The study utilized the endpoint free energy method based on MM-PBSA to evaluate the binding affinity of vitamin D. The binding free energy calculation (∆Gbind) was performed on the complexes of vitamin D2 and D3, obtained from the MD run. The g_mmpbsa tool integrated within Gromacs was used to estimate the solvation-free energy (∆Gsolv) by combining polar and non-polar components. A total of 200 frames were evenly sampled at a predetermined time of 500 ps and subjected to MM-PBSA calculations.