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Abstract The equilibrium between inorganic pyrophosphate and inorganic orthophosphate was studied by direct measurement of PPi. The reaction was catalyzed by yeast pyrophosphatase in the presence of Mg2+ ions or by alkaline phosphatase in the absence of metal ions. The concentration of PPi was determined by an isotope derivative method. Identical concentrations of PPi were obtained whether the reaction proceeded from orthophosphate or pyrophosphate provided other conditions were identical. The equilibrium concentrations were measured at varying Mg2+ and K+ ion concentrations and at different ionic strengths. All measurements were performed at pH 7.4. Equilibrium was established at different temperatures and the enthalpy changes were determined. HPP3- ⇆ 2HP2- + H+ The experimental data were used to compute an equilibrium constant (Kion ic) based on the ionic species Kion ic was found to (2.41 ± 0.20) x 10-4 m2 (zero ionic strength) corresponding to a ΔGion ic = (4.98 ± 0.05) Cal x mole-1 (20.84 kJ x mole-1). It was constant over the variety of conditions studied. From the experiments carried out in absence of metal ions and low ionic strength ΔG0'obs was found to (-5.63 ± 0.02) Cal x mole-1 (-23.56kJ x mole-1) calculated for zero ionic strength making due allowances for dissociation at pH 7.4. ΔH0'obs was found to (-1.90 ± 0.37) Cal x mole-1 (-7.95 kJ x mole-1) and TΔS0'obs to 3.87 Cal x mole-1 (16.19 kJ x mole-1) (the latter two results given for ionic strength 0.127). Increasing the Mg2+ ion concentration makes the reaction more unfavorable, thus at 26 mm free Mg2+ and 18 mm free K+ at ionic strength 0.1 ΔG0'obs was found to (-2.73 ± 0.07) Cal x mole-1 (-11.42 kJ x mole-1). The stoichiometry of the participation of H+ and metal ions in the composite reaction was calculated. The contribution of the observed enthalpy and entropy terms to the variation in ΔG0'obs is discussed in relation to the changes in production or consumption of H+ and metal ions. At free Mg2+ above 0.3 mm and low free K+ (17 to 43 mm) at ionic strengths around 0.05 the reaction was found to consume acid. The TΔS0'obs term which is negative was found to pass through a maximum negative value of -4.17 Cal x mole-1 (-17.53 kJ x mole-1) at 12.2 mm free Mg2+. The entropy term is numerically considerably larger than the enthalpy term, resulting in a less favorable reaction. ΔH0'obs values were calculated on the basis of ΔH0 for dissociation and association reactions and agreement with the experimental results was obtained in the cases where ΔH0 values reported in the literature were measured at experimental conditions similar to ours. From ΔG0'obs measured at the ionic strength and K+ and Mg2+ prevailing in the cell, ΔG for the PPi hydrolysis in the cell was calculated to -4.0 Cal x mole-1 (-16.74 kJ x mole-1) on the basis of 1 mm free Mg2+, 150 mm free K+, 2.42 mm free Pi, and concentrations of PPi measured in freeze biopsies from rat liver to (6.2 ± 0.3 (S.E. n = 6)) nmoles per g wet weight. The implication of this result for the utilization of the free energy of PPi hydrolysis in the cell is discussed. In an appendix a set of equations were introduced, which permit computation of equilibrium data also in a general case.
Flodgaard et al. (Sat,) studied this question.