中药麒麟竭的活性成分研究
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摘要
麒麟竭又名血竭(Dragon's Blood),是我国传统名贵中药,具有着活血化淤,止血定痛,生肌敛疮,补血之功效,常用于内外诸科及妇科的各种血症。本文应用高速逆流色谱法(HSCCC)分离制备了麒麟竭中的血竭素,同时利用液相色谱质谱联用技术(HPLC/ESI-MS~n)对麒麟竭主要活性成分进行了初步的分析研究。首先,利用逆流色谱法,以正己烷-乙酸乙酯-甲醇-水(2:3:2:3,v/v)为两相溶剂系统,对麒麟竭主要成分进行了分离制备,同时分离得到血竭素及其对应查尔酮结构的混合物。基于血竭素结构与其对应查尔酮结构在不同pH下可发生结构相互转换,实验通过对该混合物进行pH调控,使查尔酮结构全部转变为血竭素结构,从而建立了仅一步分离即可获得高纯度血竭素的逆流色谱制备新方法。其次,运用液质联用技术分析鉴定了麒麟竭中8个主要成分,经柱层析分离纯化,通过核磁共振,高分辨质谱分析确定了其中2个化合物的结构。通过这两个化合物在多级质谱中的特征碎裂规律,并结合文献对照推测了另外6个化合物的可能结构。
此外,本文对血竭的资源分布及麒麟竭在我国的应用进行了简要的总结。
Dragon's Blood from Daemonorops draco (D. draco) is a renowned traditional Chinese medicine. Its therapeutic applications are meant to stimulate blood circulation, control bleeding, heal wounds, promote tissue regeneration, and enrich blood. This study employed high-speed counter-current chromatography (HSCCC) to purify dracorhodin from the crude extract of D. draco. Afterward, high-performance liquid chromatography was combined with electrospray ion trap mass spectrometry (HPLC/ESI-MS~n) to investigate the major bioactive compounds in this crude extract. First, the purification procedure was performed using a two-phase solvent system composed of n-hexane/ethyl acetate/methanol/water (2:3:2:3, v/v) and a mixture of dracorhodin and its corresponding chalcone was obtained together. Since the transformation between these two compounds can be interconverted by pH changes, dracorhodin was obtained at very high purity by involving pH modulation to the isolated mixture. Thus, an HSCCC method was successfully established for purification of dracorhodin directly from the extract of D. draco. Second, the structural elucidation of the major compounds in the crude extract of D. draco have been determined by HPLC/(+) ESI-MS~n. A total of eight compounds were investigated, two of which were isolated by column chromatography and identified by NMR and ESI-FTICR-MS. Then, the structures of the other six compounds were tentatively proposed based on their mass data compared with those of the two isolated compounds, as well as those found in literature.
In addition, the distribution of Dragon's Blood and the application of its resin from D. draco were summarized briefly in this thesis.
此外,本文对血竭的资源分布及麒麟竭在我国的应用进行了简要的总结。
Dragon's Blood from Daemonorops draco (D. draco) is a renowned traditional Chinese medicine. Its therapeutic applications are meant to stimulate blood circulation, control bleeding, heal wounds, promote tissue regeneration, and enrich blood. This study employed high-speed counter-current chromatography (HSCCC) to purify dracorhodin from the crude extract of D. draco. Afterward, high-performance liquid chromatography was combined with electrospray ion trap mass spectrometry (HPLC/ESI-MS~n) to investigate the major bioactive compounds in this crude extract. First, the purification procedure was performed using a two-phase solvent system composed of n-hexane/ethyl acetate/methanol/water (2:3:2:3, v/v) and a mixture of dracorhodin and its corresponding chalcone was obtained together. Since the transformation between these two compounds can be interconverted by pH changes, dracorhodin was obtained at very high purity by involving pH modulation to the isolated mixture. Thus, an HSCCC method was successfully established for purification of dracorhodin directly from the extract of D. draco. Second, the structural elucidation of the major compounds in the crude extract of D. draco have been determined by HPLC/(+) ESI-MS~n. A total of eight compounds were investigated, two of which were isolated by column chromatography and identified by NMR and ESI-FTICR-MS. Then, the structures of the other six compounds were tentatively proposed based on their mass data compared with those of the two isolated compounds, as well as those found in literature.
In addition, the distribution of Dragon's Blood and the application of its resin from D. draco were summarized briefly in this thesis.
引文
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[1]夏鹏飞,丁里玉.血竭的研究进展.河北中医药学报,2006(1):40-42.
[2]Chen J J, Li X Y, Sun C R, Pan Y J, Schlunegger U P. Identification of polyoxypregnane glycosides from the stems of Marsdenia tenacissima by high-performance liquid chromatography/tandem mass spectrometry. Talanta, 2008,77(1): 152-159.
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[4]Cardillo G, Merlini L, Nasini G, Salvadori P. Constituents of dragon's blood .1.Structure and absolute configuration of new optically active flavans. J. Chem.Soc.C, 1971(23): 3967-3970.
[5]Jurd L. Anthocyanins and related compounds. I. Structural transformations of flavylium salts in acidic solutions. J.Org. Chem., 1963,28: 987-991.
[6]Cavaliere C, Foglia P, Gubbiotti R, Sacchetti P, Samperi R, Lagana A.Rapid-resolution liquid chromatography/mass spectrometry for determination and quantitation of polyphenols in grape berries. Rapid Commu. Mass Spectrom,2008, 22(20):3089-3099
[7]Gupta D, Bleakley B, Gupta R K. Dragon's blood: Botany, chemistry and therapeutic uses. J. Ethnopharmacol., 2008, 115(3): 361-380.
