同步荧光分析法的应用及其新进展*

同步荧光技术是解决多组分荧光物质同时测定的良好手段之一.本文从恒波长同步荧光法、恒能量同步荧光法、可变角同步荧光法、恒基体同步荧光法以及它们与导数技术、低温技术、化学计量学方法的联用等方面对同步荧光技术领域出现的新技术及应用作一评述.     

苯胺和1－萘酚的导数－可变角同步荧光法分析

苯胺和１－萘酚的导数－可变角同步荧光法分析李耀群，黄贤智，许金钩（厦门大学化学系分析化学研究所厦门３６１００５）关键词荧光，同步扫描，导数技术，苯胺，萘酚同步荧光法可同时分析多组分荧光物质［１］．可变角同步荧光法（ＶＡＳＦＳ）与恒波长同步荧光法相比，...     

β—CD存在下邻菲罗啉—二溴苯基荧光酮荧光熄灭法测定银

银的荧光分析法可分为点滴法、荧光熄灭法、直接荧光法和催化荧光法,荧光熄灭体系中已采用的试剂有曙红、荧光素、曙红加邻菲罗啉等.4,5-二溴苯基荧光酮已应用于荧光法测定其它元素,本文以β-环糊精(β-CD)为稳定剂,银与邻菲罗啉及4,5-二溴苯基荧光酮(DBPF)形成离子缔合型异配位体配合物,使荧光熄灭体系灵敏度大大提高.详细研究了异配位体配合物的最佳形成条件,在EDTA存在下可直接测定阳极泥中的银,结果满意.     

卡尔曼滤波荧光分析法用于铝,镓的同时测定

研究了SPAEC荧光螯合反应并结合卡尔曼滤波校正的高灵敏度高选择性荧光分析法.在荧光分析中首次用卡尔曼滤波以分辨铝、镓与SPAEC螯合物的重叠峰;提出了适于荧光分析的几种滤波方式,并讨论了各自的特点.该法用于铝与镓(1:40～14:1)混合物分析,改善了灵敏荧光法(及试剂)的选择性.     

鸭肉中克百威和西维因的同步荧光光谱解析及定量分析

采用同步荧光法分析了鸭肉中农药的荧光特征,其次利用同步荧光峰值法对鸭肉中农药残留进行定量分析。发现2种农药鸭肉体系中都有一个同步荧光峰,分别是克百威的荧光特征(Δλ=120 nm,λex=335 nm)和西维因的荧光特征(Δλ=140 nm,λex=322 nm);在一定浓度范围内荧光强度与浓度良好的线性关系,其克百威-鸭肉和西维因-鸭肉体系中预测样本的相关系数(R2)分别为0.991和0.994。为快速、准确检测鸭肉中农药残留含量提供了新方法。     

氢化松香酸钠与明胶蛋白质相互作用的荧光光谱法研究

在pH＝10.80及不同温度下,, 利用常规荧光法、同步荧光法、共振光散射和紫外-可见光谱研究了氢化松香酸钠(HSR)与明胶的相互作用. 结果表明HSR与明胶有一个结合位点,, 聚集体的形成引起明胶内源荧光增强.. 据此建立了体系荧光增强与HSR浓度的线性关系式,, 计算了不同温度下反应的结合常数及对应温度下结合反应的热力学参数.. 二者之间主要靠疏水作用力结合..     

稀土金属离子及pH诱导牛血清白蛋白构象变化的同步荧光光谱研究

用同步荧光法并辅以普通荧光法对不同浓度稀土离子及pH诱导的牛血清白蛋白(BSA)构象变化进行了详细研究, 发现稀土离子使色氨酸(Trp)残基的荧光蓝移, 荧光强度降低; 酪氨酸(Tyr)残基荧光峰位置不变, 稀土离子浓度较低时, 荧光峰强度降低, 而当浓度较高时, Tyr残基荧光峰强度反而增强. 据此推断了稀土离子与BSA结合反应中Trp残基微环境和Tyr残基微环境及构象的变化并与pH引起的变化进行了比较.     

