本研究主要是從台灣產軟珊瑚 Cladiella hirsuta 的乙酸乙酯溶劑萃取物中尋找具有生物活性的化學成分。本研究總共分離得到26個天然化合物 (1–26)，其中包括10個新的eunicellin類的化合物 (1–10)、2個新的steroid glycoside類的化合物 (11–12)、 2個新的α-tocopherol類的化合物 (13–14)以及12個已知的化合物；所有化合物的化學結構均由光譜資料分析(NMR﹑IR﹑MS) 以及比對文獻上已知化合物的結構和數據資料來決定。
本研究中，將所獲得的化合物 (1–10)以及(15–23及26) 進行小鼠血癌細胞 (P-388)、 人類慢性白血病细胞 (K562)、人類肺癌細胞 (HT-29) 以及人類肺腺癌細胞 (A549) 的細胞毒殺活性測試與抗發炎活性測試。其中，化合物22對於四種癌細胞株的活性最好，IC50值分別為5.4、4.0、1.6以及13.2 (μM)；抗發炎實驗的部分，化合物11及12對於elastase(彈性蛋白酶)的釋放具有抑制效果，其IC50 (μg/mL)分別為5.55 ± 1.34以及4.12 ± 0.12。

In order to discover for bioactive compounds, we studied the chemical constituents from the organic extracts of soft coral Cladiella hirsuta. This study had led to the isolation of 26 natural products, including ten eunicellin-type diterpenoids (1–10), two steroid glycosides (11–12), and two phenol glycosides (13–14), along with twelve known compounds (15–26). The structures of compounds 1–26 were established by spectroscopic methods and by comparison of the spectral data with those of the related known compounds.
The cytotoxicity of compounds 1–10, 15–23 and 26 against the P388 (murine leukemia), K562 (human erythro myeloblastoid leukemia), HT-29 (human lung adenocarcinoma epithelial cell line) and A549 (human colon adenocarcinoma grade II cell line), compound 22 showed significant cytotoxicity against the all cancer cell lines with IC50 values of 5.4、4.0、1.6 and 13.2 μM, respectively.
On the part of anti-inflammation assay, compounds 11 and 12 were found to show effectively inhibiting activity of elastase release with IC50 values of 5.55 ± 1.34 and 4.12 ± 0.12, respectively.

論文審定書 i
謝辭 ii
中文摘要 iii
英文摘要 iv

第一章、緒論 1
第一節、前言 1
第二節、研究動機 2
第三節、文獻回顧 3
第二章、生物材料與研究方法 39
第一節、 Cladiella hirsuta 樣品採集時間、地點、分類地位及研究流程 39
第二節、 Cladiella hirsuta的分離流程 42
第三章、化合物之結構證明 47
第一節：軟珊瑚Cladiella hirsuta 所分離出之化合物的結構證明 47
(一)、Hirsutalin N (1) 化合物構造之解析 47
(二)、Hirsutalin O (2) 化合物構造之解析 58
(三)、Hirsutalin P (3) 化合物構造之解析 69
(四)、Hirsutalin Q (4) 化合物構造之解析 80
(五)、Hirsutalin R (5) 化合物構造之解析 90
(六)、Hirsutalin S (6) 化合物構造之解析 101
(七)、Hirsutalin T (7) 化合物構造之解析 111
(八)、Hirsutalin U (8) 化合物構造之解析 123
(九)、Hirsutalin V (9) 化合物構造之解析 133
(十)、Hirsutalin W (10) 化合物構造之解析 143
(十一)、Hirsutosterol H (11) 化合物構造之解析 154
(十二)、Hirsutosterol I (12) 化合物構造之解析 165
(十三)、Cladophenol glycoside A (13) 化合物構造之解析 176
(十四)、Cladophenol glycoside B (14) 化合物構造之解析 187
(十五)、3-butanoyloxycladiell-11(17)-en-6,7-diol (15) 化合物構造之解析 201
(十六)、Hirsutalin D (16) 化合物構造之解析 205
(十七)、Simplexin S (17) 化合物構造之解析 209
(十八)、Palmonine D (18) 化合物構造之解析 213
(十九)、Klysimplexin S (19) 化合物構造之解析 217
(二十)、Hirsutalin F (20) 化合物構造之解析 221
