Effects of organic matter on acidic electrolysed water for reduction of foodborne pathogens on lettuce and spinach

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1 Journal of Applied Microbiology ISSN 3-57 ORIGINAL ARTICLE Effects of organic matter on acidic electrolysed water for reduction of foodborne pathogens on lettuce and spinach E.-J. Park, E. Alexander, G.A. Taylor, R. Costa and D.-H. Kang Department of Food Science and Human Nutrition, Washington State University, Pullman, WA, USA Proton Labs, Inc., Alameda, CA, USA Keywords bovine serum, electrolysed water, lettuce, organic matter, pathogen, spinach. Correspondence Dong-Hyun Kang, Department of Food Science and Human Nutrition, Washington State University, Pullman, WA 99-37, USA : received 5 December 7, revised April and accepted 3 April doi:./j x Abstract Aims: To evaluate the efficacy of acidic electrolysed water (EW) in the presence of organic matter (bovine serum) on the inoculated surfaces of lettuce and spinach. Materials and Results: Lettuce and spinach leaves were inoculated with a cocktail of three strains each of Escherichia coli O57:H7, Salmonella Typhimurium and Listeria monocytogenes and treated with deionized water, acidic EW and acidic EW containing bovine serum (5,, 5 and ml l ) ) for 5 s, 3 s, min, 3 min and 5 min at room temperature ( ± C). In the absence of bovine serum, acidic EW treatment reduced levels of cells below the detection limit (Æ7 log) in 5 min. In the presence of bovine serum, bactericidal activity of acidic EW decreased with increasing serum concentration. Conclusions: Organic matter reduces the effectiveness of acidic EW for reducing pathogens on the surfaces of lettuce and spinach. Significance and Impact of the Study: From a practical standpoint, organic matter reduces the efficacy of acidic EW. This study was conducted to confirm the effect of organic matter on the properties of acidic EW in the inactivation of foodborne pathogens on the surface of vegetables. Introduction Consumption of minimally processed and fresh-cut vegetables is on the rise in quantity and variety in recent years because of their convenience and importance. However, raw fruits and vegetables can be contaminated by pathogenic bacteria through wash water, rodents, or insects in the production plant, shredders or slicers with poor sanitization, and infected workers (Beuchat and Brackett 99; Lee and Kang ). Furthermore, pathogenic bacteria can grow on fresh fruits and vegetables during transportation and storage (Beuchat and Brackett 99). These organisms have been implicated in outbreaks of foodborne illnesses and recalls (IFPA ; Beuchat ; FDA News a,b). Escherichia coli O57:H7, Salmonella spp., Listeria monocytogenes are major foodborne pathogens linked to consumption of contaminated fresh vegetables worldwide (Beuchat 99). A variety of sanitizers, including chlorine, chlorine dioxide, ozone, hydrogen peroxide, UV light, calcinated water, organic acids and acidic electrolysed water, have been evaluated for their ability to reduce levels of pathogenic micro-organisms on fresh produce (Zhang and Farber 99; Bari et al. 999; Kim et al. a). Chlorinated water (5 ppm) is widely used to reduce levels of micro-organisms (Beuchat 99). However, most sanitizers showed a minimal microbial reduction of less log CFU g ) on the inoculated fresh fruits and vegetables, similar to that of chlorinated water (Beuchat 999; Taormina and Beuchat 999). A problem with chlorine treatment is that the efficacy of chlorinated water is reduced in the presence of organic materials (Gelinas and Goulet 93). Electrolysed water (EW) is generated from electrolysis of a Æ% NaCl solution through an electrolysing chamber. Using a two-cell chamber containing anode and cathode, acidic electrolysed water (acidic EW) and alkaline ª The Authors Journal compilation ª The Society for Applied Microbiology, Journal of Applied Microbiology 5 () 9

