臺灣博碩士論文加值系統

為了尋找海洋軟珊瑚中活性天然成分，於是研究台灣綠島海域所採集的羽珊瑚Clavularia inflata，因為綠島產羽珊瑚C. inflata之乙酸乙酯粗萃取液經in vitro細胞活性測試，發現對NUGC-3（人類胃癌細胞）及HONE-1（人類鼻咽癌細胞）兩種癌細胞有顯著的細胞毒殺活性。以癌細胞毒殺活性為指標，純化分離有機層萃取物，一共得到五個雙帖類的化合物，其中四個為已知的化合物，分別為（1R*, 12R*）- dolabella- 4（16）, 7, 10- triene- 3, 13- dione （1）、stolonidiol monoacetate （2）、stolonidiol （3）及clavutriolin （5），另外一個為新的化合物，命名為2- acetoxy- stolonidiol （4）。
上述化合物之化學結構均是以各種光譜的證據及衍生物的製備反應加以確定，核磁共振磁譜技術包括一維及二維NMR，如COSY、HMQC、HMBC、NOESY等等。化合物1-5皆為雙環dolabellane骨架雙帖類。化合物1在C-3及C-13位置上接有兩組酮基，C-4位上接有一個環外雙鍵以及在C-7及C-10位上各接有環內雙鍵。化合物2-4在C-7、C-8及C-10、C-11位置上皆接有兩組環氧基，其中化合物2之C-17位上接有一個乙醯基，C-18位上接有一個羥基；化合物3之C-17和C-18位上各接一個羥基；化合物4除了C-17和C-18位上接有羥基外，另在C-2上帶有一個乙醯基。化合物5在C-8、C-17、C-18位置上接有三個羥基，在C-7位置上接有一個氯原子，在C-10、C-11位置上接有一組環氧基。
將所獲得的天然化合物及衍生物進行KB（人類口腔上皮癌細胞）、Hepa59/VGH（人類肝癌細胞）及Hela（人類子宮頸上皮癌細胞）三種癌細胞毒殺活性測試，結果顯示化合物1-3和5對KB癌細胞具有顯著癌細胞毒殺活性，其ED50值各為0.87、3.07、1.35及0.36 mg/ml，化合物4對Hepa癌細胞有抑制活性，其ED50值為1.20 mg/ml；然而，乙醯化的衍生物6和7對三種人類癌細胞均無毒殺活性。

In order to search for bioactive substances from marine organisms,the Taiwanese soft coral Clavularia inflata,collected along the coast of Green Island,Taiwan,was studied because of its significant cytotoxicity against NUGC-3 and HONE-1 cancer cell lines in EtOAc extract. Bioassay-guided fractionation resulted in the isolation of five diterpenoids,including four known compounds, ( 1R*, 12R* ) - dolabella- 4 (16), 7, 10- triene- 3, 13- dione (1),stolonidiol monoacetate (2), stolonidiol (3),clavutriolin (5),and one new compound,2- acetoxy- stolonidiol (4).
Structures of these compounds were determined on the basis of spectroscopic evidence and chemical derivatization. The application of NMR techniques included 1D- and 2D- NMR,such as COSY, HMQC,HMBC and NOESY,etc. Compounds 1-5 are bicyclic dolabellane skeleton diterpenes. Compound 1 has two ketone groups at C-3 and C-13 positions,an exocyclic methylene at C-4 and two double bonds at C-7 and C-10 positions. Compounds 2-4 possess two epoxides at C-7,C-8,C-10,and C-11. Among them,compound 2 has an acetoxyl group at C-17,and a hydroxyl group at C-18. Compound 3 has two hydroxyl groups at C-17 and C-18 positions. Compound 4 has two hydroxyl groups at C-17 and C-18,and an acetoxyl group at C-2. Compound 5 contains three hydroxyl groups at C-8,C-17,and C-18,and a chlorine atom at C-7, and a epoxy moiety at C-10 and C-11 positions.