(二十一)、Hirsutalin J (21) 化合物構造之解析 226
(二十二)、Sarcomilasterol (22) 化合物構造之解析 231
(二十三)、9,11-Secoergosta-7,24(28)-dien-9-one (23) 化合物構造之解析 236
(二十四)、6’-O-acetyl-3β-pregna-5,20-dienyl-β-galactopyranoside (24) 化合物構造之解析 241
(二十五)、3β-Pregna-5,20-diene-β-D-xylopyranoside (25) 化合物構造之解析 246(二十六)、Apo-9'-fucoxanthinone (26) 化合物構造之解析 251
第二節: 化合物物理性質及圖譜數據整理 255
第四章、生物活性試驗
第一節、生物活性試驗方法 261
(一)、細胞毒殺活性試驗方法 261
(二)、抗發炎活性試驗方法 263
第二節、生物活性試驗結果 266
(一)、細胞毒殺活性試驗結果 266
(二)、抗發炎活性試驗結果 267
第五章、結論 270
第六章、參考文獻 275

1. Ruggieri, G. D. “Drugs from the Sea.” Sceince 1976, 194, 491–497
2. Blunt, J. W.; Copp, B.R.; Hu, W.-P.; Munro, M. H. G.; Northcotec, P. T.; Prinsep, M. R. “Marine Natural Products.” Nat. Prod. Rep. 2009, 26, 170–244
3. Blunt, J. W.; Copp, B.R.; Hu, W.-P.; Munro, M. H. G.; Northcotec, P. T.; Prinsep, M. R. “Marine Natural Products.” Nat. Prod. Rep. 2012, 29, 144–222
4. Molinski, T. F.; Dalisay, D. S.; Lievens, S. L.; Saludes, J. P. “Drug Development from Marine Natural Products.” Nat Rev. Drug Discov. 2009, 8, 69–85
5. 張書明，臺灣產軟珊瑚Cladiella hirsuta 的化學成分及生物活性之研究，國立中山大學海洋生物科技暨資源學系碩士論文，2009年
6. 陳柏維，養殖型軟珊瑚Klyxum simplex與台灣野生型軟珊瑚Cladiella hirsuta所含Eunicellin-Based雙萜化合物及生物活性研究，2011年
7. Parvataneni, R. “Chemical Constituents and Biological Activities of the Soft Corals of Genus Cladiella: A review”Biochem. Syst. Ecol. 2006, 34, 781–789.
8. Rodríguez, A. D.; Cóbar, O. M. “The Briarellins, “New Eunicellin-Based Diterpenoids from A Caribbean Gorgonian, Briareum asbestinum.” Tetrahedron, 1995, 51, 6869–6880.
9. Cai, Y.-S; Yao, L.-G; Di Pascale, A.; Irace, C.; Mollo, E.; Taglialatela-Scafati, O.; Guo, Y.-W. “Polyoxygenated diterpenoids of the eunicellin-type from the Chinese soft coral Cladiella krempfi ”Tetrahedron. 2013, 69, 2214–2219
10. Ciavatta, M. L.; Manzo, E.; Mollo, E.; Mattia, C. A.; Tedesco, C.; Irace, C.; Guo, Y.-W; Li, X.-B; Cimino,G.; Gavagnin, M. “Tritoniopsins A-D, Cladiellane-Based Diterpenes from the South China Sea Nudibranch Tritoniopsis elegans and Its Prey Cladiella krempfi”J. Nat. Prod. 2011, 74 (9), 1902–1907
11. Chen, B.-W; Chang, S.-M; Huang, C.-J; Chao, C.-H; Su, J.-H; Wen, Z.-H; Hsu, C.-H; Dai, C.-F; Wu, Y.-C; Sheu, J.-H. “Hirsutalins A－H, Eunicellin-Based Diterpenoids from the Soft Coral Cladiella hirsuta. ” J. Nat. Prod., 2010, 73 (11), 1785–1791
12. Hassan, H. M.; Khanfar, M. A.; Elnagar, A. Y.; Mohammed, R.; Shaala, L. A.; Youssef, D. T. A.; Hifnawy, M. S.; Sayed, K. A. El “Pachycladins A－E, Prostate Cancer Invasion and Migration Inhibitory Eunicellin-Based Diterpenoids from the Red Sea Soft Coral Cladiella pachyclados” J. Nat. Prod. 2010, 73, 848–853.
