Hyong Seok Park

Addition of ethanol to supercritical carbon dioxide enhances the inactivation of bacterial spores in the biofilm of Bacillus cereus

Supercritical carbon dioxide (SC-CO2) was used to inactivate Bacillus cereus spores inside biofilms, which were grown on stainless steel. SC-CO2 treatment was tested using various conditions, such as pressure treatment (10-30 MPa), temperature (35-60°C), and time (10-120 min). B. cereus vegetative cells in the biofilm were completely inactivated by treatment with SC-CO2 at 10 MPa and at 35°C for 5 min. However, SC-CO2 alone did not inactivate spores in biofilm even after the treatment time was extended to 120 min. When ethanol was used as a cosolvent with SC-CO2 in the SC-CO2 treatment using only 2-10 ml of ethanol in 100ml of SC-CO2 vessel for 60-90 min of treatment time at 10 MPa and 60°C, B. cereus spores in the biofilm were found to be completely inactivated in the colony-forming test. We also assessed the viability of SC-CO2-treated bacterial spores and vegetative cells in the biofilm by staining with SYTO 9 and propidium iodide. The membrane integrity of the vegetative cells was completely lost, while the integrity of the membrane was still maintained in most spores. However, when SC-CO2 along with ethanol was used, both vegetative cells and spores lost their membrane integrity, indicating that the use of ethanol as a cosolvent with SC-CO2 is efficient in inactivating the bacterial spores in the biofilm.

Inactivation of Alternaria brassicicola spores by supercritical carbon dioxide with ethanol entrainer.

Supercritical carbon dioxide (SC-CO(2)) was used to inactivate fungal spores of Alternaria brassicicola. The inactivation conditions were optimized using response surface methodology (RSM). When the SC-CO(2)-entrainer (ethanol) system was applied to fungal spores, the treatment time required for the complete inactivation of fungal spores was substantially reduced.

Customized optimization of cellulase mixtures for differently pretreated rice straw

Lignocellulose contains a large amount of cellulose but is recalcitrant to enzymatic hydrolysis, which yields sugars for fuels or chemicals. Various pretreatment methods are used to improve the enzymatic digestibility of cellulose in lignocellulose. Depending on the lignocellulose types and pretreatment methods, biomass compositions and physical properties significantly vary. Therefore, customized enzyme mixtures have to be employed for the efficient hydrolysis of pretreated lignocellulose. Here, using three recombinant model enzymes consisting of endoglucanase, cellobiohydrolase, and xylanase with a fixed amount of β-glucosidase, the optimal formulation of enzyme mixtures was designed for two differently pretreated rice straws (acid-pretreated or alkali-pretreated rice straw) by the mixture design methodology. As a result, different optimal compositions for the enzyme mixtures were employed depending on the type of pretreatment of rice straw. These results suggest that customized enzyme mixtures for pretreated lignocellulosic biomass are necessary to obtain increased sugar yields and should be considered in the industrial utilization of lignocellulose.

Disinfection of wheat grains contaminated with Penicillium oxalicum spores by a supercritical carbon dioxide-water cosolvent system.

Spores of the plant pathogenic fungus Penicillium oxalicum inoculated onto wheat grains were inactivated using supercritical carbon dioxide (SC-CO(2)). After the SC-CO(2) treatment at various conditions of temperature, time and amount of water cosolvent, the colony forming units (CFU) of fungal spores on wheat grains and the germination yields of wheat grains were determined. Among these SC-CO(2) treatment parameters, the inactivation of P. oxalicum spores was found to be significantly increased by adding water cosolvent. The optimal conditions determined by ridge analysis of response surface methodology were 233 μL water, 44°C, and 11 min, which resulted in a 6.41 log(10) CFU reduction of P. oxalicum. However, the germination yields of wheat grains significantly decreased when water cosolvent of 150 or 300 μL was added to the grains contained in the 100 mL SC-CO(2) treatment vessel.

Effective thermal inactivation of the spores of Bacillus cereus biofilms using microwave

Microwave sterilization was performed to inactivate the spores of biofilms of Bacillus cereus involved in foodborne illness. The sterilization conditions, such as the amount of water and the operating temperature and treatment time, were optimized using statistical analysis based on 15 runs of experimental results designed by the Box-Behnken method. Statistical analysis showed that the optimal conditions for the inactivation of B. cereus biofilms were 14 ml of water, 108°C of temperature, and 15 min of treatment time. Interestingly, response surface plots showed that the amount of water is the most important factor for microwave sterilization under the present conditions. Complete inactivation by microwaves was achieved in 5 min, and the inactivation efficiency by microwave was obviously higher than that by conventional steam autoclave. Finally, confocal laser scanning microscopy images showed that the principal effect of microwave treatment was cell membrane disruption. Thus, this study can contribute to the development of a process to control food-associated pathogens.