Pulsed Electric Field of Orange Juice
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Pulsed Electric Field Processing of Orange Juice: A Review on Microbial,Enzymatic,Nutritional, and Sensory Quality and StabilityRoman Buckow,Sieh Ng,and Stefan ToepflAbstract:During the last decades pulsed electricfield(PEF)processing received considerable attention due to its potential to enhance food products or create alternatives to conventional methods in food processing.It is generally acknowledged that PEF processing can deliver safe and chill-stable fruit juices with fresh-like sensory and nutritional properties.Relatively low-processing temperature and short residence times can achieve highly effective inactivation of microorganisms while retaining product quality.Afirst commercial application of PEF for preservation of fruit juices was launched in2006in the United States.Since then,industrial-scale processing equipment for liquid and solid products were developed and,in Europe in2009,an industrial juice preservation line was installed using20kV/cm pulses at40to 50◦C to extend the chill-stability of fruit juices,including citrus juices and smoothies,from6to21d.The related PEF processing costs are in the range of US$0.02to0.03per liter and are justified due to access to new markets and reduced return of spoiled product.However,despite its commercial success there are still many unknown factors associated with PEF processing of fruit and citrus juices and many conflicting reports in the literature.This literature review,therefore, aims to provide a comprehensive overview of the current scientific knowledge of PEF effects on microbial,enzymatic, nutritional,and sensory quality and stability of orange juices.IntroductionPulsed electricfield(PEF)processing has the ability to effectively inactivate microbial cells,when combined with low to moderate temperatures(<50◦C),which makes it a promising alternative to conventional thermal preservation processes for liquid foods that contain heat labile bioactive or volatile components such as fruit and vegetable juices.The sensitivity of microorganisms to PEF treatments depends on cell characteristics such as structure and size (T oepfland others2006).In addition,factors such as product pH, water activity(a w),soluble solids,and electrical conductivity affect the efficiency of the technology to affect biochemical reactions and inactivate microorganisms(Aronsson and R¨o nner2001). Although the underlying mechanisms are not yet fully explained on a molecular basis,PEF treatment results in changes in the membrane permeability of biological cells(G´a skov´a and others 1996).This effect can be exploited to inactivate microorganisms or to enhance mass transfer in extraction or drying processes.T ypically,food is placed between2electrodes and exposed to an electricalfield in the form of very short(a fewμs),high-voltage MS20130138Submitted1/26/2013,Accepted4/25/2013.Authors Buckow and Ng are with CSIRO,Animal,Food and Health Sciences,671Sneydes Rd.,Werribee, VIC3030,Australia.Author T oepflis with German Inst.of Food T echnologies(DIL), Prof.-von-Klitzing-Str.7,49610Quakenbr¨u ck,Germany.Direct inquiries to author Buckow(E-mail:roman.buckow@csiro.au).(kV)pulses.One electrode is connected to a high-voltage switch and the other to the ground.The electricfield strength E generated between the pair of electrodes can be estimated by dividing the applied voltage U by the distance between the electrodes d(that is,E=U/d).The electricfield strength,treatment temperature,treatment time,and specific energy input are the main processing parameters affecting the degree of microbial inactivation(Alvarez and others 2006).T o ensure effective microbial inactivation,electricfield strengths should be in the range20to50kV/cm,pulse lengths in the range of1to10μs and specific energy inputs in the range50to1000kJ/kg(T oepfland others2007).However,the multitude of variable parameters of PEF technology and the food characteristics often requires a systematic study of the individual influence of these parameters on desired and