13. Xinping, H.; Zhiwei, D.; Xiaobin, Z.; Leenvan, O.; Peter, P.; Wenhan, L.“Krempenes A-D: A Series of Unprecedented Pregnane-Type Steroids from the Marine Soft Coral Cladiella krempfi” Helv. Chim. Acta., 2006, 89, 2020–2026
14. Huang, X.-P.; Deng, Z.-W.; Ofwegen, L.V.; Li, J.; Fu, H.-Z.; Zhu, X.-B.; Lin, W.-H. “Two new Pregnane Glycosides from Soft Coral Cladiella krempfi” J. Asian. Nat. Prod. Res, 2006, 8, 287–291
15. Ahmed, A. F.; Wu, M.-H.; Wu, Y.-C.; Dai, C.-F.; Sheu, J.-H. “Metabolites with Cytotoxic Activity from the Formosan Soft Coral Cladiella australis”J. Chin.Chem. Soc. 2006, 53, 489–494
16. Ahmed, A. F.; Wu, M.-H.; Wang, G.-H.; Wu, Y.-C.; Sheu, J.-H. “Eunicellin-Based Diterpenoids, Australins A-D, Isolated from the Soft Coral Cladiella australis.” J. Nat. Prod. 2005, 68, 1051–1055
17. Zhang, G.-W.; Ma, X.-Q.; Hiroshi, K.; Zhang, C.-X.; Yao, X.-S.; Su, J.-Y.; Zhang, L.-M. “New Hemiketal Steroid from the Soft Coral Cladiella sp.” Org. Lett. 2005, 7, 991–994
18. Liat, C.; Nelly, B.; Yehuda, B.; Yoel, K. “Eunicellin Diterpenes from Two Kenyan Soft Corals.” J. Nat. Prod. 2005, 68, 19-25
19. Athar, A.; Joe, A.; Parvataneni, R. “Cladioxazole: a Novel Sesquiterpene from a Marine Soft Coral of Genus Cladiella.” Tetrahedron Lett. 2003, 44, 6951–6953
20. Christopher, A.; Gray; Michael, T.; Davies-C.; Michael, H.; Schleyer “Cembrane Diterpenes from the Southern African Soft Coral Cladiella Kashmani” J. Nat. Prod. 2000, 63, 1551–1553
21. Wang, M.-Y.; Luo, Y.; Su, J.-Y. “A New Ceramide from the Soft Coral Cladiella humesi Verseveldt” Chin. Chem. Lett. 2000, 11, 783–784
22. Koji, Y.; Narumi, O.; Kenjiro, R.; Tomofumi, M.; Tetsuya, K.; Ryuichi, H.“Bioactive Terpenoids from Octocorallia. “A New Eunicellin-Based Diterpenoid from the Soft Coral Cladiella sphaeroides.” J. Nat. Prod. 1997, 60, 393–396
23. Rao,C. B.; Rao, D. S.; Satyanarayana, C.; Rao, D. V.; Kassuhlke, K. E.; Faulkner, D. J. “New Cladiellane Diterpenes from the Soft Coral Cladiella australis of the Andaman and Nicobar Islands”J. Nat. Prod. 1994, 57, 574–580
24. Nittala, S.; Sarma, R.; Chavakula, I.; Nageswara, R.; Renuka Kadirvelraj, T.; N. Guru, R.; Isao S. “Crystal and Molecular Structure of Sclerophytin F Methyl Ether from the Soft Coral Cladiella krempfi” J. Nat. Prod. 1993, 56, 1977–1980
25. Yasuto, U.; Mitsuaki, K.; Shuji, U.; Yoshimasa F. “Three New Eunicellin-Based Diterpenoids from an Okinawan Cladiella Species of Soft Coral” Tetrahedron Lett. 1992, 33, 1317–1320
26. Yasuto, U.; Mitsuaki, K.; Nakatani, T.; Mitsuaki, K.; Shuji, U.; Yoshimasa, F. “A New Eunicellin-Based Diterpene from An Okinawan Soft Coral Cladiella sp” Tetrahedron Lett. 1989, 30, 3331–3332
27. Kazlauskas, R.; Murphy, P. T.; Wells, R. J. “Two New Diterpenes Related to Eunicellin from A Cladiella Species (Soft Coral)” Tetrahedron Lett. 1977, 52, 4643–4646
28. Bowden, B. F.; Coll, J. C.; Dai, M. C.; “Studies of Australian Soft Corals. XLIII. The Structure Elucidation of a New Diterpene from Alcyonium molle” Aust. J. Chem. 1989, 42, 665–673.
29. Su, J.; Zheng, Y.; Zeng, L.; Pordesimo, Eva O.; Schmitz, F. J.; Hossain, M. B.; Helm, D. V. D. “Patagonicol: A Diterpenoid from the Chinese Soft Coral Alcyonium patagonicum” J. Nat. Prod. 1993, 56, 1601–1604.
30. Kyeremeh, K.; Baddeley, T. C.; Stein, B. K.; Jaspars, M. “A Homologous Series of Eunicellin-Based Diterpenes from Acalycigorgia sp. Characterised by Tandem Mass Spectrometry” Tetrahedron 2006, 62, 8770–8778.
