Efficacy of lactic acid in reducing foodborne pathogens in minimally processed lotus sprouts

Food Control 30 (2013) 721e726

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Efficacy of lactic acid in reducing foodborne pathogens in minimally processed lotus sprouts Chenjie Wang a, Shuilian Wang a, Tong Chang a, Liu Shi a, Hong Yang a, *, Yanchun Shao a, Wu Feng a, Min Cui b a b

College of Food Science and Technology, Huazhong Agricultural University, Wuhan, Hubei 430070, China College of Animal Sciences and Technology, Huazhong Agricultural University, Wuhan, Hubei 430070, China

a r t i c l e i n f o

a b s t r a c t

Article history: Received 27 April 2012 Received in revised form 18 August 2012 Accepted 28 August 2012

Fresh lotus sprout is easily browned and perishable due to microbial growth and degradation. Therefore, browning and foodborne pathogen have become the most serious problems. The objective of this study was to investigate the effect of lactic acid (0.25%, 0.5%, 1% and 2% (v/v)) on reduction of foodborne pathogens, such as Escherichia coli O157:H7, Salmonella Typhimurium, and Listeria monocytogenes, in lotus sprouts. Tap water and sodium hypochlorite (200 mg/l) were used as control treatments. Results indicated that tap water caused a slight reduction (<0.5 log) in the microbial loads. The sodium hypochlorite treatment led to 1.3 log reductions of the microbial population. When treated with 0.5% and 2.0% lactic acid solutions, 1.5 and 2.3 log reductions were achieved, respectively. The effectiveness of lactic acid treatment increased with the increase of lactic acid concentration. Results showed that the lactic acid treatment at 0.5% or higher was effective to reduce foodborne pathogens in lotus sprouts. The L* values of samples treated with lactic acid decreased slightly during storage. Furthermore, the lactic acid treatment contributed to slow accumulation of red color on lotus sprouts, which was more effective than sodium hypochlorite treatment to reduce the discoloration of lotus sprouts. These results indicated that lactic acid can be used to improve the color and safety of minimally processed lotus sprouts. Ó 2012 Elsevier Ltd. All rights reserved.

Keywords: Foodborne pathogen Escherichia coli O157:H7 Salmonella Typhimurium Listeria monocytogenes Lactic acid Lotus sprouts Anti-microbial

1. Introduction China is the largest producer of Lotus (Nelumbo nucifera gaertn) in the world. Fresh lotus sprout is a seasonal vegetable available only from April to September in China. It has excellent flavor and is considered as a highly sought-after culinary item. However, with a high water content (>90% in weight), the fresh lotus sprout tends to become brown and perishable due to metabolism, physical damage and microbial growth. Therefore, minimally processed lotus sprouts could be a good choice to prolong its shelf-life to meet ever-increasing demands of consumers for high quality, fresh, nutritious, and conveniently prepared vegetables (Artés, Gómez, Aguayo, Escalona, & Artés-Hernández, 2009). However, there has been an increase in the frequency of foodborne disease outbreaks associated with consumption of fresh produce. Vegetables contain nutrients necessary for rapid growth of foodborne pathogens such as Escherichia coli, Salmonella, Listeria and Yersinia (Alegre, Abadias, Anguera, Usall, & Viñas, 2010; Issa-

* Corresponding author. Tel.: þ86 27 87671045; fax: þ86 27 87288373. E-mail address: [email protected] (H. Yang). 0956-7135/$ e see front matter Ó 2012 Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.foodcont.2012.08.024

Zacharia, Kamitani, Morita, & Iwasaki, 2010; Issa-Zacharia, Kamitani, Tiisekwa, Morita, & Iwasaki, 2010; Ruiz-Cruz, AcedoFélix, Díaz-Cinco, Islas-Osuna, & González-Aguilar, 2007). E. coli O157:H7 is a major foodborne pathogen that can cause illnesses, such as hemorrhagic colitis and hemolytic uremic syndrome (Ibrahim, Salameh, Phetsomphou, Yang, & Seo, 2006; Smigic et al., 2009; Velazquez, Barbini, Escudero, Estrada, & de Guzman, 2009). Listeria monocytogenes has caused much concern to the food industry, since it has been associated with large-scale outbreaks (Byelashov et al., 2010). Different washing chemicals have been investigated to determine their efficacy in the inactivation of pathogens on vegetables (Oms-Oliu et al., 2010; Velazquez et al., 2009). Among them, solutions of 50e200 ppm chlorine (sodium hypochlorite) are widely used at commercial scale for washing fresh cut produce (Beuchat, Nail, Adler, & Clavero, 1998; Ruiz-Cruz et al., 2007). However, the efficacy of sodium hypochlorite is limited due to the formation of toxic by-products in the presence of organic matter as a result of chlorination (Abadias, Alegre, Usall, Torres, & Viñas, 2011; Beuchat, 2000; López-Gálvez et al., 2010). Currently, consumers are paying more attention not only to the risk of foodborne pathogens but also to the safety of artificial chemical preservatives that are used to

