In this work, Δ^Rg⁺ values for the enzymatic G6P isomerization were determined as a function of the G6P equilibrium molality between 25 °C and 37 °C. The reaction mixtures were buffered at pH = 8.5. In contrast to standard literature work, Δ^Rg⁺ values were determined from activity-based equilibrium constants instead of molality-based apparent values. This yielded a Δ^Rg⁺ value of 2.55 ± 0.05 kJ mol^− 1 at 37 °C, independent of the solution pH between 7.5 and 8.5. Furthermore, Δ^Rh⁺ was measured at pH = 8.5 and 25 °C yielding 12.05 ± 0.2 kJ mol^− 1.

Accounting for activity coefficients turned out to influence Δ^Rg⁺ up to 30% upon increasing the G6P molality. This result was confirmed by predictions using the thermodynamic model ePC-SAFT.

Finally, the influence of the buffer and of potassium glutamate as an additive on the reaction equilibrium was measured and predicted with ePC-SAFT in good agreement.