3.3. Steady-State Fluorescence Spectroscopy

The fluorescence emission spectra of the studied coumarins (10 µM) were registered in aqueous solutions with the use of a Cary Eclipse Varian spectrofluorimeter (excitation and emission slits—5 nm; PMT detector voltage—600 V) in the absence and presence of 4-amino-TEMPO at concentrations up to 0.10 mM. Excitation wavelength was individually chosen for each coumarin based on its UV-Vis absorption spectrum. The fluorescence intensity values were always measured at the maximum of the emission, immediately (1–2 s) after the addition of 4-amino-TEMPO and mixing the solution (spectrofluorimeter equipped with magnetic stirrer), which caused (if any) distinct and rapid changes by leaps and bounds. The values of excitation and emission wavelengths are gathered in Table 2. The emission spectra as well as all fluorescence intensity measurements were performed at 25 °C. The possible absorption of light by 4-amino-TEMPO at the excitation and emission wavelengths of coumarins has been considered during data analysis. The extent of the inner filter effect was estimated based on the following equation:

where F_corr and F_obs are the corrected and observed fluorescence intensities, respectively, whereas A_ex and A_em are the sum of the absorbance of the appropriate coumarin and the nitroxide radical at the excitation and emission wavelength, respectively [54].

Excitation and emission wavelengths of the studied coumarins.