undesired reactions. T o establish a typical PEF system,a pulse generator is needed.A typical system comprises a high-voltage power supply,one or more(energy storage)capacitors,a high-voltage switch and a treat-ment chamber.Furthermore,a liquid handling system including heat exchangers,a product pump,and monitoring devices(oscil-loscope,temperature sensors)to control and report the conditions, are necessary(Zhang and others1995).T o deliver the energy at high voltages within a short period of time(μs)a capacitor is needed.The capacitor has the function of storing the generated energy from the pulse generator at the selected voltage.When the high-voltage switch is closed,energy is discharged from the capac-itor almost instantly to the electrodes in the form of a high-voltageC 2013CSIRO and DILdoi:10.1111/1541-4337.12026Vol.12,2013r Comprehensive Reviews in Food Science and Food Safety455PEF preservation of orange juice...pulse of a few microseconds.The relatively slow charging of the system compared to the fast high-energy discharge of the capac-itors often requires significantly longer time spans(about100to 1000times)between the pulses than the width of the pulse itself. The chamber design exhibits a significant influence on the ef-fectiveness of the process by affecting treatment uniformity,peak electricalfield strength,and product throughput(Buckow and others2011).One challenge is to design a treatment chamber capable of operating at high and uniform electricfield intensi-ties and which prevents dielectrical breakdowns(Qiu and others 1998).Batch chambers with parallel plate electrode configurations provide relatively low throughputs but high-treatment uniformity. Treatment chambers with colinear configurations of electrodes al-low continuous operation at high throughputs,but often exhibit poor treatment uniformity.Dielectrical breakdowns occur when the applied electricalfield strength exceeds the dielectric strength of the treated food product in the chamber(Zhang and others1995).Dielectrical breakdowns can also be caused by localfield enhancement and impurities(gas bubbles or solids)in liquid foods.Not less important are other rel-evant design criteria such as electrical resistance of the chamber, low-erosive electrode material(for example,platinum),improve-ment offlow behavior and temperature distribution,and of course the treatment capacity(that is,product throughput and constant power supply)(Jaeger and others2009).Nonuniform electricfield andflow velocity distributions can result in under-(often in central regions or dead spaces)or overtreated(often in boundary regions) volume elements,which lead to an increased chance of electrical sparks and system breakdown,as well as degradation of the quality of the treated product.Thus,the effectiveness and efficiency of the PEF treatment decreases.The majority of pilot and industrial-scale PEF systems comprise treatment chambers(flow cells)with cofield and colinear configu-rations of electrodes.Such chambers have2or3hollow cylindrical electrodes(2high-voltage electrodes and1grounded electrode) separated by hollow insulating spacers.The productflows through the formed tube.This electrode design and configuration provides a very nonuniform electricalfield and an undefined treatment zone (V an den Bosch2007).The nonuniformity may result in insuffi-cient treatment,dead spaces and,thus,possibly recontamination of the treated medium with microorganisms.This presents a major challenge when the technology is used for pasteurization appli-cations where a5-or6-log10reduction of pathogens has to be achieved.T o reduce the risk of under-or overprocessing,multiple PEF treatment chambers can be placed in series.For example,in a PEF system that operates continuously(for example,using colinear treatment chambers),treatment uniformity can be enhanced by adding the number of treatmentflow cells.This numbering up of treatment zones also means that the required processing time can be broken up into smaller fractions allowing intermediate cooling of the product.This can slightly reduce the effectiveness of the