Sterilization Mechanism And Application Of The Combination With Plasma-Activated Water And Organic Acids On Staphylococcus Aureus

The continuous increasing number of food poisoning caused by Staphylococcus aureus and the exposure of S.aureus to fresh vegetables and fruits constitute a serious threat to human life.Plasma-activated water（PAW）,as a cost-effective and environmentally friendly and broad-spectrum disinfectant,provides a new application possibility in the food industry.Fencing technology,as a combination of multiple advanced technologies for sterilization,may effectively inhibit food spoilage and prolong shelf life.However,"incomplete sterilization"caused by PAW can lead to the formation of sublethal damage to the strain,further limiting the inactivation efficiency of subsequent combined sterilization.As a result,optimizing the PAW sterilization method is critical to avoid the formation of cross stress states of strains during food processing.Organic acids,as a safe and environmentally friendly chemical antibacterial agent generated from fruits and vegetables,are typically approved by the FDA as"generally considered safe"and present the advantages of high sterilization efficiency and a high safety factor.At present,it is still unknown whether organic acids will significantly enhance the bactericidal ability of PAW treatment,and whether the combination of different bactericidal methods would create changes in bactericidal efficiency requires additional investigation.Therefore,this article aims to investigate the bactericidal effect of PAW in combination with organic acids in order to improve the bactericidal efficiency and the application potential of PAW（1）The optimal synergistic bactericidal ability of organic acids was selected according to the investigation of the synergistic treatment of seven organic acids and PAW,including ascorbic acid,tartaric acid,and lactic acid,et al.The optimal synergistic bactericidal combination was investigated according to the bactericidal effect of organic acid pretreatment,PAW pretreatment,and synergistic treatment on S.aureus.The results showed that the synergistic treatment of tartaric acid（TA）and PAW achieved 4.33 lg CFU/m L of inactivation within 10 minutes,significantly higher than the synergistic treatment of the other six organic acids.Secondly,in different combination treatments,the optimal combinations were selected based on the bactericidal ability at the same time were 0.75%TA pretreatment for 6 minutes combined with PAW50,PAW60 pretreatment for 6 minutes combined with 1.00%TA,and the combination with 0.75%TA and PAW50,respectively.In addition,among different pre-treatment methods,the bactericidal ability of 0.75%TA pre-treatment is higher than that of PAW60 pre-treatment.According to the fitting of sterilization kinetics curve,Log-logistic was selected as the optimal kinetic model.The time required for the removal of S.aureus（10⁴CFU/m L）of TA pretreatment and synergistic treatment is respectively 7.24,11.10,13.21 min.The sterilization efficiency of synergistic treatment is higher than that of TA pre-treatment.Therefore,the synergistic treatment of 0.75%and PAW50 showed the best bactericidal effect.（2）The bactericidal mechanism of synergistic treatment on S.aureus was elucidated according to the cell wall integrity,energy metabolism,and physicochemical properties of activated acids.The combined treatment significantly reduced the p H,increased ORA and conductivity of the activation solution,increased the NO₂^-and H₂O₂ content,resulting in more active chemicals entering the cell and providing higher bactericidal ability.Secondly,the synergistic treatment leads to the destruction of the inner membrane of cells,resulting in bacterial distortion,dents,and pores.The leakage rate of intracellular substances increases to74%,the permeability of the cell membrane increases,membrane lipid peroxidation occurs,and the activities of Ca²⁺Mg²⁺-ATPase and Na⁺K⁺-ATPase decrease to 0.45 and 0.39 U/mgprot respectively,resulting in cell membrane integrity disruption.In addition,synergistic treatment could induce a large amount of active oxygen and H₂O₂ accumulation intracellular,which disrupted cellular homeostasis and caused a substantial drop in SOD and CAT activity to 3.93and 5.31 U/mgprot（p<0.05）,hence escalating oxidative stress.The imbalance of intracellular oxidative homeostasis led to a significant decrease in ATP concentration,which affected the reduction of MDH and SDH activities in the tricarboxylic acid cycle,resulting in the inability to generate energy to resist external stress and further leading to the difficulty of maintaining normal sorting activity and death of bacteria.（3）The application of the synergistic treatment for fresh cut pineapple and fresh cut lettuce under storage conditions 4,25°C was investigated.The antibacterial application effect of combined treatment was evaluated by measuring the total number of S.aureus.The results showed that the combined treatment could reduce the microbial counts of 2.53 and 1.77 lg CFU/g on the surface of fresh-cut lettuces and pineapples.Synergistic treatment could significantly inhibit the growth of S.aureus,and effectively reduced POD and PPO activities,reduced the oxidation degree of fresh-cut lettuces and pineapples,and thereby reduced the color difference between fresh-cut lettuces and pineapples during storage.At the same time,synergistic treatment effectively reduced the content of malondialdehyde,and delayed the decline of titratable acid,which effectively delayed the quality deterioration of fresh cut lettuce and fresh cut pineapple.The shelf life of fresh cut pineapples prolonged 2 d.The shelf life of fresh cut lettuce under 4 and 25°C prolonged 2 and 3 d,respectively.