The second term in Equation 1 scales the uncertainty in the work in each λ window by the nonuniform spacing of the λ windows. the approximate contribution of window λ i to the overall PMF uncertainty. Excluding the first and last window, the average window length was 11.8 ns and 5.39 ns for GAFF v1.7 and GAFF v2.1 simulations, respectively. We took a more direct approach with the SMIRNOFF99Frosst simulations, due to changes in pAPRika that allowed us to target uncertainties of the same magnitude as in the GAFF simulations, by running each window for a constant length of 10 ns, except for the first and last window which ran for 1 μs to converge ΔH for all three force fields.