treatment but will preserve the quality of the product better than in the case where the required process intensity(that is,specific energy)is applied in one treatment zone,which often results in a significant temperature rise.In general,a turbulentflow is desirable because it results in a more uniform residence time(Heinz and others2002).The high intrinsic resistance of colinear chambers makes it possible to use several chambers in series and,in addition,enables the pulse generator to operate with smaller currents than in coaxial treatment chamber designs.Additional benefits of colinear systems include the possibility of a greater diameter enabling higher prod-uct throughput,the ease of cleaning,an effective usage of electrical energy,and goodflow dynamics(Buckow and others2010). Another way of enhancing treatment uniformity of continu-ous PEF systems is to modify the dimensions and geometry of the treatment chamber,which can either result in a more uni-form distribution of the electricalfield strength and/or enhanced flow patterns of the treated food.Modification of PEF treatment chamber design can either include shape modifications of the in-sulator(Buckow and others2011)or electrode configuration(for example,a concentric rift)(T oepfl2011)or insertion of insula-tor material or a metal grid into theflow shapes configurations (Alkhafaji and Farid2007;Jaeger and others2009).However,cur-rent PEF research and development is mainly focused on further optimizing colinear treatment chamber designs rather than look-ing into new ways of applying high-voltage pulses to food systems (Huang and W ang2009).PEF Inactivation of Microorganisms in Orange Juice Mechanisms of microbial inactivation by PEFThe inactivation of vegetative bacteria and yeasts during PEF processing is probably not due to the products of electrolysis or temperature rise alone,but rather determined by the applied elec-tricalfield strength and the treatment time(Evrendilek and Zhang 2003;Heinz and others2003).There are several hypotheses on the mechanisms involved in the rupture of the cell membrane when exposed to an electricfield.T wo hypotheses,electrical breakdown, and osmotic imbalances,are widely accepted and are based on the same principles.The theory of electrical breakdown considers the cell membrane as a capacitorfilled with a dielectric medium(Zimmermann1986). The cell cytoplasm has a greater dielectric constant(6to8times) than the membrane,as does the liquid food that surrounds the cell.The difference between dielectric constants on either side of the membrane results in a transmembrane potential(TMP)of about10mV(Jeyamkondan and others1999).The TMP is created through free charge accumulations at the inner and outer surface of the cell membrane.When an external electricalfield is applied, ions inside and outside of the cell move along thefield until they are restrained and accumulate at the membrane and an increase in the TMP occurs.The ions of opposite charge(+and–)on either side of the membrane are attracted to each other,compress the membrane and reduce its thickness.When the electricalfield strength is increased to a point where it exceeds a critical threshold value of the TMP,an electrical breakdown or pore formation is induced.The critical TMP value is around1V(Hamilton and Sale 1967).The electricalfield strength required to achieve an electrical breakdown of the cell membrane depends on cell size and shape, cell orientation in the electricfield,dielectric characteristics of liquid food,cytoplasm and membrane,and temperature. Breakdown and pore formation can be reversible or irreversible, depending on the treatment intensity.If the intensity of the PEF is such that the energy deposited on the membrane does not result in considerable Joule heating the cell can often recover from the damage done to its membrane(Kolb and others2006). In this case,the cell membrane can remain permeable for some minutes depending on the temperature after exposure to mild PEF treatment(Rols and T eissie1998).However,when pores in the membrane surface become numerous and/or large in