This study addresses cell membrane permeabilization phenomena over microorganisms induced by high pressure CO2: a real-time measurement has been set-up and performed by an innovative cell fluorescent staining technique. A common yeast, S. cerevisiae was used as test-microorganism. Different sets of experiments were carried out in batch mode at 100 bar, 36 degrees C and different treating time (10-30 min). The change of optical signal intensity collected by an optical spectrometer indicates a progressive permeabilization of the cell envelopes during the treatment. This new analytical method provides a real-time indication of cell membrane integrity during the pasteurization process. Both the permeabilization and inactivation kinetic rates were calculated and compared; the reported data evidence a correlation between the cellular death and the CO2 permeabilization inside the cell, which, in turn, can be considered the key factor of microbial inactivation. (C) 2008 Elsevier B.V. All rights reserved.