Employing the microhydration model that involves two to five explicit water molecules, two plausible dissociative hydrolysis pathways of 2’,3’-didehydro-2’,3’-dideoxyuridine (d4U), α-path with configuration-inversion and β-path with configuration-retention, have been investigated by M06-2X(CPCM)/6-31++G(d,p) method. Using this model, inclusion of three explicit water molecules (n=3) is shown to be the smallest system that gives the minimal activation free energy for α-path and β-path. Our results suggest that the glycoside cleavage is the RDS, and α-path is more favorable kinetically than β-path. Whereas β-path with exergonic formation of β-dihydrofuran-like sugar with keto pyrimidine complex possesses thermodynamic preference over α-path, where the formation of α-dihydrofuran-like sugar with enol pyrimidine complex is endergonic. The free energy barriers of RDSs for d4U (24.8 kcal mol^-1) and d4T (27.3 kcal mol^-1) suggest that the glycosidic bond in d4T is more stable than that in d4U. The relative lower stability of d4U is probably an important factor for less antiviral activity of d4U. The small free energy barrier differences of ~1 kcal mol^-1 for β-path over α-path, and the reaction free energy differences of ~ -12 kcal mol^-1 for β-path lower than α-path in d4T and d4U suggest a competitive β-path in pyrimidine d4Ns. The higher free energy barriers of RDSs in ddU (27.6 kcal mol^-1) and ddT (29.0 kcal mol^-1) indicate that the saturated sugar moiety increases the stability of glycosidic bond in contrast to the unsaturated counter parts in d4U and d4T. NBO analysis also shows the kinetic preference of α-path over β-path. Our results provide an exploration for the less antiviral activity of d4U and the influence of saturated ribose on the glycosidic bond stability of pyrimidine d4Ns.