| Food safety problems caused by foodborne pathogens,such as Salmonella,threaten human health.Radio frequency(RF)technology can rapidly heat large volume samples and achieve efficient pasteurization of both high moisture foods(HMFs)and low moisture foods(LMFs)due to its volumetric heating and deep penetration depth.However,RF heating technology has caused the degradation of heat-sensitive food quality due to its thermal effect and local overheating caused by non-uniformity heating,limiting its wide applications in the food industry.Natural antimicrobials are green and safe with little damage to food quality,but the complex ingredients in food inhibit bactericidal effects,resulting in low bactericidal efficiency when used alone.Therefore,to enhance the bactericidal effect and ensure food quality,a combined pasteurization technology of natural antimicrobials and RF heating was developed on the basis of the analysis of bactericidal effects of natural antimicrobials and heat treatment in high and low moisture substrates alone and in combination,and the applicability of the combined pasteurization technology in HMF(pork)and LMF(alfalfa seeds)was analyzed.The synergistic bactericidal mechanism of RF heating and natural antimicrobials was explained at levels of cell and metabolism,providing theoretical basis for applications of the combined pasteurization technology in practical production.The specific research contents and results are summarized as follows:(1)High-lipid and high-moisture fluid substrate(diluted tahini),high-lipid and low-moisture fluid substrate(undiluted tahini)and granular low-moisture substrate(sesame seeds)were constructed,and the bactericidal effects of natural antimicrobials and heat treatment alone and in combination in different substrates were studied.The results showed that oregano essential oil(OEO)andε-PL had good antibacterial effect on Salmonella in vitro,and the minimum inhibitory concentration(MIC)of OEO orε-PL against Salmonella Montevideo CICC 21588 was 0.03125%or 0.0625%(w/w),respectively.However,the antibacterial effect decreased in food substrate.The antibacterial effect of essential oil(EO)was severely weakened in the high-lipid fluid substrates.EO vapor had obvious antibacterial effect on bacteria on the low-moisture particles.ε-PL had poor antibacterial effect in high-lipid and low-moisture fluid substrate,but had stronger antibacterial effect than EO in high-lipid and high-moisture fluid substrate.Weibull model was suitable to fit the thermal lethal curves of S.Montevideo in three different substrates(R~2=0.973-0.998,RMSE=0.021-0.279).In high moisture substrate,the thermal lethal curves of bacteria were concave-downward(p>1),a great number of sublethal damage cells were generated;In low moisture substrate,the thermal lethal curves of bacteria were concave-upwards(p<1),and few sublethal cells were generated after heating.In addition,heat treatment and OEO orε-PL showed a synergistic bactericidal effect on S.Montevideo in high-moisture fluid substrate,but no synergistic bactericidal effect in low-moisture fluid substrate.The synergistic bactericidal effect was related to the sublethal damage of bacteria cells after heat treatment.(2)Based on the previous studies on the bactericidal effects of natural antimicrobials and heat treatment alone and in combination,the combined pasteurization technology of RF heating and natural antimicrobials was developed,and its applicability was analyzed in high moisture food--pork.Cinnamon essential oil nanoemulsion(CEON)was prepared by ultrasound methods and characterized.The effects of CEON andε-PL on microbial population,microbial structure and quality of raw pork during cold storage were investigated,and the effects of RF heating combined with CEON andε-PL on microbial inactivation and quality of pork were analyzed.The results showed that prepared CEON had good physical stability and antibacterial effect.CEON andε-PL had the additive antibacterial effect on S.Montevideo CICC 21588 and Pseudomonas lundensis in vitro.Soaking in the solution containing CEON andε-PL could both inhibit the growth of total bacterial count(TBC)and reduce S.Montevideo population in raw pork during cold storage.High-throughput sequencing(HTS)indicated that 0.250%CEON and 0.125%CEON+0.250%ε-PL exhibited better antimicrobial effects on Photobacterium than Pseudomonas.0.5%ε-PL had a weak antimicrobial effect on Photobacterium.CEON and CEON+ε-PL treatment slowed down the accumulation of total volatile base nitrogen(TVB-N)and p H rise rate in raw pork.RF cooking saved cooking time compared with water bath(WB)cooking and there were no significant differences in quality between RF and WB cooked pork samples.The combination of RF heating and CEON+ε-PL had synergistic bactericidal effect on S.Montevideo and TBC in pork.CEON+ε-PL had no significant effect on TVB-N content,p H,cooking loss and appearance of RF cooked pork,but reduced TBARS formation and enhanced the hardness,springiness and chewiness.CEON+ε-PL inhibited the TBC growth of RF cooked pork during cold storage.High concentration of CEON(0.25%)affected the sensory properties and volatile components of RF cooked pork.