臺灣博碩士論文加值系統

論文名稱： 使用高效率液體層析法和毛細管電泳法來分析人參和靈芝的有效成分 頁數：139


校系(所)組別： 淡江大學化學學系碩士班

畢業時間及提要別： 八十七學年度第一學期 碩士學位論文提要

研究生： 吳淑惠 指導教授： 薛文發 博士

論文提要內容：

自古以來人參和靈芝在漢藥中佔有極重要的地位，具有各種療效，甚至作為抗癌治療試劑，探討人參、靈芝的有效成分對其醫藥應用上有很大的幫助。
傳統人參、靈芝有效成分的抽取方法包含有溶劑的多次萃取或吸附劑萃取，此種萃取方法相平衡需時甚久且步驟繁雜。本論文利用Sep-Pak C18 Cartridges的固相萃取方法來進行人參、靈芝樣品分析的前處理，可以達到分離、純化及濃縮的作用，是一種簡單、快速且具有再現性的萃取法。
經Sep-Pak C18 Cartridges萃取所得的人參有效成分人參皂和靈芝有效成分Ganoderic acid應用高效率液體層析法及毛細管電泳法進行分析。毛細管電泳是一種簡單、分析時間短、分離效率高、樣品用量少的分析方法。使用高效率液體層析法已可有效分離人參皂Re/Rg1、Rb1、Rc、Rb2、Rd及Ganoderic acid A、B、C、D。膠束電動毛細管層析法亦可在10分鐘內完成人參皂的部分分離，至於Ganoderic acid的分離則尚需進一步的探討。

Title of Thesis: The Analyses of Ginsenosides from Ginseng Drug Preparations and Ganoderic Total Pages:139
Acid from Ganoderma lucidum by High-performance Liquid Chromatography
and Capillary Electrophoresis

Key Word: Sep-Pak C18，Ginseng，Ginseng drug preparations，Ganoderma lucidum，High-performance liquid chromatography，Capillary electrophoresis

Name of Institute:Graduate Institute of Chemistry, Tamkang University

Graduate Date: January, 1999 Degree Conferred: Master

Name of Student: Shu-Hui Wu Advisor: Dr. Wen-Fa Sye
吳淑惠 薛文發

Abstract:
Ginseng and Ganoderma lucidum play important roles in Chinese medicine. Ginseng has widely been used in traditional Chinese medicine. There are many reports of pharmacological and chemical studies on ginseng saponins. The fungus Ganoderma lucidum has been used in traditional oriental medicine for the treatment of various diseases and as an antitumor agent. The study of the effective components contained in Ginseng and Ganoderma lucidum helps a lot in their medical applications. Common preliminary procedures for the isolation of ginsenosides from ginseng drug preparations and ganoderic acids from Ganoderma lucidum involve solvent extraction or extraction by adsorbents. Such methods demand a great deal of working time because of the phase separation and experimental procedures. In this thesis, solid phase extractions of Sep-Pak C18 Cartridges were used to the extractions of ginsenosides and ganoderic acids. An extraction using Sep-Pak C18 Cartridges is a simple, rapid and reproducible methods. Sep-Pak C18 Cartridges provide a superior technique for the sample cleanup and concentration. After solid phase extractions, the extracts were analyzed by high-performance liquid chromatography and capillary electrophoresis. Capillary electrophoresis is becoming increasingly recognized as an important analytical separation technique as a result of its speed, high reproducibility and high efficieny. As a result, ginsenosides Re/Rg1、Rb1、Rc、Rb2、Rd from ginseng drug preparations and ganoderic acids A、B、C、D from Ganoderma lucidum can be effectively separated by high-performance liquid chromatography. The use of micellar electrokinetic capillary chramatography can also partly separate ginsenosides in ten minutes. As for the separation of ganoderic acids, we need further research work.

