Assessment of accurate drug binding affinity to a protein remains a challenge for in silico drug development. In this research, we used the smooth reaction path generation (SRPG) method to calculate binding free energies and determine potential of mean forces (PMFs) along the smoothed dissociation paths of influenza A neuraminidase and its variants with oseltamivir (Tamiflu) and zanamivir (Relenza) inhibitors. With the gained results, we found that the binding free energies of neuraminidase A/H5N1 in WT and two mutants (including H274Y and N294S) with oseltamivir and zanamivir show good agreement with experimental results. Additionally, the thermodynamic origin of the drug resistance of the mutants was also discussed from the PMF profiles.