Auramine O as a fluorescent probe for the binding of basic drugs to human α1-acid glycoprotein ( α1-AG) The development of a simple fluorometric method for the determination of α1-AG in human serum

Abstract

A cationic fluorescent dye, auramine O (AO), exhibited an intense increase in fluorescence after binding to human α 1-acid glycoprotein ( α 1-AG). The interaction between AO and the protein was studied by fluorescence spectroscopy and by equilibrium dialysis. AO binds to the protein via a single site with a dissociation constant of 24 μM. Various basic drugs such as chlorpromazine, imipramine, desipramine, quinidine, propranolol and lidocaine, which are known to bind to the protein, competitively inhibited the AO binding to the protein. The dissociation constants of these basic drugs obtained from such inhibitory experiments were comparable to those obtained with other methods (equilibrium dialysis, quenching of protein intrinsic fluorescence, and the difference spectrophotometric method) and from the literature. It is concluded that AO may be a useful fluorescent probe that binds to a single basic drug binding site on α 1-AG. In addition, a simple fluorometric method for the determination of α 1-AG in serum was developed using AO, and the validity of this method was confirmed by comparing it with the conventional radial immunodiffusion method.