五爪金英（Tithonia diversifolia (Hemsl.) A. Gray）是廣泛被民間使用的草藥，最常做為養肝、退火的苦茶成分之一，曾有文獻指出五爪金英對多種癌細胞株有抗癌的效果；五爪金英其主要活性成分為Tagitinin C。
本研究將中草藥五爪金英甲醇萃取物(TDM)純化出來的Tagitinin C，經細胞實驗發現對於人類腦癌細胞U373及人類肝癌細胞Hep-G2具有毒殺效果。實驗結果顯示，針對U373及Hep-G2，加入TDM後24小時後結果發現其IC50分別為59.2 ± 3.7µg/mL和40.0 ± 2.0µg/mL。針對具有抗癌效果之TDM經過管柱分離，分離出各種不同極性的0~5% EtOAc/Hexane (TDM-EA-a), 10~20% EtOAc/Hexane (TDM-EA-b), 20~30% EtOAc/Hexane (TDM-EA-c), 30~60% EtOAc /Hexane (TDM-EA-d), 60~100% EtOAc/Hexane (TDM-EA-e), 1~100% MeOH/ EtOAc (TDM-EA-f)，最後將TDM-EA-b、c、d利用半製備級HPLC進行分離與純化，純化後的物質重複以上細胞實驗，確認所純化的物質為抗癌有效成份，接著以核磁共振光譜(NMR)結構鑑定，結果發現有效成份為Tagitinin C其對於U373及Hep-G2的IC50分別約為6.1 ± 0.1µg/ mL和2.0±0.1µg/mL。
在確定有效成分為Tagitinin C後，在U373發現caspase 3並沒有活化，另一方面經由流式細胞儀分析細胞凋亡也是沒有細胞凋亡的現象出現，然而在poly(ADP-ribose) polymerase-1 (PARP-1), ULK1, LC3-I, LC-II, p-p38則是有表現，其死亡的方式可能是走autophagy的模式；在Hep-G2發現caspase 3 和caspase 8皆有活化，流式細胞儀分析的結果：Tagitinin C處理後的Hep-G2其細胞週期中的次G14期(Sub-G1) 比例增加，故其殺死Hep-G2的方式可能是走細胞凋亡的模式。在以上的實驗可推斷Tagitinin C對於不同的癌細胞造成毒殺的方式會有所不同，未來有助於我們探討癌細胞被Tagitinin C殺死的機制。

Tithonia diversifolia are used in bitter tea and traditional medicine in Taiwan. Pharmacological studies of Tithonia diversifolia showed that it has anti-diabetic, anti-malarial, anti-inflammatory, analgesic, and cancer chemopreventive activity. They are particularly rich in sesquiterpenoids and flavonoids and these sesquiterpenoids were evaluated for their potential as cancer chemopreventive agents in cell culture. Tagitinin C is the major sesquiterpenoid compound in Tithonia diversifolia. But their mechanisms of anti-cancer activity are still unknown. The Tithonia diversifolia methanolic extract (TDM), which showed anti-proliferative activity against human glioblastoma U373 and human hepatoma Hep-G2 cells with an IC50 value 59.2 ± 3.7µg/mL and 40.0 ± 2.0µg/mL, individually, was passed through silica gel chromatography and successively eluted with 0~5% EtOAc/Hexane (TDM-EA-A), 10~20% EtOAc/Hexane (TDM-EA-B), 20~30% EtOAc/Hexane (TDM-EA-C), 30~60% EtOAc/Hexane (TDM-EA-D), 60~100% EtOAc/Hexane (TDM-EA-E), 1~100% MeOH/ EtOAc (TDM-EA-F). Cytotoxicity- guided subfractions TDM-EA-B, TDM-EA-C, and TDM-EA-D which exhibited a comparatively higher anti-proliferative activity were isolated by semipreparative HPLC and then were proceeded structural identification and determination with 1H NMR. The isolated active compound was Tagitinin C, a kind of sesquiterpenoids. The IC50 was 6.1 ± 0.1µg/mL in U373 and 2.0±0.1µg/mL in Hep-G2 separatively treated with the isolated Tagitinin C from TDM. In flow cytometric analysis and caspase 3 expression, the results showed anti-proliferative effect was not induced by apoptosis in U373 after treatment of the isolated Tagitinin C. In, and poly(ADP-ribose) polymerase-1 (PARP-1), ULK1, LC3-I, LC-II, p-p38 expression, the anti-proliferation in U373 induced by Tagitinin C isolated from TDM was likely via autophagy. In caspase 3, caspase 8 protein expression and flow cytometric analysis , the results showed anti-proliferative effect in Hep-G2 was induced by apoptosis after treatment of the isolated Tagitinin C.
In conclusion, the results showed anti-proliferative effect was not induced by apoptosis but induced by autophagy in U373, but the anti-proliferation in Hep-G2 was via apoptosis after treatment of the isolated Tagitinin C. Although many components in anti-cancer and their mechanism were further studied in the future, this is the first study to reveal that Tagitinin C isolated from TDM have anti-glioblastoma U373 and anti-hepatoma Hep-G2 potential .