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control these foodborne pathogens (Ibrahim, Yang, & Seo, 2008; López-Gálvez et al., 2010). The effect of organic acids on reducing populations of microorganisms on fresh vegetables has been explored, while lactic acid with different concentrations (from 0.2% to 2%) has shown promising results against common pathogens such as E. coli O157:H7, S. Typhimurium, L. monocytogenes and Yersinia enterocolitica (Akbas & Olmez, 2007; Huang & Chen, 2011; Ibrahim et al., 2008; Sagong et al., 2011; Tajkarimi & Ibrahim, 2011; Uyttendaele, Neyts, Vanderswalmen, Notebaert, & Debevere, 2004; Velazquez et al., 2009). The inhibiting effect of weak organic acid is based on their pKa, antimicrobial activity of their non-dissociated form, and specific effects of each acid (Giannuzzi & Noem, 1996; Velazquez et al., 2009). Up to now, most research on lotus sprout has been directed to prevent its browning and softening (Du, Fu, & Wang, 2009; Li et al., 2010). Little information is available regarding the microbiological quality. Effective methods for assuring the microbiological safety of minimally processed lotus sprout have not been extensively explored. Therefore, the main objective of this study was to assess the effectiveness of lactic acid on E. coli O157:H7, S. Typhimurium, and L. monocytogenes inoculated on minimally processed lotus sprouts, and compare its efficacy with tap water and sodium hypochlorite solution. In addition, the color of minimally processed lotus sprout was determined to evaluate the antibrowning effect of lactic acid on lotus sprouts during storage at 4  C. 2. Materials and methods 2.1. Bacterial strains The bacterial cultures of E. coli O157:H7, S. Typhimurium and L. monocytogenes strains were obtained from the Food Microbiology Laboratory, College of Food Science and Technology, Huazhong Agricultural University (Wuhan, China). The cultures were preserved at 25  C with 50% (v/v) glycerin (Sinopharm Chemical Reagent Co., Ltd, Shanghai, China) solution at a ratio of 1:1. 2.2. Inoculum preparation The E. coli O157:H7, S. Typhimurium and L. monocytogenes suspension were prepared by transferring 0.1 ml of each culture to a 10 ml of 0.1% peptone water (PW, Qingdao Hope Bio-Technology Co., Ltd, Qingdao, China) and each sample was incubated for 24 h at 37  C (DNP-9272E, Shanghai Jing Hong laboratory Instrument Co., Ltd, Shanghai, China) and transferred at 24 h intervals. Bacterial cells were obtained by centrifugation (J-E, BACKMAN, California, USA) at 9100 g for 10 min at 4  C. The supernatant of each bacterial suspension was decanted, while the cell pellet was resuspended in 100 ml of 8.5 g/l sterile physiological saline (Sanshen YM30F, Shanghai Sanshen Medical Instrument Co., Ltd, Shanghai, China). Three selective media were used to enumerate the bacteria. For E. coli O157:H7, Sorbitol-MacConkey agar supplemented with Cefixime-Tellurite. For S. Typhimurium and L. monocytogenes, Xylose Lysine Desoxycholate and Trypticase Soy-Yeast Extract Agar were used, respectively. All selective media were obtained from Qingdao Hope Bio-Technology Co., Ltd (Qingdao, China). The initial concentration was approximately109 cfu/ml. The population of each bacteria was confirmed by plating 0.1 ml portions of appropriately diluted suspension on duplicated plates (90 mm) for 24 h at 37  C. 2.3. Sample preparation Lotus (Nelumbo nucifera Gaertn cv.) sprouts were purchased from a local wholesale market (Wuhan, Hubei, China) on the day of

