多糖抗菌活性初步研究[J].南开大学学报,1996,20(4):305-308.
[42] 秦华珍,夏新华,李钟文.黄花倒水莲多糖的抗应激作用[J].广西中医药,1996,14(3):52-54.
[43] 田庚元.植物多糖的研究进展[J].中国中药杂志,1998,20(7):441.
[44] 黄泰康.党参水煎液及多糖的药理研究[J].中国中药杂志,1997,16(7):3.
[45] Czop J K, Austen K F. Properties of glycans that activate the human alternative complement pathway and interact with human monocyte fl-glucan receptor[J]. Immunol, 1985, 135(1): 3388-3393.
[46] Willian L, Polysaccharides from lichems[J]. Characteristics and biological activity, 2001, 208(3): 199.
[47] Demleitner S, Kraus J, Franz C.Synthesis and anti-tumor activity of derivatives of curdlan and Iichenan branched[J]. Carbohydrate Research, 1992, 226(2): 26-29.
[48] Matsuzaki K, Yamamoto I, Sato T, etal. C-NMR investigation of synthetic branched polysaccharides[J]. Carbohydrate Polymers, 1986, 6(2): 155-163.
[49] Perret J, Bruneteau M, Michel G, et al. Effect of growth conditions on the structure of β-D-glucans from Phytophthora parasitica Dastur[J]. Carbohydrate Polymer, 1991, 17(2): 231-236.
[50] 胡文祥,王文曦.多糖及其衍生物的医药学研究[J].多糖药理学研究,1994,187(3):4-8.
[51] Gao Y, Fukuda A, Katsuraya K, et al. Synthesis of regioselective substituted curdlan sulfates with medium molecular weights and their specific activi- ties[M]. Macrom, 1997, 30(11): 3224-3228.
[52] Groth T, Wagenknect W. Anticoagulant potential of regioselective derivatized cellulose[M]. Biomaterials, 2001, 22(8): 2719-2729.
[53] Misaki A, Kawagnchi K. Structure of pestalotan-a highly branched (1,3)-β-D-glucan elaborated by pestlotia, and the enhancement of its antitumor activity by polyol modification of the side chains[J]. Carbohydrate Researchs, 1984, 129(1): 209-227.
[54] Mischnick P.Determination of the substitution pattern in the polymer chain of cellulose sulfates[J]. Carbohydr Res, 1998, 309(1): 109-110.
[55] Godes M, Mischnick P. Determination of the substitution pattern in the polymer chain of cellulose sulfates[J]. Carbohydr Res, 1998, 309(1): 109-115.
[56] 张丽萍,汉丽萍,王月秋等.硫酸化高山红景天多糖(RSASL)的置备及鉴定[J].分子科学学报,1999,15(3):205-210.
[57] 陈惠黎,王克夷.糖复合物的结构和功能[M].上海医科大学出版社,1998,353-354.
[58] 田庚元.天然多糖的研究和应用(下)[M].上海化工,2000,11:23-25.
[59] 陈毓,丁安伟,杨星昊,张丽.小蓟化学成分药理作用及临床应用研究述要[J].中医药学刊,2005,4:614-615.
[60] 曾再新.漫话刺儿菜[J].上海蔬菜,1998,2:52.
[61] 李清华.小蓟止血成分的研究[J].中草药,1982,13(9):393.
[62] 姚乾元,赵渤年,吴保杰等.小蓟升压成分的化学研究[J].中草药,1992,23(10):517.
[63] 顾玉诚,屠呦呦.小蓟化学成分研究[J].中国中药杂志,1992,17(9):547.
[64] 周文序,田珍.中药大、小蓟中柳穿鱼甙和芦丁的薄层扫描法定量分析[J].药物分析杂志,1994,6:35.
[65] 张来新.小蓟中提取分离小蓟类化合物的实验研究[J].中成药,2004,26(6):503-504.
[66] 张来新.从小蓟中提取止血药成分咖啡酸[J].中成药,2002,24(10):807-808.
[67] 顾玉诚.小蓟化学成分研究[J].中国中药杂志,1992,(9):547-548.
