摘要
海洋生物生存环境特殊,其次级代谢产物化学结构新颖、种类繁多,是新活性先导化合物的重要来源。为了寻找抗肿瘤、抗氧化活性先导化合物,本论文采用活性追踪的方法开展了4株海洋微生物和2种海洋底栖动物活性次级代谢产物的研究工作。内容包括:不同培养条件下微生物次级代谢产物的比较和目标菌株的选择;活性成分的追踪分离;单体化合物的结构解析;单体化合物的活性初步评价。
在对本实验室已研究过次级代谢产物的20株海洋活性真菌进行培养条件改变摸索研究,发现了2株真菌的粗提物仍然保持抗肿瘤活性,但TLC薄层色谱和高压液相指纹图谱显示其次级代谢产物与原菌株的明显不同,发现两株菌的代谢产物中主要成分极性明显增大以及紫外吸收明显发生改变,于是决定将其作为本论文的主要研究对象。此外,本论文还对2种具有抗肿瘤活性的海洋底栖动物和2株采自韩国沿海具有抗氧化活性的海洋真菌次级代谢产物进行了研究。
对4株目标活性菌进行了大量发酵,对2种海洋动物进行了匀浆和浸提。并对发酵产物和浸提物运用萃取,薄层层析,正相、反相硅胶柱层析,LH-20凝胶柱层析,反相高压液相等化学的分离纯化手段进行活性追踪分离,从土壤青霉菌(Penicillium terrestre)的代谢产物中分离得到29个化合物单体(1-29);从匍匐曲霉(Aspergillus repens)中分离得到了7个单体化合物(30-36);从韩国沿海真菌B382中分离得到6个单体化合物(37-42);从韩国沿海真菌B523中分离得到了6个单体化合物(43-48);从哈氏刻肋海胆(Temnopleurus hardwickii)中分离得到了3个单体化合物(49-51);从星座美洲海鞘(Amaroucium constellatum)中分离得到了9个单体化合物(52-60),共计60个化合物。
继而,利用理化性质和波谱学方法(IR,UV,MS,NMR,X-ray)结合化学反应的方法阐明了这60个化合物的化学结构(化合物结构参见Figure 1),其中发现新化合物20个,包括8个聚gentisyl alcohol类化合物(1-8)(首次报道该类单体的三聚物),3个gentisyl alcohol单体的衍生物(12,18,19),5个Sorbicillin类衍生物(14-17,20),1个玉米赤霉烯酮类化合物(37),3个硫酸酯铵盐类化合物(49-51)。此外还有1个化合物(13)为首次从天然来源中分离得到。
另外发现的其他已知化合物的结构类型还涉及17个苯的衍生物(9-11,22-29,43,47-48,56-58),1个Sorbicillin类衍生物(21),3个甾醇类化合物(33,34,53),2个脑苷酯类化合物(31-32),3个生物碱类化合物(30,35-36),5个玉米赤霉烯酮类化合物(38-42),3个蒽醌类衍生物(44-46),4个核苷类衍生物(52、55、59-60),1个甘油醚类化合物(54)。
利用流式细胞术结合形态学检测、MTT和SRB法,对所分得的新化合物的体外抗肿瘤活性进行了初步评价。新化合物1-8对HL-60、MOLT-4、A-549和BEL-7402四种细胞株都表现出不同程度的抑制活性,其中化合物6对四种癌细胞的活性都较强。在对6的进一步研究中发现,该化合物在10μM浓度下对酪氨酸激酶Src和KDR,抑制率分别为35.9%和31.8%。同时还发现新化合物14、15对HL-60有较弱的抑制活性(IC50分别为9.2和37.8μM),14对A-549有较弱的抑制活性(IC50为39.7μM)。此外,在以DPPH为模型的抗氧化试验中,发现聚酚类化合物(1-8)和单体(12,13)均显示较强的活性(IC50分别为4.3, 4.6, 4.4, 6.2, 5.2, 4.1, 6.3, 2.6, 8.5和9.8μM),明显强于阳性对照物维生素C(IC50为17.4μM)。
综上所述,本文对4株海洋真菌和2种海洋底栖动物的次级代谢产物进行了系统研究,共分离鉴定了60个化合物的结构,包括新化合物20个并推测了其中12个新化合物可能的生合成途径;发现了一类结构新颖的具有抗肿瘤、抗氧化等多重活性gentisyl alcohol聚合体(1-8);最值得一提的是,通过对2株真菌改变培养条件所分离得到的36个化合物与原条件所分得的22个化合物无一重复。因此上述研究既为抗肿瘤新药提供了先导结构,也为今后充分发掘活性菌株次级代谢的潜力,从而获得大量结构新颖的活性次级代谢产物提供实验依据。
Marine microorganisms and invertebrates continue to be an important source of bioactive secondary metabolites. A study on four strains of marine microorganisms and two invertebrates was carried out to investigate the potential anti-tumor and antioxidant compounds. Studies include comparison of microbial metabolites under different culture conditions, selecting aimed strains, fermentation studies, bioassay-guided fractionation, structural elucidation and preliminary evaluation for activities of pure compounds.