导数-可变角同步荧光法用于1-萘酚和2-萘酚的同时测定

同步扫描和导数光谱法是两类很有用的荧光分析新技术。固定波长或恒能量同步荧光法和导数技术的结合可进一步提高光谱分辨率和排除基体干扰。本文将导数技术和可变角同步荧光法结合起来,用于1-萘酚和2-萘酚的同时分析,效果良好。研究表明,导数-可变角同步荧光法可望发展为荧光分析复杂样品的新技术。     

同步荧光法同时测定苏丹红Ⅱ和苏丹红Ⅲ

开展了苏丹红的荧光分析研究, 为快速检测苏丹红提供了新的方法. 所建立的恒波长同步荧光法可同时对苏丹红Ⅱ和Ⅲ进行定性定量检测. 荧光方法具有很大的应用前景, 有望成为苏丹红的常规、快速、 简便的检测方法.     

中药注射剂荧光光谱法的快速鉴别和热稳定性研究

红外光谱指纹图谱技术应用中药注射剂的快速鉴别中，某些试样因难于成膜造成制样困难，为此，直接或仅经水稀释后测定了12种21批次的常用中药注射剂的荧光光谱。结果表明：由于中药注射剂常含荧光性物质，但因不同注射剂所含具体荧光性物质不同，同种注射剂又因厂家的具体配方的差异或制备工艺条件的波动，均会使其特征的荧光激发、发射光谱不同或其强度呈现差异；从而可充分利用荧光分析的高灵敏度，使其作为中药整体红外指纹识别的一种辅助手段，根据注射剂的荧光图谱的差异达到快速鉴别、认定和控制配方、工艺的目的。比较而言，荧光法表现出来的差异更为一目了然，易于判断。此外，荧光光谱法还可用于中药注射液的热稳定性的研究。     

Monofluorination and Trifluoromethylation of BODIPY Dyes for Prolonged Single‐Molecule Detection

Electrophilic monofluorination with Selectfluor and nucleophilic trifluoromethylation with the Ruppert–Prakesh reagent of dimethyl‐, tetramethyl‐ and pentamethyl‐substituted boron dipyrromethenes (BODIPY) are investigated. Monofluorinated dyes are synthesized with low yields (<30 %), however trifluoromethyl derivatives are obtained in moderate to high yields (≈40–90 %). All compounds are characterized by steady‐state and time‐resolved fluorescence spectroscopy, the photostability is investigated with fluorescence correlation spectroscopy (FCS) and total internal reflection fluorescence microscopy (TIRF). Monofluorination hardly affects the spectroscopic parameters of the unsubstituted parent compounds, but distinctly enhances the photostability, whereas trifluoromethylation leads to a hypsochromic shift by up to 17 nm in both absorption and emission, slightly enhanced intersystem crossing, and higher photostability. Further development of soft fluorination and trifluoromethylation methods is therefore highly desired.     

Time-resolved fluorescence spectroscopy and imaging of proteinaceous binders used in paintings

The differentiation of proteins commonly found as binding media in paintings is presented based on spectrally resolved and time-resolved laser-induced fluorescence (LIF) and total emission spectroscopy. Proteins from eggs and animal glue were analysed with pulsed laser excitation at 248 nm (KrF excimer) and 355 nm (third harmonic of Nd:YAG) for spectrally resolved measurements, and at 337 nm (N₂) and 405 nm (N₂ pumped dye laser) for spectrally resolved lifetime measurements and fluorescence lifetime imaging (FLIM). Total emission spectra of binding media are used for the interpretation of LIF spectra. Time-resolved techniques become decisive with excitation at longer wavelengths as fluorescence lifetime permits the discrimination amongst binding media, despite minimal spectral differences; spectrally resolved measurements of fluorescence lifetime have maximum differences between the binding media examined using excitation at 337 nm, with maximum observed fluorescence at 410 nm. FLIM, which measures the average lifetime of the emissions detected, can also differentiate between media, is non-invasive and is potentially advantageous for the analysis of paintings. Figure The fluorescence of solid ox glue extracted from collagen can be visualised in this Total Fluorescence Spectrum; three different peaks from multiple fluorophores are present and allow the discrimination between collagen- and non-collagen proteinaceous binding media found in paintings     