目 錄

壹、緒論 ………………………………………………………………………………………………………1
一、人參、靈芝之簡介……………………………………………………………………………………1
二、人參、靈芝有效成分之抽取方法 …………………………………………………………………19
三、人參、靈芝有效成分的分析 ………………………………………………………………………28
四、毛細管電泳原理 ……………………………………………………………………………………32
貳、實驗 ………………………………………………………………………………………………………49
一、藥品 …………………………………………………………………………………………………49
二、儀器 …………………………………………………………………………………………………50
三、人參、靈芝樣品的製備方法 ………………………………………………………………………51
(一)人參樣品的製備 …………………………………………………………………………………51
(二)靈芝樣品的製備 …………………………………………………………………………………51
四、高效率液體層析法(HPLC)之操作條件 ……………………………………………………………52
五、毛細管電泳法(CE)之操作條件 ……………………………………………………………………53
(一)緩衝溶液之製備及改變緩衝溶液濃度之操作條件……………………………………………53
(二)改變分離電壓……………………………………………………………………………………54
(三)改變分離緩衝溶液pH值 ………………………………………………………………………55
(四)改變界面活性劑SDS濃度 ………………………………………………………………………55
(五)添加第二種界面活性劑Brij-35…………………………………………………………………56
(六)添加不同比例的有機修飾劑(Organic Modifier) ………………………………………………57
(七)GINSANAO-G115之人參皂成分鑑定 ………………………………………………………57
參、結果與討論 ………………………………………………………………………………………………59
一、利用Sep-Pak C18 Cartridges抽取人參、靈芝的有效成分…………………………………………59
二、應用HPLC分離人參、靈芝萃取液中成分…………………………………………………………59
三、以HPLC分離人參之成分鑑定………………………………………………………………………60
四、使用傳統有機溶劑與Sep-Pak C18 Cartridges萃取法抽取人參、靈芝有效成分的比較…………61
五、比較GINSANAO-G115人參膠囊之新舊產品的人參有效成分……………………………………63
六、應用毛細管區帶電泳法(CZE)分離GINSANAO-G115人參膠囊的萃取液 ………………………63
(一)改變分離緩衝溶液濃度之分離結果……………………………………………………………63
(二)改變分離電壓之分離結果 ……………………………………… ……………………………65
(三)改變分離緩衝溶液pH值之分離結果 ……………………………………… …………………65
七、應用膠束電動毛細管層析法(MEKC)分離GINSANAO-G115人參膠囊的萃取液 ………………66
(一)比較不同濃度的界面活性劑SDS對分離結果的影響 …………………………………………66
(二)添加第二種界面活性劑Brij-35對分離結果的影響……………………………………………66
八、添加有機修飾劑(Organic modifier)以改善MEKC對GINSANAO-G115人參成分之分離 ………67
九、MEKC對GINSANAO-G115人參成分分離結果之人參皂成分鑑定……………………………67
十、應用CZE分離台灣靈芝與森の靈芝的萃取液 ……………………………………………………68
十一、應用MEKC分離台灣靈芝與森の靈芝的萃取液 ………………………………………………69
肆、結論 ………………………………………………………………………………………………………70
伍、參考資料…………………………………………………………………………………………………134