After careful comparison of microbial metabolites under different culture conditions, the metabolite patterns of two fungal strains exhibited TLC and HPLC profiles very distinct from those of the original strains, such as higher polarity and different UV absorbance. However, their anti-tumor activity remained. So they were chosen as main objects of this thesis. Furthermore, two invertebrates with anti-tumor activity and two marine fungus of Korean coast with antioxidant activity were selected.
Large-scale fermentations of the active strains and smash of the invertebrates were performed. Following the bioactivity, compounds were isolated and purified by using solvent extraction, silica gel column, Sephadex LH20, PHPLC and etc. From fungus Penicillium terrestre, 29 compounds (1-29) were isolated; from fungus Aspergillus repens, 7 compounds (30-36) were isolated; from fungus B382, 6 compounds (37-42) were isolated; from fungus B523, 6 compounds (43-48) were isolated; from echinus Temnopleurus hardwickii, 3 compounds (49-51) were isolated; from ascidian Amaroucium constellatum, 9 compounds (52-60) were isolated.
By means of physico-chemical properties and spectral analysis (IR, UV, MS, NMR, X-ray, etc.), structures of 60 pure compounds were respectively determined, Among them there are 20 new compounds, including 8 new gentisyl alcohol polymers (1–8), 3 gentisyl alcohol derivatives (12, 18, 19), 5 sorbicillin derivatives (14-17, 20), 1 zearalenone derivative (37), 3 sulfated alkenes (49-51). Furthermore, 1 new natural product (13) was also isolated.
In addition, the types of compounds are involved in benzene derivates (9-11,22-29,43,47-48,56-58), sorbicillin derivative (21), steroides (33,34,53), cerebrosides (31-32), alkaloids (30, 35-36), zearalenone derivatives (38-42), anthraquinones (44-46), nucleosides (52, 55, 59-60), glycerol derivative (54). The antitumor activity against several cancer cell lines of the new compounds was assayed by MTT, SRB and flow cytometry methods. Compounds 1-8 all showed inhibitive activity against HL-60, MOLT-4, A-549, and BEL-7402, and compound 6 showed stronger activity than others. In the continuous research, 6 showed moderate inhibitive activity against protein tyrosine kinases (Src and KDR) by ELISA (inhibitive rate 35.9% and 31.8% on concentration 10μM). In addition, compounds 14 and 15 have weak cytotoxicity against HL-60 with IC50= 9.2 and 37.8μM; 14 also has weak cytotoxicity against A-549 with IC50=39.7μM. When evaluated for their radical scavenging activity against DPPH, compounds 1–8, 12 and 13 showed moderate activity(IC50=4.3, 4.6, 4.4, 6.2, 5.2, 4.1, 6.3, 2.6, 8.5 and 9.8μM respectively. They were all stronger than the positive control (ascorbic acid, IC50=17.4μM).
