【作者】 刘向伟；

【导师】 张前前；

【作者基本信息】 中国海洋大学 ， 分析化学， 2010， 硕士

【摘要】 关于铜／锌-生命配体配合物的研究是过渡金属配合物研究的最活跃的课题之一,其药用性一直是科研人员关注的重要方面。本文通过合成新型铜／锌-生命配体配合物,与结构已知的同系列的配合物进行比较研究,采用多种方法研究其生物活性,即以电子吸收光谱法、荧光光谱分析法研究了它与DNA作用,采用琼脂糖凝胶电泳法研究了该配合物对DNA的切割作用,首次以斑马鱼胚胎作为模式生物研究了配合物的生物毒性,旨在为高效低毒药物研发提供理论依据。主要研究结果如下：(1)合成了一种新的锌(Ⅱ)-色氨酸-邻菲啰啉混配配合物[Zn(Trp)(phen)2] Cl·7H2O(1)(Trp代表色氨酸离子,phen代表邻菲啰啉),通过元素分析、红外光谱、电导及热重-差热分析对其进行表征。(2)对锌(Ⅱ)-色氨酸-邻菲啰啉三元配合物[Zn(Trp)(phen)2] Cl·7H2O(1)及两种已知结构的锌-邻菲啰啉二元配合物[Zn(phen)2] Cl2·5H2O(2)和锌-邻菲啰啉-氯三元配合物[Zn(phen)Cl2](3)进行了比较研究。电子吸收光谱法和荧光光谱法实验结果表明：锌-生命配体配合物与DNA发生作用强弱顺序为配合物1>配合物2>配合物3,但它们与DNA作用模方式有所不同；琼脂糖凝胶电泳实验结果显示：在抗坏血酸存在条件下,锌配合物1与pBR322 DNA作用10 min时已经对pBR322 DNA产生显著作用,配合物2和配合物3与pBR322 DNA作用10 min时无明显作用,从作用时间来看,配合物1的切割活性比另外两个更强。从浓度来看,当浓度为2.5μmol·L-1时,配合物2将约50%Ⅰ型DNA转变为Ⅱ型DNA,而在配合物3体系中,DNA主要以Ⅰ型存在；当浓度为10μmol·L-1时,配合物2将Ⅰ型DNA完全转变为Ⅱ型DNA,并且切割出Ⅲ型,配合物3也将Ⅰ型DNA完全转变为Ⅱ型,但没有切割出Ⅲ型,说明配合物2切割活性比配合物3强。综上,锌-生命配体配合物对pBR322 DNA切割活性顺序为：配合物1>配合物2>配合物3。在斑马鱼胚胎毒性实验中,综合24hpf时的致死率和孵化期72hpf时的半致死浓度和孵化率三个指标数据比较结果可以得到：锌的三种配合物对斑马鱼胚胎毒性作用顺序为：配合物2>配合物1>配合物3,说明相对于配合物2,配合物1因引入一个色氨酸离子配位,降低了配合物的生物毒性。(3)选取两种已知结构的铜(Ⅱ)-生命配体配合物即[Cu(SF)phen Cl]·4H2O(4)(SF代表司帕沙星)和[Cu(SF)2]·4H2O(5),以电子吸收光谱法、荧光光谱分析法研究了它与DNA的作用,采用琼脂糖凝胶电泳法研究了该配合物对DNA的切割作用,并考察了它们对斑马鱼胚胎的生物毒性。两种铜-生命配体配合物与DNA发生作用强弱顺序为配合物4>配合物5,其与DNA作用模方式也是不同的。在抗坏血酸存在条件下,铜配合物4与pBR322 DNA作用10 mmin时已经对pBR322 DNA有显著作用,而配合物5与pBR322 DNA作用10 min时无明显作用,从作用时间来看,配合物4的切割活性更强。铜的两种配合物对斑马鱼胚胎毒性作用顺序为：配合物4>配合物5。上述结果表明,相对于配合物5而言,配合物4中一个配体司帕沙星换成邻菲啰啉后,与DNA作用增强的同时,生物毒性也明显增强。更多还原