2 E.-J. Park et al. Effects of organic matter on acidic electrolysed water electrolysed water (alkaline EW) is produced in anode and cathode compartments, respectively. Acidic EW has a low ph (<Æ5) and a strong oxidizing potential (approx. mv). Conversely, alkaline EW has a high ph (>Æ) and a strong reducing potential (approx. mv) (Anon 997). Acidic EW is reported to have strong bactericidal activity against most pathogenic microorganisms (Izumi 999; Kim et al. a). The decontaminative effects of acidic EW on the surfaces of lettuce, tomatoes, strawberries, cucumbers and spinach have been reported (Koseki et al., ; Bari et al. 3; Guentzel et al. ). Acidic EW effectively inactivated E. coli O57:H7, Salmonella, L. monocytogenes and Bacillus cereus on the surfaces of vegetables (Venkitanarayanan et al. 999; Kim et al. b; Park et al. ). In general, aerobic bacteria grow in a ph range of 9, and at an ORP range of + to mv. Low ph may reduce the intensity of the bacterial outer membrane and allow entry of chlorine compounds through the inner membrane. Some mechanisms of bactericidal activity of chlorine compounds that have been proposed include killing the bacteria through glucose oxidation, disruption of protein synthesis, or destruction of a key enzyme (Huang et al. ). Liao et al. (7) proposed a bactericidal theory based on the high oxidation potential of acidic EW causing damage to cell membranes. Therefore, the bactericidal activities of acidic EW may be dependent on the kinds of bacterial species and can be seen by using a scanning electron microscope (Liao et al. 7). Wrinkled cell membranes are not an easy target for attack by acidic EW compared to smooth ones (Osafune et al. ). Park et al. () demonstrated that acidic EW is very effective for inactivating and killing E. coli O57:H7 and L. monocytogenes. Several factors such as the variety vegetable surfaces, kinds of contaminating micro-organisms, and soils present on vegetables, affect the inactivating effect of sanitizers on pathogens during food processing. Organic matter reduces the efficacy of sanitizers on pathogens in fresh produce. The presence of organic matter reduced the bactericidal effect of acidic EW. Nutrient broth, proteose peptone, glycine, glucose, sucrose, corn oil and chicken serum react with free available chlorine, reducing the effect of acidic EW (Oomori et al. ; Ayebah et al. 5). Protein matter reacts with chlorine to form organochloramine and could reduce the oxidizing effects of acidic EW (White 999). Since there are currently no reports on the efficacy of acidic EW against foodborne pathogens on the surfaces of fresh vegetables in the presence of organic matter, it must be established in order to optimize sanitization procedures. In practical usage, the effect of organic matter on the sanitizing effect of acidic EW is the challenge to the food processing industry. Other researchers reported that proteins such as proteose peptone and chicken serum reduced the effectiveness of acidic EW (Oomori et al. ; Ayebah et al. 5). Therefore, bovine serum was chosen to simulate organic matter contamination of produce since it can be easily controlled and quantified, whereas natural soiling would be difficult to replicate and quantify under laboratory conditions. In the present study we examined the effects of organic matter, such as bovine serum, on the properties of acidic EW and efficacy of acidic EW in the inactivation of foodborne pathogens such as E. coli O57:H7, S. Typhimurium, and L. monocytogenes on the surfaces of lettuce and spinach. Materials and methods Bacterial strains Three strains each of E. coli O57:H7 (ATCC 355, ATCC 39 and ATCC 39), S. Typhimurium (ATCC 955, ATCC 33 and ATCC ), and L. monocytogenes (ATCC 93, ATCC 9 and ATCC 7) were obtained from the Food Science and Human Nutrition culture collection at Washington State University (Pullman, WA, USA). These strains are of both human and veterinary clinical origin. All strains of E. coli O57:H7, S. Typhimurium or L. monocytogenes were grown in 9 ml Tryptic Soy Broth (TSB; Difco, Chicago, IL, USA) at 37 C for h, collected by centrifugation at g for 3 min at C, washed three times with 5 ml of Buffered Peptone Water (BPW; Difco), and resuspended to the original volume in 5 ml BPW (Difco), corresponding to approx. 9 CFU ml l ). To inoculate lettuce and spinach, all pathogen strains were combined to construct culture cocktails and maintained at ± C. These culture cocktails were used in subsequent experiments within h of preparation. Preparation of inoculated sample Iceberg lettuce and baby spinach were purchased at a local supermarket (Pullman, WA, USA). Lettuce and spinach leaves were trimmed to g, washed for min with deionized water, drained, separated and placed on sterile aluminium foil in a laminar flow biosafety hood. For inoculation, Æ ml of each pathogen cocktail was applied as droplets onto leaf surfaces with a micropipettor. Inoculated leaves were air dried in the hood for 3 h with the fan running for bacteria to attach to leaf surfaces. To determine the initial number of the three pathogens inoculated onto surfaces of lettuce and spinach (Æ ml), cell concentrations of culture cocktails were enumerated by conventional plating on selective media. ª The Authors Journal compilation ª The Society for Applied Microbiology, Journal of Applied Microbiology 5 () 9 3

