Relevance of the approximately hyperbolic relationship between fluorescence and concentration to the determination of quantum efficiencies

Abstract

An absorptivity-related constant and a quantum efficiency-related constant can be derived, by a simple graphical procedure, from data obtained in a standard fluorimeter. The quantum efficiency of an unknown fluorophore can be determined by comparison of its quantum efficiency-related constant with that of a fluorophore of known quantum efficiency. The absorptivity can similarly be determined using the absorptivity-related constant. This method relies on the approximately hyperbolic relationship that exists between light absorbed and chromophore concentration. The approximation may be derived from the Beer-Lambert equation and the limits of its validity have been tested using theoretical and experimental data.