【作者】 魏玉霞；

【导师】 董川；

【作者基本信息】 山西大学 ， 分析化学， 2004， 硕士

【摘要】 第一章：本章综述了核酸探针的种类，探针与核酸相互作用的方式，研究探针与核酸作用的手段，以及核酸探针的应用；并综述了蛋白质光谱探针的种类以及常用的研究蛋白质与小分子探针作用的光谱方法。 第二章：考察了二碘荧光素和四溴荧光素的流体荧光和固体基质室温磷光特性。发现这两种染料都可以产生流体荧光和固体基质室温磷光，对影响它们发光强度的各种因素，包括滤纸基质类型，pH条件，以及点样时机和稳定性等进行了详细的研究。测得了一些光谱参数如：荧光和磷光偏振值，磷光寿命等，并建立了这两种荧光素的流体荧光和固体基质室温磷光分析法。 第三章：用荧光光谱法和固体基质室温磷光光谱法研究了四溴荧光素与DNA的相互作用。实验表明：DNA会使四溴荧光素的荧光和固体基质室温磷光强度增强。阴离子猝灭实验表明，DNA的存在使得阴离子对四溴荧光素的猝灭程度增大，这说明四溴荧光素与DNA发生了沟槽键合。偏振实验表明，在DNA的存在下，四溴荧光素的磷光偏振值增加，这表明四溴荧光素嵌插在DNA碱基对之间。可能是四溴荧光素中的二苯并吡喃环嵌入了DNA的碱基对之间，其侧链的苯环可能键合在DNA的沟槽中。DNA的存在使乙醇对四溴荧光素荧光强度的影响减小，离子强度的大小也会影响四溴荧光素与DNA的相互作用。 第四章：本文考察了四氯四溴荧光素的流体荧光和固体基质室温磷光特性，并对影响其发光的条件如：pH条件，固体基质，重原子效应，烘烤条件等，进行了详细考察。结果表明，四氯四溴荧光素的荧光激发和发射波长分别为540nm/560nm；磷光的激发和发射波长为556nm/699nm，发射处于长波区。光谱法研究四氯四溴荧光素与DNA的相互作用的实验表明：DNA的存在会使四氯四溴荧光素的荧光和固体基质室温磷光强度发生变化。阴离子猝灭实验表明，四氯四溴荧光素可以嵌插在DNA的碱基对之间，受DNA的保护作用，使得阴离子对四氯四溴荧光素的猝灭程度减小。荧光偏振实验表明，在DNA的存在下，四氯四溴荧光素的偏振值增加，这也支持四氯四溴荧光素嵌插在DNA碱基对之间的实验结果。离子强度的大小会影响四氯四溴荧光素和DNA的相互作用。用Scatchard方程求得二者的结合常数为2×10~6L/mol，结合位点数为0.16。 第五章：用荧光光谱法研究了四碘荧光素与牛血清白蛋白的结合反应。测得该反应的结合常数为5.7×10~4 L·mol，结合数n=0.86。并探讨了它们的相互作用机理，结果表明四碘荧光素主要以疏水作用力与牛血清白蛋白相互作用。同时，研究表明，牛血清白蛋白的存山西大学2以抖届硕士研究生学位论文在会增强四腆荧光素的发光，且强度的变化在一定范围内与牛血清白蛋白的浓度成正比，以此为墓础在pH=8，64的条件下建立了讨定牛血清白蛋白的方法。该法的线性范围为8.0x10一moFL.9.0 xlo石咖比，精密度为3.钱，检出限9.88x lrmo比.第六章:本文考察了丽丝胺罗丹明RB.Zoo的流体荧光和固体基质室温磷光特性，并对影响其发光的条件如:PH条件，固体墓质，重原子效应，烘烤条件等，进行了详细考察.结果表明丽丝胺罗丹明RB一oo的荧光徽发和发射波长分别为5研nml586.rn，磷光的傲发和发射波长为570r助刀02lun。光谱法研究贾丝胺罗丹明RB一00与DNA相互作用的实验表明:DNA的存在会使丽丝胺罗丹明RB.200的吸收光谱产生明显的减色效应，荧光碎灭。阴离子碎灭实验表明，丽丝胺罗丹明RB一oo与DNA作用后受DNA的保护作用，使得阴离子对丽丝胺罗丹明RB.20()的碎灭程度减小，这说明器丝胺罗丹明曲一00可以嵌插在DNA的碱墓对之间。偏振实验表明，在DNA的存在下，丽丝胺罗丹明RB一oo的偏振值增加，这也支持丽丝胺罗丹明RB一oo嵌插在DNA孩基对之间的实脸结果。且离子强度的大小会影响丽丝胺罗丹明RB.20()和DNA的相互作用。第七幸:研究了次甲基紫的流体荧光和固体荃质室温磷光特性。次甲基紫的荧光傲发和发射波长分别为538nml615lun;以滩纸作为固体荃质，以.敌铅为重原子徽扰剂，次甲基紫可以产生磷光发射确光的橄发和发射波长分别为538nmI714nm.用Britton.Robinson缓冲溶液调节体系的酸度，考查酸度对次甲基紫发光强度的形响结果表明:溶液的团值会影响次甲基紫的流体荧光和固体基质室沮礴光强度，但形响不是很大.体系中加入D琳会导致次甲墓紫的荧光碎灭，磷光稗灭和共振光散射光谱的增强。阴离子碎灭实脸表明，DNA的加入使得碘化钾对次甲基紫的碎灭程度减小。这是因为次甲基紫插入洲A的碱基对之间受到有效的保护从而使琪化钾对次甲墓紫的稗灭程度减小。荧光伯振实验表明，在洲A的存在下，次甲墓紫的荧光偏振值会增大。DNA热变性实脸表明:与双链洲人相比，单链渊人对次甲基紫荧光碎灭的程度小。离子强度的大小会形响次甲墓紫和侧A的相互作用。用Se副tCh田吐方程求得二者的结合常数为lxlo勺山。三，结合位点数为1.0.随着侧A浓度的增加，s嘛hard方程的共振光散射光谱一开始会产生明显的增强效应，增强到一定程度后不再变化。明显的增强效应说明次甲基紫可以以侧A为模板进行长距组装.由此可见，次甲基紫和D以之间存在嵌入式结?更多还原

