钌多吡啶类核酸分子“光开关”配合物的分析应用研究
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摘要
荧光法在DNA的研究中应用非常广泛,但是由于DNA的内源荧光很弱,在研究DNA的结构或测定其含量时,必须引入一定的探针分子。在众多的探针分子中,具有核酸分子“光开关”性质的钌(II)多吡啶类配合物是最令人瞩目的。这类荧光探针在水溶液中没有荧光,而当溶液中有双链DNA存在时其荧光急剧增强,因此它们被称为核酸分子“光开关”。本论文以核酸分子“光开关”配合物Ru(phen)_2dppx~(2+)(phen =1,10-二氮杂菲,dppx=7,8-二甲基吡啶并[3,2-a:2’,3’c]吩嗪)为探针,分别采用荧光法和近年来新兴的共振瑞利散射光谱的方法研究了“光开关”配合物在核酸研究中的应用。主要工作包括以下两个方面:
1.以核酸分子“光开关”配合物Ru(phen)_2dppx~(2+)为探针,分别采用紫外-可见分光光度法、荧光光谱法研究了蒽醌类抗肿瘤药物道诺霉素(DNM)与小牛胸腺DNA之间的相互作用。结果表明DNM是以插入的方式与DNA相互作用。DNM对Ru(phen)_2dppx~(2+)-DNA体系Scatchard方程的影响结果也表明,DNM主要是以插入方式与DNA结合;而且核酸分子“光开关”Ru(phen)_2dppx~(2+)与小牛胸腺DNA的结合比在1:4~1:5之间。与常规核酸荧光探针溴化乙锭相比,Ru(phen)_2dppx~(2+)具有灵敏度高、毒性低、稳定性好、选择性好、使用方便等优点。
2.在水溶液中,带正电荷的Ru(phen)_2dppx~(2+)在阴离子表面活性剂十二烷基苯磺酸钠(SDBS)的表面聚集后,产生很强的共振瑞利散射。由于Ru(phen)_2dppx~(2+)与DNA之间有较大的结合常数(KB≥106 L/mol),DNA的加入会促使体系中“光开关”Ru(phen)_2dppx~(2+)与SDBS发生分离,导致其散射光强度大大降低,且体系散射光的降低程度与DNA的浓度呈线性关系。基于散射光强度的降低,建立了一种DNA的定量分析新方法。
Fluorescence analysis is one of the widest methods in DNA study. But spectroscopic probe is need to analysis the structure or quantity of DNA for weekly luminescent itself. One of the attractive probes is a kind of polypyridine ruthenium complexes which act as nuclear acids molecular“light-switch”. They have no fluorescence in water solution, but have much strong fluorescence in the presence of double helix DNA. In this paper, one of the“light-switch”complex Ru(phen)_2dppx~(2+) was applied as a spectroscopic probe to study the application of these complexes by flourescence and Rayleigh resonance light scattering methods. The main contents of this study could be described as follows:
1. The interaction between daunomycin and ctDNA was studied using Ru(phen)_2dppx~(2+) as an interacting mode spectroscopic probe. And the way for Ru(phen)_2dppx~(2+) action on DNA is intercalation. The study of the influence of DNM on the ultraviolet spectra and fluorescence spectra of Ru(phen)_2dppx~(2+)-DNA system indicate that DNM intercalates the base pairs of DNA. And the results of the influence of DNM on the Scatchard equation of Ru(phen)_2dppx~(2+)-DNA system also indicate that DNM intercalates the base pairs of DNA.Compared with normal DNA interacting mode spectroscopic probe Ethidium Bromide (EB), Ru(phen)_2dppx~(2+) has the advantages of higher sensitivity, lower toxicity, higher stability and simplicity.
2. In water solution, positive Ru(phen)_2dppx~(2+) could assemble on the surface of negative Sodium dodecylbenzene sulfonate (SDBS) and resulting in strong resonance rayleigh scattering. Because the strong affinity between Ru(phen)_2dppx~(2+) and DNA (KB≥106 L/mol), Ru(phen)_2dppx~(2+) will separated with SDBS while DNA was added, and resulting scattering light reduce greatly. The decrease of resonance Rayleigh scattering light is linear to the concentration of DNA. Based on which, a novel method is proposed for the determination of DNA.
1.以核酸分子“光开关”配合物Ru(phen)_2dppx~(2+)为探针,分别采用紫外-可见分光光度法、荧光光谱法研究了蒽醌类抗肿瘤药物道诺霉素(DNM)与小牛胸腺DNA之间的相互作用。结果表明DNM是以插入的方式与DNA相互作用。DNM对Ru(phen)_2dppx~(2+)-DNA体系Scatchard方程的影响结果也表明,DNM主要是以插入方式与DNA结合;而且核酸分子“光开关”Ru(phen)_2dppx~(2+)与小牛胸腺DNA的结合比在1:4~1:5之间。与常规核酸荧光探针溴化乙锭相比,Ru(phen)_2dppx~(2+)具有灵敏度高、毒性低、稳定性好、选择性好、使用方便等优点。
2.在水溶液中,带正电荷的Ru(phen)_2dppx~(2+)在阴离子表面活性剂十二烷基苯磺酸钠(SDBS)的表面聚集后,产生很强的共振瑞利散射。由于Ru(phen)_2dppx~(2+)与DNA之间有较大的结合常数(KB≥106 L/mol),DNA的加入会促使体系中“光开关”Ru(phen)_2dppx~(2+)与SDBS发生分离,导致其散射光强度大大降低,且体系散射光的降低程度与DNA的浓度呈线性关系。基于散射光强度的降低,建立了一种DNA的定量分析新方法。
Fluorescence analysis is one of the widest methods in DNA study. But spectroscopic probe is need to analysis the structure or quantity of DNA for weekly luminescent itself. One of the attractive probes is a kind of polypyridine ruthenium complexes which act as nuclear acids molecular“light-switch”. They have no fluorescence in water solution, but have much strong fluorescence in the presence of double helix DNA. In this paper, one of the“light-switch”complex Ru(phen)_2dppx~(2+) was applied as a spectroscopic probe to study the application of these complexes by flourescence and Rayleigh resonance light scattering methods. The main contents of this study could be described as follows:
1. The interaction between daunomycin and ctDNA was studied using Ru(phen)_2dppx~(2+) as an interacting mode spectroscopic probe. And the way for Ru(phen)_2dppx~(2+) action on DNA is intercalation. The study of the influence of DNM on the ultraviolet spectra and fluorescence spectra of Ru(phen)_2dppx~(2+)-DNA system indicate that DNM intercalates the base pairs of DNA. And the results of the influence of DNM on the Scatchard equation of Ru(phen)_2dppx~(2+)-DNA system also indicate that DNM intercalates the base pairs of DNA.Compared with normal DNA interacting mode spectroscopic probe Ethidium Bromide (EB), Ru(phen)_2dppx~(2+) has the advantages of higher sensitivity, lower toxicity, higher stability and simplicity.
2. In water solution, positive Ru(phen)_2dppx~(2+) could assemble on the surface of negative Sodium dodecylbenzene sulfonate (SDBS) and resulting in strong resonance rayleigh scattering. Because the strong affinity between Ru(phen)_2dppx~(2+) and DNA (KB≥106 L/mol), Ru(phen)_2dppx~(2+) will separated with SDBS while DNA was added, and resulting scattering light reduce greatly. The decrease of resonance Rayleigh scattering light is linear to the concentration of DNA. Based on which, a novel method is proposed for the determination of DNA.
引文
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