Fluorescence Interaction and Determination of Calf Thymus DNA with Two Ethidium Derivatives

In this paper, we reported the syntheses and investigation of the modes of binding to DNA of the two new ethidium derivatives containing benzoyl and phenylacetyl groups of both amines at 3-and 8- positions. The interactions between calf thymus DNA (ct-DNA) and the two derivatives, 3,8-dibenzoylamino-5-ethyl-6-phenylphenantridinium cloride (E2) and 3,8-diphenylacetylamino-5-ethyl-6-phenylphenantridinium chloride (E3), were investigated by fluorescence quenching spectra and UV-vis absorption spectra. The Stern-Volmer quenching constants, binding constants, binding sites and the corresponding thermodynamic parameters ΔH, ΔS and ΔG were calculated at different temperatures. The results indicated the formation of E2 and E3-DNA complexes and van der Waals interactions as the predominant intermolecular forces in stabilizing for each complex. In addition, increasing nucleophilicity of the functional groups at 3- and 8- positions exhibited the respectable increment the DNA binding affinities of derivatives. The results of absorption, ionic strength and iodide ion quenching suggested that the interaction mode of E2 and E3 with ct-DNA was intercalative binding. The limit of detection (LOD) of ct-DNA were 7.49 × 10⁻⁸ (n = 4) and 4.18 × 10⁻⁸ mol/l (n = 7) in presence of E2 and E3, respectively.     

血卟啉及金属血卟啉与甲氧卞氨嘧啶相互作用的荧光光谱研究

荧光光谱法研究了甲氧卞氨嘧啶与血卟啉及金属血卟啉的作用情况 ,考察了酸度对甲氧卞氨嘧啶荧光强度的影响。在 p H=11.0的 KH2 PO4- Na2 HPO4缓冲液中 ,甲氧卞氨嘧啶有较强的荧光峰 ,血卟啉及金属血卟啉对甲氧卞氨嘧啶有明显的识别作用 ,结合常数大于 10 5,结合比均为 1∶ 1。金属血卟啉与甲氧卞氨嘧啶的结合常数小于血卟啉与甲氧卞氨嘧啶的结合常数 ,并对识别机理进行了探讨。     

荧光法研究左氧氟沙星与人血清白蛋白作用机理及含量测定

利用荧光及紫外光谱法研究了水溶液体系中左氧氟沙星(LVFX)与人血清白蛋白(HSA)的相互作用机制。结果表明左氧氟沙星对人血清白蛋白的荧光有较强的猝灭作用,其猝灭类型主要为静态猝灭。在不同温度下求得了左氧氟沙星与人血清白蛋白的结合常数k,发现随反应温度上升k值下降。由热力学参数,确定了左氧氟沙星与人血清白蛋白的结合作用主要为色散力。用同步荧光技术考察了左氧氟沙星对人血清白蛋白构象的影响,又根据Foerster理论,测得了左氧氟沙星与人血清白蛋白结合的能量转移效率,相互结合距离。进一步证明了该反应是单一静态猝灭过程,阐述了其猝灭机理是通过能量转移产生的。研究发现人血清白蛋白荧光猝灭程度的对数与左氧氟沙星含量有良好的线性关系,25℃时线性范围为:2.0×10-6—2.4×10-5mol·L-1,检出限为1.2×10-7mol·L-1。用于片剂中左氧氟沙星含量的测定结果满意,相对标准偏差为:1.5%—2.1%,回收率为:94.8%—97.7%。     

Interaction of Imidacloprid with Hemoglobin by Fluorescence and Circular Dichroism

Imidacloprid belongs to a major new class of insecticides, called neonicotinoids, which are accounting for 11–15% of the total insecticide market. The binding characteristics of insecticide imidacloprid with hemoglobin (Hb) have been studied by employing different spectroscopic techniques. The results proved the formation of complex between imidacloprid and Hb. Hydrophobic interaction and hydrogen bond dominated in the association reaction. Hydrophobic probe 8-anilino-1-naphthalenesulfonic acid (ANS) competitive experiments indicated that the binding of imidacloprid to Hb primarily took place in hydrophobic regions. The distance between Hb donor and acceptor imidacloprid was 4.88 nm as derived from Förster’s theory. Alternations of Hb secondary structure in the presence of imidacloprid were confirmed by synchronous fluorescence, circular dichroism (CD) and three-dimensional fluorescence spectra. This study enriches our understanding of toxic effect of imidacloprid to the physiologically important protein Hb.     

