金莲花多糖的分离纯化及其抗氧化活性研究

神经药理学报 ›› 2012, Vol. 2 ›› Issue (6): 28-35.

金莲花多糖的分离纯化及其抗氧化活性研究

饶娜，王树林，曹欣欣，安芳，王书华

河北北方学院药学系，张家口, 075000,中国

出版日期:2012-12-26 发布日期:2014-06-27
通讯作者: 王书华，教授，硕士生导师。主要从事中药有效成分的提取及药理研究。Email：wangshyh@126.com
作者简介:饶娜，河北北方学院2009级药理学硕士研究生， Email：raonaomi@126.com
基金资助:
张家口市科技局资助项目（11110015D）河北北方学院重大项目(ZD1314)

Purification and Anti-oxidant Activity of the Polysaccharide Isolated from Trolliouschinensis

RAO Na，WANG Shu-lin，CAO Xin-xin，AN Fang，WANG Shu-hua

College of Pharmacy, Hebei North University, Zhang jiakou，075000, China

Online:2012-12-26 Published:2014-06-27
Contact: 王书华，教授，硕士生导师。主要从事中药有效成分的提取及药理研究。Email：wangshyh@126.com
About author:饶娜，河北北方学院2009级药理学硕士研究生， Email：raonaomi@126.com
Supported by:
张家口市科技局资助项目（11110015D）河北北方学院重大项目(ZD1314)

摘要/Abstract

摘要： 目的：对金莲花粗多糖进行分离纯化，研究纯化多糖的体外抗氧化作用，为进一步开发金莲花中药资源提供研究资料。方法：1常规回流提取、乙醇沉淀得到金莲花粗多糖，经AB-8大孔吸附树脂脱色、脱蛋白后得精制的金莲花多糖（trollius chinensis bunge polysaccharide, TCBP），以蒸馏水、不同浓度的氯化钠和氢氧化钠溶液为洗脱剂，应用DEAE-52纤维素离子交换柱层析，Sephadex G-150凝胶过滤柱层析对得到的主要组分多糖进一步分离纯化，采用高效凝胶过滤色谱（HPGPC）鉴定所得组分纯度并测定其相对分子质量，通过高效液相色谱（HPLC）分析多糖的单糖组成。 2 通过水杨酸-Fe2+-H2O2体系（·OH）、二苯代苦味肼基自由基(DPPH)体系和邻苯三酚自氧化（·O2－）法对分离纯化的金莲花多糖组分的抗氧化能力进行研究。结果： 1得到TCBP1，TCBP2，TCBP3，TCBP4四个多糖组分均为杂多糖，其中 TCBP1分子量为3283220 Da； TCBP2分子量为156819 Da； TCBP3分子量为10949 Da； TCBP4分子量为1068 Da；2 TCBP1，TCBP2，TCBP3，TCBP4均具有清除·OH的能力，其强弱顺序为：TCBP2 > TCBP3 > TCBP1 > TCBP4， TCBP2和TCBP3对·DPPH 具有一定清除作用， TCBP1，TCBP2，TCBP3，TCBP4均没有清除·O2－的能力。相反，还能诱导·O2－的产生，且随着它们浓度的增大，邻苯三酚自氧化程度也增强。结论金莲花多糖为复杂的杂聚多糖，且具有一定的抗氧化活性。

关键词: 金莲花, 多糖, 分离纯化, 抗氧化

Abstract: Objective: To study purification and anti-oxidant activity of the polysaccharide Isolated from Trolliouschinensis. This study provides basical information to develop and utilize Trollious chinensis resource efficiently. Methods: 1 The methods of reflux extraction was adapted to extract polysaccharide from Trollius chinensis, AB-8 resin can be used as a high effective one to romove pigment and protein of Trollius chinensis polysaccharide(TCBP). A DEAE-52 cellulose ion exchange column ，a Sephadex G-150 gels filter column were successively used to further separate crude polysaccharides, and then determining the product purity and molecular weight with high performance gels filtration chromagraphy(HPGPC). High performance liquid Chromatography(HPLC) was applied to study the momosaccharide composition of Trollius chinensis polysaccharide. 2 Antioxidative capability of polysaccharide was determined by salicylic-Fe2+-H2O2 system(·OH), 1,1-Diphenyl-2-picrylhydrazyl radical 2, 2-Diphenyl-1-(2, 4, 6-trinitrophenyl)hydrazyl(DPPH) system, and means of pyrogallol oxidation（·O2－）. Results: 1 Our results showed that four polysaccharides are complex polysaccharide with a variety of monosaccharides. Molecular weight of TCBP1 is 3283220 Da, TCBP2, 156819 Da, TCBP3, 10949 Da and TCBP4, 1068 Da. 2 Four polysaccharides had significant effects on scavenging hydroxyl radical, The sequence from strong to weak of this effects is: TCBP2 > TCBP3 > TCBP1 > TCBP4. TCBP2 and TCBP3 had certain effect on 1, 1-Diphenyl -2- picrylhydrazyl radical. They all induced superoxide radical instead of scavenging. Conclusion: TCBP1, TCBP2, TCBP3 and TCBP4 are complex polysaccharide with a variety of monosaccharides，they have the certain antioxidant activity.