(3)The application of the combined pasteurization technology of RF heating and natural antibacterial substances in low moisture food--alfalfa seed was analyzed.The effects of EO vapor and RF heating on Salmonella population on alfalfa seed,seed germination rate and sprout quality were investigated.The results indicated that cinnamon essential oil(CEO)vapor had good bactericidal effect on S.Montevideo CICC 21588 on alfalfa seeds.The germination rate and heat resistance of S.Montevideo decreased with the increase of moisture content,target temperature and holding time during RF heating.Weibull model well fitted the thermal lethal curves of S.Montevideo on seeds under RF heating(R~2=0.952-0.999,RMSE=0.025-0.231,p<1).According to the bactericidal effect of EO vapor,the appropriate treatment condition was selected(Seeds were treated by CEO vapor at the concentration of 0.83μL/m L for 3 d).According to the germination rate change,bacteria inactivation curve and temperature distribution after RF heating,the appropriate RF heating conditions were determined.RF heating conditions were as follows:seeds with 7.52%w.b.,9.53%w.b.and 11.45%w.b.were intermittently heated to 65℃(RF55-60-65),60℃(RF50-55-60)and 55℃(RF45-50-55)of the cold spots,respectively,and holding for 20 min.The CEO vapor sequentially combined with RF heating had additive bactericidal effect on S.Montevideo and caused about 4 log CFU/g reductions in the cold spot.There was no significant increase of sublethal cells of Salmonella in single treatment and combined treatments.After combined treatments,the seed germination rates were more than 90%and there was no significant effect on the length,fresh weight(FW),total phenol content,ascorbic acid content and DPPH radical scavenging activity of alfalfa sprouts.The volatile components of CEO did not remain in alfalfa sprouts and volatile components of alfalfa sprouts did not change after combined treatments.(4)The synergistic bactericidal mechanisms of natural antimicrobials(CEON orε-PL)and RF heating on Salmonella were explored at the cellular level.The results indicated that RF heating caused sublethal damages to cells and promoted the interaction of antimicrobials(CEON orε-PL)with bacterial cells,thus leading to a synergistic bactericidal effect on S.Montevideo CICC 21588 of RF heating and antimicrobials.The synergistic bactericidal mechanisms of RF heating andε-PL were mainly promoting the cell morphological alterations,increasing cell membrane permeability,increasing ATP leakage,reducing intracellular ATP content,increasing intracellular ROS level and reducing intracellular nucleic acid substances level.Compared with single RF heating,although the cell membrane permeability and intracellular ROS level increased after the combined treatment of RF heating and CEON,the increasing level was less than that of the combination of RF heating andε-PL.In addition,the extracellular ATP level in the combined group of RF heating and CEON was lower than that of RF heating group,indicating that the main synergistic bactericidal mechanism of RF heating and CEON was to significantly inhibit intracellular ATP synthesis.(5)The synergistic bactericidal mechanisms of RF heating and CEON orε-PL on Salmonella were explored based on cell metabolomics.Results of the analysis of the numbers of significantly different metabolites(SDMs)between control and treatment groups,and fold change(FC)of metabolites level indicated that the combined treatment of RF heating andε-PL or CEON had increased impacts on cell metabolism compared to single treatment,thus leading to synergistic bactericidal effect against Salmonella.KEGG(Kyoto Encyclopedia of Genes and Genomes)enrichment results illustrated that the combination of RF heating and antimicrobials(ε-PL and CEON)enhanced the disruptions to nucleotide metabolism,amino acid metabolism,carbohydrate metabolism,metabolism of cofactors and vitamins,biosynthesis of cofactors,lipid metabolism and membrane transport pathways compared to single treatment.In addition,the impact of the combination of RF heating andε-PL on cell metabolites was different from that of the combination of RF heating and CEON. |
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