SYBR Green I: Fluorescence Properties and Interaction with DNA

In this study, we have investigated the fluorescence properties of SYBR Green I (SG) dye and its interaction with double-stranded DNA (dsDNA). SG/dsDNA complexes were studied using various spectroscopic techniques, including fluorescence resonance energy transfer and time-resolved fluorescence techniques. It is shown that SG quenching in the free state has an intrinsic intramolecular origin; thus, the observed >1,000-fold SG fluorescence enhancement in complex with DNA can be explained by a dampening of its intra-molecular motions. Analysis of the obtained SG/DNA binding isotherms in solutions of different ionic strength and of SG/DNA association in the presence of a DNA minor groove binder, Hoechst 33258, revealed multiple modes of interaction of SG inner groups with DNA. In addition to interaction within the DNA minor groove, both intercalation between base pairs and stabilization of the electrostatic SG/DNA complex contributed to increased SG affinity to double-stranded DNA. We show that both fluorescence and the excited state lifetime of SG dramatically increase in viscous solvents, demonstrating an approximate 200-fold enhancement in 100?% glycerol, compared to water, which also makes SG a prospective fluorescent viscosity probe. A proposed structural model of the SG/DNA complex is compared and discussed with results recently reported for the closely related PicoGreen chromophore.     

蝎毒素LmKTT-1a 与胰蛋白酶相互作用研究

LmKTT-1a 是最近发现的一个蝎毒素,该多肽分子不仅具有钾离子通道调节剂的功能,还具有胰蛋白酶（Trypsin）抑制剂的活性,是第一个被发现的双功能蝎毒素．虽然前期工作已对LmKTT-1a 的溶液结构和钾离子通道调节剂的功能进行了研究,但未阐明LmKTT-1a 和Trypsin 的作用方式和位点．文中利用液体NMR 的手段,采用化学位移扰动分析的方法,确定了LmKTT-1a 的loop 区域V10~F17 等氨基酸位点可能参与了与Trypsin的相互作用．进一步采用定点突变的方法验证了NMR 实验的结果,并确认了LmKTT-1a与Trypsin 相互作用最关键的氨基酸位点是K14．该研究结果有助于进一步理解LmKTT-1a具有双功能活性的结构基础．     

Fluorescence Property of ZnO Nanoparticles and the Interaction with Bromothymol Blue

We synthesized ZnO quantum dots (QDs) simply in alcoholic solution, and investigated the interaction between ZnO QDs and bromothymol blue. The structural, morphological, size and spectral properties of ZnO QDs were studied. It was found that ZnO QDs were spherical nanoparticles in the crystal structure, and the average diameter of ZnO QDs was about 4.8 nm. The excitation and emission peaks were located at 346 nm and 520 nm, respectively, which were obtained on a common fluorophotometer. The quantum yield of ZnO QDs was obtained by using quinine sulfate as a reference reagent. In addition, the fluorescence of ZnO QDs can be quenched by bromothymol blue, and the quenching mechanism was proposed in a dynamic quenching mode.     

水溶性富勒醇的荧光特性及其与各种金属离子结合的研究

本文考察了C60 水溶性衍生物 富勒醇的荧光特性。观测到富勒醇的荧光自猝灭现象以及不同酸度对其荧光强度的影响。在 pH =6 5～ 7 5条件下 ,富勒醇的荧光发射最强 ,并以此为最佳酸度条件 ,详尽研究了富勒醇同各种金属离子的作用 ,发现金属离子Cu2 + ,Pb2 + ,Co2 + ,Fe2 + ,Fe3 + ,Al3 + ,Cr3 + 和Cr(Ⅵ )均能有效猝灭富勒醇的荧光。其中Cu2 + ,Fe2 + ,Fe3 + ,Al3 + 和Cr3 + 在猝灭富勒醇荧光的同时 ,还使富勒醇的荧光激发和发射峰发生不同程度的红移和蓝移 ,表明这五种金属离子与富勒醇之间同时形成了基态和激发态的络合物 ;而Pb2 + ,Co2 + 和Cr(Ⅵ )则只在激发态下同富勒醇发生作用。各金属离子与富勒醇的结合能力为 :Cu2 + >Fe2 + >Pb2 + >Co2 + ;Fe3 + >Al3 + >Cr(Ⅵ ) >Cr3 + ,同时估测了各金属离子与富勒醇的结合常数KA 及结合数n。     

光谱法结合分子对接模拟技术研究头孢他啶与胰蛋白酶的相互作用机理

本文主要通过荧光光谱法与分子对接技术研究了在298,303,310 K温度下头孢他啶(CFD)与胰蛋白酶(TRP)之间的作用机制。研究结果表明,CFD与TRP之间是通过1∶1的静态猝灭方式相互作用。依照双对数方程处理荧光猝灭数据得到了CFD与TRP作用的结合常数Ka和结合位点数n。通过热力学方程求得了不同温度下CFD与TRP作用的热力学参数。实验数据表明,它们之间的作用力主要是疏水作用和氢键作用,这与分子对接技术所得的结果是一致的。     

