Fluorescent probes polarity-sensitive

There are two important drawbacks of such an approach (1) a polarity scale based on a particular class of probes, in principle, does not account, for example, sizes of probes, which should strongly effect the interactions (2) betain dyes do not fluoresce, which restrict essentially the field of application of this approach, because in many cases, absorption spectrum could not be measured accurately (small volumes of samples, study of cells, and single molecules spectroscopy). Therefore, polarity-sensitive fluorescent dyes offer distinct advantage in many applications. 

Sackett, D. L. and Wolff, J. (1987). Nile red as a polarity-sensitive fluorescent-probe of hydrophobic protein surfaces. Anal. Biochem. 167, 228-234. 

Another advantage to examine these polyaers is that characterizations of ablated materials can be Bade possible by fluorescence spectroscopy. Fluorescence is very sensitive, and such surrounding aicroenvironaental conditions around the it -chromophore as polarity and viscosity and chroaophore aggregation can be probed. 

In Section 3.4, structural effects were often discussed in conjunction with the nature of the solvent. As emphasized in the introduction to this book, the fluorescence emitted by most molecules is indeed extremely sensitive to their microenvironment (see Figure 1.3), which explains the extensive use of fluorescent probes. The effects of solvent polarity, viscosity and acidity deserves much attention because these effects are the basis of fluorescence probing of these microenvironmental characteristics and so, later chapters of this book are devoted to these aspects. The effects of polarity and viscosity on fluorescence characteristics in fluid media and the relevant applications are presented in Chapters 7 and 8, respectively. The effect of acidity is discussed in Sections 4.5 and 10.2. This section is thus mainly devoted to rigid matrices or very viscous media, and gases. 

Polarity plays a major role in many physical, chemical, biochemical and biological phenomena. This chapter aims to describe how local polarity can be estimated using a fluorescent probe. But what is polarity This apparently simple question deserves some attention before describing the methodologies based on polarity-sensitive fluorescent probes. 

The polarization properties of light in combination with fluorescence can be used as a powerful tool for determining motional properties of membranes. This is possible due to the fact that the time scale of interest for membrane lipids falls within the time frame of the fluorescence decay phenomena (0-100+ ns). This, coupled with high sensitivity, low perturbing properties of fluorescent probes, and the large number of available probes, makes the fluorescence approach the method of choice for membrane motional studies. 

Fluorescent compounds are sensitive to changes in their chemical environment. Alterations in media pH, buffer components, solvent polarity, or dissolved oxygen can affect and quench the quantum yield of a fluorescent probe (Bright, 1988). The presence of absorbing components in solution that absorb light at or near the excitation wavelength of the fluorophore will have the effect of decreasing luminescence. In addition, noncovalent interactions of the probe with other components in solution can inhibit rotational freedom and quench fluorescence. 

Steady-state and multifrequency phase and modulation fluorescence spectroscopy are used to study the photophysics of a polar, environmentally-sensitive fluorescent probe in near- and supercritical CF3H. The results show strong evidence for local density augmentation and for a distribution of cluster sizes. These results represent the first evidence for lifetime distributions in a "pure solvent system. 

T. Parasassi, E.K. Krasnowska, L. Bagatolli, E. Gratton, Laurdan and prodan as polarity-sensitive fluorescent membrane probes. J. Fluor. 8,365 (1998)... 

FDCD measurements, and a basic theoretical formalism for this technique, were first reported by Turner, Tinoco and Maestre in 1974 [5]. In this experiment one uses the selectivity and sensitivity of luminescence measurements to probe the local chiral environment of fluorescent chromophores. The ultimate goal in many applications of FDCD is to relate the observed differential fluorescence signal to the conventional CD measurement. In certain multi-component absorbing systems this procedure may be difficult. This technique is sometimes applied to systems for which CD measurements are impossible or very difficult. FDCD, like CPL and other polarization sensitive techniques, is not immune to troublesome background and noise problems, and these will be discussed in Section 3. The only detailed discussion of the applicability of FDCD measurements, and other characteristics of the technique has been presented by Turner in 1978 [6]. In this chapter we will also list some of the more recent applications of FDCD. 

Obviously the dipole characteristics of the fluorescent probe and the solvent together with the rigidity of the solvent environment will determine the possibility and extent of any reorientation process. The sensitivity of these fluorescent probes to environmental relaxation processes emphasise that care is required in interpreting the ftuo-res ence behaviour in biodiemical sterns purely in terms of the polarity of the bonding site. The possibility that intramolecular diarge-transfer may occur in the excited states of ANS or that specific solute-solvent complexes may form further com-... 

To study the aggregation of the synthesized polymers, 8-anilino-l-naphthalin-sulfonacid ammonium (ANS) was used as the fluorescence probe. ANS has a minimum solubility of 28gL 1 in water but a higher solubility in hydrophobic media. With increasing polarity of the media its fluorescence intensity increases sensitively, and a simultaneous blue shift of the fluorescence band was also... 

Fluorescence methods are used to study chemical equilibria and kinetics in much the same way as absorption spectrophotometry is used. Often it is possible to study chemical reactions at lower concentrations becau.se of the higher sensitivity of fluorescence methods. In many cases where fluorescence monitoring is ordinarily not feasible, fluorescent probes or tags can be bound covalently to specific sites in molecules such as proteins, thus making them detectable via fluorescence. These tags can be used to provide information about energy transfer processes, the polarity of the protein, and the distances between reactive sites (see, for example. Feature 27-1). 

Novel fluorescence probes for nitric oxide (DAMBO-P ) and for Zn " (ZnAB) have been developed based on BODIPY structure (Fig. 3). DAMBO-P is a pH-independent and more highly sensitive fluorescence probe for nitric oxide than DAF-2. ZnAB has the advantages of less sensitivity to solvent polarity and pH than ZnAF-2, fluorescein-based Zn " probe, and is also not influenced by other cations such as Na", K, Ca and Mg, which exist at high concentration under physiological conditions. 

Another extremely useful method for cac determination, especially in the light of high sensitivity, is fluorescence emission spectroscopy [15]. Some aromatic water-insoluble dyes that are present in trace amounts in mixed polyelectrolyte-surfactant solutions have an ability to solubilize within the self-assembled surfactant aggregates and to change their photophysical properties because of the change of environmental polarity. Through this, they offer a very sensitive method for the determination of cac values. A typical and lately frequently used compound is pyrene, which is used as a fluorescence probe to assess various micellar properties. Pyrene exhibits a polarity dependent fluorescence spectrum with the ratio /,//3 (the ratio of the intensity... 

Fluorescence is very sensitive to the chemical environment and may be utilized to provide information about the microenvironment surrounding the probe (5, 6). The fluorescence intensity (quantum yield) (1-3), the maximum emission wavelength (1-3), the fluorescence lifetime (6-9), or the polarization (10-12) may all be monitored for specific changes that are induced as a result of changes in polarity, pH, ion concentration, membrane potential, or ligand binding. 

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