Asako NARAI-KANAYAMA

For the purpose of clarifying the relationship between pasteurization and inactivation of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) in Saccharomyces pastorianus cells induced by pressurized carbon dioxide microbubbles (CO2MB) treatment, a storage test of S. pastorianus cells after CO2MB treatment was conducted to ascertain their recovery, and the treatment condition in the inactivation of GAPDH in S. pastorianus cells by CO2MB was investigated. Each population of S. pastorianus for 48, 96, and 144 h at 25°C was decreased significantly by CO2MB treatment at 35°C for 3 min (MB35-3 and MB35-5) or at 40°C and 45°C for 1 and 3 min (MB40-1, MB40-3, and MB45-1). In the storage test, recovery of treated cells was not observed after storage for 144 h at 25°C. The denaturation of GAPDH in the S. pastorianus cells caused by the same treatment as the storage test was detected by using sodium dodecyl sulphate polyacrylamide gel electrophoresis. While the activities at MB35-1, MB35-3, and MB40-1 were significantly higher than those at non-treatment, and those at MB35-5, MB40-3, and MB45-1 were lower. Therefore, GAPDH denaturation, but not the activity, was associated with the inactivation of S. pastorianus cells.

To clarify the relationship between irreversible inactivation and intracellular protein denaturation of Saccharomyces pastorianus by low-pressure carbon dioxide microbubbles (CO2MB) treatment, a storage test of S. pastorianus cells treated with CO2MB was performed, and the effect on the intracellular protein was investigated. In the storage test, the S. pastorianus population, which decreased below the detection limit by CO2MB treatment at a temperature of 45 and 50 degrees C (MB45 and MB50), and thermal treatment at a temperature of 80 degrees C (T80), remained undetectable during storage for 3 weeks at 25 degrees C. However, 4.1 and 1.3-logs of the S. pastorianus populations, which survived after CO2MB treatment at temperatures of 35 and 40 degrees C (MB35 and MB40), increased gradually during storage for 3 weeks at 25 degrees C. Insolubilization of intracellular proteins in S. pastorianus increased with increasing the temperature of CO2MB treatment. Activity of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) identified as one of the insolubilized proteins increased at MB35 and MB40 than non-treatment but disappeared at MB45 and MB50, and T80. Therefore, it was revealed that S. pastorianus cells inactivated below the detection level by CO2MB treatment did not regrow and that the denaturation of intracellular proteins of S. pastorianus was caused by CO2MB and thermal treatments. Furthermore, it was suggested that denaturation of intracellular vital enzymes was an important factor for achieving irreversible inactivation of S. pastorianus by CO2MB and thermal treatments.