31. Fusetani, N.; Nagata, H.; Hirota, H.; Tsuyuki, T. “Astrogorgiadiol and Astrogorgin, Inhibitors of Cell Division in Fertilized starfish Eggs, from a Gorgonian Astrogorgia sp.” Tetrahedron Lett. 1989, 50, 7079–7082.
32. Rodríguez, A. D.; Cóbar, O. M. “Studies on the Minor Constituents of the Caribbean Gorgonian Octocoral Briareum asbestinum Pallas. Isolation and Structure Determination of the Eunicellin-Based Diterpenoids Briarellins E–I” Chem. Pharm. Bull. 1995, 43, 1853–1858.
33. Ospina, C. A.; Rodríguez, A. D.; Ortega-Barria, E.; Capson, T. L. “Briarellins J–P and Polyanthellin A: New Eunicellin-Based Diterpenes from the Gorgonian Coral Briareum polyanthrs and Their Antimalarial Activity” J. Nat. Prod. 2003, 66, 357–363.
34. Ospina, C. A.; Rodríguez, A. D. “Bioactive Compounds from the Gorgonian Briareum polyanthes. Correction of the structures of Four Asbestinane-Type Diterpenes” J. Nat. Prod. 2006, 69, 1721–1727.
35. Ketzinel, S.; Rudi, A.; Schleyer, M.; Benayahu, Y.; Kashman, Y. “Sarcodictyin A and Two Novel Diterpenoid Glycosides, Eleuthosides A and B, from the Soft Coral Eleutherobia aurea” J. Nat. Prod. 1996, 59, 873–875.
36. De Rosa, S. ; Cimino, G.; De Giulio, A.; Milone, A.; Crispino, A.; Iodice, C. “A New Bioactive Eunicellin-Type Ditepene from the Gorgonian Eunicella cavolini” Nat. Prod. Lett. 1995, 7, 259–265
37. Mancini, I.; Guella, G.; Zibrowius, H.; Pietra, F. “Configuration, Conformation, and Reactivity of Highly Functionalized Eunicellane Diterpenes Isolated from the Gorgonians Eunicella cavolinii and Eunicella singularis from Marseille” Helv. Chim. Acta. 2000, 83, 1561–1575.
38. Ortega, M. J.; Zubía, E.; Salvá, J. “A New Cladiellane Diterpenoid from Eunicella labiata” J. Nat. Prod. 1997, 60, 485–487.
39. Roussis, V.; Fenical, W.; Vagias, C.; Kornprobst, J.-M.; Miralles, J. “Labiatamides A, B, and other Eunicellan Diterpenoids from the Senegalese Gorgonian Eunicella labiata” Tetrahedron 1996, 52, 2735–2742.
40. Kakonikos, C.; Vagias, C.; Roussis, V.; Roussakis, C.; Kornprobst, J.-M. “New Eunicellin Type Diterpenoids from the Gorgonian Coral Eunicella labiata” Nat. Prod. Lett. 1999, 13, 89–95.
41. Kennard, O.; Watson, D. G.; Sanseverino, L. R. D. “The Crystal and Molecular Structure of Eunicellin Dibromide, C28H42O9Br2” Acta Cryst. 1970, 26, 1038.
42. Ortega, M. J.; Zubía, E.; He, H.-Y.; Salvá, J. “New Eunicellin-Based Diterpene from the Gorgonian Eunicella verrucosa” Tetrahedron 1993, 49, 7823–7828.
43. Ortega, M. J.; Zubía, E.; Salvá, J. “Structure and Absolute Configuration of Palmonine F, A New Eunicellin-Based Diterpene from the Gorgonian Eunicella verrucosa” J. Nat. Prod. 1994, 57, 1584–1586.
44. Ochi, M.; Yamada, K.; Futatsugi, K.; Kotsuki, H.; Shibata, K. “Litophynin D and E, Two new Diterpenoids from a Soft Coral Litophyton sp.” Chem. Lett. 1990, 12, 2183–2186.
45. Miyamoto, T.; Yamada, K.; Ikeda, N.; Komori, T.; Higuchi, R. “Bioactive Terpenoids from Octocorallia, I. Bioactive Diterpenoids: Litophynols A and B from the Mucus of the Soft Coral Litophyton sp.” J. Nat. Prod. 1994, 57, 1212–1219.
46. Ochi, M.; Yamada, K.; Futatsugi, K.; Kotsuji, H.; “Litophynin F, G and H, Three new Diterpenoids from a soft coral Litophyton sp.” Heterocycles, 1991, 1, 29–31.
