The effect of hydration on the fluorescent properties of protonated and deprotonated ribonucleotides and their constituents

Our previous studies have demonstrated the antiviral activity of protonated 5′-ribonucleotides. To elucidate the molecular mechanisms of their biological action, it is important to determine the physicochemical properties of these compounds in an aqueous environment. It is known that hydration significantly influences the electronic states and fluorescence characteristics of nucleotides; however, the comparative characteristics of protonated ribonucleotide forms and their salts have not been sufficiently studied.

The aim of the study was to investigate the effect of hydration on the fluorescence characteristics of protonated 5′-ribonucleotides and their disodium salts. Three-dimensional excitation–fluorescence spectra were recorded in the dry state and in aqueous solution using a HORIBA FluoroMax-4P+ spectrofluorometer. 

It was shown that hydration induced changes in the fluorescence spectra of the studied compounds in different ways, with the exception of protonated UMP. A significant enhancement of fluorescence was observed for protonated purine ribonucleotides. In the case of AMP, hydration was accompanied by a shift in the fluorescence maximum of 21 nm and an eightfold increase in intensity. For AMPNa₂, the corresponding changes were 25 nm and a 1.3-fold increase. For protonated GMP, a spectral shift of 55 nm and a 25-fold increase in intensity were observed. At the same time, two bands appeared in the spectrum of GMPNa₂ upon hydration. For CMP and CMPNa₂,  shifts were 5 and 92 nm, respectively. Meanwhile, protonated UMP showed no response to hydration, whereas the fluorescence intensity of its sodium salt increased approximately threefold.

Hydration affects the fluorescence properties of protonated ribonucleotides and their disodium salts differently. The particularly pronounced effect observed for protonated AMP and GMP indicates the significant influence of the aqueous environment on their electronic-energy characteristics. These differences must be taken into account when investigating the interaction of ribonucleotides with biological targets and development of new medicines.