size,an irreversible breakdown of the membrane occurs,which leads to a mechanical destruction of the membrane and subsequent cell death.456Comprehensive Reviews in Food Science and Food Safety r Vol.12,2013C 2013CSIRO and DILPEF preservation of orange juice...The theory of osmotic imbalance describes the loss of cell mem-brane functionality through formation of hydrophilic pores in the membrane and the forced opening of protein channels.The elec-tricalfield causes changes in the conformation of phospholipids, leading to rearrangement of the membrane and formation of hy-drophilic pores.The opening and closing of protein channels em-bedded in the membrane also depends on the TMP.There is ev-idence that charged H+protons rapidly move along a lipid-water interface(for example,a biological cell in a suspension)and can af-fect transmembrane proton conduction(T eissie and others1985). The gating potentials of protein channels that must be overcome for solutes(for example,ions)toflow through are approximately 50mV,which is150to500mV less than the breakdown potential of the lipid bilayer of the membrane(Tsong1991).During appli-cation of an electricfield,protein channels might not only open, but also become denatured(irreversible pores)by local Joule heat-ing or electric modification of functional groups(Tsong1991). There are reports that the conductivity of the media surround-ing the biological cell will affect the permeabilization process dur-ing PEF treatment(Ivorra and others2010;Moisescu and others 2013).This is because the time of charge accumulation on the cell surface is smaller in high-conductive media.This means that mi-crobial inactivation is likely to be enhanced in high-conductive media than in a low-conductive environment(for example, <1mS/cm).However,increasing the electrical conductivity also increases theflow of electrical current leading to pronounced Joule heating and increased energy consumption during PEF processing. Factors affecting microbial inactivationMany process parameters affect the effectiveness of the PEF treatment for inactivating microorganisms.Changing one param-eter might affect another.These circumstances make it difficult to compare the results of different studies.Nevertheless,the main process parameters influencing the effectiveness of PEF treat-ments and,thus,microbial inactivation are the applied electric field strength,pulse length and shape,total treatment time,treat-ment temperature,and specific energy input(Heinz and others 2002).T ypically,the greater the electricfield strength,higher the tem-perature or longer the treatment time,the greater the microbial inactivation(W outers and others2001).For example,a study by McDonald and others(2000)demonstrated the effects of PEF on microorganisms in orange juice applying electricfield strengths of 30and50kV/cm for12μs and at55◦C outlet temperature.PEF treatment at30or50kV/cm inactivated4.75-log10and6.2-log10 CFU/mL Leuconostoc mesenteroides in orange juice,respectively.Es-cherichia coli O157:H7inoculated in orange juice was inactivated 1.0,2.4,and3.4-log10through75μs PEF treatment at13.1, 19.7,and23.7kV/cm at55◦C,respectively(Gurtler and others 2010).´Alvarez and others(2003)reported a“Z PEF-value”(that is, a10-fold increase of the inactivation rate)for PEF inactivation of different Salmonella serovars in citrate–phosphate buffer(pH7.0) of15.8kV/cm(in the range of19to28kV/cm).The impact of temperature on the PEF lethality of E.coli in apple juice was systematically studied by Heinz and others(2003) at temperatures ranging from35to70◦C.Energy requirements (which correlates with processing time)to achieve a7-log10inacti-vation of E.coli during PEF processing of apple juice at24kV/cm decreased from160to100kJ/kg when the process temperature was increased from40to50◦C clearly indicating enhanced ef-ficiency of PEF at elevated temperature.E.coli O157:H7sus-pended in apple juice is inactivated approximately0.5,1.5,and 2.8-log10by PEF treatments at20kV/cm and20,30,and40◦C (initial temperature),respectively(Saldana and others2011).Simi-larly,inactivation of Salmonella