In the present work the effect of High Pressure Carbon Dioxide (HPCD) on the activity and structure of a commercial pectin methylesterase (PME) was studied and the results were compared to the conventional thermal inactivation technology. The use of supercritical CO2 (pressure 6-18 MPa, temperature 40-55 degrees C and time up to 75 min) increased dramatically the PME inactivation rate. The experimental data fitted the first order model and the inactivation kinetic study of PME was completed with the calculations of the activation energy and volume of activation. HPCD treatment induced important changes in the tertiary structure of the enzyme, as revealed the intrinsic fluorescence, KI quenching and ANS binding analyses. They showed that HPCD treatment rearranged the structure of the enzyme. These changes induced by HPCD were significantly different from those induced by mild thermal treatment. Finally, the structural changes detected correlated with the enzyme activity losses observed.