荧光光谱法研究胆红素与牛血清白蛋白的相互作用

在模拟生理条件下,利用荧光光谱法研究了胆红素BR和牛血清白蛋白的相互作用。实验结果表明胆红素BR对牛血清白蛋白有较强的荧光猝灭作用,两者形成了新的复合物,属于静态荧光猝灭,发生分子内的非辐射能量转移。根据荧光猝灭法得到胆红素与牛血清白蛋白的结合常数和结合位点数。根据Foster非辐射能量传递理论确定了胆红素BR和牛血清白蛋白间的结合距离。此外,利用同步荧光光谱分析胆红素BR对牛血清白蛋白构象的影响。     

荧光光谱法研究咖啡因与肌红蛋白的相互作用

采用荧光光谱法在生理pH 7.4条件下研究了药物咖啡因与肌红蛋白分子间的相互作用,表明这种相互作用能使肌红蛋白的内源荧光猝灭。通过猝灭常数,结合常数和结合位点数的计算,证明此猝灭为静态猝灭机制。咖啡因和肌红蛋白形成1∶1稳定配合物,形成常数(18 ℃)K_A=1.82×104 L·mol-1;根据热力学参数确定了它们之间的主要作用力为疏水力和静电力。利用同步荧光光谱法研究了咖啡因对肌红蛋白构象的影响。咖啡因能使肌红蛋白的构象发生改变,导致蛋白质分子中色氨酸和酪氨酸残基所处微环境由原来的疏水环境不同程度地向亲水环境转变。     

荧光光谱法研究2-硝基苯胺与牛血清白蛋白的相互作用

采用荧光光谱法研究了不同酸度下,2-硝基苯胺与牛血清白蛋白(BSA)间的相互作用。实验结果表明,2-硝基苯胺对BSA的猝灭机制属于静态猝灭过程。经研究得到了不同酸度和不同温度下2-硝基苯胺与BSA反应的结合常数、结合位点数。根据热力学常数确定了二者间的作用力类型为氢键和疏水作用力。同步荧光结果表明,2-二硝基苯胺的存在改变了牛血清白蛋白的分子构象。     

Investigation of the Interaction Between Rutin and Trypsin in Solution by Multi-Spectroscopic Method

ABSTRACT

The interactions between rutin and trypsin were investigated by UV-Vis absorption, CD, fluorescence, resonance light-scattering spectra, synchronous fluorescence, and three-dimensional fluorescence spectra techniques under physiological pH 7.40. Rutin effectively quenched the intrinsic fluorescence of trypsin via static quenching. The enthalpy change and entropy change were estimated to be ?8.23 kJ·mol^?1 and 53.66 J·mol^?1·K^?1 according to the van't Hoff equation. The process of binding rutin to trypsin was a spontaneous molecular interaction procedure. This result indicates that hydrophobic and electrostatic interactions played a major role in stabilizing the complex. The conformation of trypsin was discussed by CD, synchronous, and three-dimensional fluorescence techniques.     

荧光光谱法研究盐酸胍浓度不同时变性胰蛋白酶的构象变化

蛋白变性过程中间体的存在是蛋白变性及复性动力学研究中不可缺少的证据。以胰蛋白酶为模型蛋白 ,用荧光光谱法系统地研究了在不同浓度变性剂盐酸胍存在时胰蛋白酶构象的变化 ,并与活性数据进行了对比。发现胰蛋白酶荧光光谱发射波长随变性剂盐酸胍浓度增大而逐渐增大 ,并且当盐酸胍浓度达到 2mol·L- 1 时胰蛋白酶的最大发射波长达到最大值 ,其后随盐酸胍浓度的增大最大发射波长反而逐渐减小 ,当盐酸胍浓度大于 3mol·L- 1 呈现不变的趋势。也就是说 ,在低浓度变性剂环境下 ,胰蛋白酶存在着一个与天然态和完全变性态的分子构象都不同的中间体状态 ,这个中间体状态的荧光发射波长最大 ,荧光发射强度也最大 ,而以此状态为复性起点 ,最终得到的复性产率也最低。对此原因从分子结构的基础上进行了探讨。     

TNS与牛血清白蛋白相互作用的荧光光谱研究

在pH 7.0、20mmol·L-1磷酸盐缓冲溶液(PB)及室温条件下,利用荧光光谱研究了药物模拟物TNS与牛血清白蛋白的作用.结果表明TNS与牛血清白蛋白有一个强结合位点,其条件结合常数为 (4.86±0.14)×105L·mol-1;结合主要以疏水作用为主,静电结合也起一定作用;依据Forster能量转移机制得到授体-受体间距离r=2.10nm;该条件下被环糊精包结的TNS也可以转化为TNS-牛血清白蛋白复合物.     