typhimurium inoculated into citrate–phosphate buffer(pH3.5)was inactivated1.5,2.9,4.0,and5.0 log10CFU/mL after90μs PEF treatment at30kV/cm and15, 27,38,and50◦C,respectively(Saldana and others2010).Hence, an increased treatment temperature during PEF processing gener-ally corresponds to increased inactivation of microorganisms at a particular electricfield strength.Finally,pulse shape and pulse width can affect the microbial killing efficiency of PEF.The efficiency of PEF to permeabilize biological cells is largely dependent on the time during which the electricfield strength exceeds a certain critical value(Kotnik and others2003).Therefore,it is generally accepted that exponential decay pulses require more energy than rectangular or square wave pulses since the critical electricalfield strength required to kill microorganisms is only exceeded during thefirst third of the total pulse width.De Haan and others(2002)concluded that exponential decay pulses will not exceed an energy efficiency of 38%compared to square wave pulses at similar peak voltages. Increasing the pulse width from0.05to3μs at50kV/cm enhanced inactivation of Salmonella enteritidis in28mM sodium sulfate with glucose solution(Korolczuk and others2006).The estimated decimal reduction energy required for PEF destruction of S.enteritidis decreased from44to32kJ/kg when the pulse width was increased from0.05to1μs during PEF treatment at 50kV/cm and15◦C.Increasing the pulse width from1to3μs (and taking variations in temperature increase into account)did not further enhance the microbial inactivation efficiency of PEF (Korolczuk and others2006).Abram and others(2003)examined the effect of pulse width on inactivation of Lactobacillus plantarum in sodium phosphate buffer.Pulse widths of1,2,and5μs applied at25kV/cm were tested for the same treatment times.The re-sults clearly showed that higher levels of L.plantarum inactivation were obtained using a longer pulse width for the same treatment time.On the other hand,increasing the pulse width from2.5 to4μs did not enhance inactivation of L.plantarum in an orange milk beverage during PEF processing at40kV/cm(Sampedro and others2007).These contradictory results can be consequence of temperature variations due to noncontrolled evolution of the tem-perature during treatments using different PEF systems or pulses. Furthermore,it should be noted that pulse shape,rise and decay time,or polarity have not been found to influence the efficiency of electro-permeabilization of biological cells(Kotnik and others 2003).Another factor affecting microbial inactivation by PEF is the pH of the treatment medium(Alvarez and others2006).The low pH of fruit juices acts as a stressor for many bacteria and several studies have investigated its effect on the sensitivity of microorganisms to PEF.For example,E.coli inactivation was drastically increased from 1.7to5.7log10after PEF treatment at30kV/cm and30◦C for 80μs when the pH was reduced from7.0to4.0whereas for Sac-charomyces cerevisiae,the pH effect was less pronounced(Aronsson and R¨o nner2001).S.typhimurium and E.coli O157:H7in citrate–phosphate buffer at pH3.5,5.2,and7.0showed highest resistance to PEF treatment at30kV/cm for99μs at pH5.2independently of the process temperature(4to50◦C isothermal conditions) when enumerated straight after the treatment(Salda˜n a and others 2012).However,another factor to consider is the occurrence of cell damage and microbial survival after PEF treatments.There is evidence that PEF treatment induces sublethally injury in bacte-rial cells depending on the processing conditions(electricalfieldC 2013CSIRO and DIL Vol.12,2013r Comprehensive Reviews in Food Science and Food Safety457PEF preservation of orange juice...Table 1–Summary of PEF inactivation of microorganisms in orange juice.Log 10MicroorganismJuice pH PEF conditions (E,t,T max )areduction ReferenceStaphylococcus aureus 3.740kV/cm,150μs,56◦C 5.5(Walkling-Ribeiro and others 2009b )Listeria innocua n.d.c 30kV/cm,12μs,50◦C 6.0(McDonald and others 2000)Listeria innocua 3.540kV/cm,100μs,56◦C 3.8(McNamee and others 