harvest and transported to the laboratory without delay. Prior to experimental studies, tap water was used to remove the impurities (such as mud and withered leaves) from the surface of lotus sprouts. All the samples were submerged in a big container with enough tap water and stored at room temperature over night before processing. In the second day, top and tail of lotus sprouts were removed and discarded while central parts were cut into 8e10 cm in length (10  0.2 g each) with a stainless steel knife previously disinfested with 70% (v/v) alcohol. 2.4. Sample inoculation Fresh-cut lotus sprouts (10  0.2 g each, about 8e10 cm in length) were submerged in a container with inocula of each microorganism and gently agitated for 5 min at room temperature to obtain an initial level of above 107 cfu/g. Samples were removed and then air-dried in a biosafety chamber (AIRTECH, SW-CJ-1FD, Jiangsu Suzhou Purification Group, Suzhou, China) for 30 min to facilitate bacterial adhesion before exposure to various disinfection treatments. Appropriate selective medium as mentioned above was used to determine the initial population of each pathogen inoculated onto the surface of samples. 2.5. Procedures for treatment In the study, all treatment solutions were prepared within 1 h before each experiment. Lactic acid (LA) (Shanghai Chemicals Reagent Co., Ltd, Shanghai, China) was dissolved in distilled water to obtain solutions containing 0.25%, 0.5%, 1% and 2% (v/v). Sodium hypochlorite (NaClO) (Sinopharm Chemical Reagent Co., Ltd, Shanghai, China) was used to prepare the solution at 200 mg/l of free chlorine. NaCl solution (8.5 g/l) was prepared to dilute each bacterial suspension in advance. As stated above, inoculated samples were individually placed in different washing treatments after removing excess water. During each experiment, two samples were assayed after each treatment for 10 min. Three untreated samples inoculated with E. coli O157:H7, L. monocytogenes and S. Typhimurium were used to determine the initial bacterial loads on the sample surface. Two untreated samples were immersed in tap water and sodium hypochlorite (200 mg/l) for 10 min as control treatments. All samples were gently agitated during the treatment period. After washing, each sample was transferred to a sterile stomacher blender bag (Qingdao Hope Bio-Technology Co., Ltd, Qingdao, China) to determine the microbial populations. For storage, samples were packaged after washing treatments by a vacuum packaging machine (0.08 MPa) (Jiesheng, DZQ 400-2D, Suzhou, China) and stored at 4  C (Haier Pharmacy Refrigerators, Qingdao, China) until microbiological analysis. 2.6. Bacterial enumeration For enumeration, 25 g of lotus sprout sample was transferred into sterile stomacher blender bags (Qingdao Hope Bio-Technology Co., Ltd, Qingdao, China), combined with 225 ml of sterile physiological saline (8.5 g/l NaCl), and shaken vigorously by hand for 1 min. The solution (1 ml) was serially diluted in test tubes containing 9 ml of sterile physiological saline. Then, properly diluted bacterial solutions were transferred to appropriate selective media and then poured to sterile petri dishes. Finally, the solidified plates were incubated for 24 h at 37  C (DNP-9272E, Shanghai Jing Hong Laboratory Instrument Co., Ltd, Shanghai, China), while colonies were counted following the incubation in the next day.

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The surface color of samples was measured with a HunterLab Ultra Scan XE colorimeter (HunterLab Co., Ltd, Reston, VA, USA). The L*, a* and b* values were measured, where L*, a* and b* indicates luminosity, chromaticity on a green () to red (þ) axis, and chromaticity on a blue () to yellow (þ) axis, respectively (Du et al., 2009). Measurement was carried out with 10 sites on the surface of each sample. The chroma meter was calibrated on a standard white plate before each series of measurements.

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All experiments were performed in triplicate on different treatments. Prior to ANOVA, averages of duplicate plate counts from three replications of each treatment were converted to log cfu/g. SAS software (SAS 9.0 for windows, SAS (Shanghai) Software Co., Ltd, Shanghai, China) was used for data analysis. Significant differences between treatments were analyzed by least significant difference (LSD) at a significance level of P < 0.05. The mean values as shown in this study were calculated as the mean  SD (n ¼ 3).