47. Seo, Y.; Rho, J.-R.; Cho, K. W.; Shin, J. “Isolation of Diterpenoids of Cladiellane Class from Gorgonians of the Genus Muricella” J. Nat. Prod. 1997, 60, 171–174.
48. Wang, G.-H.; Sheu, J.-H.; Chiang, M. Y.; Lee, T.-J. “Pachyclavulariaenines A–C, Three Novel Diterpenoids from the Soft Coral Pachyclavularia violacea” Tetrahedron Lett. 2001, 42, 2333–2336.
49. Wang, G.-H.; Sheu, J.-H.; Duh, C.-Y.; Chiang, M. Y. “Pachyaclavulariaenones D–G, New Diterpenoids from the Soft Coral Pachyclavlaria violacea” J. Nat. Prod. 2002, 65, 1475–1478
50. Alam, M.; Sharma, P.; Zektzer, A. S.; Martin, G. E.; Ji, X.; Helm, D. V. D. “Sclerophytin C–F: Isolation and Structures of Four New Diterpenes from the Soft Coral Sclerophytum capitalis” J. Org. Chem. 1989, 54, 1896–1900.
51. Sharma, P.; Alam, M. “Sclerophytin A and B. Isolation and Structures of Novel Cytotoxic Diterpenes from the Marine Coral Sclerophytum capitalis” J. Chem. Soc., Perkin Trans. 1 1988, 2537–2540.
52. Kusumi, T.; Uchida, H.; Ishitsuka, M. O.; Yamamoto, H.; Kakisawa, H. “Alcyonin, a New Cladiellane Diterpene from the Soft Coral Sinularia flexibilis” Chem. Lett. 1988, 6, 1077–1078.
53. Bloor, S. J.; Schmitz, F. J.; Hossain, M. B.; Helm, D. V. D. “Diterpenoids from the Gorgonian Solenopodium stechei” J. Org. Chem. 1992, 57, 1205–1216.
54. Su, J.-H.; Huang, H.-C.; Chao, C.-H.; Yan, L.-Y.; Wu, Y.-C.; Wu, C.-C.; Sheu, J.-H. “Vigulariol, a New Metabolite from the Sea Pen Vigularia juncea” Bull. Chem. Soc. Jpn. 2005, 78, 877–879.
55. Wu, S.-L; J Su, J.-H; Huang, H.-C; Tai, C.-J; Sung, P.-J; Liaw, C.-C; Sheu, J.-H. “Simplexins P–S, Eunicellin-Based Diterpenes from the Soft Coral Klyxum simplex” Mar. Drugs 2012, 10, 1203–1211
56. Chau, V.-M; Cuong, N.-X; Tuan, T.-A; Choi, E.- M; Kim, Y.-H; Van Phan, V. K. “A New 9,11-secosterol from the Vietnamese Sea Soft Coral, Sarcophyton mililatensis, Increases the Function of Osteoblastic MC3T3-E1 cells”. Nat. Prod. Commun. 2007, 11, 1095–1100
57. Anna, M.; Vincenzo, P.; Donato, S. “Two New 9,11-secosterols from the Marine Sponge Spongia officinalis. Synthesis of 9,11-seco-3β,6 α,11-trihydroxy-5 alpha-cholest-7-en-9-one” Steroids. 1992, 57, 344–7.
58. Marville, Kelly I.; Reynolds, William F.; Sealy, R. Lynn; Tinto, Winston F. “Terpenoid Metabolites of the Marine Octocoral Eunicea laciniata.” Heterocycles. 2004, 63, 107–113
59. Shi, Y.-P; Rodriguez, A.D.; Barnes, C. L.; Sanchez, J. A.; Raptis, R. G.; Baran, P. “ New Terpenoid Constituents from Eunicea Pinta.” J. Nat. Prod. 2002, 65, 1232–1241.
60. Yukiko, D.; Masami, I.; Naoko,Y.; Kobayashi, J. “ Isolation of Apo-9'-fucoxanthinone from the Cultured Marine Dinoflagellate Amphidinium sp.” J . Nat. Prod. 1995, 58, 1097–9
61. Charles, L. C.; Tiansheng, L.; Frank, R. F.; Nikolaus, H. F. “Antimycobacterial Cycloartanes from Borrichia frutescens” J. Nat. Prod. 1996, 59, 1131–1136
62. Hwang, T.-L.; Leu, Y.-L.; Kao, S.-H.; Tang, M.-C.; Chang, H.-L. “Viscolin, a New Chalcone from Viscum coloratum, Inhibits Human Neutrophilsuperoxide Anion and Elastase Release Via a cAMP-dependent Pathway” Free Radical. Biol. Med. 2006, 41, 1433–1441.