The Fungicidal Effect Of Low-temperature Plasma On Penicillium Expansum

China is the largest apple producer in the world,and the apple's annual output in China accounted for 1/3 of the world.However,mechanical damage and some pathogenic mould infection caused heavy losses of apple production in the process of picking,storage,transportation and processing.Of all the microorganism to cause apples rot,infection of Penicillium expansum for apples and other fruits is the most serious.P.expansum could cause entirely spoilage of fruit within ten days due to its stronger adaptability,fecundity and patulin-production ability.Patulin,a secondary metabolites,produced by many species of moulds has extensive poisonous,carcinogenic and mutagenic effects on animal tissues and cells.Therefore,it is essential to develop a safe and efficient method to kill P.expansum for the fruit processing industry.In this study,P.expansum,isolated from rot apples,were used as model microorganism.The aims of this work were:(1)to study the growth and patulin-producing characteristics of three P.expanusm strains,(2)to investigate the factors,influencing the fungicidal efficiency of low-temperature plasma generated by gas phase surface discharge on Penicillium expansum,(3)to elucidate the fungicidal mechanism of lowtemperature plasma.The research provided a theoretical basis for application of low-temperature plasma sterilization in the fruit processing industry.The main research results are as follows:(1)Three strains(F-LPH10-6,CICC40658 and CGMCC3.3703)of P.expansum were used to investigate their growth and patulin-producing characteristics in the experiment.The biomass(dry weight)of F-LPH10-6,CICC40658 and CGMCC3.370 were 128.7,86.8 and 28.6 mg,respectively after incubation for 14 days at 25 ?,and the corresponding concentration of patulin were 147.97?0.91 and 0.51 mg/L respectively,under the same conditions.Moreover,P.expansum F-LPH10-6 produced much more patulin than that of P.expansum CICC40658 and CGMCC3.3703.(2)The fungicidal effect was studied by self-made low-temperature plasma generated by gas phase surface discharge reactor.Influencing factors of fungicidal effect by plasma included: treatment time,power supply voltage,initial spores concentration,sample medium and concentration of ascorbic acid.These factors showed much significant influence(p<0.01)on fungicidal effeciency except for the sample medium.(3)Furthermore,the changes in morphology of P.expansum spores before and after treatment were characterised by using scanning lectron microscope and transmission electron microscope.The observation showed that spores before plasma treatment were regular round or oval,possessed full morphology,complete the cellular structure and clearly visible intracellular organelles.However,spores after plasma treatment were surface shrinkage or sag,cell wall and membrane rupture,cytoplasm spillover,some cells even broken into pieces.(4)The alteration of pH and concentration of nitrite in medium during plasma treatment were studied.Meanwhile,the effect of different pH on fungicidal effect of P.expansum F-LPH10-6 was also investigated.The results showed that pH of the sample reduced with treatment time and a little of nitrite was generated in samples,which was probably related to the reactive particle interactions during discharge.Meanwhile,it turned out that the infulence of pH on fungicidal effect was not significant,which proved that the acidification may be not the primary mechanism of plasma sterilization and may only play a supplementary role in the process of sterilization.(5)The reactive oxygen species,heat effect and membrane damage during plasma sterilization were determinated.The results showed that concentrations of ozone and hydrogen peroxide in P.expansum F-LPH10-6 sample increased with treatment time.Concentrations of ozone and hydrogen peroxide were 22 ?mol/Land 0.41 mg/L respectively,after 30 min treatment,which suggested that reactive oxygen species played a crucial role in plasma sterilization.Meanwhile,it turned out that the change of temperature in sample was low 3?(close to room temperature),which proved that heat effect was not the main reason of spores death.The results also showed that concentration of MDA increased with treatment time in the samples with the content as 134 nmol/L after 18 min.The scanning results of samples before and after plasma treatment from 200 to 400 nm wavelength showed that the leak of nucleic acids and proteins existed in the sample after plasma treatment.