Excitation wavelength-dependent fluorescence anisotropy of 3-hydroxyflavone: revisiting the solvation processes and high-energy state excitation in ESIPT-active compounds.

To gain a more comprehensive understanding of the phenomenon of high-fluorescence anisotropy of the normal form emission of ESIPT-active compounds in protic solvents, excitation wavelength dependence of emission anisotropy was investigated for 3-hydroxyflavone (3HF) using steady-state spectroscopic technique and quantum chemical calculations. It was shown for the first time that the anisotropy of 3HF normal form emission is characterized by significant dependence on excitation energy. Experimental results indicate that the fluorescence anisotropy of 3HF in methanol (at 20 °C) changes abruptly from about 0.18 to about 0.10 with a decrease in excitation wavelength. This spectroscopic phenomenon can be explained by two factors: (1) breaking of intermolecular solute-solvent hydrogen bonds upon photoexcitation and (2) excitation of ESIPT-active fluorophores to the second singlet state (S). The results of quantum chemical calculations clearly indicate that specific hydrogen bonding solvation interactions can lead to the formation of 3HF-methanol complexes with larger molecular volumes than the volume of free 3HF molecule. High excitation energy can reform and break solute-solvent bonds, which leads to a decrease in molecular system volume. This results in a decrease in rotational correlation time and fluorescence anisotropy. As is known, the fluorescence lifetime of small-sized molecules is closely correlated with the conformational changes in the excited state, and in the case of ESIPT-active compounds, the lifetime of normal form emission is almost fully determined by the ultrafast ESIPT process. Therefore, although in general, fluorescence lifetime is considered independent of excitation energy, but because the timescale of ESIPT processes is of the same order as the timescale of internal conversion, in the case of ESIPT-active compounds, fluorescence lifetime changes caused by high-energy state excitation cannot be neglected. The emission anisotropy of the normal form of an ESIPT-active compound will decrease with an increase in fluorescence lifetime caused by an increase in excitation energy sufficient to excite molecules to higher electronic states. In this work, both hypotheses are discussed and verified using experimental data and quantum chemical calculations for 3HF in methanol.