Isomerization and fluorescence depolarization of merocyanine 540 in polyacrylic acid. Effect of p H

Dynamics of isomerization and fluorescence depolarization of merocyanine 540 (MC540) in an aqueous solution of polyacrylic acid (PAA) have been studied using picosecond time resolved fluorescence spectroscopy. It is observed that the dynamics of isomerization and depolarization are sensitive enough to monitor the uncoiling of PAA at high pH (> 6). At low pH (< 3), when the polymer remains in a hypercoiled form, polymer bound MC540 experiences very high microscopic friction and, hence, the isomerization and depolarization processes are very slow. At high pH (> 6) a polyanion is formed and the polymer assumes an extended configuration due to electrostatic repulsion. At high pH (> 6), the anionic probe MC540 is expelled from the polyanion to bulk water and the dynamics of isomerization and fluorescence depolarization become faster by 12 and 5 times respectively, compared to those at low pH.     

Fluorescence behavior of individual charge-transfer complexes revealed by single-molecule fluorescence spectroscopy: Influence of the host polymer matrix

Extremely pure polymer matrices were used for elucidating the fluorescence properties of singly isolated charge-transfer (CT) complexes formed between the donor N-ethylcarbazole and the acceptor 1,2,4,5-tetracyanobenzene. Simultaneous measurements (time traces of CT fluorescence intensities and lifetimes) using single-molecule fluorescence spectroscopy showed three patterns: (1) fluctuations in the fluorescence intensities and lifetimes seldom occurred, (2) the fluorescence intensities frequently fluctuated together with the lifetimes, or (3) in addition to the above fluctuations with time, blinking and/or off-states longer than 1 s were observed. For methacrylate polymers, both the degree of fluctuations in the CT fluorescence lifetimes and the percentage of the CT complexes showing off-states increased with the free volume of the host polymers. These results suggest that the degree of fluctuations in the relative geometrical arrangements of the donor and acceptor molecules is related to the availability of space in the host polymer, and that the free volume provides the necessary space for formation of non-fluorescent donor-acceptor geometries of the CT complexes and/or temporal dissociation of the CT complexes. Survival times of the CT fluorescence were also closely related with the free volume of the host polymers.     

Nanowires Formed by the Co‐Assembly of a Negatively Charged Low‐Molecular Weight Gelator and a Zwitterionic Polythiophene

Conjugated organic nanowires have been prepared by co‐assembling a carboxylate containing low‐molecular weight gelator (LMWG) and an amino acid substituted polythiophene derivative (PTT). Upon introducing the zwitterionic polyelectrolyte PTT to a basic molecular solution of the organogelator, the negative charges on the LMWG are compensated by the positive charges of the PTT. As a result, nanowires form through co‐assembly. These nanowires are visualized by both transmission electron microscopy (TEM) and atomic force microscopy (AFM). Depending on the concentration and ratio of the components these nanowires can be micrometers long. These measurements further suggest that the aggregates adopt a helical conformation. The morphology of these nanowires are studied with fluorescent confocal laser scanning microscopy (CLSM). The interactions between LMWG and PTT are characterized by steady‐state and time‐resolved fluorescence spectroscopy studies. The steady‐state spectra indicate that the backbone of the PTT adopts a more planar and more aggregated conformation when interacting with LMWG. The time‐ resolved fluorescence decay studies confirm this interpretation.     