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3. Results and discussion

3.1. Effect of different LA concentration on bacteria Fig. 1 showed the surviving populations of E. coli O157:H7, S. Typhimurium and L. monocytogenes on lotus sprouts as a result of treating with tap water, NaClO (200 mg/l) and different LA concentrations (0.25%, 0.5%, 1% and 2%). Tap water and NaClO (200 mg/l) were used as control treatments. The initial populations of E. coli O157:H7, S. Typhimurium and L. monocytogenes inoculated on lotus sprouts samples were 7.30, 7.34 and 7.43 log cfu/g, respectively. Slight reductions (0.40, 0.25, 0.23 log, respectively) were achieved by washing inoculation samples with tap water. Similar results were observed on organic fresh lettuce (<0.5 log cfu/ g) (Sagong et al., 2011). The result was mainly due to physical removal of pathogen cells from the surfaces of lotus sprouts. The NaClO (200 mg/l) treatments reduced the three pathogen counts by 1.29, 1.30 and 1.15 log cfu/g, respectively, which were similar to the results observed in treating baby spinach (1.2e1.6 log cfu/g) (Huang & Chen, 2011) and fresh-cut carrots (1e2 log cfu/g) (Ruiz-Cruz et al., 2007). The efficacy of the wash treatments could be related to surface characteristics of lotus sprouts that were also noted on other vegetables (Velazquez et al., 2009). For three pathogens, there were significant differences between two control treatments (tap water and NaClO) (P < 0.05). As shown in Fig. 1, the reduction of pathogens increased with increasing LA concentration from 0.25% to 2.0%, while 2.0% LA was the most effective (P < 0.05). Except for tap water and the lowest concentration LA (0.25%) treatments, other treatments were effective to reduce three pathogens (E. coli O157:H7, S. Typhimurium and L. monocytogenes), while the reduction was approximately in the order of 2% LA > 1% LA > 0.5% LA > NaClO. There were significant differences in the effectiveness of different LA treatments against E. coli O157:H7 (P < 0.05) (Fig. 1a), while no significance was obtained between 0.5% LA and 1% LA in S. Typhimurium (Fig. 1b) and L. monocytogenes (Fig. 1c) (P > 0.05). Similar to this present study, it has been found that 2.0% LA showed significant

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inactivation on E. coli O157:H7, S. Typhimurium, and L. monocytogenes inoculated on fresh lettuce comparing with low concentrations (0.3%, 0.5%, 0.7% and 1%) (Sagong et al., 2011). Furthermore, 0.2% and 1% LA were effective to prevent the growth of E. coli O157:H7 and Y. enterocolitica inoculated onto lettuce and tomatoes (Velazquez et al., 2009), while 0.2% LA was used to inactivate E. coli O157:H7 in carrot juice (Ibrahim et al., 2008; Tajkarimi & Ibrahim, 2011). In the present study, LA (0.5%) was found to reduce E. coli O157:H7, S. Typhimurium and L. monocytogenes effectively. There was no significant difference with NaClO solution for commercial use (P > 0.05). Therefore, 0.5% LA was selected in the following study finally. 3.2. Comparison of LA sanitization potency with that of NaClO and tap water Fig. 2 showed the populations of E. coli O157:H7, S. Typhimurium and L. monocytogenes on minimally processed lotus sprouts before and after washing with tap water, NaClO (200 mg/l), LA (0.5% v/v) and its survival during storage at 4  C up to 8 days. The initial microbial loads were 8.05, 8.07 and 8.29 log cfu/g, respectively.

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significance in S. Typhimurium (P > 0.05) among Day 3 tap water (8.49 log), Day 8 NaClO (8.60 log) and Day 8 LA (8.76 log). LA (0.5%) was found to reduce three common pathogens effectively (Fig. 2). In washing solution, LA exists in a pH-dependent equilibrium between the undissociated and dissociated state. Lactic molecule diffuses into the cell until equilibrium is reached in accordance with the pH gradient across the membrane, resulting in the accumulation of anions and protons inside the cell and ultimate death of microbial cells (Brula & Coote, 1999). The diffusion of lactic molecules into microbial cells caused a series of interference including membrane disruption, resulting in leakage, disruption of outer membrane permeability, influence on macromolecular synthesis, stress on intracellular pH homeostasis, and accumulation of toxic anions (Ibrahim et al., 2008; Tajkarimi & Ibrahim, 2011; Velazquez

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Fig. 2. Survival and growth of E. coli O157:H7 (a), S. Typhimurium (b) and L. monocytogenes (c) on minimally processed lotus sprouts stored at 4  C. LA: lactic acid; BT: before treatments.