參 考 資 料

(1)張志純譯, 人參的科學研究 , P. 1-6, 徐氏基金會, 台北 (1983)
(2)大韓民國專賣廳著, 劉華亭譯, 高麗人參 , P. 45-48, 青春出版社 (1987)
(3)許溢輯, 高麗人參功效摘要集 , P. 11, 韓國人參煙草研究所, 韓國 (1983)
(4)甘偉松, 藥用植物學 , P. 408-411, 國立中國醫藥研究所, 台北 (1975)
(5)張志純譯, 人參的科學研究 , P. 141-151, 徐氏基金會, 台北 (1983)
(6)顏正華, 中藥學 , P. 729, 知音出版社 (1991)
(7)張志純譯, 人參的科學研究 , P. 31-32, 徐氏基金會, 台北 (1983)
(8)顏正華, 中藥學 , P. 726, 知音出版社 (1991)
(9)顏正華, 中藥學 , P. 730-731, 知音出版社 (1991)
(10)田明, 人參是百補之王 , P. 210-211, 元氣齋出版社 (1992)
(11)大韓民國專賣廳著, 劉華亭譯, 高麗人參 , P. 82-116, 青春出版社 (1987)
(12)木崎國嘉著, 陳鶴芳譯, 藥效驚人的高麗人參 , P. 31-71, 永大書局 (1982)
(13)大韓民國專賣廳著, 劉華亭譯, 高麗人參 , P. 82-116, 青春出版社 (1987)
(14)太田富藏著, 劉華亭譯, 鍺可以改善癌體質 , P. 42-58, 青春出版社 (1986)
(15)丹羽芳男著, 梁惠珠譯, 鍺可以治療現代病 , P. 82-91, 青春出版社 (1986)
(16)Carrigues, S., Ann. Chem. Pharm. , 90, 231 (1854)
(17)張志純譯, 人參的科學研究 , P. 33-35, 徐氏基金會, 台北 (1983)
(18)章關德著, 人參的研究及栽培 , P. 169-170, 淑馨出版社 (1989)
(19)M. Nagai, T. Ando, O. Tanaka, and S.Shibata, Tetrahedron Letters , 40, 4797 (1966)
(20)M. Nagai, T. Ando, O. Tanaka, and S.Shibata, Tetrahedron Letters , 37, 3579 (1967)
(21)S. Shibata, O. Tanaka et al., Tetrahedron Letters , 3, 207 (1965)
(22)章關德著, 人參的研究及栽培 , P. 171-178, 淑馨出版社 (1989)
(23)J. Zhou, O. Tanaka et al., Chem. Pharm. Bull. , 29(10), 2844-2850 (1981)
(24) 許溢輯, 高麗人參功效摘要集 , P. 53-55, 韓國人參煙草研究所, 韓國 (1983)
(25)W. F. Sye and P. F. Tsai, J. Chin. Chem. Soc. , 34, 1-6 (1987)
(26)T. Kaku, and Y. Kawashima, Arzneim-Forsch./Drug Res. , 30(1), 936-943 (1980)
(27)F. Soldati and O. Sticher, Planta Med. , 39, 348-357 (1980)
(28)H. R. Schulten and F. Soldati, J. Chromatogr. , 212, 37-49 (1981)
(29)E. M. Martinelli, Recent Developments in Mass Spectro-metry in Biochemistry, Medicine and Environmental Research , 7, 307-314 (1981)
(30)S. E. Chen, E. J. Staba, S. Taniyasu, R. Kasai, and O Tanaka, Planta Med. , 42, 406-411 (1981)
(31)S. YaHara, K. Kaji, and O. Tanaka, Chem. Pharm. Bull. , 27(1), 88-92 (1979)
(32)顏正華, 中藥學 , P. 734-736, 知音出版社 (1991)
(33)H. Matsunaga, M. Katano, H. Yamamoto, M. Mori, and K. Takata, J. Chromatogr. , 481, 368-372 (1989)
(34)劉國柱編著, 現代科學看靈芝 , P. 14-15, 雙利實業有限公司, 台北 (1990)
(35)蔡宗統, 廖淑女編著, 靈芝之臨床應用與市場的現況 , P. 41-52, 參雲出版社, 台中 (1982)
(36)蔡宗統, 廖淑女編著, 靈芝之臨床應用與市場的現況 , P. 37-41, 參雲出版社, 台中 (1982)
(37)有地滋博士著, 高陽堂譯, 靈芝的神妙效用 , P. 199-200, 王家出版社, 台南 (1985)
(38)有地滋博士著, 高陽堂譯, 靈芝的神妙效用 , P. 4-6, 王家出版社, 台南 (1985)
(39)劉國柱編著, 現代科學看靈芝 , P. 25-26, 雙利實業有限公司, 台北 (1990)
(40)蔡宗統, 廖淑女編著, 靈芝之臨床應用與市場的現況 , P. 69-72, 參雲出版社, 台中 (1982)
(41)H. Kohda, W. Tokumoto, K. Sakamoto, M. Fujii, Y. Hirai, K. Yamasaki, Y. Komoda, H. Nakamura, S. Ishihara, and M. Uchida, Chem. Pharm. Bull. , 33, 1367 (1985)
(42)J. O. Toth, B. Luu, and G. Ourisson, Tetrahedron Lett. , 24, 1081 (1983)
(43)T. Nishitoba, H. Sato, S. Shirasu, and S. Sakamura, Argic. Biol. Chem. , 50(8), 2151 (1986)
(44)W. F. Sye, J. Chin. Chem. Soc. , 38(2), 179 (1991)
(45)劉國柱編著, 現代科學看靈芝 , P. 109-195, 雙利實業有限公司, 台北 (1990)
(46)劉國柱編著, 現代科學看靈芝 , P. 207-227, 雙利實業有限公司, 台北 (1990)
(47)T. Namba and J. Hase et al., Planta Med. , 25, 28 (1974)
(48)SEP-PAKO C18 Cartridges－biological applications, Waters Associates, Milford, MA (February 1981)
(49)SEP-PAKO C18 Cartridges Application Bibliography, Waters Associates, Milford, MA (November 1981)
(50)SEP-PAKO C18 Cartridges Streamline Sample Preparation, Waters Associates, Milford, MA (February 1981)