醇对水中DDAHPS分子聚集行为的影响

分别用稳态荧光猝灭技术和时间分辨荧光方法研究醇对两性表面活性剂 (十二烷基羟基磺化甜菜碱 ,DDAHPS)在水中聚集行为的影响 .结果表明 :在 4.0 0× 1 0 - 2 mol·L- 1DDAHPS水溶液中 ,随着正丁醇浓度的逐渐增大 ,表面活性剂的聚集数 (N)逐渐降低 .恒定醇的浓度为 2 .1 4× 1 0 - 2mol·L- 1时 ,醇碳链越长 ,N值越大 .与无醇体系相比 ,正丙醇和正丁醇使N值变小 ,正戊醇、正己醇和正庚醇使N值变大 .本文同时还测定了醇对微环境的极性 ,芘的荧光寿命及胶束内芘的激基缔合物形成效率的影响 .     

半花菁衍生物LB膜的光致荧光特性研究

通过改变半花菁的亲水基团与疏水基团而得到了四种半花菁衍生物 ,利用稳态和时间分辨荧光研究了不同亲水基团与疏水基团对半花菁衍生物光学特性的影响.主要包括半花菁衍生物在LB膜中的聚集特性;亲水基团与疏水基团对半花菁衍生物激发态寿命的影响等.     

Detection of Explosive Vapors: The Roles of Exciton and Molecular Diffusion in Real‐Time Sensing

Time‐resolved quartz crystal microbalance with in situ fluorescence measurements are used to monitor the sorption of the nitroaromatic (explosive) vapor, 2,4‐dinitrotoluene (DNT) into a porous pentiptycene‐containing poly(phenyleneethynylene) sensing film. Correlation of the nitroaromatic mass uptake with fluorescence quenching shows that the analyte diffusion follows the Case‐II transport model, a film‐swelling‐limited process, in which a sharp diffusional front propagates at a constant velocity through the film. At a low vapor pressure of DNT of ≈16 ppb, the analyte concentration in the front is sufficiently high to give an average fluorophore–analyte separation of ≈1.5 nm. Hence, a long exciton diffusion length is not required for real‐time sensing in the solid state. Rather the diffusion behavior of the analyte and the strength of the binding interaction between the analyte and the polymer play first‐order roles in the fluorescence quenching process.     

Theoretical study of determining orientation and alignment of symmetric top molecule using laser-induced fluorescence

General expressions used for extracting the orientation and alignment parameters of a symmetric top molecule from laser-induced fluorescence (LIF) intensity are derived by employing the density matrix approach. The molecular orientation and alignment are described by molecular state multipoles. Excitation and detection are circularly and linearly polarized lights, respectively. In general cases, the LIF intensity is a complex function of the initial molecular state multipoles, the dynamic factors and the excitation-detection geometrical factors. It contains a population, ten orientation and fourteen alignment multipoles. The problem of how to extract the initial molecular state multipoles from the resolved LIF intensity is discussed.     

Effect of Surface Modification on Semiconductor Nanocrystal Fluorescence Lifetime

Semiconductor nanocrystals, namely, quantum dots (QDs), present a set of unique photoluminescence properties, which has led to increased interest in using them as advantageous alternatives to conventional organic dyes. Many applications of QDs involve surface modification to enhance the solubility or biocompatibility of the QDs. One of the least exploited properties of QDs is the very long photoluminescence lifetime that usually has complex kinetics owing to the effect of quantum confinement. Herein, we describe the effect of different surface modifications on the photoluminescence decay kinetics of QDs. The different surface modifications were carefully chosen to provide lipophilic or water‐soluble QDs with either positive or negative surface net charges. We also survey the effect on the QD lifetime of several ligands that interact with the QD surface, such as organic chromophores or fluorescent proteins. The results obtained demonstrate that time‐resolved fluorescence is a useful tool for QD‐based sensing to set the basis for the development of time‐resolved‐based nanosensors.