Summarily, this work obtained sixty compounds from four strains of marine fungus and two invertebrates. Among them, 20 compounds were identified new and the possible biogenetic pathways of 12 new compounds were given. A series of novel gentisyl alcohol polymers with antitumor and antioxidant activity was reported. The most important, none of the 36 compounds isolated from the new cultural condition was the same as the 22 compounds isolated from the original condition. Studies mentioned above provided novel structures for searching leading antitumor compounds, and gave a good example for the deep development of the active strains.
引文
1. Xu X L, Song F H, Wang S J, et al. Dibenzyl Bromophenols with Diverse Dimerization Patterns from the Brown Alga Leathesia nana. J. Nat. Prod. 2004, 67, 1661–1666.
2. Fan X, Xu N J, Shi J G. Bromophenols from the Red Alga Rhodomela confervoides J. Nat. Prod. 2003, 66, 455–458.
3. Ma M, Zhao J L, Wang S J, et al. Bromophenols Coupled with Methyl -Ureidobutyrate and Bromophenol Sulfates from the Red Alga Rhodomela confervoides J. Nat. Prod. 2006, 69, 206–210.
4. Suh S B, Kim J C, Choi Y C, et al. Nature of One-Dimensional Short Hydrogen Bonding: Bond Distances, Bond Energies, and Solvent Effects J. Am. Chem. Soc. 2004, 126, 2186–2193.
5. Li Y, Li X F, Son B W. Antibacterial and radical scavenging epoxycyclohexenones and aromatic polyols from a marine isolate of the fungus Aspergillus. Nat. Prod. Sci. 2005, 11, 136–138.
6. O’Shea K E, Fox M A. Pulse radiolytic kinetic study of the decay of alpha. -methyl-substituted benzoquinone radical anions: a possible mechanistic model for bioreductive alkylation. J. Am. Chem. Soc. 1991, 113, 611–615.
7.赵天增.核磁共振氢谱.北京大学出版社,第一版,1983,96.
8. Li D, Wang F, Cai S, et al. Two New Bisorbicillinoids Isolated from a Deep-sea Fungus, Phialocephala sp. FL30r. J. Antibiot., 2007, 60, 317-320.
9. Andrade R, Ayer W A, Mebe P P. The metabolites of Trichoderma longibrachiatum. PartⅠ. Isolation of the metabolites and the structure of trichodimerol. Can. J. Chem. 1992, 70: 2526-2535.
10. Trifonov L S, Bieri J H, Prewo R, et al. Isolation and structure elucidation of three metabolites from Verticillium intertextum: sorbicillin, dihydrosorbicillin and bisvertinoquinol. Tetrahedron, 1983, 39 (24): 4243-4256.
11. Abe N, Sugimoto O, Hirota A, et al. Novel DPPH radical scavengers, bisorbicillinol anddemethyltrichodimerol, from a fungus. Biosci. Biotechnol. Biochem. 1998, 62 (4): 661-666.
12. Abe N, Yamamoto K, Hirota A. Novel fungal metabolites, demethylsorbicillin and oxosorbicillinol, isolated from Trichoderma sp. USF-2690. Biocsi. Biotechnol. Biochem., 2000, 64 (4): 620-622.
13. Trifonov L S, Floersheim H H, Dreiding A S, et al. Bisvertinols: a new group of dimeric vertinoids from Verticillium intertextum. Tetrahedron, 1986, 42 (12): 3157-3179.
14. Maskey R, Grün-Wollny I, Laatsch H. Sorbicillin analogues and related dimeric compounds from Penicillium notatum. J. Nat. Prod., 2005, 68: 865-870.