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Similar to previous study, slight reductions (0.32, 0.30, 0.33 log cfu/g on E. coli O157:H7, S. Typhimurium and L. monocytogenes, respectively) were obtained by washing inoculated samples with tap water, while significantly higher reduction levels (about 1.5 log cfu/g or even more) were obtained by treating with NaClO (200 mg/l) and LA (0.5%, v/v) (P < 0.05). NaClO and LA treatments showed better sanitizing effects than tap water treatments on three pathogens tested after washing and during storage (P < 0.05) (Fig. 2). However, in all treatments, three microbial loads increased during storage. There was no significant difference in microbial loads of lotus sprouts treated with 0.5% LA and NaClO (200 mg/l) during storage (P > 0.05), while the microbial load of lotus sprouts treated with tap water kept the highest (P < 0.05). Furthermore, the microbial loads of samples treated with NaClO and LA at Day 8 were close to that of tap water at Day 3, while the trend of three pathogens were similar. For example, there was no

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Fig. 3. Effects of different treatments on L* of minimally processed lotus sprouts treated with E. coli O157:H7 (a), S. Typhimurium (b) and L. monocytogenes (c) during storage at 4  C.

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et al., 2009; Virto, Sanz, Alvarez, & Condon, 2005). Consequently, LA has been found to have a strong inhibitory effect against gramnegative (Fooks & Gibson, 2002a, 2002b) and gram-positive bacteria (Markas & Vuyst, 2006). 3.3. Color evaluation To determine the antibrowning effect of LA on minimally processed lotus sprouts, changes in color parameters (L*, a* and b*) were evaluated during the storage (Figs. 3e5). Among three pathogens, all samples of LA treatments retarded enzymatic browning on minimally processed lotus sprouts during storage (Fig. 3). The L* values (lightness) in all treatments decreased during storage.

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However, it was indicated that LA treatments had significantly effective antibrowning (P < 0.05). It might be due to the low pH value of organic acid solution which can inactivate the polyphenol oxidase (PPO) activity of minimally processed lotus sprouts. A factor linking pH and a main quality marker, browning, is the activity of PPO, which catalyzes cut surface discoloration most effectively at a neutral pH of 7.0 approximately (Arslan & Dogan, 2005; Rico, Martin-Diana, Barat, & Barry-Ryan, 2007). PPO is widely distributed in plants and is rich in lotus. It can result in lotus sprouts browning easily and even lead produce not to be accepted by consumers (Arslan & Dogan, 2005; Du et al., 2009; Li et al., 2010). On the other hand, changes of the a* values of lotus sprouts during storage were different from those of the L* values (Figs. 3

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and 4). Results of different treatments decreased sharply during the first 2 days with significant differences (P < 0.05). Then, the a* values of lotus sprouts inoculated with L. monocytogenes increased gradually (Fig. 4c). There was a similar trend for E. coli O157:H7, S. Typhimurium when treated with LA and tap water (Fig. 4a and b). But, the a* values of NaClO treatments of E. coli O157:H7, S. Typhimurium changed not so regularly (Fig. 4a and b). However, there were no significant differences among three treatments (Fig. 4) after Day 2 (P > 0.05). Among all treatments, LA always exhibited the lowest a* values while tap water was the highest and NaClO was the middle level. The a* values of tap water treatment kept the highest value during storage, which had the least effective to reduce the discoloration of lotus sprouts (Du et al., 2009; Li et al., 2010). As a result, LA treatment contributed to the slowest accumulating red color on lotus sprouts, which was more effective than NaClO to improve the color of minimally processed lotus sprouts. Unlike the a* values, there was no considerable change in the b* values during storage. The yellowness values (b*) increased during storage in all treatments with three pathogens inoculation, but no significant difference (P > 0.05) was found between LA and NaClO treatments, which might be attributed to the nature of lotus sprout (Fig. 5).

