Polymers such as polyethylene (PE) and ethylene vinyl alcohol (EVOH) are primary constituents of multilayer films used in the food and biopharmaceutical packaging. In order to explore the film behavior after gamma-sterilization, several tests are performed (tensile tests, differential scanning calorimetry (DSC), oxygen transmission rate (OTR) and water vapor transmission rate (WVTR). The investigation of the PE/EVOH/PE multilayer film is done at several gamma-irradiation doses up to 270 kGy. For irradiation dose up to SO kGy, mechanical properties are not altered. Whatever the irradiation dose, water barrier in PE/EVOH/PE multilayer film is not altered whereas the oxygen barrier property is increased by 300%. This enhancement is shown not to occur due to reorientation, closer chain packaging and restriction of chain mobility of polymer chains in the amorphous phase of the different polymers. This enhancement rather comes from a change of the chemical environment of the EVOH layer.