After the validation of the modified force field parameters, we show how the buffer particles that maintain the neutrality of the simulation box, have to be parameterized to prevent finite-size effects on proton affinities due to periodicity. Sampling Klimovic and Mobley have shown that calculated hydration free energies of single amino acids depend on the starting conformation. Because a few picoseconds typically suffice to sample bond and angle degrees of freedom in the amino acid, as well as the rotational degrees of freedom of the water molecules, we speculate that their observation implies a lack of sampling in the dihedral degrees of freedom in the amino acid side chain. Therefore,