Aminofluoresceins Versus Fluorescein: Peculiarity of Fluorescence

In this article, we examined the fluorescent properties of 4′- and 5′-aminofluorescein, unsubstituted fluorescein, and its 4′-nitro derivative in a set of solvent systems. Fluorescence lifetimes, quantum yields, time-resolved fluorescence spectra, and quantum chemical calculations allowed clarifying the reasons of the emitting properties in this dye series. In water, the dianions R2– of aminofluoresceins are practically nonfluorescent; in alcohols, the quantum yields are low. In dimethylsulfoxide (DMSO), acetonitrile, and other non-hydrogen bond donor solvents, the bright fluorescence of R2– ions is quenched either on adding small amounts of water which hydrate the carboxylate group or under conditions of protonation of this group (COO– → COOH). The last observation is possible owing to the peculiarities of the tautomerism of the 5′-aminofluorescein monoanion, HR–, which exists in DMSO as an equilibrium mixture of a colorless lactone and colored "phenolate" tautomer with an ionized xanthene moiety and unionized carboxylic group. In contrast, the R2– anion of 4′-nitrofluorescein demonstrates spectral behavior different from that of the amino derivatives. It practically does not emit in aprotic solvents; however, in alcohols or water media, its quantum yield increases to some extent. Such changing spectral properties are explained in terms of the excited-state interfragmental charge transfer.