Fluorescent probe molecule

The sample cells for molecular fluorescence are similar to those for optical molecular absorption. Remote sensing with fiber-optic probes (see Figure 10.30) also can be adapted for use with either a fluorometer or spectrofluorometer. An analyte that is fluorescent can be monitored directly. For analytes that are not fluorescent, a suitable fluorescent probe molecule can be incorporated into the tip of the fiber-optic probe. The analyte s reaction with the probe molecule leads to an increase or decrease in fluorescence. 

F. Mulders, H. van Langen, G. van Ginkel, and Y. K. Levine, The static and dynamic behaviour of fluorescent probe molecules in lipid bilayers, Biochim. Biophys. Acta 859, 209-218 (1986). 

Spectroscopic measurements of solvatochromic and fluorescent probe molecules in room temperature ILs provide an insight into solvent inter-molecular interactions, although the interpretation of the different and generally uncorrelated polarity scales is sometimes ambiguous [23]. It appears that the same solvatochromic probes work in ILs as well [24], but up to now only limited data are available on the behavior of electronic absorption and fluorescence solvatochromic probes within ILs and IL-organic solvent mixtures. 

To further explore the environment inside such hexameric hosts, Atwood and coworkers performed structural characterization and spectrofluorometric studies of the capsule-bound fluorescent probe molecule pyrene butyric acid (PBA) 52 [71]. Single-crystal X-ray diffraction studies on 52 (50b)6 show not only that encapsulated guests interact with the host walls in the solid state, but also that the n surfaces of the guest molecules are well separated from one another within the capsule. The spectroscopic studies in solution corroborated this finding and revealed an average of 1.5 molecules of 52 per capsule. The assembly remains intact over four weeks in the solution phase, suggesting that the carboxylic acid groups and the polyaromatic nature of 52 do not destabilize the overall supramolecular assembly. 

The nitrobenzofurazans used as fluorescent probe molecules in cell membranes were studied by NMR spectroscopy [774-777],... 

Fig. 22. Correlation of percent increase in fluorescence intensity emitted by a rotor-fluorescent probe molecule (p-dimethylaminobenzylidene malononitrile) with the corresponding logarithm of the number, a, of residual sorbed molecules (acetone, CHC13, or THF) per phenyl group of (Sty)97(DVB)3 as described in the text...

Solvatochromic fluorescent probe molecules have also been used to establish solvent polarity scales. The solvent-dependent fluorescence maximum of 4-amino-V-methylphthalimide was used by Zelinskii et al. to establish a universal scale for the effect of solvents on the electronic spectra of organic compounds [80, 213], More recently, a comprehensive Py scale of solvent polarity including 95 solvents has been proposed by Winnik et al. [222]. This is based on the relative band intensities of the vibronic bands I and III of the % - n emission spectrum of monomeric pyrene cf. Section 6.2.4. A significant enhancement is observed in the 0 0 vibronic band intensity h relative to the 0 2 vibronic band intensity /m with increasing solvent polarity. The ratio of emission intensities for bands I and III serves as an empirical measure of solvent polarity Py = /i/Zm [222]. However, there seems to be some difficulty in determining precise Py values, as shown by the varying Py values from different laboratories the reasons for these deviations have been investigated [223]. 

The oxazine dyes are Nile blue A perchlorate (NB) and oxazine 1 perchlorate (0X1), which are ionic dyes, and coumarin 102 is chosen as a fluorescent probe molecule in the dynamic Stokes shift measurements. The electron donating solvents are aniline (AN) and N,N-dimethylaniline (DMA). 

This review is concerned with the use of the time-reK)lved behaviour of fluorescence probe molecules in synthetic and biopolymers, and it will be useful to conader briefly the nature of fluorescence, and phenomena which can influence decay rates and induce spectral changes. 

The binding of various fluorescent probe molecules to biolo cal systems is often accompanied by changes in decay time and emissnn characteristics of the probe ... 

To illustrate the capabilities of the system shown in Fig. 16.12, we present new, previously unpublished results from our laboratory. In Fig. 16.13, we illustrate results for three fluorescent probe molecules (pyrene, DCM [4-(dicyanomethylene)-2-methyl-6-(p-dimethylaminostyryl)-4H-pyran], and PRODAN [6-propionyl-2-(/V,/V-dimethylamino)naphthalene]) that were doped within a series of PFFA/Pluronic P104 BP blends. In the initial experiments, 16 BP formulations were prepared manually using micropipettes, while subsequent experiments utilized the ALHS to prepare 21 BP formulations. These formulations were spun cast into thin films employing quartz microscope slides as substrates. To characterize the local microenvironment surrounding each probe within a given formulation, steady-state fluorescence measurements using a conventional spectrofluorometer were performed. 

Figure 2.27 Some common fluorescence probe molecules...

The polarities of different solvents can be compared using fluorescence probe molecules or solvatochromic dyes. When such molecules are dissolved in a solvent, the frequency of the maximum fluorescence or absorption, respectively, changes as a function of the polarity of the medium, and this property can then be compared with the result for standard solvents. The polarities of several ionic liquids were estimated using 5-dimethylamino-isoindole-l,3-dione 34 as a fluorescence probe and Reichardt s dye 35 for visible absorbance measurements, allowing the determination of the normalized parameter ( . (tetramethylsilane) = 0,... 

To avoid the rather large perturbation introduced by fluorescent probe molecules, the use of water-soluble and small ions is recommended to probe the polarity of the inner water cores [22,69-71]. 

Another type of micelle formation has also attracted the attention of researchers, a brief mention of which will be made below. Gravsholt [93], in an early work, reported viscoelasticity in highly diluted aqueous solutions of some cationic surfactants, namely, certain derivatives of the base structure of cetyltrimethylammonium bromide (CTAB), CTA-X (where X=salicylate, m-chlorobenzoate, p-chlorobenzoate). He came to the conclusion that the viscoelastic behavior indicates a micellar shape other than spheres and rods. Later it has been shown [e.g. 94] by fluorescence anisotropy using fluorescent probe molecules that CTAB and sodium p-toluenesulfonate, NapTS (with suitably weak fluorescence) in aqueous solution could produce long, threadlike micelles with network structure in which the cross-points had finite lifetime. Sodium salicylate was another useful additive for synthesis, but the intensity of its fluorescence was found unsuitable for examining the behavior of the probes. 

Studies of the microenvironment of fluorescent probe molecules. Many environmental factors modify the fluorescent properties of a molecule, e.g., solvent polarity, pH, proximity, and concentration of quenching species. The changes experienced by the fluorophore, sometimes subtle, can be used to obtain information about the specific region in which the fluorophore is localized. 

The use of fluorescent probe molecules results in information on the molecular environment in the vicinity of the chromophore, interpreting spectral shifts, changes in fluorescence lifetimes and fluorescence polarization. 

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