[68] 李德华.小蓟临床研究[J].山东医学院学报,1959,(7):42.
[69] 魏彦,邱乃英,欧阳青.大蓟、小蓟的鉴别与临床应用[J].北京中医杂志,2002,21(5):296-297.
[70] 中国医学科学院药物研究所抗菌工作组[J].中药通报,1960,(2):59.
[71] 赵学建.小蓟药理研究进展[J].中国医药学刊,2005,23(4):614-615.
[72] 余冰宾(主编).生物化学实验指导[M].清华大学出版社,2004,136-137.
[73] 陈业高主编.《植物化学成分》[M].化学工业出版社,2004,4:41-44.
[74] 北京医学院编.《中草药化学成分》[M].人民卫生出版社,1983,5:32-35
[75] 陈业高主编.《植物化学成分》[M].化学工业出版社,2004,4:49-50.
[76] 李俊,韩向晖,李钟洪等.茯苓多糖的提取及含量测定[J].中国现代应用药学杂志,2000,17(2):49-50.
[77] 马如复主编.《生物化学试验》[M].武汉大学出版社,1998,1:45-46.
[78] 方积年.多糖的分离纯化及其纯度鉴别与分子量测定[J].药学通报,1984,19(10):46-49.
[79] 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.
[80] 欧阳平凯主编.《生物分离原理及技术》[M].化学工业出版社,1999,2.
[81] 郭玫,余晓晖.多糖的研究概况[J].甘肃中医学院学报,1994,11(2):55-56.
[82] 赵永芳主编.《生物化学技术原理及其应用》[M].武汉大学出版社,1991,2.
[83] 任金山,吴梧桐,李颖等.花粉多糖的分离纯化及其组成单糖分析[J].中国药科大学学报,1990,21(3):173-175.
[84] 张林维.当归水溶性多糖级分As-Ⅲa和As-Ⅲb的纯瘾定与结构研究[J].激光生物学报,1999,8(2):123-126.
[85] 陶乐平,丁在富.气相色谱在多糖结构测定中的应用[J].色谱,1994,12(5):351-354.
[86] 孟令芝,何永炳.有机波谱分析[M].武汉大学出版社,1997,10:71-73.
[87] 凌关庭.抗氧化食品与健康[M].北京:化学工业出版社,2004,38-39.
[88] 方允中等.自由基生物学地理论与应用[M].科学出版社,2002,42-43.
[89] 金鸣,蔡亚欣,李金荣等.邻二氮菲-Fe~(2+)氧化法检测H_2O_2/Fe~(2+)产生的羟自由基[J].生物化学与生物物理进展,1996,23(6):553-555.
[90] 张尔贤,俞丽君,周意琳.Fe~(2+)诱发脂蛋白PUFA过氧化体系及对若干天然产物抗氧化作用的评价[J].生物化学与生物物理学报,1996,28(2):218-222.
[91] 郑健仙编著.《功能性食品》(第三卷)[M].中国轻工业出版社,1999,181-435.
[92] Criqui, M.H. and Ringel, B.L. Does diet or alcohol explain the French paradaox[J]. Lancet 1994, 344: 1719-1723.
[93] 黄益丽,廖鑫凯等.香菇多糖的生物活性[J].生命化学,2001,21(5):371-373.
[94] 施新酋犬.医学动物实验方法[M].北京:人民卫生出版社,1980,103-104.
[95] 陈钧辉著.生物化学实验[M].科学出版社,2003(3):63-64.
[96] 何来英,严卫星,楼密密等.保健食品抗疲劳作用试验方法研究[J].中国食品卫生杂志,1997,9(4):1-6.
[97] 卫生部卫生监督司.保健食品检验与评价技术规范[M].北京:中科多媒体电子出版社,2003,87-93
© 2004-2018 中国地质图书馆版权所有 京ICP备05064691号 京公网安备11010802017129号 地址：北京市海淀区学院路29号 邮编：100083 电话：办公室：(+86 10)66554848；文献借阅、咨询服务、科技查新：66554700 |