对硝基苯酚与牛血清白蛋白相互作用的荧光光谱研究

在模拟生理条件下应用荧光光谱和紫外-可见吸收光谱技术研究了牛血清白蛋白(BSA)与对硝基苯酚的相互作用.通过修正Stern-Volmer方程求算出在不同温度下(304,307,310K)的淬灭常数为1.186,1.030,0.940×105 L·mol-1,证实了BSA与对硝基苯酚相互结合作用为单一的静态猝灭过程,并且得到对硝基苯酚与BSA相互作用的热力学参数△H0和△S0分别为-28.061kJ·mol-1和-14.331J·mol-1·K-1,推出两者的相互作用以氢键为主.根据非辐射能量转移理论,计算出对硝基苯酚与BSA相互结合时其供体-受体间的结合距离(r=5.94nm)和能量转移效率(E=0.09).     

Interaction of Thyroxine with 7 Hydroxycoumarin: A Fluorescence Quenching Study

The interaction between thyroxine hormone and 7 hydroxycoumarin (7HC) was investigated using fluorescence quenching method. The experimental results showed that thyroxine could quench the fluorescence of 7HC by forming the 7HC–thyroxine complex with static quenching. The apparent binding constants (K) between 7HC and thyroxine were determined to be 1.51 × 10⁴ (297 K) and 9.06 × 10³ (310 K). The binding sites (n) 0.98 ± 0.1. The thermodynamic parameters showed that the interaction between 7HC and thyroxine was driven mainly by hydrogen bonding interactions and van der Waals force. Calibration for thyroxine, based on quenching titration data, was linear in the concentration range 2.0 × 10⁻⁸ to 3.0 × 10⁻⁷ mol/l. The relative standard deviation was 2.58% for 2.0 × 10⁻⁷ mol/l thyroxine (n = 4) and the 3σ limit of detection was 3.42 × 10⁻⁸ mol/l in cationic surfactant CTAB medium.     

Fluorescence Study on the Interaction of Bovine Serum Albumin with P-Aminoazobenzene

In this paper, the interaction between p-aminoazobenzene (PAAB) and BSA was investigated mainly by fluorescence quenching spectra, circular dichroism (CD) and three-dimensional fluorescence spectra under simulative physiological conditions. It was proved that the fluorescence quenching of BSA by PAAB was mainly a result of the formation of a PAAB-BSA complex. The modified Stern-Volmer quenching constant K _a and the corresponding thermodynamic parameters ΔH, ΔG and ΔS at different temperatures were calculated. The results indicated that van der Waals interactions and hydrogen bonds were the predominant intermolecular forces in stabilizing the complex. The distance r?=?4.33 nm between the donor (BSA) and acceptor (PAAB) was obtained according to Förster’s non-radioactive energy transfer theory. The synchronous fluorescence, CD and three-dimensional fluorescence spectral results showed that the hydrophobicity of amino acid residues increased and the losing of α-helix content (from 63.57 to 51.83%) in the presence of PAAB. These revealed that the microenvironment and conformation of BSA were changed in the binding reaction.     

苏木精与DNA作用机理的荧光光谱

采用荧光光谱法,在生理条件下(pH=7.40)对苏木精与DNA的作用机理进行了研究,通过对体系盐效应和KI效应的实验以及体系的荧光猝灭所求得的热力学参数,证明苏木精通过静电作用和疏水作用两种方式与DNA相互作用.     

Fluorescence Study of Sinapic Acid Interaction with Bovine Serum Albumin and Egg Albumin

The mechanism of interaction of protein with compounds used for preparation of matrices for matrix-assisted laser desorption ionization–mass spectrometry (MALDI-MS) methods is unknown. This paper reports the investigation of this mechanism for sinapic acid and bovine serum albumin and egg albumin. To examine these interactions in water a fluorescence method was applied. Sinapic acid can exist in three different forms, depending on pH: undissociated and with one or two deprotonated groups. pK_as of these states are: 4.47 for the COOH group and 9.21 for the OH group [1]. Therefore the interactions were examined at pH: 2.0, 6.4, and 10.5. The results show that sinapic acid at pH 10.5, being a bivalent anion, does not form any complex with these two proteins. At pH 2.0, sinapic acid, being undissociated, interacts weakly with egg albumin. Sinapic acid does not interact with bovine serum albumin at this pH. At pH 6.4, sinapic acid interacts only with bovine serum albumin. Parameters of the sinapic acid and bovine serum albumin complex were calculated based on the theory of multiple equlibria: the total number of binding sites, N = 15; the binding constant, K = 600 M ^–1; and the Hill's coefficient, j = 0.97. These parameters indicate (but not definitively because a large saturation was not obtained) that this is a simple binding of sinapic acid to bovine serum albumin with the binding sites of the same type.