Key words: trollius chinensis, polysaccharide, isolation and purification, anti-oxidation

饶娜, 王树林, 曹欣欣, 安芳, 王书华. 金莲花多糖的分离纯化及其抗氧化活性研究[J]. 神经药理学报, 2012, 2(6): 28-35.

RAO Na, WANG Shu-lin, CAO Xin-xin, AN Fang, WANG Shu-hua. Purification and Anti-oxidant Activity of the Polysaccharide Isolated from Trolliouschinensis[J]. ACTA NEUROPHARMACOLOGICA, 2012, 2(6): 28-35.

参考文献

[1] 刘洁, 李文香, 王文亮, 等. 生物多糖研究进展[J]. 山东农业科学, 2011, 5: 98-101.

[2] Bum Chun Lee, Jun Tae Bae, Hyeong Bae Pyo, et al. Biological activities of the polysaccharides produced from submerged culture of the edible Basidiomycete Grifola frondosa[J]. Enzyme and Microbical Technology, 2003, 32(5): 574-581.

[3] Du Xiu-ju, Zhang Jing-song, Yan Yan, et al. Purification, chemical modification and immunostimulating activity of polysaccharides from Tremella aurantialba fruit bodies[J]. Biomed and Biotechnol, 2010, 11(6): 437- 442.

[4] 沈志平. 中药金莲花药源考证与应用[J]. 时珍国医国药, 2000, 11 (12):1110.

[5] 刘召阳, 罗都强. 金莲花的化学成分研究[J].中草药, 2010, 41(3): 370-373.

[6] Fang An, Yang Guo-dong, Tian Jia Mimg , et al. Antioxidant effects of the orientin and vitexin in Trollius chinensis Bunge in D-galactose-aged mice [J]. Neural Regeneration Research,2012,7（33）:2565-2575.

[7] 朱登祥, 安芳, 王书华. 牡荆苷对人食管癌EC-109细胞生长及凋亡的影响[J].中草药, 2012, 43(9): 1781-1784.

[8] 付晓春, 王希, 郑辉, 等. 荭草苷对缺血缺氧心肌的保护作用[J]. 南方医科大学学报, 2007, 27(8): 1173-1175.

[9] 张丽娟, 张贵君, 李仁伟. 金莲花蛋白超声提取工艺优化及其抑菌活性的初步测定[J]. 天津中医药, 2007, 24(1): 63-65.

[10] 颜娟, 曲彩红, 安芳, 等. PHPLC法分离纯化金莲花中荭草苷和牡荆苷[J]. 中成药, 2011, 33(4): 655-658.

[11] 田灵敏, 邱雪梅, 潘子敬, 等. 新型糖组成高效液相色谱分析技术的构建及其在水飞蓟多糖质控中的应用[J]. 药学学报, 2010, 45(4): 498-504.

[12] Sudha G, Sangeetha Priya M, Indhu Shree R, et al. Antioxidant activity of ripe pepino fruit (solanum muricatum aiton)[J]. Int J Pharm Pharm Sci, 2011, 3(3): 257- 261.

[13] Zeng P Y, Wu J G.,Liao L M, et al. In vitro antioxidant activities of endophytic fungi isolated fro m the liverwort Scapania verrucosa[J]. Helvetica Chimica Acta, 2011, 10(4): 3169- 3179.

[14] 贺寅. 龙眼多糖制备工艺及其抗氧化活性研究[D]. 北京：中国农业科学院,中国优秀硕士学位论文全文数据库,2011.

[15] Sun Hai-hong, Mao Wen-jun, Chen Yin, et al. Isolation, chemical characteristics and antioxidant properties of the polysaccharides from marine fungus Penicillium sp. F23-2[J]. Carbohydrate Polymers, 2009, 78(1): 117- 124.

[16] 侯飞, 高华. 冬凌草多糖的提取、分离及抗氧化活性研究[J]. 中国生化药物杂志, 2012, 32(5): 619-622.

[17] 程超, 李伟. 几种植物水溶性多糖的体外抗氧化作用[J]. 食品工业科技, 2006, 27(9): 63- 65.