Dissociation and tautomerism of some amino-, nitro-, and aminonitrofluoresceins in DMSO: Either resolution or inversion of the Ka1 and Ka2 values.

Fluorescein compounds are diprotic acids prone to tautomeric transformations. Due to their valuable spectral properties, they are among the most commonly used dyes. However, nitro- and aminofluoresceins have been studied much less frequently than other fluorescein derivatives. At the same time, these compounds exhibit behavior that is different from that of other fluoresceins. Studying complex equilibria in organic solvents allows to identify molecular structures and equilibrium constant relationships that are impossible to observe in aqueous solutions. This study examines the acid-base equilibria of amino, nitro, and mixed aminonitro derivatives of fluorescein in DMSO. Basing on our previous results, we selected eight dyes, to clarify further the scheme of protolytic equilibria in this series. Among them are newly synthesized by us 3'-nitro-, 3'-amino-, 4,5-dinitro-4'-amino-, and 4,5-dinitro-5'-aminofluorescein. The pKa values were determined using the spectrophotometric method. Tautomerism is considered using the absorption spectra of molecular and anionic forms of the dyes and their esters and ethers that are used as model (auxiliary) compounds. All fractions of the tautomers, α, and the microscopic dissociation constants, pk, of the dyes are determined. For the neutral form, H2R, the colorless lactone predominates. Depending on the nature and position of the substituents, colorless tautomers were observed for the HR- and R2- anions, along with ordinary intensively colored species, which is atypical for aqueous solutions. This feature leads to very different ratios of stepwise dissociation constants: from resolution of pKa1 and pKa2 in the range from 1.5 to 4.1 to inversion, pKa1 > pKa2, for 4'-aminofluorescein. The results were compared with those obtained in water and an aqueous ethanol solution. A summary of all α and pk values of these and previously studied dyes, a total of 25 compounds, provides a quantitative description of the detailed protolytic equilibrium scheme of all amino-, nitro-, and aminonitrofluoresceins in solution.