2010)Escherichia coli n.d.c 30kV/cm,12μs,50◦C 6.0(McDonald and others 2000)Escherichia coli k123.540kV/cm,100μs,56◦C 6.3(McNamee and others 2010)Eschericia coli O157:H7(EHEC) 3.422kV/cm,59μs,45◦C 1.59(Gurtler and others 2010)20kV/cm,75μs,55◦C 2.0228kV/cm,75μs,55◦C 3.79Salmonella typhimurium3.422kV/cm,59μs,45◦C 2.05(Gurtler and others 2010)20kV/cm,70μs,55◦C 2.81–3.54Samonella typhimurium n.d.c 90kV/cm,50μs,50◦C5.9(Liang and others 2002)Total aerobic count 440kV/cm,97μs,approximately 60◦C6.2(Min and others 2003)Yeasts and molds5.8Aerobic microorganisms,yeasts and molds b3.85b 25kV/cm,280μs,T not reported >3(Rivas and others 2006)Aerobic plate count n.d.c 29.5kV/cm,60μs,T not reported4.2(Qiu and others 1998)Aerobic plate count 3.7835kV/cm,59μs,60◦C 4.0(Yeom and others 2000b )Yeasts and molds4.0Zygosaccharomyces bailii 2.934.3kV/cm,4μs,20◦C3.5(spores)(Raso and others 1998b )5(veg.cells)Saccharomyces cerevisiae (ascospores)n.d.c 50kV/cm,approximately 16μs,50◦C 2.5(McDonald and others 2000)Saccharomyces cerevisiae 3.412.5kV/cm,800μs,approximately 10◦C 5.8(Molinari and others 2004)Saccharomyces cerevisiae 3.635kV/cm,1000μs,39◦C 5.1(Elez-Martinez and others 2004)Pichia fermentans3.540kV/cm,100μs,56◦C4.7(McNamee and others 2010)Byssochlamys fulva (conidiospores) 3.934.3kV/cm,30μs,20◦C 5(Raso and others 1998a )Neosartoria fischeri (ascospores) 3.942.6kV/cm,20μs,34◦C<0.1(Raso and others 1998a )Lactobacillus plantarum b 4.1935.8kV/cm.,46.3μs,T not reported 2.5(Rodrigo and others 2001)Lactobacillus plantarum 3.422kV/cm,59μs,45◦C 2.57(Gurtler and others 2010)20kV/cm,70μs,55◦C 3.07Lactobacillus lactis 3.422kV/cm,59μs,45◦C 4.15(Gurtler and others 2010)20kV/cm,70μs,55◦C 4.53Lactobacillus fermentum 3.422kV/cm,59μs,45◦C 2.11(Gurtler and others 2010)20kV/cm,70μs,55◦C 3.22Lactobacillus casei 3.422kV/cm,59μs,45◦C 0.43(Gurtler and others 2010)20kV/cm,70μs,55◦C 0.60Lactobacillus brevis 3.625kV/cm,150μs,32◦C 1.4(Elez-Mart ´ınez and others 2005)35kV/cm,1000μs,32◦C 5.8Leuconostoc mesenteroidesn.d.c30kV/cm,15μs,60◦C5.1(McDonald and others 2000)a E,electrical field strength;t,total treatment time;Tmax ,maximum (outlet)temperature.b suspended in an orange-carrot blend.c n.d.,not determined.strength,temperature,treatment time)and pH of the cell.For example,survival of Salmonella or E.coli at acidic pH (that is,3.5)was clearly decreased post-PEF treatment at 25to 35kV/cm for 100μs resulting in an additional 1to 3log 10reduction of bacte-ria count within 24-h refrigerated storage (Somolinos and others 2008;Salda˜n a and others 2010).PEF inactivation of pathogens in orange juicesPathogenic strains of Escherichia coli,Salmonella,Staphylococcus,and Listeria monocytogenes continue to cause serious outbreaks of foodborne illness and frequently occur in nonpasteurized orange juices (Ghanshyam and others 2011).The USA federal juice haz-ard analysis critical control point (HACCP)rule was set up in 2001after encountering numerous microbiological outbreaks of pathogens in juice.Among other things,the rule stipulates that the juice processors must apply treatments to reduce “pertinent”microorganisms by 99.999%or 5-log 10CFU/mL.The pertinent pathogen in citrus juice is generally regarded as Salmonella (Parish 1998).Thus,for PEF acceptance as an alternative to conventional thermal pasteurization of foodstuffs,regulators often demand the successful destruction of greater than 5-log 10of pathogenic mi-croorganisms in fruit juices.An overview of published data on PEF inactivation of pathogenic microorganisms in citrus juices is presented in Table 1.The efficacy of PEF against E.coli in fruit juice was demon-strated by Evrendilek and others (1999).A 5-log 10and 5.4-log 10inactivation of E.coli O157:H7and E.coli 8739,respectively ,were achieved in apple juice by applying 29kV/cm for 172μs at approximately 25◦C (<35◦C outlet temperature)in a con-tinuous PEF treatment system.Gurtler and others (2010)deter-mined inactivation levels of 1strain of Enterohemorrhagic E.coli O157:H7(EHEC),2strains of S.typhimurium ,and 20strains of nonpathogenic bacteria in orange juice after PEF treatment