Dye-protein interactions between Rhodamine B and whey proteins that affect the photoproperties of the dye

In fluorescence labeling, proteins are usually assayed to minimize the impact of fluorophores on their intrinsic structure and functions, however, the effect of the protein itself on the fluorescence properties of its attached fluorophore has not been studied at length. In this paper, the fluorescence properties of the ubiquitously used Rhodamine B (RhB) were investigated in the presence of whey proteins, particularly at alkaline conditions. Results reveal the existence of dye-protein interactions whether the dye is free or covalently linked to proteins. Alkali was able to destroy these interactions, while base-denaturation of proteins also occurred that complicates the dye-protein interactions. In addition, RhB was found able to quench the tryptophan (Trp) fluorescence of whey proteins, leading to the formation of nonfluorescent Trp-RhB complex. This stable Trp-RhB complex might explain the irreversible binding of RhB upon thermal-induced whey protein aggregates. This finding could suggest a cheaper alternative of labeling on proteins.