4. Conclusion As a highly seasonable vegetable, fresh lotus sprout is susceptible to browning and is easily perishable. In this study, the LA concentration of 0.5% or higher was effective to reduce foodborne pathogens in lotus sprouts. When treated with 0.5% and 2.0% lactic acid,1.5 and 2.3 log reductions were achieved, respectively. The effectiveness of LA treatment on pathogens increased with the increase of its concentration. The L* values of LA treatment decreased slightly during storage. Furthermore, LA treatment contributed to slow accumulation of red color on lotus sprouts, which was more effective than NaClO treatment to improve the color of minimally processed lotus sprouts. Results indicated that LA treatment had significantly effective antibrowning on lotus sprout (P < 0.05). Acknowledgments The authors would like to thank for the financial support of the Start-up Research Fund from Huazhong Agricultural University. References Abadias, M., Alegre, I., Usall, J., Torres, R., & Viñas, L. (2011). Evaluation of alternative sanitizers to chlorine disinfection for reducing foodborne pathogens in freshcut apple. Postharvest Biology and Technology, 59, 289e297. Akbas, M. Y., & Olmez, H. (2007). Inactivation of Escherichia coli and Listeria monocytogenes on iceberg lettuce by dip wash treatments with organic acids. Letters in Applied Microbiology, 44(6), 619e624. Alegre, I., Abadias, M., Anguera, M., Usall, J., & Viñas, I. (2010). Fate of Escherichia coli O157:H7, Salmonella and Listeria innocua on minimally-processed peaches under different storage conditions. Food Microbiology, 27, 862e868. Arslan, O., & Dogan, S. (2005). Inhibition of polyphenol oxidase obtained from various sources by 2,3-diaminopropionic acid. Journal of the Science of Food and Agriculture, 85, 1499e1504. Artés, F., Gómez, P., Aguayo, E., Escalona, V., & Artés-Hernández, F. (2009). Sustainable sanitation techniques for keeping quality and safety of fresh-cut plant commodities. Postharvest Biology and Technology, 51, 287e296. Beuchat, L. R. (2000). Use of sanitizers in raw fruit and vegetable processing. In S. M. Alzamora, M. S. Tapia, & A. López-Malo (Eds.), Minimally processed fruits and vegetables (pp. 63e77). Beuchat, L. R., Nail, B. U., Adler, B. B., & Clavero, M. R. S. (1998). Efficacy of spray application of chlorinated water in killing pathogenic bacteria on raw apples, tomatoes and lettuce. Journal of Food Protection, 61, 1305e1311.

