两株海洋来源微生物次级代谢产物抗肿瘤活性成分研究

【作者】 李冬；

【导师】 崔承彬； 顾谦群；

【作者基本信息】 中国海洋大学 ， 生药学， 2005， 硕士

【摘要】 为有效地从海洋微生物中寻找抗肿瘤活性成分,本文采用小鼠乳腺癌细胞tsFT210,以细胞周期抑制、细胞凋亡诱导及细胞坏死为活性指标,从采自青岛近海海域的24个海泥样品中共筛选出110株放线菌,其中具有抗肿瘤活性的菌株10株,活性率为9.1%。并采用活性追踪分离方法对一株放线菌和一株真菌的代谢产物活性成分进行了初步研究 采用减压硅胶柱层析、凝胶柱层析（Sephadex LH-20）、反相硅胶柱层析和反相制备高效液相（HPLC）等分离手段,从具有细胞坏死活性的放线菌H2003分离得到22个单体化合物。利用理化性质和光谱学方法（UV、IR、MS、1D-NMR和2D-NMR）,21个单体化合物分别鉴定为:邻苯二甲酸二丁酯（1）;丁二烯二甲酯（2）;3,4-二羟基-2-甲基-吡啶（3）;对羟基苯甲酸（5）;3,4-二羟基苯甲酸甲酯（6）;环-（脯-异亮）（7）;环-（脯-亮）（8）;环-（脯-缬）（9）;4,4’硝基—联苄基（10）;环-（脯-苯丙）（11）;环-（脯-丙）（12）;环-（丙-缬）（13）;N-[2-（4-羟基苯基）乙基]乙酰胺（14）;环-（羟脯-亮）（15）;环-（丙-异亮）（16）;环-（丙-亮）（17）;环-（缬-亮）（18）;环-（脯-酪）（19）;环-（甘-苯丙）（20）;环-（丙-苯丙）（21）;环-（甘-脯）（22）。化合物4初步鉴定为环-（脯-脯-亮-亮）,其结构有待进一步确定。从具有周期抑制活性的真菌Aspergillus terreus Thom中分离得到13个单体化合物分别鉴定为:麦角甾醇（23）;十八碳二烯酸单甘油酯（24）;大黄酚（25）;大黄素（26）;芦荟大黄素（27）;苯甲酸（28）;邻苯二甲酸二异丁酯（29）;3,4-二甲氧基苯酚（30）;二硫甲基胶霉毒素（31）;3-甲氧基苯酚（32）;对羟基苯乙腈（33）;反-对羟基苯乙烯酰氨（34）;顺-对羟基苯乙烯酰氨（35）。 生物活性测试结果表明,化合物1、23、24、26、29、30、32对tsFT210小鼠乳腺癌细胞具有显著的坏死性细胞毒活性,化合物14、34具有显著的诱导细胞调亡活性,化合物3、25具有G₂/M周期抑制活性,化合物4、5、11具有G₀/G₁周期抑制活性。更多还原

【Abstract】 We have undertaken the screening for novel cell cycle inhibitors and apoptosis inducers from the marine-derived actinomycetes H2003 and Aspergillus terreus Thom by use of a mouse cdc2 mutant cell line, tsFT210. In anticipation of achieving the above objectives, marine-derived actinomycetes, H2003 and Aspergillus terreus Thom have been screened by using this model, the EtOAC extracts of strain H2003 and Aspergillus terreus Thom showed bioactivities of inhibiting the cell cycle progression and cytoclasis of tsF210 cells. Thus, we selected them for further investigation.Eleven compounds were isolated from the bioactive part, EtOAC extract of strain H2003, by repeated column chromatography on Sephadex LH-20, silica gel and RP18, followed by reverse-phase high performance liquid chromatragraphy. The structures of compounds were elucidated mainly by use of spectroscopic method (UV, IR, MS, 1D-NMR, 2D-NMR) and according to their physicochemical properties: dibutyl phthalate (1); Acetic acid 4-acetoxy-buta-l,3-dienyl ester (2); 3,4-Dihydroxy-2-methylpyridine (3); 4-hydroxybenzoic acid (5); Protocatechuic acid methyl ester (6); cyclo-(Pro-Ile) (7); cyclo-(Pro-Leu) (8); cycol-(Pro-Val) (9); 4,4’-Dintrobibenzyln (10) ;cyclo-(Pro-Phe) (11); cyclo-(Pro-Ala) (12); cyclo-(Ala-Val) (13); N-(4-hydroxyphenethyl)acetamide (14); cyclo-(4-hydroxy-Pro-Leu) (15); cyclo- (Ala-Ile) (16) ; cyclo-(Ala-Leu) (17); cyclo-(Val-Leu) (18); cyclo-(Tyr-Pro) (19); cyclo-(Gly-Phe) (20); cyclo-(Ala-Phe) (21); cycol-(Gly-Pro) (22). The structure of compound (4) is still in affirming.By the same activity-guided method, thirteen compounds were isolated from the bioactive part, EtOAC extract of Aspergillus terreus Thom, by physicochemical evidence and spectroscopic methods, the structures of the eleven compounds were elucidated as: ergosterol (23); glyceride (24); chrysophanol (25); emodin (26); 1,8-dihydroxy-3-(hydroxymethyl)anthracene-9,10-dione (27); benzoic acid (28); diisobutyl phthalate (29); 3,4-dimethoxyphenol (30); dithiol gliotoxin (31); 3-methoxyphenol (32); 2-(4-hydroxyphenyl)acetonitrile (33);(E)-N-(4-hydroxystyryI)formamide (34); (Z)-N-(4-hydroxystyryl)formamide (35).Bioassay results indicated that 1,23,24,26,29,30,32 show cell cycle inhibition and cytoclasis of tsFT210 cells. Compound 14,34 showed a strong apoptosis of tsFT210 cells. Compound 3,4,5,11,25 show cell cycle inhibition of tsFT210 cells.Bioassay results indicated that we isolated the main antitumor active compounds of strain H2003 and Aspergillus terreus Thorn successfully which showed that this antitumor model and the method were exact and available in achieving bioactive components from marine microbes.更多还原