15. Levsen K, Preiss A, Godejohann M. Application of high-performance liquid chromatography coupled to nuclear magnetic resonance and high-performance liquid chromatography coupled to mass spectrometry to complex environmental samples. TrAC. Trends in Analytical Chemistry. 2000, 19(1), 27-48
16.杜康平,姜蓉,李宝义,等.微生物来源白细胞介素1受体拮抗剂的研究II.链霉素660代谢产物的化学研究.中国抗生素杂志,2001,26: 410
17. McCloskey P J, Reilly W W, Shanklin E W, et al. Poly(arylene ether)-polycarbonate block copolymer compositions, methods, and articles. PCT Int. Appl., 2007, 109.
18. Patel A V, Blunden G, Crabb T A. Transformations of solasodine and derivatives of hecogenin by Cunninghamella elegans. Phytochemistry 1994, 35(1), 125-133
19.于德泉,杨峻山等.分析化学手册(第二版)北京:化学工业出版社. 1999
20. Kawai H, Katoono R, Nishimura K, et al. Positive Homotropic Allosteric Binding of Benzenediols in a Hydrindacene-Based Exoditopic Receptor: Cooperativity in Amide Hydrogen Bonding. Journal of the American Chemical Society 2004, 126(16), 5034-5035.
21. Du M, Zhang Z, Wang X, et al. Configuration flexibility of 2,5-bis(3-pyridyl) -1,3,4 -oxadiazole in controllable cocrystallization with 3-hydroxybenzoic acid. Journal of Molecular Structure 2006, 791(1), 131-136.
22. Liu W, Gu Q, Zhu W, et al. Penicillones A and B, two novel polyketides with tricyclo [5.3.1.03, 8] undecane skeleton, from a marine-derived fungus penicillium terrestre. Tetrahedron Lett. 2005, 46: 4993–4996.
23. Liu W, Gu Q, Zhu W, et al. Two new benzoquinone derivatives and two new bisorbicillinoids were isolated from a marine-derived fungus Penicillium terrestre. J.Antibiot., 2005, 58: 441–446.
24. Maskey R P, Grün-wollny I, Laatsch H. Sorbicillin analogues and related dimeric compounds from Penicillium notatum. J. Nat. Prod., 2005, 68: 865-870.
25. Bringmann G, Lang G, Gulder T, et al. The first sorbicillinoid alkaloids, the antileukemic sorbicillactones A and B, from a sponge-derived Penicillium chrysogenum strain. Tetrahedron, 2005, 61: 7252–7265.
26. Neumann K, Abdel-Lateff A, Wright A D, et al. Novel sorbicillin derivatives with an unprecedented carbon skeleton from the sponge-derived fungus Trichoderma Species. Eur. J. Org. Chem. 2007, 2268–2275
1. Nozawa K, Nakajima S. Isolation of radicicol from penicillium luteo-aurantium, and meleagrin, a new metabolite, from penicillium meleagrinum. J. Nat. Prod. 1979, 42: 374
2. Kawai K, nozawa K, Nakajima S, et al. Studies on fungal products.Ⅶ. The structures of meleagrin and 9-O-p-bromobenzoylmeleagrin. Chem. Pharm. Bull. 1984, 32 (1): 94
3. Konda Y, Onda M, Hirano A, et al. Oxaline and Neoxaline. Chem. Pharm. Bull. 1980, 28 (10): 2987
4.刘仕平,曾松荣,郭仕平,等。中华山荷叶内生真菌的分离及其发酵产生药用成分的初步研究.中国药物与临床,2003, 3 (3): 227-228
5.粟晓黎.中药鬼臼毒性成分HPLC/UV指纹图谱分析方法研究及与威灵仙、龙胆HPLC图谱比较.中成药,2000, 22(12): 819-824
6.陈雪松,陈迪华,刘柯.脑甙类化合物的研究概况.天然产物研究与开发,2001,13(2): 63-68
7.高锦明,沈杰,杨雪,等.黄白红菇的化学成分.云南植物研究,2001,23(3): 385-393
