Intrinsic chromophores and fluorophores

Intrinsic Chromophores and Fluorophores in Synthetic Molecular Receptors... 

An 1,8-naphthyridine q-aminonitrile moiety serves both as an effective donor-acceptor array for complexation of creatinine and as an intrinsic chromophore and fluorophore. In the pH range of 4.1-4.6 the monoprotonated form apparently predominates in 70 % aqueous methanol, producing the absorption spectrum shown in Figure 14. Under these conditions creatinine exists as a mixture of protonated and unprotonated forms, since its pK is approximately 4.2 in this solvent mixture. Such proton-transfer equilibria complicate the calculation of specific stability constants, but under buffered conditions absorption and emission changes result only from complexation, not from proton transfer. As shown in Figure 14, addition of creatinine to a buffered solution decreases the intensity of the 442 nm absorption band attributed to the protonated receptor. Creatinine complexation also quenches the yellow-green fluorescence of the protonated receptor and titration experiments in progress may yield the effective stability constant of the complex. This receptor exemplifies the manner in which intrinsic chromophores and fluorophores may be incorporated into hosts for reversible complexation of clinically important analytes (26). 

Figure 1. Cartoons illustrating potential differences in the optical responses of intrinsic and extrinsic chromophores and fluorophores in synthetic receptors.

A suitable fiuorescent probe is an organic molecule, which must change its characteristic parameters with changes in its microenvironment and the parameter must be measurable when the probe is added to the system [54]. The fluorescent probes are categorized as either extrinsic, intrinsic, or covalently bound probes. The intrinsic probes allow a system to be observed without any chemical perturbation. This occurs when the system to be characterized has an in-built fluorescent chromophore unit like tryptophan, tyrosine and phenyl alanine in protein. In some cases the fluorophore is covalently... 

For chromophores that are part of small molecules, or that are located flexibly on large molecules, the depolarization is complete—i.e., P = 0. A protein of Mr = 25 kDa, however, has a rotational diffusion coefficient such that only limited rotation occurs before emission of fluorescence and only partial depolarization occurs, measured as 1 > P > 0. The depolarization can therefore provide access to the rotational diffusion coefficient and hence the asymmetry and/or degree of expansion of the protein molecule, its state of association, and its major conformational changes. This holds provided that the chromo-phore is firmly bound within the protein and not able to rotate independently. Chromophores can be either intrinsic—e.g., tryptophan—or extrinsic covalently bound fluorophores—e.g., the dansyl (5-dimethylamino-1-naphthalenesulfonyl) group. More detailed information can be obtained from time-resolved measurements of depolarization, in which the kinetics of rotation, rather than the average degree of rotation, are measured. For further details, see Lakowicz (1983) and Campbell and Dwek (1984). 

Certain chromophore systems are intrinsically predisposed for ultrafast single molecule microscopy. Among these, emitters coupled to metal surfaces stand out as exceptionally well-suited subjects. Numerous observations of substantial radiative rate enhancement at the surface or in the vicinity of the surface of a metal were reported. Radiative rate enhancements as large as 10 have been predicted for molecular fluorophores and for semiconductor quantum dots coupled to optimized nanoantennae.Such accelerated emission rates put these systems well within the reach of the emerging femtosecond microscopy techniques. As a result, we decided to apply the Kerr-gated microscope to study of fluorescence dynamics of individual core-shell quantum dots in contact with smooth and nanostructured metal surfaces. 

A fluorophore is a component of a protein or small molecule that exhibits fluorescence. It can also be called a fluorescent label, chromophore, or fluorescent probe. Each fluorophore has characteristic excitation and emission wavelengths. In other words, a fluorophore will fluoresce when the light of a particular energy, corresponding to the excitation wavelength, is used. Examples of fluorophores include GFP and related emissive proteins, and small molecules like fluorescein and coumarin. Many biomolecules have intrinsic fluorescence. For instance, tryptophan, an amino acid, fluoresces in the ultraviolet region of the electromagnetic spectrum. 

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