here ∆G alch solvent/complex are the ∆Gs calculated in equations 4 and 2 in the solvent/complex simulations (transforming L A into L B ). Where the solvent simulation is the ligand in solvent and the complex simulation is the ligand in complex with the solvated protein. ∆G binding L A /L B is the binding free energy of ligand A/B to the protein. The difference of these alchemical free energies is equal to the difference of binding free energy of the ligands A and B which allows for the final ∆∆G to be calculated.