(51)W. F. Sye and C. T. Cheng, J. Chin. Chem. Soc. , 32, 59 (1985)
(52)Simplify Concentration of Trace Organics from Water, Waters Associates, Milford, MA (October 1979)
(53)L. Needham, D. Paschal, Z. J. Rollen, J. Liddle, and D. Bayse, Journal of Chromatographic Science , 17, 87-90 (1979)
(54)R. M. Riggin and C. C. Howard, Anal. Chem. , 51(2), 210-214 (1979)
(55)R. E. Subden, R. G. Brown, and A. C. Noble, Journal of Chromatography , 166, 310-312 (1978)
(56)G. J. Diebold, N. Karny, and R. N. Zare, Anal. Chem. , 51(1), 67-69 (1979)
(57)M. J. Fasco, M. J. Cashin, and L. S. Kaminsky, Journal of Liquid Chromatography , 2(4), 565-575 (1979)
(58)R. A. Hartwick, D. Van Haverbeke, M. Mckeag, and P. R. Brown, Journal of Liquid Chromatography , 2(5), 725-744 (1979)
(59)A. Tiselius, Faraday Soc. , 33, 524 (1937)
(60)S. Hjerten, Chromatogr. Rev. , 9, 122 (1967)
(61)J. W. Jorgenson and K. D. Lukacs, Anal. Chem. , 53, 1298 (1981)
(62) J. W. Jorgenson and K. D. Lukacs, Science , 222, 266 (1983)
(63)S. Terabe, K. Otsuka, K. Ichikawa, A. Tsuchiya, and T. Ando, Anal. Chem. , 56, 111 (1984)
(64)S. F. Y. Li, Capillary Electrophoresis - Principle, practice and application , P. 1-4, Journal of Chromatography Library, Elsevier Scientific Publishers, Amsterdam (1992)
(65)P. Jandik and G. Bonn, Capillary electrophoresis of small molecules and ions , VCH, New York (1993)
(66)S. J. Sheu, Chemistry , 54(3), 55 (1996)
(67) J. H. Paul Yu, Chemistry , 54(3), 49 (1996)
(68)J. W. Jorgenson and K. D. Lukacs, J. Chromatogr. , 218, 209 (1981)
(69)D. N. Heiger著, 孫亦梁等譯, High Performance Capillary Electrophoresis - An Introduction , P. 11-17, Hewlett-Packard GmbH, Germany (1992)
(70)J. W. Jorgenson and K. D. Lukacs, Journal of High Resolution Chromatography & Chromatography Communications , 4, 230 (1981)
(71)A. G. Ewing, R. A. Wallingford, and T. M. Olefirowicz, Anal. Chem. , 61(4), 292A (1989)
(72)D. N. Heiger著, 孫亦梁等譯, High Performance Capillary Electrophoresis - An Introduction , P. 74-80, Hewlett-Packard GmbH, Germany (1992)
(73)D. N. Heiger著, 孫亦梁等譯, High Performance Capillary Electrophoresis - An Introduction , P. 89-101, Hewlett-Packard GmbH, Germany (1992)
(74)D. N. Heiger著, 孫亦梁等譯, High Performance Capillary Electrophoresis - An Introduction , P. 1-5, Hewlett-Packard GmbH, Germany (1992)
(75)D. A. Skoog and J. J. Leary, Principles of Instrumental Analysis, fourth edition, P. 675-679, Saunders College Publishing, U. S. A. (1992)
(76)S. F. Y. Li, Capillary Electrophoresis - Principle, practice and application , P. 4-12, Journal of Chromatography Library, Elsevier Scientific Publishers, Amsterdam (1992)
(77) L. J. Cline Love, J. G. Habarta, and J. G. Dorsey, Anal. Chem. , 1133a (1984)
(78)K. Altria, T. Kelly, and B. Clark, LC‧GC Asia Pacific , 1(2), 24 (1998)
(79)T. de Boer and K. Ensing, J. Chromatogr. A , 788, 212 (1997)
(80)D. N. Heiger著, 孫亦梁等譯, High Performance Capillary Electrophoresis - An Introduction , P. 42, Hewlett-Packard GmbH, Germany (1992)
(81)M. -L. Riekkola et al., J. Chromatogr. A , 792, 13 (1997)
(82)H. T. Rasmussen, L. K. Goebel and H. M. McNair, J. Chromatogr. , 517, 549 (1990)
(83)E. S. Ahuja, E. L. Little, K. R. Nielsen and J. P. Foley, Anal. Chem. , 67, 26 (1995)