3 Effects of organic matter on acidic electrolysed water E.-J. Park et al. Preparation of acidic EW Acidic EW was generated using a Super Oxide Series P electrolysed water generator (Proton Lab., Portland, OR, USA). This apparatus generates electrolysed water by the electrolysis of a Æ% sodium chloride solution for 5 min. The initial available chlorine content and ph of the mixture were determined by chlorine test kit (Bio-Lab Co., GA, USA) and a Corning Instruments ph meter (Corning, NY, USA), respectively. Acidic EW was prepared on the day of experiments and used within h of production. The temperature of acidic EW was ± C for the entire experiment. Effect of organic matter on the ph and available free chlorine content of acidic EW Different volumes (,,,,, 5 and ml l ) ) of sterile bovine serum ( mg ml l ) protein, Sigma, St Louis, MO, USA) were added to acidic EW. These mixtures were stirred 5 min using a Corning Instruments magnetic stirrer (Corning, NY, USA). The residual available free chlorine content and ph of the mixture were measured using previously described methods. Treatment of E. coli with acidic EW in the presence of organic matter Different volumes (,,, and ml l ) ) of sterile filtered bovine serum (Sigma) were added to acidic EW, and mixed for 5 min as described previously. A volume of Æ ml of a three strain E. coli culture cocktail ( 9 CFU ml ) ) was added to 9Æ9 ml of deionized water (DW, control), acidic EW and acidic EW containing different concentrations bovine serum for 3 s, min, 3 min and 5 min. Samples ( ml) were taken at treatment time intervals, neutralized with 9 ml of D E broth (Difco). The neutralized mixture was serially diluted in 9 ml of sterile buffered peptone water (Difco), and Æ ml of sample or diluent was spread-plated onto each selective medium. All tests were performed at room temperature ( ± C). Treatment of inoculated lettuce or spinach with acidic EW in the presence of organic matter Inoculated lettuce or spinach leaves were immersed in 5 ml of DW, acidic EW, and acidic EW with different concentrations (5,, 5 and ml l ) ) of sterile filtered bovine serum, and agitated for <s (momentary dispersal), followed by stationary exposure in the treatment solutions at one-minute intervals for 5 s, 3 s, min, 3 min and 5 min. At the selected time intervals, lettuce or spinach leaves ( g) were removed from treatment solutions and immediately placed in a stomacher bag containing 5 ml of neutralizing D E broth (Difco) and homogenized for min with a Seward stomacher ( Circulator, Seward, London, UK). After homogenization, ml of neutralized mixture was serially diluted in 9 ml of sterile buffered peptone water (Difco), and Æ ml of sample or diluent was spread-plated onto each selective medium. All tests were performed at room temperature ( ± C). Microbiological analysis Escherichia coli O57:H7, S. Typhimurium, and L. monocytogenes were enumerated on Sorbitol MacConkey agar (SMAC; Difco), Xylose Lysine Desoxycholate agar (XLD; Difco), and modified Oxford Agar Base (OAB; Difco) with antimicrobic supplement (Bacto Ô Oxford antimicrobic supplement, Difco), respectively. Where low levels of surviving cells were anticipated, ml of undiluted aliquots was equally distributed between four plates of each selective medium and spread-plated. All inoculated enumeration media were incubated at 37 C for h, and then the presence of typical colonies were enumerated. Statistical analysis Three replicate trials for each experiment were performed. The data were analysed by analysis of variance using the anova procedure of sas (SAS Institute, Cary, NC, USA). Means were separated using Duncan s multiple range tests. Significant differences between mean values are presented at a level of P =Æ5. Results The initial ph and free available chlorine concentration of acidic EW were Æ and 37Æ5 ±Æ5 mgl ), respectively. The detection limit for free chlorine analysis was 5mgl ). Fig. shows the changes of ph and free available chlorine of acidic EW mixed with different volumes of bovine serum. As the bovine serum concentration increased from to ml l ), the free available chlorine disappeared from 35 to mg l ). The addition of bovine serum up to ml l ) did not significantly affect the ph of acidic EW. The population of E. coli O57:H7 was reduced to below the detection limit (Æ7 log) with acidic EW treatment in the presence of bovine serum from to ml l ) after 5-s treatment. With bovine serum at a concentration of and ml l ) in acidic EW, the survival of E. coli ª The Authors Journal compilation ª The Society for Applied Microbiology, Journal of Applied Microbiology 5 () 9