The ctytotoxicity of these compounds and their derivatives against KB（human oral epidermoid carcinoma）,Hepa（human liver carcinoma）and Hela（human cervix epitheloid carcinoma）cancer cell lines were also measured. Compounds 1-3 and 5 exhibited significant cytotoxicity against KB cancer cell lines with ED50 values of 0.87,3.07, 1.35 and 0.36 mg/ml,respectively,meanwhile, compound 4 exhibited cytotoxicity against Hepa cancer cells with ED50 value of 1.20 mg/ml. However,the acetylated derivatives 6 and 7 were inactive against these cell lines.

壹、緒論
一、前言 1
二、相關化合物之研究回顧 3
三、研究動機與目的 20
貳、實驗程序與方法
一、採樣、分離與純化 22
二、衍生物之製備 25
三、生物活性之篩選 26
四、使用儀器與材料 28
參、結果與討論
一、化合物之結構鑑定 30
化合物（1）之結構解析 30
化合物（2）之結構解析 42
化合物（3）之結構解析 55
化合物（4）之結構解析 67
化合物（5）之結構解析 80
化合物（6）之結構解析 93
化合物（7）之結構解析 106
二、活性測試結果 116
肆、結論 117
伍、參考文獻 119

1.方力行 “珊瑚學-兼論台灣的珊瑚資源” 教育部大學聯合出版委員會，中華民國78年。2.Bowden, B. F.; Braekman, J. C.; Coll, J. C.; Mitchell, S. J.; “A New Sesquiterpene Furan from Soft Corals of the Family Xeniidae and an Examination of Clavularia inflata from North Queensland Waters”, Aust. J. Chem., 1980, 33, 927-932.3.Amico, V.; Oriente, G.; Piattelli, M.; Tringali, C.; Fattorusso, E.; Magno, S.; Mayol, L.; “Diterpenes Based on the Dolabellane Skeleton from Dictyota dichotoma”, Tetrahedron, 1980, 36, 1409-1414.4.Look, S. A.; Fenical, W.; “New Biocyclic Diterpenoids from the Caribbean Gorgonian Octocoral Eunicea calyculata”, J. Org. Chem., 1982, 47, 4129-4134.5.González, A. G.; Martín, J. D.; Norte, M.; Pérez, R.; Weyler, V.; “A New Diterpene from Aplysia dactylomela”, Tetrahedron Lett., 1983, 24, 1075-1076.6.De Rosa, S.; De Stefano, S.; Macura, S.; Trivellone, E.; Zavodnik, N.; “New Dolabellane Diterpenes from the Brown Alga Dilophus fasciola from North Queensland Waters”, Tetrahedron, 1984, 40, 4991-4995.7.Tringali, C.; Piattelli, M.; Nicolosi, G.; “Structure and Conformation of New Diterpenes Based on the Dolabellane Skeleton from a Dictyota Species”, Tetrahedron, 1984, 40, 799-803.8.Rao, C. B.; Pullaiah, K. C.; Surapaneni, R. K.; Sullivan, B. W.; Albizati, K. F.; Faulkner, D. J. J.; “The Diterpenes Dictyota dichotoma from the Indian Ocean”, Org. Chem., 1986, 51, 2736-2742.9.Mori, K.; Iguchi, K.; Yamada, N.; Yamada, Y.; Inouye, Y.; “Stolonidiol, a New Diterpenoid with a Strong Cytotoxic Activity from the Japanese Soft Coral”, Tetrahedron Lett., 1987, 28, 5673-5676.10.Kelecom, A.; Teixeira, V. L.; “New Dolabellane Diterpenes from the Marine Brown Alga Dilophus cervicornis”, Phytochemistry, 1988, 27, 2907-2909.11.Matsuo, A.; Kamio, K.-I.; Uohama, K.; Yoshida, K.