Bergamini E., Cavallini G., Donati A., Gori Z. The anti-ageing effects of caloric restriction may involve stimulation of macroautophagy and lysosomal degradation, and can be intensified pharmacologically.
Biomed Pharmacother 2003; 57: 203-208.

Bras M., Queenan B., Susin S A. Programmed cell death via mitochondria: different modes of dying. Biochemistry(Mosc) 2005;70(2):231-239.

Chiang LC., Cheng HY., Chen CC., Lin CC. In vitro anti-leukemic and antiviral activities of traditionally used medicinal plants in Taiwan. The American Journal of Chinese Medicine 2004;32(5):695-704.

Cos P., Hermans N., De BT., Apers S., Sindambiwe JB., Witvrouw M., De CE., Vanden BD., Pieters L., Vlietinck AJ. Antiviral activity of Rwandan medicinal plants against human immunodeficiency virus type-1 (HIV-1).Phytomedicine 2002; 9: 62-68.

Degterev A., Boyce M., Yuan J .A decade of caspases. Oncogene 2003; 22: 8543-8567.

do Céu de Madureira M, Martins AP, Gomes M, Paiva J,Proença da Cunha A, do Rosário V.Anti-malarial activity of medicinal plants used in traditional medicine in S. Tomé and Príncipe islands. J Ethnopharmacol 2002; 81: 23-29.

Dorn BR., Dunn WA Jr., Progulske-Fox A. Bacterial interactions with the autophagic pathway.Cell Microbiol 2002; 4: 1-10.

Dunn W A., Jr.Autophagy and related mechanisms of lysosome-mediated protein degradation. Trends Cell Biol 1994; 4: 139-143.

Elufioye TO., Agbedahunsi JM. Antimalarial activities of Tithonia diversifolia (Asteraceae) and Crossopteryx febrifuga (Rubiaceae) on mice in vivo.Ethnopharmacol 2004; 93: 167-171(2004).

Elufioye, TO., Agbedahunsi JM., Adesanya SA. 8-(2-methlybutanoyl) -3,10 epoxy-, 8-dihydroxyl-4, 11(13)-germacridien-12, 6-olide, a new sesquiterpene lactone from Tithonia diversifolia. Nigerian Journal of Natural Products and Medicine 2004;8: 74-76.

Goffin E., da Cunha AP., Ziemons E., Tits M., Angenot L., Frederich M. Quantification of tagitinin C in Tithonia diversifolia by reversed-phase high-performance liquid chromatography.Phytochem Anal 2003; 14(6):378-80.

Goffin E., Ziemons De Mol, P., De Madureira MD., Martins AP., Da Cunha AP.,Philippe G., Tits,M., Angenot, I., Fredrich, M., . In vitro antiplasmoidal activity of Tithonia diviersifolia and identification of its main active constituents: tagitinin C. Planta Medica 2002; 68:543-545.

Gu JQ., Gills JJ., Park EJ., Mata-Greenwood E., Howthorne ME., Axelrod F., Chavez PI., Fong HHS., Kinghorn AD. Sesquiterpenoids from Tithonia diversifolia with potential cancer chemopreventive activity . Journal of natural products 2002; 65: 532-536.

Hetts SW.To die or not to die: an overview of apoptosis and its role in disease. The Journal Of the American Medical Association 1998; 279:1699-1700.

Kabeya Y., Mizushima N., Ueno T., Yamamoto A., Kirisako T., Noda T., Kominami E,. Ohsumi Y., Yoshimori T. LC3, a mammalian homologue of yeast Apg8p, is localized in autophagosome membranes after processing. The EMBO Journal 2000;19(21):5720-5728.

Kerr JF., Searle J. A suggested explanation for the paradoxically slow growth rate of basal-cell carcinomas that contain numerous mitotic figures. The Journal of Pathology 1972 ;107(1):41-44.

Kirkegaard K., Taylor MP., Jackson WT. Cellular autophagy: surrender, avoidance and subversion by microorganisms. Nature Reviews Microbiology 2004;2: 301-314.

Kuroda M., Yokosuka A., Kobayashi R., Jitsuno M., Kando H., Nosaka K., Ishii H., Yamori T., Mimaki Y.Sesquiterpenoids and flavonoids from the aerial parts of Tithonia diversifolia and their cytotoxic activity. Chemical & Pharmaceutical Bulletin 2007; 55(8): 1240-1244 .