4 E.-J. Park et al. Effects of organic matter on acidic electrolysed water ph Bovine serum concentration (ml l ) 5 Figure The ph and free available chlorine content of acidic electrolyse water (acidic EW) containing varying levels of bovine serum.d, ph; m, free available chlorine content. 3 Chlorine concentration (mg l ) Log CFU g Figure 3 Survival curves for Escherichia coli O57:H7 on lettuce concentrations of bovine serum. The error bars indicate 95% confidence intervals. Values below detection limit (<Æ7 log CFU g ) ) not shown. h, water;, acidic EW; s, acidic EW with 5 ml l ) serum; d, acidic EW with ml l ) serum;, acidic EW with 5 ml l ) serum;, acidic EW with ml l ) serum. Log CFU g 7 Log CFU g 3 5 Figure Survival curves for Escherichia coli O57:H7 exposed to acidic electrolysed water (acidic EW) with different concentrations of bovine serum. h, acidic EW with ml l ) serum; s, acidic EW with mll ) serum; d, acidic EW with ml l ) serum;, acidic EW with mll ) serum water;, acidic EW with ml l ) serum; h, acidic EW with 5 ml l ) serum;, acidic EW with ml l ) serum. O57:H7 after treatment generally increased (Fig. ). As bovine serum concentration increased from up to ml l ), there were no significant reductions in the population of E. coli O57:H7 after treatment with acidic EW. Reductions calculated are values exceeding those of the water control treatment observed at each exposure time. Shown in Figs 3 5 are the surviving cells of E. coli O57:H7, S. Typhimurium, and L. monocytogenes from Figure Survival curves for Salmonella Typhimurium on lettuce concentrations of bovine serum. The error bars indicate 95% confidence intervals. Values below detection limit (<Æ7 log CFU g ) ) not shown. h, water;, acidic EW; s, acidic EW with 5 ml l ) serum; d, acidic EW with ml l ) serum;, acidic EW with 5 ml l ) serum;, acidic EW with ml l ) serum. lettuce leaves treated with DW, acidic EW, and acidic EW with various concentration of bovine serum (5,, 5 and ml l ) ), respectively. Treatment with only acidic ª The Authors Journal compilation ª The Society for Applied Microbiology, Journal of Applied Microbiology 5 () 9 5

5 Effects of organic matter on acidic electrolysed water E.-J. Park et al. Log CFU g Figure 5 Survival curves for Listeria monocytogenes on lettuce leaves exposed to acidic electrolysed water (acidic EW) with different concentrations of bovine serum. The error bars indicate 95% confidence intervals. Values below detection limit (<Æ7 log CFU g ) ) not shown. h, water;, acidic EW; s, acidic EW with 5 ml l ) serum; d, acidic EW with ml l ) serum;, acidic EW with 5 ml l ) serum;, acidic EW with ml l ) serum. EW reduced levels of E. coli O57:H7 on lettuce to below the detection limit (Æ7 log) after 3 min (Fig. 3). The levels of surviving cells treated with acidic EW containing serum at 5 ml l ) were significantly reduced by Æ7, Æ7, Æ, Æ9 and 3Æ log CFU g ) after 5 s, 3 s, min, 3 min and 5 min treatment (P <Æ5), respectively, compared to the deionized water control. At the ml l ) containing serum, the numbers of E. coli O57:H7 reduced by Æ, Æ59, Æ, Æ and 3Æ7 log CFU g ) at the selected time intervals, respectively (P < Æ5). However, there were no significant differences between levels of E. coli O57:H7 cells treated with acidic EW containing serum at 5 and ml l ) compared with the water control (DW). For S. Typhimurium (Fig. ), there were no significant differences between treatments with DW and acidic EW containing serum at, 5 and ml l ) at all treatment times (P >Æ5). However, acidic EW treatment alone showed significant reduction of Æ53 log CFU g ) after 3 s and more than Æ7 log CFU g ) (detection limit Æ7 log) after 3 min treatment (P < Æ5). At 5 ml l ) concentration, levels of S. Typhimurium were significantly reduced after 5 s, 3 s, and min; however, there was no significant difference compared with DW after 3 min. Fig. 5 shows surviving cells of L. monocytogenes enumerated on OAB agar after treatment with test solutions. The number of L. monocytogenes cells experienced significant reductions of Æ9, 3Æ9, Æ and Æ9 log CFU g ) after 5 s, 3 s, min, and 3 min of treatment with acidic EW alone, respectively (P <Æ5). After 5 min of treatment, levels of cells were reduced to below the detection limit (Æ7 log). At all treatment solution containing different volume of bovine serum, however, there was no statistically significant difference up to 3 min treatment (P > Æ5). After 5 min, the surviving