at 22kV/cm for 59μs at approximately 45◦C (outlet temperature)and at 20kV/cm for 70μs and 55◦C (outlet temperature).The level of inactivation for the E.coli O157:H7strain under each treatment condition was 1.59and 2.22log 10CFU/mL,respec-tively .Nonpathogenic E.coli strain 35218had a similar level of resistance to PEF treatment under both treatment conditions and is a potential surrogate for pilot plant challenge studies (Gurtler and others 2010).In the same study ,Gurtler and others (2010)detected reductions of 2.8and 3.5-log 10CFU/mL of S.typhimurium strains UK-1and 14028in orange,respectively ,after 59μs treatment at 22kV/cm and 55◦C.The efficacy of PEF against Salmonella in freshly squeezed orange juice was also demonstrated by Liang and others (2002).PEF treatment in a coaxial chamber with 90kV/cm for 50μs at 55◦C,achieved 5.9-log 10inactivation of S.typhimurium in orange juice.458Comprehensive Reviews in Food Science and Food Safety r Vol.12,2013C 2013CSIRO and DILPEF preservation of orange juice...Studies on PEF inactivation of other bacterial pathogens(andtheir surrogates)in orange juice are limited(Table1).PEFinactivation kinetics(approximately10◦C inlet temperature)ofStaphylococcus aureus in orange juice indicated D-values of80,49.5,and26.8μs at20,30,and40kV/cm,respectively(W alkling-Ribeiro and others2009b).McDonald and others(2000)reportedsusceptibility of Listeria innocua in orange juice to PEF processing.Treatments at30kV/cm for only5μs(42◦C)or12μs(50◦C)re-sulted in3.5and6.0-log10inactivation of L.innocua,respectively.In contrast,McNamee and others(2010)only found3.9-log10inactivation of L.innocua in orange juice after more intense PEFprocessing at40kV/cm and56◦C for100μs.This differencein treatment efficiency might be explained by differences in PEFtreatment chamber design,pulse shape(exponential decay com-pared with rectangular),chemical properties of the orange juice,and variations of bacteria cultivation methods.L.monocytogenes in apple juice was inactivated by5and6.5log10CFU/mL after treatments at25kV/cm for31.5μs(outlet tem-perature50◦C)and37μs(outlet temperature55◦C),respectively.In sour cherry juice,L.monocytogenes was inactivated by3-log10CFU/mL upon application of PEF of27kV/cm for131μs andapproximately20◦C(Altuntas and others2011).Shelf-life extension of orange juices by PEFPEF is effective at inactivating spoilage microorganisms in liquidfoods such as juices thereby extending shelf life and often improv-ing quality(Min and others2007).A summary of selected paperson microbial inactivation and shelf-life extension of orange andother juices by PEF is provided in Table1.PEF inactivation of spoilage microorganisms in orange juicewas demonstrated by Timmermans and others(2011)where PEFtreatment(23kV/cm,36μs,inlet temperature38◦C,outlet tem-perature58◦C)resulted in microbial counts at levels less than thedetection limit for up to2mo of refrigerated(4◦C)storage.Atthe end of the shelf life(58d at4◦C),PEF-treated orange juiceexhibited microbial counts of<10CFU/mL for Enterobacteriaceaeand yeast and mold,<100CFU/mL for lactic acid bacteria and, <1000CFU/mL for total plate count(that is,all counts were less than the respective detection limits).Hence,the study demon-strated that PEF is an effective process to increase the microbialshelf life of orange juice from9(fresh juice)to up to2mo whenstored at4◦C.Orange juice treated at29.5kV/cm for60μs at30◦C inlettemperature and aseptically bottled was shelf stable at4◦C for7mo whereas untreated juice spoiled(that is,total plate count>106CFU/mL)after30d(Qiu and others1998).PEF was employed in inactivating spoilage microorganisms injuice-milk blends.The combined effect of thermal and PEF pro-cessing on the shelf life of an orange juice-milk beverage(OJMB)was studied by Sampedro and others(2009).Thermal treatmentconditions were85◦C for66s whereas PEF treatment conditionswere30kV/cm for50μs,inlet temperature65◦C,and outlettemperature80◦C(estimated5s until cooling).The reductions inbacterial as well as yeast and mold counts were similar after PEF orthermal treatments(4.5and4.1-log10CFU/mL