Double-bond character of phosphates in solid and liquid phases probed by oxygen K-edge X-ray absorption spectroscopy

The double-bond character of phosphates in solid and liquid phases was investigated using the P=O π* peaks in oxygen K-edge X-ray absorption spectroscopy (XAS). In the solid phase, the double-bond character of phosphates increased with increasing negative charge; NaH₂PO₄ exhibited weak P=O π* peaks, whereas Na₃PO₄ exhibited prominent P=O π* peaks. In an aqueous Na₃PO₄ solution, the double-bond character was reduced owing to interactions between phosphate and Na⁺ as well as the formation of hydration structures. These findings provide insights into the influence of chemical environment on the electronic structure and reactivity of phosphate compounds involved in various biological processes such as adenosine triphosphate hydrolysis.

High-energy phosphoric compounds such as adenosine triphosphate are vital for energy production and transport in living cells. To elucidate the underlying mechanisms of biological processes, the reactivity of biochemical reactions must be correlated with the double-bond character of phosphate groups in high-energy phosphate compounds. Oxygen K-edge X-ray absorption spectroscopy (XAS) is ideal for probing the double-bond character of phosphate groups via the electronic transitions of O 1s electrons from occupied states to unoccupied P=O π* orbitals. Herein, the double-bond character of phosphates in solid and liquid phases was investigated based on P=O π* peaks observed in the corresponding O K-edge XAS spectra. Variations in the double-bond character among different phosphates were analyzed by combining molecular dynamics (MD) simulations and inner-shell calculations.

The XAS measurements of solid phosphates were performed at the soft X-ray beamline BL3U of the UVSOR-III synchrotron facility using the total electron yield method, which measured sample drain currents generated after soft X-ray absorption. The XAS measurements of phosphates in the liquid phase were conducted at the soft X-ray beamline BL-7A of the Photon Factory (KEK-PF) using a transmission-type liquid cell. Figure 1 shows the O K-edge XAS spectra of solid NaH₂PO₄ and Na₃PO₄, exhibiting a weak P=O π* peak at 532.6 eV and a prominent P=O π* peak at 532.3 eV, respectively. These results reveal that the double-bond character of phosphates is enhanced with increasing negative charge in the solid phase, consistent with the inner-shell calculations. Compared with solid Na₃PO₄, no distinct P=O π* peaks were observed in the O K-edge XAS spectrum of the aqueous Na₃PO₄ solution. The O K-edge inner-shell spectrum of this solution was obtained using 1100 representative molecular structures containing PO₄³⁻, Na⁺, and water molecules extracted from snapshots obtained via MD simulations. The intensities of the P=O π* peaks in the calculated inner-shell spectrum of aqueous Na₃PO₄ solution were considerably reduced compared with those of solid Na₃PO₄. MD simulations revealed that the distances between Na⁺ and PO₄³⁻ in the aqueous solution were shorter than those in the solid phase. In addition, solvent water molecules approached the PO₄³⁻ owing to the presence of hydration structures surrounding the Na⁺. These findings indicated that the double-bond character of phosphates was reduced by interactions between phosphates and Na⁺ and the formation of hydration structures.

The O K-edge XAS measurements of solid diphosphates revealed that the double-bond character of phosphate groups increased with increasing negative charge. The O K-edge XAS spectrum of Na₂H₂P₂O₇ does not show a distinct P=O π* peak, whereas that of Na₄P₂O₇ shows the P=O π* peak. These results indicate that variations in the double-bond character of phosphate groups are typically observed in high-energy phosphate compounds. The reactivity of biological processes such as adenosine triphosphate hydrolysis is related to changes in the double-bond character of phosphate groups under the influence of pH, interaction of phosphate and Na⁺, and hydration. Therefore, O K-edge XAS is an effective technique for investigating biochemical reaction mechanisms based on changes in the double-bond character of phosphate groups associated with P=O π* orbitals.