cells of L. monocytogenes were significantly reduced by 3Æ log CFU g ) at5mll ) serum containing acidic EW compared with DW (P < Æ5). The surviving cells of E. coli O57:H7, S. Typhimurium, and L. monocytogenes after exposure to DW, acidic EW, and acidic EW containing bovine serum on spinach leaves are shown in Figs., 7 and, respectively. Figure shows the population of E. coli O57:H7 after 5 s, 3 s, min, 3 min and 5 min treatment with selected solutions. In the absence of serum, the numbers of cells were reduced to below the detection limit (Æ7 log) at 3 min exposure. With serum at a concentration of 5 ml l ), Æ7, Æ5, Æ9, 3Æ and Æ log CFU g ) significant reductions of E. coli O57:H7 were detected after 5 s, 3 s, min, 3 min and 5 min treatment, respectively (P < Æ5). As the serum concentration increased to ml l ), the survival of E. coli O57:H7 after treatment with acidic EW containing ml l ) serum was reduced by Æ7, Æ, Æ9, 3Æ and 3Æ9 log CFU g ) after each exposure time, respectively. There were significant differences in numbers of surviving cells between DW and acidic EW containing ml l ) serum after 3 min treatment (P <Æ5). The populations of E. coli O57:H7 after treatment with acidic Log CFU g Figure Survival curves for Escherichia coli O57:H7 on spinach concentrations of bovine serum. The error bars indicate 95% confidence intervals. Values below detection limit (<Æ7 log CFU g ) ) not shown. h, water;, acidic EW; s, acidic EW with 5 ml l ) serum; d, acidic EW with ml l ) serum;, acidic EW with 5 ml l ) serum;, acidic EW with ml l ) serum. ª The Authors Journal compilation ª The Society for Applied Microbiology, Journal of Applied Microbiology 5 () 9

6 E.-J. Park et al. Effects of organic matter on acidic electrolysed water Log CFU g Log CFU g Figure 7 Survival curves for Salmonella Typhimurium on spinach concentrations of bovine serum. The error bars indicate 95% confidence intervals. Values below detection limit (<Æ7 log CFU g ) ) not shown. h, water;, acidic EW; s, acidic EW with 5 ml l ) serum; d, acidic EW with ml ) serum;, acidic EW with 5 ml l ) serum;, acidic EW with ml l ) serum. 3 5 Figure Survival curves for Listeria monocytogenes on spinach concentrations of bovine serum. The error bars indicate 95% confidence intervals. Values below detection limit (<Æ7 log CFU g ) ) not shown. h, water;, acidic EW; s, acidic EW with 5 ml l ) serum; d, acidic EW with ml l ) serum;, acidic EW with 5 ml l ) serum;, acidic EW with ml l ) serum. EW in the presence of 5 and ml l ) serum showed no significant differences compared to DW. Fig. 7 shows levels of surviving cells of S. Typhimurium after treatment with solutions. Acidic EW treatment alone reduced microbial levels by Æ and Æ log CFU g ) after 5 s and 3 s, respectively. And after min, this pathogen was reduced to below the detection limit (Æ7 log). The survival of S. Typhimurium after treatments with acidic EW containing bovine serum is a result lowered bactericidal activity. No significant reductions were observed in acidic EW containing 5 and ml l ) serum compared with DW at all exposure times. L. monocytogenes (Fig. ) experienced significant reductions of Æ, Æ3 and 5Æ log CFU g ) after 5 s, 3 s and min of treatment, respectively, with acidic EW in the absence of serum (P <Æ5). After treatment with acidic EW containing serum at 5 ml l ), levels of L. monocytogenes were significantly reduced by Æ, Æ5, 3Æ and Æ log CFU g ) after 3 s, min, 3 min and 5 min, respectively (P <Æ5), compared to the deionized water control. However, no significant differences in survival were detected with acidic EW amended with serum at, 5 and ml l ) (P >Æ5). Discussion In this study, we evaluated the effects of acidic EW in reducing foodborne pathogens in the presence of organic matter. No significant reductions (< log) occurred when inoculated lettuce and spinach were subjected to the water control treatment. This may be the result of physical removal of bacterial cells from vegetable surfaces. Investigations of the bactericidal activity of acidic EW have been conducted for lettuce, tomatoes, cucumbers, and strawberries (Koseki et al. ; Park et al. ; Bari et al. 3). However, there were differences in removal of surface micro-organisms depending on the vegetable studied. Because lettuce and tomatoes have relatively smoother surfaces than cucumbers and strawberries, acidic EW may more easily contact surface micro-organisms on those vegetables (Koseki et al. ). In this study, there were no significant difference in acidic EW activity between lettuce and spinach. Perhaps the smoother surface of spinach