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The effects of intracellular acidification and damage of cellular membrane on the inactivation of Saccharomyces pastorianus by a two-stage method of low-pressure carbon dioxide microbubbles (two stage MBCO2) were investigated. Intra/extra cellular pH (pH(in) and pH(ex)) and propidium iodide (PI) uptake of S. pastorianus cells treated with the two-stage MBCO2 were measured by fluorescence analysis, and were compared with the inactivation efficiency. The pHex and pHin of S. pastorianus cells treated with only mixing vessel at 10 degrees C of two-stage MBCO2 was lower than those that were untreated. However, the surviving number and the pHin of S. pastorianus cells treated with the two-stage MBCO2 containing the mixing vessel at 1.0 MPa and the heating coil of 40 degrees C decreased concomitant with the exposure time, and the increase of the PI uptake ratio was confirmed. In addition, when the pressure of the mixing vessel was 1.0 MPa, the surviving number and the pHin of S. pastorianus cells significantly decreased, and the PI uptake ratio increased by increasing the temperature to 45 and 50 degrees C with the heating coil of the two-stage MBCO2. These results suggested that the inactivation of S. pastorianus by two-stage MBCO2 might be induced by the damage to the cellular membrane at 45 and 50 degrees C, and by the lowering of the pHin at 40 degrees C. (C) 2016 Elsevier Ltd. All rights reserved.
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