Brula, S., & Coote, P. (1999). Preservative agents in foods. Mode of action and microbial resistance mechanisms. International Journal of Food Microbiology, 50, 1e17. Byelashov, O. A., Daskalov, H., Geornaras, I., Kendall, P. A., Belk, K. E., Scanga, J. A., et al. (2010). Reduction of Listeria monocytogenes on frankfurters treated with lactic acid solutions of various temperatures. Food Microbiology, 27, 783e790. Du, J. H., Fu, Y. C., & Wang, N. Y. (2009). Effects of aqueous chlorine dioxide treatment on browning of fresh-cut lotus root. LWT e Food Science and Technology, 42(2), 654e659. Fooks, L. J., & Gibson, G. R. (2002a). In vitro investigations of the effect of probiotics and prebiotics on selected human intestinal pathogens. FEMS Microbiology Ecology, 39, 67e75. Fooks, L. J., & Gibson, G. R. (2002b). Mixed culture fermentation studies on the effects of synbiotics on the human intestinal pathogens Campylobacter jejuni and Escherichia coli. Anaerobe, 9, 231e242. Giannuzzi, L., & Noem, E. Z. (1996). Effect of ascorbic acid in comparison to citric and lactic acid on Listeria monocytogenes inhibition at refrigeration temperatures. Lebensmittel-Wissenschaft und-Technologie, 29, 278e285. Huang, Y. X., & Chen, H. Q. (2011). Effect of organic acids, hydrogen peroxide and mild heat on inactivation of Escherichia coli O157:H7 on baby spinach. Food Control, 22, 1178e1183. Ibrahim, S. A., Salameh, M. M., Phetsomphou, S., Yang, H., & Seo, C. W. (2006). Application of caffeine, 1,3,7-trimethylxanthine, to control Escherichia coli O157: H7. Food Chemistry, 99, 645e650. Ibrahim, S. A., Yang, H., & Seo, C. W. (2008). Antimicrobial activity of lactic acid and copper on growth of Salmonella and Escherichia coli O157:H7 in laboratory medium and carrot juice. Food Chemistry, 109(1), 137e143. Issa-Zacharia, A., Kamitani, Y., Morita, K., & Iwasaki, K. (2010). Sanitization potency of slightly acidic electrolyzed water against pure cultures of Escherichia coli and Staphylococcus aureus, in comparison with that of other food sanitizers. Food Control, 21(5), 740e745. Issa-Zacharia, A., Kamitani, Y., Tiisekwa, A., Morita, K., & Iwasaki, K. (2010). In vitro inactivation of Escherichia coli, Staphylococcus aureus and Salmonella spp. using slightly acidic electrolyzed water. Journal of Bioscience and Bioengineering, 110(3), 308e313. Li, X. H., Xing, Y. G., Xu, Q. L., Jiang, Y. H., Yun, J. A., & Li, W. L. (2010). Effects of chitosan-based coating and modified atmosphere packaging (MAP) on browning and shelf life of fresh-cut lotus root (Nelumbo nucifera Gaerth). Innovative Food Science & Emerging Technologies, 11(4), 684e689. López-Gálvez, F., Allende, A., Truchado, P., Martínez-Sánchez, A., Tudela, J. A., Selma, M. V., et al. (2010). Suitability of aqueous chlorine dioxide versus sodium hypochlorite as an effective sanitizer for preserving quality of fresh-cut lettuce while avoiding by-product formation. Postharvest Biology and Technology, 55, 53e60. Markas, L., & Vuyst, L. D. (2006). The in vitro inhibition of Gram-negative pathogenic bacteria by bifidobacteria is caused by the production of organic acids. International Dairy Journal, 16, 1049e1057. Oms-Oliu, G., Rojas-Graü, A., González, L. A., Varela, P., Soliva-Fortuny, R., Hernando, M. I. H., et al. (2010). Recent approaches using chemical treatments to preserve quality of fresh-cut fruit: a review. Postharvest Biology and Technology, 57(3), 139e148. Rico, D., Martin-Diana, A. B., Barat, J. M., & Barry-Ryan, C. (2007). Extending and measuring the quality of fresh-cut fruit and vegetables: a review. Trends in Food Science & Technology, 18, 373e386. Ruiz-Cruz, S., Acedo-Félix, E., Díaz-Cinco, M., Islas-Osuna, M. A., & GonzálezAguilar, G. A. (2007). Efficacy of sanitizers in reducing Escherichia coli O157:H7, Salmonella spp. and Listeria monocytogenes populations on fresh-cut carrots. Food Control, 18, 1383e1390. Sagong, H. G., Lee, S. Y., Chang, P. S., Heu, S., Ryu, S., Choi, Y. J., et al. (2011). Combined effect of ultrasound and organic acids to reduce Escherichia coli O157:H7, Salmonella Typhimurium, and Listeria monocytogenes on organic fresh lettuce. Postharvest Biology and Technology, 145(2011), 287e292. Smigic, N., Rajkovic, A., Antal, E., Medic, H., Lipnicka, B., Uyttendaele, M., et al. (2009). Treatment of Escherichia coli O157:H7 with lactic acid, neutralized electrolyzed oxidizing water and chlorine dioxide followed by growth under sub-optimal conditions of temperature, pH and modified atmosphere. Food Microbiology, 26, 629e637. Tajkarimi, M., & Ibrahim, S. A. (2011). Antimicrobial activity of ascorbic acid alone or in combination with lactic acid on Escherichia coli O157:H7 in laboratory medium and carrot juice. Food Control, 22(6), 801e804. Uyttendaele, M., Neyts, K., Vanderswalmen, H., Notebaert, E., & Debevere, J. (2004). Control of Aeromonas on minimally processed vegetables by decontamination with lactic acid, chlorinated water, or thyme essential oil solution. International Journal of Food Microbiology, 90(3), 263e271. Velazquez, L. D., Barbini, N. B., Escudero, M., Estrada, C., & de Guzman, A. M. S. (2009). Evaluation of chlorine, benzalkonium chloride and lactic acid as sanitizers for reducing Escherichia coli O157:H7 and Yersinia enterocolitica on fresh vegetables. Food Control, 20(3), 262e268. Virto, R., Sanz, D., Alvarez, I., & Condon, J. R. (2005). Inactivation kinetics of Yersinia enterocolitica by citric and lactic acid at different temperatures. International Journal of Food Microbiology, 103, 251e257.