facilitated better contact with acidic EW. The bactericidal effects of acidic EW on foodborne pathogens are well documented by numerous studies. Koseki et al. (3) investigated the effect of spot inoculated lettuce leaves treated with acidic EW containing ppm free chlorine for 3 min. They observed an approximately Æ and Æ log CFU g ) reduction of E. coli O57:H7 and S. Typhimurium, respectively. Ayebah et al. (5) showed that acidic EW produced a > log CFU ml ) reduction in L. monocytogenes after a min treatment. These results show that acidic EW is effective at killing pathogens on vegetables and resulted in above Æ log CFU g ) reduction for min treatment. However, there are several studies that showed that acidic EW was less effective at killing pathogens on vegetables than previous studies. Park et al. () demonstrated that a min treatment of ª The Authors Journal compilation ª The Society for Applied Microbiology, Journal of Applied Microbiology 5 () 9 7

7 Effects of organic matter on acidic electrolysed water E.-J. Park et al. acidic EW containing 5 ppm free chlorine significantly reduced E. coli O57:H7 and L. monocytogenes by Æ and Æ log CFU g ) on lettuce, respectively. Izumi (999) reported that immersing lettuce in acidic EW containing 5 ppm free chlorine for min results in reduction of Æ7 and Æ3 log CFU g ) of E. coli O57:H7 and L. monocytogenes, respectively. In practical usage, however, acidic EW generally is used in the presence of organic matter, such as soils or vegetable debris. In our study, treatment with acidic EW containing bovine serum as a form of organic matter resulted in a lower bactericidal effect on foodborne pathogens, including E. coli O57:H7, S. Typhimurium, and L. monocytogenes. In the absence of serum, acidic EW completely inactivated all pathogens within 5 min. As the serum concentration increased, the free available chlorine content decreased rapidly. The effect of organic matter on chlorinated water has been demonstrated by several researchers. White (999) demonstrated that proteins can react with chlorine and results in formation of organichloramines. The neutralization of chlorine in acidic EW may due to the addition of organic matter (Ayebah et al. 5). El-Kest and Marth (9) reported that as organic matter present increased, free available chlorine content decreased. Tanaka et al. (99) reported that organic materials convert free available chlorine in acidic EW to the combined form. Also, Oomori et al. () concluded that the bactericidal activity of acidic EW declines in the presence of organic materials, such as amino acids and proteins. In their study, materials containing amino acids or proteins were quickly transformed from free available chlorine into N-chloro compounds (combined available chlorine). They also observed that free available chlorine in acidic EW was removed through the reaction of oxidation reduction with corn oil, vitamins, minerals or lipids. Ayebah et al. (5) studied that the effect of acidic EW on the inactivation of planktonic cells and biofilms of L. monocytogenes in the presence of chicken serum. They reported that acidic EW treatment with 5 and ml l ) serum for 5 min reduced the population of L. monocytogenes by only Æ and Æ log CFU g ) ml ), respectively, compared to deionized water. Also, the populations of L. monocytogenes biofilms exposed to acidic EW alone and acidic EW with increasing serum concentration up to 7Æ5 mll ) for s experienced reductions of Æ37 and Æ95 log CFU g ), respectively. The results of our study show that acidic EW is an effective and convenient sanitizer for reduction of foodborne pathogens. However, several factors affect the bactericidal activity of acidic EW, including the type of food product and degree of organic matter contamination. Thus, for effective usage in the food processing industry, it is suggested that a pretreatment or preremoval step of organic debris, before use of sanitizers, is need to improve their activity. References Anon (997) Electrolyzed Water. Central Laboratory Report. Toyoake, Aichi, Japan: Hoshizaki Electric Co.. Ayebah, B., Hung, Y.C. and Frank, J.F. (5) Enhancing the bactericidal effect of electrolyzed water on Listeria monocytogenes biofilms formed on stainless steel. J Food Prot, Bari, M.L., Kusunoki, H., Furukawa, H., Ikeda, H., Isshiki, K. and Uemura, T. (999) Inhibition of growth of Escherichia coli O57:H7 in fresh radish (Raphanus sativas L.) sprout production by calcinated calcium. J Food Prot, 3. Bari, M.L., Sabina, Y., Isobe, S., Uemera, T. and Isshiki, K. 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