Kuwana T., Mackey MR., Perkins G., Ellisman MH, Latterich M., Schneiter R., Green DR., Newmeyer DD. Bid., Bax, and lipids cooperate to from superamolecular openings in the outer mitochondrial membrane. Cell 111: 331-342, 2002 .

Levine B., Kroemer G. Autophagy in the pathogenesis of disease. Cell 2008; 132: 27-42.

Levine B., Klionsky DJ. Development by self-digestion: molecular mechanisms and biological functions of autophagy. Developmental cell 2004;6 (4):463~477.

Longo VD., Finch CE. Evolutionary medicine: from dwarf model systems to healthy centenarians? Science 2003; 299:1342-1346.

Majeski A E., Dice JF. Mechanisms of chaperone-mediated autophagy. The International Journal of Biochemistry & Cell Biology 2004;36 (12):2435-2444.

Massey A., Kiffin R.,Cuervo AM. Pathophysiology of chaperone -mediated autophagy. The International Journal of Biochemistry & Cell Biology 2004 2420-2434.

Meléndez A., Tallóczy Z., Seaman M., Eskelinen EL., Hall DH., Levine B. Autophagy genes are essential for dauer development and life-span extension in C. elegans. Science 2003; 301:1387-1391.

Miura T., Furuta K., Yasuda A., Iwamoto N., Kato M., Ishihara E., Ishida T., Tanigawa K. Antidiabetic effect of nitobegiku in KK-Ay diabetic mice. The American Journal of Chinese Medicine 2002; 30: 81-86.

Miura T., Nosaka K., Ishii H., Ishida T. Antidiabetic effect of Nitobegiku, the herb Tithonia diversifolia, in KK-Ay diabetic mice. Biological & Pharmaceutical Bulletin 2005;28:2152-2154.

Nabin C Baruah., Ram P Sharma.,K. P. Madhusudanan., Gopalakrishna Thyagarajan.,Werner Hem., Ramaswamy Murari. Sesquiterpene Lactones of Tithonia diversifolia. Stereochemistry of theTagitinins and Related Compounds'. The Journal of Organic Chemistry 1979; 44: 1831-1835.

Owoyele VB., Wuraola CO., Soladoye AO., Olaleye SB. Studies on the anti-inflammatory and analgesic properties of Tithonia diversifolia leaf extract. The Journal of Ethnopharmacology 2004; 90: 317-321.

Pereira PS., Dias DA., Vichnewski W., Nasi AM., Herz W. Sesquiterpene lactones from Brazilian Tithonia diversifolia.Phytochemistry 1997; 45: 1445-1448.

Raghwendra Pal ., Kulshreshtha DK., Rastogi RP. Antileuckemic and other constituents of Tithonia tagitiflora desf 〔J〕. Journal of Pharmaceutical Science 1976a ;65 (6) : 918-920.

Reggiori F., Klionsky DJ. Autophagy in the eukaryotic cell. Eukaryot Cell 2002; 1: 11-21.

Rungeler P., Lyss G., Castro V., Mora G., Pahl H. L., Merfort I. Study of three sesquiterpene lactones from Tithonia diversifolia on their anti-inflammatory activity using the transcription factor NF-kappa B and enzymes of the arachidonic acid pathway as targets. Planta Medica 1998; 64: 588-593.

Taiwo LB.,Makinde JO. Influence of water extract of Mexican sunflower (Tithonia diversifolia) on growth of cowpea (Vigna unguiculata). African Journal of Biotechnology 2005;4:355-360.

Tona L., Kambu K., Mesia K., Cimanga K., Apers S., De Bruyne T., Pieters L., Totté J., Vlietinck AJ. Biological screening of traditional preparations from some medicinal plants used as antidiarrhoeal in Kinshasa, Congo.Phytomedicine 1999;6:59-66.

Tona L., Kambu K., Ngimbi N., Mesia K., Penge O., Lusakibanza M., Cimanga K., De Bruyne T., Apers S., Totte J., Pieters L., Vlietinck AJ. Antiamoebic and spasmolytic activities of extracts from some antidiarrhoeal traditional preparations used in Kinshasa, Congo.Phytomedicine 2000;7(1):31-38.

Vellai T., Takacs-Vellai K., Zhang Y., Kovacs AL., Orosz L., Müller F. Genetics: influence of TOR kinase on lifespan in C. elegans. Nature2003; 426: 620.

Ziemons E., Goffin E., Lejeune R., da Cunha AP., Angenot L., Thunus L. Supercritical carbon dioxide extraction of tagitinin C from Tithonia diversifolia. Journal of Supercritical Fluids 2005;33:53-59.

陳德順 胡大維 台灣外來觀賞植物名錄 川流出版社印刷所發行，p.321，(1976)。

詹宗憲，中草藥五爪金英在抗癌和抗氧化上的研究。元培科學技術學院，影像醫學研究所，碩士論文，095YUST7770004，(2007)。