8. Jinichiro K, Toyozo Y, Masaru S, et al. Cerebrosides A and C, sphingolipid elicitors of hypersensitive cell death and phytoalexin accumulation in rice plants. The Journal of Biological Chemistry. 1998, 48(27), 31985
9.张鞍灵,刘国强,高锦明.脑苷脂B的结构鉴定.西北植物学报,2001, 21(4): 684-688
10.乐长高,林永成,姜广策,等. Julellaavicenniae两个代谢产物结构的测定. Journal of East China Geological Institute, 1999, 21(4): 64-66
11. Wright J L. Identification of C-24 Alkyl Epimers of marine Sterols by 13C nuclear magnetic resonance spectroscopy. Can. J. Chem., 1978, 56(14): 1898-1901
12.严小红,宋国强,周秀红,等.一种南海海绵(Acanthella sp.)的化学成分研究.天然产物研究与开发, 2003, 15(3): 199-202
13.蒋亭,田黎,李果.黄海葵附生真菌Aspergillus flavipes化学成分的研究.哈尔滨商业大学学报, 2002, 18(1): 30-33
14. Lin C, Tome W. Novel Cytotoxic Principles of Formosan ganoderma lucldum. Journal ofNatural Products, 1998, 54(4): 998-1002
15. Nam K S, Jo Y S, Kim Y H. Cytotoxic activities of acetoxyscirpenediol and ergosterol peroxide from Paecilomyces tenuipes. Life Sciences, 2001, 69: 229–237
16. Yue J, Chen S, Lin Z. Sterols from the fungus Lactarium volemus. Phytochemistry, 2001, 56: 801-806
17. Takaishi Y, Uda M, Ohashi T, et al. Glycosides of ergosterol derivatives from Hericum erinacens. Phytochemistry 1991; 30(12):4117–220.
18. Takaishi Y, Adachi R, Murakami Y, et al. A polyoxygenated steroid from Lasiosphaera nipponica. Phytochemistry 1992, 31(1): 243–246.
19. Yagi R, Doi. M. Isolation of an antioxidative substance produced by Aspergillus repens. Biosci. Biotechnol. Biochem., 1999, 63: 932-933.
20. Dossena A, Marchelli R, Pochini A. New metabolites of Aspergillus amstelodami related to the biogenesis of neoechinulin. J. Chem. Soc. Chem. Comm., 1974: 771-772.
21. Nagasawa H, Isogai A, Ikeda K, et al. Isolation and structure elucidation of a new indole metabolite from Aspergillus rubber. Agr. Biol. Chem., 1975, 39 (9): 1901-1902.
22. Houghton E, Saxton J E. Echinuluin series.Ⅱ. Synthesis of (+)-alanyl- tryptophan anhydride and L-alanyl-2-(1,1-dimethylallyl)tryptophan anhydride. J. Chem. Soc. [section] C: Organic, 1969, (6): 1003-1012.
23. Stipanovic R D, Schroeder H. Preechinulin, a metabolite of Aspergillus chevalieri. Transactions of the British Mycological Society, 1976, 66: 178-179.
1. Mass spectra database, Korea Basic Science Institute.
2. El-Sharkawy S H, Abul-Hajj Y J. Microbial transformation of zearalenone. J.Org.Chem., 1988, 53: 515-519
3. Peters C A, Hurd R N. A stereoselective synthetic route to (R)-zearalanone. J.Med.Chem., 1975, 18(2): 215-217.
4. Urry W H, Wehrmeister H L, Hodge E B, et al. The structure of zearalenone. Tetrahedron Lett., 1966, 27: 3109-3114.
5. Burckhardt S, Ley S V. The use of allyltricarbonyliron lactone complexs in the synthesis of the resorcylic macrolides and zearalenol. J.Chem.Soc.,Perkin Trans.1, 2002: 874-882
6. Li X, Kim S, Kang, J S, et al. Radical scavenging hydroxyphenyl ethanoic acid derivatives from a marine-derived fungus. Journal of Microbiology and Biotechnology, 2006, 16(4): 637-638.
