| Usually,microbial cells will be affected by a variety of stresses in their surrounding environment during their natural habitats or industrial training process,such as the light,heat,pH,pressure,electromagnetic field and so on.Under those stresses conditions,the cellular damage phenomenon will be produced inevitably,including sublethal injury cells(SICs)which still have a certain physiological activity and could repair and restore their ability to reproduce under the appropriate condition.Because of their special physiological properties,if the SICs are not be killed completely during the process of sterilization,the huge economic loss will be very likely to be brought into the food industry.Therefore,it is of important significance and broad application prospects in the future to carry out related research on the sublethal injury effect of microbial cells exposed to the external stress environments.Saccharomyces cerevisiae is one kind of the ideal model of eukaryotic organisms,and commonly used as the biological materials in industrial production.Take the S.cerevisiae cells as the research object,the detection methods of SICs induced by pulsed electric fields(PEF)and their physiological behavior after PEF-treated were discussed within this paper in detail.Using NaCl as the osmotic pressure modifier,the NaCl concentration of a selective medium which was applied for the quantitative analysis of the SICs induced by PEF was confirmed as 4.25% by exploring the critical osmotic pressure in which the growth and reproduction of SICs would be inhibited completely.Based on the selective medium,the S.cerevisiae cells were treated by PEF.The results showed that the critical electric field parameters influencing the induction of SICs were electric field strength and the total treatment time.Comparatively speaking,the influential effect of frequency and pulse width seemed to be not so important.And the results also indicated that the PEF with a low electric field strength and a short treatment time could induce a large number of SICs.The sterilization power and the heating power causing the temperature rise of treatment fluid for the PEF treatment system were analyzed during the processing.The results showed that the material conductivity seriously affected the heating power,and then affected the thermal effect.Under a low electric field intensity,there was a higher-energy utilization rate for the high conductivity materials when they were treated by PEF.In addition,with the help of image analysis on the colonies,the presence of SICs could be identified quickly,and the different energy dissipation had a significant influence on the induction rate of sublethal injury,but the induction of the SICs during PEF treatment was a combined result from the multiple factors.With the help of static headspace gas chromatography,the release process of carbon dioxide(CO2)from the situ culture of S.cerevisiae cells was monitored and analyzed.The results showed that the overall growth of the yeast cells during their growing process could be indirectly reflected by the changes of CO2 which was generated during the growth of cells and detected by static headspace gas chromatography.Furthermore,the growth process of yeast cells in situ cultivating was predicted by the four common S-shaped models.The results showed that the Weibull model could be well applied to predict the growth process of yeast cells which were in the lag phase and cultivated in a non-selective medium,but not suitable for the cells in the exponential phase.The fitting degree was relatively decreased when the four kinds of prediction models were implemented to predict the growth process of yeast cells which were cultivated in a selective medium,especially,the R-squared was below 0.8000 for all the models when the inoculation amount was 10.0%.Relatively speaking,Logistic model showed the best predictive effect and had the good fitting degree for yeast cells cultivated in all the conditions when the yeast cells were in the exponential phase,which implied that the Logistic model could be suitable for the prediction of growth process of the SICs induced from the yeast cells in logarithmic phase.But for the yeast cells in the stationary phase,only when the inoculation amount was relatively low(e.g.5.0%),there was a good fitting degree for all the four kinds of prediction models.By means of static headspace gas chromatography,the growth process of yeast cells in exponential phase and treated by PEF before cultivated in a different medium was monitored,and the Logistic model was used to predict all the growth processes,and the fitting degree was very good.In addition,the changes of specific growth rate in the growth process for the yeast cells cultivated in different conditions were analyzed in detail,and the results showed that the maximum specific growth rates of yeast cells were all increasing and the time of achieving the maximum specific growth rate was significantly longer when the cells were treated by PEF.The growth processes of yeast cells were contrastively analyzed when the cells were cultivated in non-selective medium and selective medium respectively,and the results indicated that the quantitative proportion of SICs induced by PEF treatments was closely connected with their population growth process.The preliminary analysis for the ion metabolic processes of yeast cells during stationary culture under starvation or no starvation after treated by PEF showed that the instant changing law of pH and conductivity in the suspension after PEF-treated was similar.In other words,the value of pH and conductivity were reduced when the electric field strength applied was low,but with the increase of electric field strength,the values of pH and conductivity of the suspension increased slowly.And when the electric field strength sequentially increased to a certain value,the value of pH and conductivity of suspension could exceed the untreated finally.In addition,the concentration changes of sodium and chloride ions showed a trend of decrease,but relatively speaking,with the increase of electric field strength,the chloride ion concentration reduced faster.There were a number of changes for the value of pH,conductivity,chlorine ion and sodium ion in suspension when the yeast cells were in stationary culture under starvation or no starvation after treated by PEF.For example,the value of pH and conductivity increased and the chloride ion and sodium ion were decreased when the suspension was in stationary culture,and when the presence of carbon and energy source material such as sugar,there were obvious differences for the degree of reduce or increase.These results demonstrated that the dynamic change of the ions in a certain sense also was a kind of result for cells adapting to the nutritional environment,and the more obvious for the dynamic change,the cell vitality was stronger and their physiological metabolism was stronger too.In addition,all sorts of extracellular cation had an important effect on the osmotic pressure outside and inside of yeast cells.In addition,the presence of sugars is helpful for the sublethal injury yeast cells to restore their actives,which indicated that the available sugar may have a reliable protective effect on the SICs.The purpose of this paper is that,on the one hand,the in-depth understanding of microbial sublethal injury could help to strengthen the extent of liquid food sterilization in food-industry,and on the other hand,it would open a new route for some important raw materials in the food industry production. |
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