for thermal com-pared with4.5and5.0-log10CFU/mL for PEF).The estimatedshelf lives were2and2.5wk at8to10◦C for the thermally andPEF processed OJMB.Therefore,PEF-treated OJMB achieveda slightly higher shelf life than the thermally treated OJMB.Foran OJMB,treatment with PEF achieved the equivalent enzymeinactivation as the thermal treatment,but better preserved colorandflavor compounds(Sampedro and others2009).However,it should be noted that thermal processing times(66s)used in this study are slightly greater than typically used in industry and should have yielded better microbial kill and stability.A study by Sharma and others(1998)produced a PEF-treated protein-fortified orange juice with a microbiologically stable shelf life of5mo at4◦C.Jia and others(1999)achieved a6-wk shelf life at4◦C for PEF-treated(30kV/cm for240μs at25◦C initial temperature)orange juice.This PEF treatment resulted in a total aerobic plate count and yeast and mold count below the detection limit(<1CFU/mL).Y eom and others(2000b)achieved a112-d shelf life at4◦C for orange juice treated by PEF.The PEF treatment applied was 35kV/cm for59μs at25(inlet)to59◦C(outlet)temperature and the inactivation was comparable to a thermal treatment at 94.6◦C for30s.The PEF-treated orange juice maintained low levels of microorganisms(approximately1log10total plate count) indicating that the PEF treatment resulted in irreversible damage of cells.Min and others(2003)demonstrated that commercial-scale (500L/h)PEF treatment of orange juice at40kV/cm for97μs at45to65◦C(inlet compared with outlet temperature)reduced the total aerobic plate count and yeast and mold count by6log10 CFU/mL,achieved a shelf life of196d at4◦C.PEF was also successfully applied for refrigerated shelf-life ex-tension of apple cider,cranberry juice,and fruit smoothies.For example,Evrendilek and others(1999)reported an increase of more than67d of microbial shelf life for apple cider stored at both 4◦C and22◦C following a PEF treatment at35kV/cm for94μs and a thermal treatment of60◦C for30s.Jin and Zhang(1999) demonstrated more than4-log10inactivation of total aerobic bac-teria,yeasts,and molds in cranberry juice with a PEF treatment of 35kV/cm for195μs(at15to25◦C T max).The spoilage microor-ganisms were inhibited and the treated and aseptically packaged cranberry juice had a shelf life of8mo,37d and30d at4,22and 37◦C,respectively.A combination of mild heat(55◦C for60s)and PEF treatment (34kV/cm for60μs at55◦C[outlet])successfully extended the refrigerated(4◦C)shelf life of a tropical fruit juice smoothie (50%pineapple,28%banana,12%apple,7%coconut,3%orange juice)to21d(W alkling-Ribeiro and others2010).This treatment reduced the total plate count of the smoothie by approximately 4.4-log10which was similar to microbial reductions found after15 s thermal processing at72◦C.Interestingly,the thermally treated fruit smoothie exhibited only14d refrigerated shelf life and had similar sensory quality to the smoothie treated under combined mild heat and PEF.Lactic acid bacteria are the predominant microorganisms,in spoiled orange juice and L.plantarum is one of the most common species.G´o mez and others(2005)developed a model describing PEF inactivation of L.plantarum in McIlvaine buffer of varying pH(3.5to7.0)and tested the validity for PEF-treated orange and apple juices in a batch PEF system.L.plantarum was more sensitive to PEF at higher electricfield strengths and in media of low pH. For example,treatment at22kV/cm for400μs(temperature was kept at<35◦C)inactivated around5.0,2.8,2.6,and1.1-log10 CFU/mL of L.plantarum at pH3.5,5.0,6.5,and7.0,respectively.A W eibull-type model accurately described upward concave sur-vival curves of L.plantarum.The model satisfactorily predicted the inactivation of L.plantarum in apple and orange juices but is not suitable for commercial application of PEF processing due to the relatively long treatment time(400μs)used.A compara-tive study,which included a range of bacterial species,indicatedC 2013CSIRO and DIL Vol.12,2013r Comprehensive Reviews in Food Science and Food Safety459。