7.许颂,梁华清.刺人参的蒽醌成分研究.中草药, 1998, 29(4): 222-223
8.毕志明,张勉,王峥涛.流苏石斛化学成分的研究.中国药科大学学报, 2001, 32(6): 421-422
9.卢艳花主编,中药有效成分提取分离技术.化学工业出版社,163-164.
10.胡海峰,朱宝泉,龚炳勇.微生物来源的胆固醇生物合成醇抑制剂研究:抗生素SIPI-8926-111的研究.中国医药工业杂志,1995, 26 (11): 484-486.
11. Stob M, Baldwin R S, Tuite J, et al. Isolation of an anabolic, uterotrophic compound from corn infected with Gibberella zeae. Nature, 1962, 196: 1318
12. Taub D, Girotra N N, Hoffsommer R D, et al. Total synthesis of the macrolide, zearalenone. Tetrahedron, 1968, 24: 2443-2461
13. Shier W T, Shier A C, Xie W, et al. Structure-activity relationships for human estrogenic activity in zearalenone mycotoxins. Toxicon, 2001, 39: 1435-1438.
1. Fujita M, Nakao Y, Matsunaga S, et al. Sodium 1-(12-hydroxy)octadecanyl sulfate, an MMP2 inhibitor, isolated from a tunicate of the family Polyclinidae. J. Nat. Prod. 2002, 65, 1936-1938.
2. Roccatagliata A J, Maier S M, Seldes A M, et al. A new sulfated alkene from the ophiuroid Ophiocoma echinata. J. Nat. Prod. 1997, 60, 285-286.
3. Tsukamoto S, Kato H, Hirota H, et al. Antibacterial and antifungal sulfated alkane and alkenes from the hepatopancreas of the ascidian Halocynthia roretzi. J. Nat. Prod. 1994, 57, 1606-1609.
4. Aiello A, Carbonelli S, Esposito G, et al. Novel bioactive sulfated alkene and alkanes from the Mediterranean ascidian Halocynthia papillosa. J. Nat. Prod. 2000, 63, 1590-1592.
5. Aristoff P A, Johnson P D, Harrison A W. Total synthesis of a novel antiulcer agent via a modification of the intramolecular Wadsworth-Emmons-Wittig reaction. J. Am. Chem. Soc. 1985, 107, 7967-7974.
6. Aldridge D C, Armstrong J J, Speake R. N, et al. Structures of cytochalasins A and B. J. Chem. Soc. C. 1967, 17: 1667-1676.
7. Giovanni G, Paolo M, Piero S G, et al. The circular dichroism of (-)-(S)-3-methylthian: a study of the electronic transitions and stereochemistry of cyclic sulfur derivatives. J. Chem. Soc. Perkin Trans. 2. 1981, 12: 1529-1533.
8. Vlastimil H. Detection of purines, pyrimidines, and their derivatives. Biokhimiya (Moscow), 1960, 25: 891-896.
9. Kyosuke T, Saburo A, Yukichi K, et al. Sargasterol from Sargassum ringgoldianum. Pharmaceutical Bulletin, 1957, (5): 85-86.
10. Ryoichi H. Steroid studies. VII. Synthesis of sargasterol, fucosterol, and 20-isocholesterol. Pharmaceutical Bulletin, 1957, (5): 452-459.
11. Taguchi H, Paal B, Armarego W. Glyceryl-ether monooxygenase [EC 1.14.16.5]. Part VIII.Probing the nature of the active site. Pteridines, 1995, 6(2): 45-57.
12. Li H J, Lin Y C, Liu X H, et al. Vrijmoed, Studies on the secondary metabolites of marine fungus Fusarium sp. (#2489) from the south china sea. Marine sci, 2002, 26: 57-59.
13. Yadav J S, Reddy B V, Subba G, et al. Gallium(III) chloride catalyzed stereoselective synthesis of E-configured, unsaturated ketones. Synlett 2007, 5: 809-811.
