Intracellular pH measurement during high-pressure CO 2 pasteurization evaluated by cell fluorescent staining

Abstract

This study addresses bacterial cell inactivation phenomena induced by high-pressure CO 2, particularly intracellular pH decrease. A common bacteria, Listeria monocytogenes, was used as test microorganism. In order to monitor intracellular pH decrease, the bacterial cells were loaded with the specific fluorescent probes 5-(and-6)-carboxyfluorescein diacetate succinimidyl ester (cFDASE) and 5-(and-6)-carboxy-2′7′-dichlorofluorescein diacetate succinimidyl ester (cDCFDASE). Different sets of experiments were carried out in batch mode at 25 °C and 6.8 MPa at different treatment times. The intensity change of two fluorescent emission signals, obtained respectively at 490 and 510 nm excitation wavelengths, was monitored with a spectrofluorometer at peak emission wavelengths (525 and 550 nm). The data analysis indicated a progressive intracellular pH decrease inside the cell caused by the action of CO 2, followed by an increase after cellular death. This innovative analytical method provides a promising technique for monitoring cell behavior during high-pressure CO 2 processes.