-I.; Connolly, J. D.; Sim, G. A.; “Dolabellane Diterpenoids from the Liverwort Odontoschisma denudatum”, Phytochemistry, 1988, 27, 1153-1159.12.Cáceres, J.; Rivera, M. E.; Rodríguez, A. D.; “Palominol, a New Diterpene Based on the Dolabellane Skeleton from the Caribbean Gorgonian Octocorals Eunicea calyculata and Eunicea laciniata”, Tetrahedron, 1990, 46, 341-348.13.Su, J.; Zhong, Y.; Shi, K.; Cheng, Q.; Snyder, J. K.; Hu, S.; Huang, Y. J.; “Clavudiol A and Clavirolide A, Two Marine Dolabellane Diterpenes from the Soft Coral Clavularia viridis”, Org. Chem., 1991, 56, 2337-2344.14.Shin, J.; Fenical, W.; “Structures and Reactivities of New Dolabellane Diterpenoids from the Caribbean Gorgonian Eunicea laciniata”, J. Org. Chem., 1991, 56, 3392-3398.15.Rodríguez, A. D.; Acosta, A. L.; Dhasmana, H.; “The Structure of Palominol by Interpretation of Two-Dimensional NMR Shift Correlation Experiments: Isolation and Structure Determination of Isopalominol”, J. Nat. Prod., 1993, 56, 1843-1849.16.Mohamed, K. M.; Ohtani, K.; Kasai, R.; Yamasaki, K.; “Dolabellane Diterpene Glucosides from Chrozophora obliqua”, Phytochemistry, 1994, 37, 495-500.17.König, G. M.; Wright, A. D.; “New Dolabellanes from the Marine Alga Dictyota pardalis f. pseudohamata”, Tetrahedron, 1994, 50, 8011-8018.18.Rao, C. B.; Trimurtulu, G.; Sreedhara, C.; Rao, D. V.; Bobzin, S. C.; Faulkner, D. J.; “Diterpenes from the Brown Alga Dictyota bartayresiana”, Phytochemistry, 1994, 37, 509-513.19.König, G. M.; Wright, A. D.; “Extensive 1D and 2D NMR and X-ray Studies of Diterpenes Isolation from the Marine Alga Dictyota pardalis f. pseudohamata”, J. Nat. Prod., 1994, 57, 1529-1538.20.Govindan, M.; Govindan, G. N.; Kingston, D. G. I.; “Mechanism-Based Antitumor Screening of Caribbean Marine Organisms: Isolation and Strcture Determination of Novel Diterpenoids from the Gorgonian Eunicea tourneforti”, J. Nat. Prod., 1995, 58, 1174-1184.21.Rodríguez, A. D.; González, E.; González, C.; “Additonal Dolabellane Diterpenes from the Caribbean Gorgonian Octocoral Eunicea laciniata”, J. Nat. Prod., 1995, 58, 226-232.22.Durán, R.; Zubía, E.; Ortega, M. J.; Salvá, J.; “New Diterpenoids from the Alga Dictyota dichotoma”, Tetrahedron, 1997, 53, 8675-8688.23.Hashimoto, T.; Toyota, M.; Koyama, H.; Kikkawa, A.; Yoshida, M.; Tanaka, M.; Takaoka, S.; Asakawa, Y.; “Novel ent-Vibsane- and Dolabellane-type Diterpenoids from the Liverwort Odontoschisma denudatum”, Tetrahedron Lett., 1998, 39, 579-582.24.Duh, C. Y.; Chia, M. C.; Wang, S. K.; Chen, H. J.; EI-Gamal, A. A. H.; Dai, C. F.; “Cytotoxic Dolabellane Diterpenes from the Formosan Soft Coral Clavularia inflata”, J. Nat. Prod., 2001, 64, 1028-1031.25.Alley, M. C.; Scudiero, D. A.; Monks, A.; Hursey, M. L.; Czerwonski, M. J.; Fine, D. L.; Abbott, B. J.; Mayo, J. G.; Shoemark, R. H.; Boyd, M. R.; “Feasibility of Drug Screening with Panels of Human Tumor Cell Lines Using a Microculture Tetrazolium Assay”, Cancer Res., 1988, 48, 589-601.26.Mosmann, T.; “Rapid Colormetric Assay for Celluar Growther and Survival: Application to Proliferation and Cytotoxicity Assays”, J. Immu. Method., 1983, 65, 55-63.27.Xia, Z.; Zhang, Z.; Huang, J. Jie Gou Hua Xuemu, 1986, 5, 263.