14. Onizawa Y, Kusama H, Iwasawa N. Efficient control of alkyne and vinylidene complex pathways for the W(CO)5(L)-catalyzed synthesis of two types of nitrogen-containing bicyclic compounds. Journal of the American Chemical Society 2008, 130(3): 802-803.
15. Regnier J, David R. Some physicochemical properties of aqueous solutions, equivalent in content of base (p-aminobenzoate of diethylaminoethanol, procaine base) prepared with different acids. I. Determination of the surface tension at the liquid-air interface. Anesthesie et analgesie 1938, 4: 483-488.
16.于德泉,杨峻山.分析化学手册(第二版).北京:化学工业出版社, 1999
17. Aldrich Library of 13C and 1H FT NMR Spectra, 1992, 3, pp371B
1. Liu W, Gu Q, Zhu W, et al. Penicillones A and B, two novel polyketides with tricyclo [5.3.1.03, 8] undecane skeleton, from a marine-derived fungus penicillium terrestre. Tetrahedron Lett. 2005, 46: 4993–4996.
2. Liu W, Gu Q, Zhu W, et al. Two new benzoquinone derivatives and two new bisorbicillinoids were isolated from a marine-derived fungus Penicillium terrestre. J. Antibiot., 2005, 58: 441–446.
3. Liu W, Gu Q, Zhu W, et al. Dihydrotrichodimerol and tetratrichodimerol, two new bisorbicillinoids, from a marine-derived Penicillium terrestre. J. Antibiot. 2005, 58: 621–624.
4. Duan X, Li X, Wang B. Highly brominated mono- and bis-phenols from the marine red alga Symphyocladia latiuscula with radical-scavenging activity. J. Nat. Prod., 2007, 70: 1210-1213
1. Solis P N, Wright C W, Anderson M M, et al. A microwell cytotoxicity assay using Artemia salina (brine shrimp). Planta Med. 1993, 59 (3), 250-252.
2. Meyer B N, Ferrigni N R, Putnam J E, et a1. Brine Shrimp: A convenient general bioassay for active plant constituents . P1anta Med. 1982, 45, 31-34.
3.张永杰,王建锋,黄耀坚,等. 4种裸子植物内生真菌抗肿瘤菌株的筛选.厦门大学学报,2002, 41 (6), 804-809.
4.麦克劳林,顾哲明.两种简易的抗肿瘤活性初筛方法.中国中药杂志, 1997, 22 (10), 617-619.
5. Skehan P, Storeng R, Scudiero D, et al. New colorimetric cytotoxicity assay for anticancer drug screening. J. Natl. Cancer Inst. 1990, 82 (13), 1107-1112.
6. Voigh W. Sulforhodamine B assay and chemosensitivity. Methods Mol. Med. 2005, 110, 39-48.
7. Mossman T. Rapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assay. J.Immunol. Meth., 1983, 65, 55-63.
8. Patricia L, Judson M D, Linda V L, et al. Flow cytometry. Oncology Update. 1997, 4, 87-91.
1. Cueto M, Jensen P R, Kauffman C, et al. Pestalone, a new antibiotic produced by a marine fungus in response to bacterial challenge. J. Nat. Prod., 2001, 64: 1444-1446.
2. Bringmann G,Lang G, Steffens S,et al. Evariquinone, isoemericellin, and stromemycin from a sponge derived strain of the fungus Emericella variecolor. Phytochemistry, 2003,63: 437-443.
3. Kang H S, Chuang H Y, Jung J H, et al. A new phlorotannin from the brown alga Ecklonia stolonifera, Chem. Pharm. Bull., 2003, 51(8): 1012-1014.
4. Xu X, Song F, Wang S, et al. Dibenzyl bromophenols with diverse dimerization patterns from the brown alga Leathesia nana, J. Nat. Prod., 2004, 67: 1661-1666.
