KINETICS AND IDENTIFIABILITY OF AN INTERMOLECULAR 2-STATE EXCITED-STATE PROCESS IN THE PRESENCE OF A FLUORESCENT IMPURITY

This paper presents a study analyzing the conditions for the recovery of the rate constants and the spectral parameters from the fluorescenc e decay surface due to three-state excited-state processes. The studie d system models a Situation when, in addition to a fluorescence indica tor which undergoes an excited-state reaction with a co-reactant, a fl uorescent impurity is present. It is demonstrated theoretically that, if the fluorescence decay surface contains traces measured in the abse nce of co-reactant and additionally at two nonzero coreactant concentr ations, all five rate constants can always be recovered. In some cases the measurement in the absence of co-reactant is not necessary. If th e fluorescence decay surface is collected at three different emission wavelengths and two co-reactant concentrations, the ratios of the abso rbances of the free and bound form of the fluorescent indicator can be determined. If two different emission wavelengths are used in combina tion with three excitation wavelengths (or three co-reactant concentra tions), the ratios of the emission weighting factors of the free and b ound form of the fluorescent indicator can be obtained. Under both pre vious conditions no spectral information about the fluorescent impurit y can be acquired. The ground-state dissociation constant can be deter mined from the fluorescence decay traces excited at an isosbestic poin t.