[8]Merlini L, Nasini G. Constituents of Dragon's Blood. 2. Structure and oxidative conversion of a novel secobiflavonoid. J. Chem. Soc. Perkin Trans. I, 1976, (14):1570-1576.
[9]Arnone A, Nasini G, Merlini L. Constituents of Dragon's Blood. 3. Dracooxepine,a novel type of biflavonoid. Heterocycles, 1989, 29(6): 1119-1125.
[10]Arnone A, Nasini G, dePava O V, Merlini L. Constituents of Dragon's Blood .5.Dracoflavans B-1, B-2, C-1, C-2, D-1, and D-2, new A-type deoxyproanthocyanidins. J. Nat. Prod., 1997, 60(10): 971-975.
[2] Gupta D, Bleakley B, Gupta R K. Dragon's blood: Botany, chemistry and therapeutic uses. J. Ethnopharmacol., 2008, 115(3): 361-380.
[3] Milburn, M. Dragon's Blood in East and West Africa, Arabia and the Canary Islands. Africa, 1984, 39(3): 486-493.
[4] Cai, X T, Xu, Z F. Studies on the plant origin of Chinese Dragon's blood. Acta Botanica Yunnanica, 1979,1(2): 1-9.
[5] Bensky, D, Gamble, A, Kaptchuk, T J. Chinese Herbal Medicine: Materia Medica,Eastland Press, Seattle, Wash. (USA), 1993.
[6] Carlson, T J S, King, S R. Sangre de Drago (Croton lechleri Muell.-Arg.)-A phytomedicine for the treatment of diarrhoea. Health Notes. Review of Complementary and Integrative Medicine, 2000, 7: 315-320.
[7] Chen, Z P, Cai, Y, Phillipson, J D. Studies on the anti-tumor, antibacterial, and wound-healing properties of Dragon's blood. Planta Medica, 1994, 60(6):541-545.
[8]Pieters, L, De Bruyne, T, Van Poel, B, Vingerhoets Tott'e, J, Vanden Berghe, D,Vlietinck, A. In vivo wound healing activity of Dragon's blood (Croton spp.), a traditional South American drug, and its constituents. Phytomedicine, 1995,2(1):17-22.
[9]Zheng Q A, Chen J T, Zhang Y J, Yang C R, Origin and diffusion of Dragon's Blood, a famous folk herb. Acta Botanica Yunnanica, 2003, suppl, ⅩⅣ: 102-107.
[10]中国药典委员会.中国药典2005版.北京:化学工业出版社,2005,96.
[11]国家中医药管理局.中华本草.上海:上海科学技术出版社,1999.
[12]郑宵蓓,陈科力.麒麟血竭的研究概况.亚太传统医药,2007,(2):35-38.
[13]Brockmann, H, Junge, H. The constitution of the dracorhodin, a new pigment from the "dragon blood". Berichte Der Deutschen Chemischen Gesellschaft,1943,76:751-763.
[14]Brockmann, H, Junge, H, Eckhardt, I. Benzopyrylium compounds, Ⅲ Announcement - Comparison between dracorhodin with similarly built compounds. Berichte Der Deutschen Chemischen Gesellschaft, 1944, 77(5):347-353.
[15]Robertson, A, Whalley, W B. The Pigments of Dragon's Blood Resin .2. A Synthesis of Dracorhodin. Journal of the Chemical Society, 1950: 1882-1884.
[16]Bhrara, S C, Goel, R N, Seshadri, T R, Jain, A C. Constitution + Synthesis of Strobopinin + Cryptostrobin+ Synthesis of Dracorhodin. Indian Journal of Chemistry, 1964, 2(10): 399.
[17]Cardillo G, Merlini L, Nasini G, Salvadori P. Constituents of dragon's blood .1.Structure and absolute configuration of new optically active flavans. J. Chem.Soc. C, 1971(23): 3967-3970.
[18]Rao G S R, Gerhart M A, Lee Ⅲ R T, Mitscher L A, Drake S, Antimicrobial agents from higher plants. Dragon's blood resin. J. Nat. Prod., 1982,45(5):646-648.
[19]纪淑芳,周亚青.血竭素的合成.沈阳药学院学报,1988,5(2):104-106.
[20]高文芳,郑红,王宇双,张志宏,吕角川,张守芳.血竭素的合成.中国医药工业杂志,1989,20(6):247-249.
[21]Xia, M Y, Wang, D, Wang, M W, Tashiro, S, Onodera, S, Minami, M, Ikejima, T.Dracorhodin perchlorate induces apoptosis via activation of caspases and generation of reactive oxygen species. Journal of Pharmacological Sciences,2004, 95(2): 273-283.
[22]Xia, M, Wang, M W, Tashiro, S I, Onodera, S, Minami, M, Ikejima, T.Dracorhodin perchlorate induces A375-S2 cell apoptosis via accumulation of p53 and activation of caspases. Biological & Pharmaceutical Bulletin, 2005, 28(2):226-232.
[23]Xia, M, Wang, M W, Cui, Z, Tashiro, S I, Onodera, S, Minami, M, Ikejima, T.Dracorhodin perchlorate induces apoptosis in HL-60 cells. Journal of Asian Natural Products Research, 2006, 8(4): 335-343.
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