【Abstract】 The research of Cu(Ⅱ)/Zn(Ⅱ)-life ligand complexes is one of the most active areas in transition metal complexes scientific research.The biological activities of these complexes play an important part in lives’research. Its medical effect is concerned of the important aspects. In this paper, new Cu(Ⅱ)/Zn(Ⅱ)-life ligand complexes are synthesized, a series of known complexes were chosen to study as comparison reference. A variety of methods are used to research the biological activities:The interaction of the complexe with sperm DNA was investigated by electric absorption spectrscopy,ethidium bromide(EB) fluorescence spectrscopy and gel electrophoresis measurements.The bio-toxicity of the complexe was determined by zebrafish embryos. With the expection of providing theoretical basis for drug development of new metal complexes. Main research results are as follows:(1) A novel complex of [Zn(Trp)(phen)2]Cl·7H2O (1) (Tip=L-Tryptophan, phen=Phenanthroline) was synthesized and characterized by elemental analysis, IR spectrscopy and TG-DTA.(2) [Zn(Trp)(phen)2] Cl·7H2O is studied compared with two known complexes [Zn(phen)2] Cl2·5H2O(2) and [Zn(phen)Cl2](3) The results of electronic absorption spectrometry and fluorescence spectrometry indicated that the three Zn(Ⅱ)-life ligand complexes bond to DNA by different modes,with the order of the binding ability of the complexes to DNA:complex 1> complex 2> complex 3. The results of agarose gel electrophoresis experiment indicated that after 10 min of the action between complexes and pBR322 DNA in the presence of vitamin C as a reducing regent,complex 1 has significant effect on pBR322 DNA, but complex 2 and complex 3 have no obvious effect on pBR322 DNA.According to time,the activity of complex 1 cleaving pBR322 DNA is stronger. Concerning the concentration, when the concentration of complex is 2.5μmol·L-1,nearly a half of pBR322 DNA is cleaved from typeⅠinto typeⅡwith complex 2,while there is only typeⅠwith complex 3. When the concentration of complex is 10μmol·L-1,pBR322 DNA is completely cleaved from typeⅠinto typeⅡwith complex 2,what is more,some typeⅢappears, while there is no typeⅢappears with complex 3 although pBR322 DNA is also completely cleaved from typeⅠinto typeⅡ. The results indicated that complexes 2 cutting activity is stronger than complexes 3. To sum up,the three complexes cleave pBR322 DNA in the presence of vitamin C as a reducing regent with the order of cleavage activity of the complexes to DNA:complex 1> complex 2> complex 3. The bio-toxicity of the complexes on zebrafish embryos is studied by studying the mortality of 24hpf,median lethal dose and hatching rate of 72hpf of zebrafish embryos. The results indicated that the bio-toxicity of the Zn(Ⅱ)-life ligand complexes on zebrafish embryos is increased in the order:complex 3<complex 1<complex 2.1t can be said that the tryptophan ion reduces the bio-toxicity of complex 1,compared with complex 2.(3) Two known copper complexes,[Cu(SF)phen Cl]·4H2O(4)(SF=Sparfloxacin)and [Cu(SF)2]·4H2O(5),are also studied. The interaction of the complexe with sperm DNA was investigated by electric absorption spectrscopy,ethidium bromide(EB) fluorescence spectrscopy and gel electrophoresis measurements.The bio-toxicity of the complexe was determined by zebrafish embryos too.The results of electronic absorption spectrometry and fluorescence spectrometry indicated that the two Cu(Ⅱ)-life ligand complexes bond to DNA by different modes too, with the order of the binding ability of the complexes to DNA:complex 4> complex 5. The results of agarose gel electrophoresis experiment indicated that after 10 min of the action between complexes and pBR322 DNA in the presence of vitamin C as a reducing regent,complex 4 has significant effect on pBR322 DNA,but complex 5 have no obvious effect on pBR322 DNA.According to time,the activity of complex 4 cleaving pBR322 DNA is stronger than complex 5. The bio-toxicity of the complexes on zebrafish embryos is studied by studying the mortality of 24hpf,median lethal dose and hatching rate of 72hpf of zebrafish embryos. The results indicated that the bio-toxicity of the Cu(Ⅱ)-life ligand complexes on zebrafish embryos is increased in the order:complex 4< complex 5. It can be said that the 1,10-phenanthroline increases the bio-toxicity of complex 4. To sum up,the novel zinc complex:[Zn(Trp)(phen)2] Cl·7H2O is more suitable to be studied as candidates of highly effective and low poisonous drugs.更多还原