Mechanism Of High Voltage Electric Field Cold Plasma On Damage Of Salmonella Enteritidis And Listeria Monocytogenes

Cold plasma bactericidal technology has been a new bactericidal method in recent years.Compared with traditional bactericidal method,it has the advantages of simple operation,low price of equipment and high bactericidal efficiency.Moreover,dielectric barrier discharge system has become one of the most widely used device for cold plasma excitation.S.enteritis and L.monocytogenes are common pathogenic bacteria in food,which can cause great harm to human beings,livestock and poultry.The inhibition and killing of these two pathogenic bacteria have always been the focus of researchers.In this paper,cold plasma cold sterilization is taken as a new sterilization method and high voltage electric field dielectric barrier discharge system is adopted to generate reactive oxygen species.S.enteritis and L.monocytosa are taken as the research objects to study the sterilization effect and damage mechanism of cold plasma treatment.The specific contents and results are as follows:1.Study on bactericidal effect of CPCS conditions on bacteria and morphological damageThe effects of different treatment voltage,time and oxygen concentration on bactericidal efficacy of L.monocytogenes and S.enteritidis were studied.The damage effects of cold plasma on S.enteritidis and L.monocytogenes were studied from the perspective of morphological changes of bacteria.The results showed that treatment time,voltage and oxygen concentration have significant impact on bactericidal effect.L.monocytogenes and S.enteritidis could be absolutely killed by 80 kV for 150 s.When the voltage was not higher than 70 kV,there was no significant difference in the bactericidal efficiency of plasma treatment between the two bacteria.At 80 kV,the bactericidal effect against S.enteritidis was significantly better than that of L.monocytogenes.According to SEM,the bacteria showed shrinkage and deformation,the morphology was seriously damaged and perforating after cold plasma treatment.With the increase of treatment time,the damage of L.monocytosa was more serious than that of S.enteritidis.2.Study on effect of CPCS for changes of ROS content and damage to cell membraneBased on the change of the species and concentration of reactive oxygen components with the treatment time,the correlation between different ROS components and the total number of colonies was established.The reactive oxygen species with high correlation with sterilization efficiency were determined.In addition,the conductivity and leakage of proteins and nucleic acids in the supernatant were used to characterize the damage of bacterial cell stracture after CPCS treatment.Finally,the degree of oxidative damage of membrane lipids was used to characterize the membrane damage.The results showed that the intracellular ROS content of L.monocytogenes and S.enteritidis increased significantly after CPCS treatment.The intracellular ROS content of L.monocytogenes was significantly higher than that of S.enteritidis.The highest concentration of O3 in the packaging systerm can reach about 5000 ppm.Among the active oxygen components involved in bacteriostasis,H2O2 and·OH have the best oxidation and bactericidal effect.Cold plasma treatment would destroy the integrity of the cells and the permeability would increase,which was manifested in the increase of conductivity and the leakage of proteins and nucleic acid macromolecules.3.Effect of CPCS on protein structure and enzyme activityNa+/K+--ATPase and malate dehydrogenase were used as the rescarch object to study the effect of cold plasma treatment on enzyme activity.At the same time,the changes of molecular weight of total protein was observed by SDS-PAGE electrophoresis.Moreover,the changes of the protein secondary structure of malate dehydrogenase after treatment were explored.The results showed that after plasma treatment,the activities of key enzymes such as Na+/K+--ATPase and malate dehydrogenase decreased gradually with the increase of treatment time.The residual activity of Na+/K+--ATPase in S.enteritis and L.monocytogenes decreased by 85%and 93%,while the residual activity of malate dehydrogenase decreased by 97%and 85%,respectively.What's more,for different bacteria,the correlation between enzyme activity and colony number of the two enzymes was also different.Na+/K+--ATPase had higher correlation with S.enteritidis in colony.While malate dehydrogenase had higher correlation with L.monocytogenes.The molecular weight and structure of the protein also changed after being treated by CPCS.The a-helix was decreased,while the ?-folding was increased.The stable secondary structure tends to be irregular after treated by cold plasma.