Analysis of the effect of polyanionic phosphates on the solvent features of aqueous media

Adenosine triphosphate (ATP) has emerged as a critical regulator of liquid-liquid phase separation (LLPS), protein aggregation, and cytoplasmic fluidity, yet the molecular basis of its effects remains incompletely understood. In this study, attenuated total reflectance Fourier-transform infrared (ATR-FTIR) spectroscopy combined with Gaussian decomposition of the OH-stretch region was used to investigate the effects of ATP on hydrogen-bond organization in aqueous phosphate-buffered solutions. Four water subpopulations corresponding to distinct hydrogen-bond arrangements were analyzed to characterize ATP-induced restructuring of the solvent network. Increasing ATP concentrations produced a reproducible increase in the contribution of the strong tetrahedrally hydrogen-bonded water subpopulation (3080 cm^-1), while the relative contributions of more weakly hydrogen-bonded water populations remained largely unchanged. Similar but stronger effects were observed for sodium triphosphate, whereas DNA and longer-chain polyphosphates did not exhibit comparable behavior. These findings suggest that ATP and related triphosphates modulate aqueous hydrogen-bond organization through selective stabilization of highly coordinated water structures rather than broad disruption of the hydrogen-bond network. The results provide new insight into solvent restructuring mechanisms that may contribute to ATP-mediated solubilization, condensate regulation, and cytoplasmic fluidization.