| Resonance light scattering (RLS) technique, proposed by Pasternack et al in the beginning of 1990's, is a sensitive means to detect light scattering signals of aggregates with a conventional fluorescence spectrophotometer. After that, RLS technique attracted many researchers' much interest and extensive concern not only for its simple manipulation but also for its high sensitivity and convenient experimental conditions, which cannot be achieved by traditional light scattering technique. From the 1996 year to now, the short period of 7 years, RLS technique has been widely and successfully applied to determine biomacromolecules and Pharmaceuticals. During these investigations, it was found that RLS technique has the advantage of possessing more extensive area of probe reagents as well as much better sensitivity, than spectrophotometry and spectrofluorimetry, which are often used as a routine way to assay biomacromolecules and pharmaceuticals. In this contribution, based on the significantly enhanced RLS signals of proteins in the presence of sodium dodecyl benzene sulfonate (SDBS) and water blue (WB), respectively, we established simple, rapid and sensitive assays of proteins although neither obvious color nor fluorescence change occurs in these interactions, further displaying the particular advantage of RLS technique in expanding the area of probe reagents.At pH 1.98 and ionic strength 0.001 M, the interactions of proteins with sodium dodecyl benzene sulfonate (SDBS) give rise to greatly enhanced RLS spectra with the maximum light scattering peak located at 470.0 nm. The enhanced RLS signals are in good proportional to the concentrations of proteins in the range of microgramme levels. With this method, proteins, including bovine serum albumin (BSA), human serumalbumin (HSA),α-chymotrypsin (Chy), lysozyme (Lys), can be determined with the limit of detection below 58.1 ng ml"'. Artificial samples and human urine samples were analyzed satisfactorily.At pH 2.09 and ionic strength 0.001, strongly enhanced RLS spectra of the mixture of proteins and WB with the maximum light scattering peak at 346.0 nm can be observed and the enhanced RLS signals at 346.0 nm are in good proportion to the concentrations of proteins in the range of microgramme levels. With the enhanced RLS signals at 346.0 nm, proteins, including bovine serum albumin (BSA), human serum albumin (HSA), γ-globulin (γ-IgG), a-chymotrypsin (Chy), lysozyme (Lys) and cellulose (Cel), can be determined with the limit of detection below 1.5 ng ml-1. Synthetic samples and human urine samples were analyzed satisfactorily.However, a regretting case is that RLS technique still suffers from several disadvantages, such as failing to distinguish light scattering signals of analytes with those of their coexisting foreign substances, which are not beneficial to improve further selectivity and sensitivity. One way to solve this problem, in our opinion, is to introduce an oil/water system. In an oil/water system, amphiphilic species, involving both hydrophilic moieties and hydrophobic moieties, are repelled from both the water and oil phases, but can be well adsorbed at the oil/water interface. As a result, an analyte can be well separated from its coexisting substances and be enriched to the oil/water interface by means of developing an amphiphilic species with appropriate reagents, and thus determination of the analyte with better selectivity and sensitivity can be available. Lately, we successfully combined RLS method with total internal reflected (TIR) light to propose a total internal reflected resonance light scattering (TIR-RLS) method to analyze amphiphilic species at an oil/water interface by simply constructing a TIR optical arrangement in the sample compartment of a conventional fluorescence spectrophotometer. In the present method, RLS signals are obtained from the amphiphilic species in the oil/water interfacial region with the excitation of an evanescent field coupled with total internal reflection of light at the interface. Since the inte...
|
PDF Full Text Request