5. Glombitza K W, Schmidt A. Nonhalogenated and halogenated phlorotannins from the brown alga Carpophyllum angustifolium. J. Nat. Prod., 1999, 62: 1238-1240.
6. Carte B K, Troupe N, James A, et al. Rawsonol, an inhibitor of HMG-CoA reductase from the tropical green alga Avraznvillea rawsoni. Phytochemistry, 1989, 28:2917-2919.
7. Chen J L, Gerwick W H. Isorawsonol and related imp dehydrogenase inhibitors from the tropical green alga Avraznvillea rawsoni. J. Nat. Prod., 1994, 57: 947-952.
8. Kurihara H, Mitani T, Kawabata J, et al. Two new bromophenols from the red alga Odonthalia corymbifera. J.Nat.Prod., 1999, 62: 882-884.
9. Xu N, Fan X, Yan X , et al. Antibacterial bromophenols from the marine red alga Rhodomelaconfervoides. Phytochemistry, 2003, 62: 1221-1224.
10. Fan X, Xu N, Shi J. Bromophenols from the Red Alga Rhodomela confervoides. J.Nat.Prod. 2003, 66: 455- 458.
11. Zhao J, Fan X, Wang S. Bromophenol derivatives from the red alga Rhodomela confervoides. J.Nat.Prod. 2004, 67:1032-1035.
12. Zhao J, Ma M, Wang S, et al. Bromophenols coupled with derivatives of amino acids and nucleosides from the red alga Rhodomela confervoides. J.Nat.Prod. 2005, 68: 691-694.
13. Ma M, Zhao J, Wang S, et al. Bromophenols Coupled with Methylγ-Ureidobutyrate and Bromophenol Sulfates from the Red Alga Rhodomela confervoides. J.Nat.Prod. 2006, 69: 206-210.
14. Ma M, Zhao J, Wang S, et al. Bromophenols Coupled with Nucleoside Bases and Brominated Tetrahydroisoquinolines from from the Red Alga Rhodomela confervoides. J.Nat.Prod. 2007, 70: 337-341.
15. Duan X, Li X, Wang B. Highly Brominated Mono- and Bis-phenols from the Marine Red Alga Symphyocladia latiuscula with Radical-Scavenging Activity. J.Nat.Prod. 2007, 70: 1210-1213.
16. Pham N B, Butler M S, Quinn R J. Isolation of psammaplin a 11’-sulfate and bisaprasin
11’-sulfate from the marine sponge Aplysinella rhax. J.Nat.Prod. 2000, 63: 393-395.
17. Utkina N K, Denisenko V, Scholokova O V, et al. Spongiadioxins A and B, two new polybrominated dibenzo-p-dioxins from an Australian mmarine sponge Dysidea dendyi. J.Nat.Prod. 2001, 64: 151-153.
18. Handayani D, Edrada R A, Proksch P, et al. Four new bioactive polybrominated diphenyl ethers of the Sponge Dysidea herbacea from West Sumatra, Indonesia.J.Nat.Prod. 1997, 60: 1313-1316.
19. Miao S, Andersen R J. Rubrolides A-H, metabolites of the colonial tunicate Ritterella rubra. J. Org. Chem., 1991, 56: 6275-6280.
20. Davis R A, Sandoval I T, Concepcion G P, et al. Lissoclinotoxins E and F, novel cytotoxic alkaloids from a philippine didemnid ascidian. Tetrahedron, 2003, 59: 2855-2859.
21. Liu H, Pratasik S B, Nishikawa T, et al. Lissoclibadin 1, a novel trimeric sulfur-bridged dopamine derivative, from the tropical ascidian Lissoclinum cf. badium. Tetrahedron Letters,2004,45: 7015-7017.
22. Eisenbarth S, Gehling M, Harder A, et al. Pentaporins A, B and C: disulfides from the marine bryozoan Pentapora fascialis. Tetrahedron, 2002, 58: 8461-8464.
