High fluorescence method

Note that in liquid phase chromatography there are no detectors that are both sensitive and universal, that is, which respond linearly to solute concentration regardless of its chemical nature. In fact, the refractometer detects all solutes but it is not very sensitive its response depends evidently on the difference in refractive indices between solvent and solute whereas absorption and UV fluorescence methods respond only to aromatics, an advantage in numerous applications. Unfortunately, their coefficient of response (in ultraviolet, absorptivity is the term used) is highly variable among individual components. 

A highly sensitive method for the determination of anionic surfactants, particularly sodium dodecyl sulfate, has been described [275]. The method is based on the formation of fluorescent ionic complexes of the anionic surfactant with acridine red and acridine yellow. The complexes are extracted with dichloro-... 

In current practice the fluorescence assay is often followed by the use of hybridization techniques when more selectivity is required. We have for instance used the fluorescence techniques to obtain data on the nucleic acid content of malaria vaccine proteins produced in Escherichia coli. The rapid turnaround time of the fluorescence assay is particularly useful during the early stages of purification to determine the optimal process conditions. After the final process has been arrived at and a variety of methods used to assess the nucleic acid content (including the hybridization techniques), the fluorescence method can be developed for routine quality-control purposes. In certain cases, particularly at high protein concentrations, the dye may bind to the protein with... 

Franco, C. M., Fente, C. A., Vazquez, B., Cepeda, A., Lallaoui, L., Prognon, P., and Mahuzier, G., Simple and sensitive high-performance liquid chromatography-fluorescence method for the determination of citrinin. Application to the analysis of fungal cultures and cheese extracts, /. Chromatogr. A, 723, 69, 1996. 

Somsen et al. [796] have reported the use of SERR spectroscopy for the in situ selective determination and semi-quantitative analysis of structurally similar dyes separated by TLC. The limits of identification of the TLC-SERRS method (ca. 5ng applied) were sufficient for acquisition of spectra of impurities present in the certified dye standards. SERRS may also be used for in situ identification of highly fluorescent molecules on HPTLC plates. 

This method is perfectly suitable for low concentrations of fluorescent materials. However, in order to study factors which affect the fluorescence quantum yield, such as molecular association or photochemical reactions, much higher concentrations than can be used in the right-angle fluorescence method are required. This follows from the fact that the 0 - 0 vibrational bands in the absorption and emission spectra often overlap. Therefore at relatively high concentrations light emitted at these overlapping wavelengths will be reabsorbed. 

Giauque, R. D., F. Asaro, F. H. Stross, and T. R. Hester (1993), High-precision nondestructive X-ray fluorescence method applicable to establishing the provenance of obsidian artifacts, X-Ray Spectrom. 22, 44—53. 

Refaat et al. [24] used a spectrophotometric method for the determination of primaquine, and 16 other tertiary amine drugs, in bulk or in pharmaceuticals. The method involved the condensation of malonic acid with acetic anhydride in the presence of a tertiary amine in an aliphatic or a heterocyclic system. The condensation product is highly fluorescent and allows the spectrofluorimetric determination of the drug in the ng/mL ranges (Xcx = 415 nm and >.em = 455 nm). 

Most studies of the physical binding of hydrocarbon metabolites and metabolite model compounds have measured the effect of DNA binding on hydrocarbon fluorescence intensities, fluorescence lifetimes and UV absorption spectra Radioactive labelling has also been used, but less frequently. Spectroscopic methods are particularly convenient. These methods, especially fluorescence methods, are also very sensitive. All of the hydrocarbons in Figure 1 except the epoxides have high fluorescence quantum yields, which permit routine detection in the 10 -10 7 M concentration range. 

Samples in highly rigid or viscous media (e.gglass) is examined frequently in phosphorescence methods and also in some fluorescence methods. 

The design of fluorescent sensors is of major importance because of the high demand in analytical chemistry, clinical biochemistry, medicine, the environment, etc. Numerous chemical and biochemical analytes can be detected by fluorescence methods cations (H+, Li+, Na+, K+, Ca2+, Mg2+, Zn2+, Pb2+, Al3+, Cd2+, etc.), anions (halide ions, citrates, carboxylates, phosphates, ATP, etc.), neutral molecules (sugars, e.g. glucose, etc.) and gases (O2, CO2, NO, etc.). There is already a wide choice of fluorescent molecular sensors for particular applications and many of them are commercially available. However, there is still a need for sensors with improved selectivity and minimum perturbation of the microenvironment to be probed. Moreover, there is the potential for progress in the development of fluorescent sensors for biochemical analytes (amino acids, coenzymes, carbohydrates, nucleosides, nucleotides, etc.). 

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