【Abstract】 Chapter 1: The progress of DNA probe had been reviewed, including the kinds of DNA probes, the binding mode of DNA probe to DNA, the methods applied in this field and the applications of probe in DNA study; the progress of protein spectral probe had also been reviewed, including the kinds of protein spectral probes and the spectral analysis methods applied in the study on the interaction of the probe and protein.Chapter 2: The luminescence behavior of diiodofluorescein and tetrabromofluorescein had been investigated including the solid surface room temperature phosphorescence (SS-RTP) and the room temperature fluorescence (RTF). The luminescence intensities of the two compounds were strongest in alkaline solution. RTP lifetime of the two compounds are in the range of 130-140ms. The RTP and RTF polarity was in the range of 0.01-0.05. The two analytical methods-SS-RTP and RTF, of the two compounds had been established.Chapter 3: The interaction of tetrabromofluorescein (TBF) and DNA was studied by the fluorescence and SS-RTP. Deduced from the fluorescence and SS-RTP, there were two binding mode in the whole interaction: intercalation binding and groove binding. The benzopyranyl ring of the TBF intercalated between the ctDNA base pairs supported by potassium ferrocyanide quenching study, and the side chains of the benzene ring extended along the groove of DNA supported by phosphorescence polarization experiment. The effect of ethanol on the fluorescence intensity was smaller in the presence of DNA, compared with that in absence of DNA. The ion strength influenced the binding of TBF and DNA.Chapter 4: The fluorescence and solid surface room temperature phosphorescence (SS-RTP) spectral properties of tetrachlorotetrabromofluorescein (TTF) were investigated. And the factors influencing the fluorescence and phosphorescence emission were discussed. The interaction of TTF and ctDNA was studied by the fluorescence and SS-RTP. The fluorescence and phosphorescence emission changed by the addition of DNA.TTF intercalated into the ctDNA helix supported by fluorescence polarization experiment and potassium iodide quenching study. The ion strength can influence the interaction of TTF and DNA. Based on the fluorescence quench experiment of TTF by ctDNA, the binding constant of TTF binding to ctDNA was 2×106L/mol, the binding site was 0.16.Chapter 5: The binding of tetraiodofluorescein (TIF) to Bovine Serum Albumin (BSA) inaqueous was studied by Fluorimetry. The binding constant was 5.7×104L/mol, and the binding sites were 0.86.The research results of binding model indicated that the hydrophobia force was the main binding force. The presence of BSA could increase the fluorescence intensity of TIF. And the AF was linear to the concentration of BSA in certain range and the first analytical application was proposed. The linear dynamic range (LDR) was 8×10-7mol. L~9× 10-6mol. L, the limit of detection (LOD) was 9.88×10-8mol/L, the relative standard deviation is 3.4%.Chapter 6: The spectral properties of Lissamine Rhodamine RB-200 (LSR) were investigated, including fluorescence and SS-RTP. The excitation and emission wavelength of LSR is 568nm/586nm for fluorescence, and 570nm/702nm for phosphorescence. And the factors influencing the fluorescence and phosphorescence emission were discussed. The emission wavelength of the two compounds were located in longer wavelength which will avoid the interference of the bio-samples. The first analytical method was proposed by SS-RTP. The interaction of LSR and DNA was studied by absorption and fluorescence spectroscopy. The absorption and fluorescence intensity of LSR decreased by the addition of DNA. LSR intercalated into the DNA helix supported by fluorescence polarization experiment and potassium ferrocyanide quenching study. The ion strength influenced the interaction of LSR and DNA.Chapter 7: The fluorescence and SS-RTP spectral properties of methylene violet (MV) were investigated. And the factors influencing the phosphorescence emission were discussed. Th更多还原