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Combined Effects of Alkaline Electrolyzed Water and Citric Acid with Mild Heat to Control Microorganisms on Cabbage

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Effects of alkaline electrolyzed water (AlEW), acidic electrolyzed water (AcEW), 100 ppm sodium hypochlorite (NaClO), deionized water (DIW), 1% citric acid (CA) alone, and combinations of AlEW with 1% CA (AlEW + CA), in reducing the populations of spoilage bacteria and foodborne pathogens on cabbage were investigated at various dipping times (3, 5, and 10 min) with different dipping temperatures (1, 20, 40, and 50 degrees C). Inhibitory effect of the selected optimal treatment against Listeria monocytogenes and Escherichia coli O157 : H7 on cabbage were also evaluated. Compared to the untreated control, AlEW treatment most effectively reduced the numbers of total bacteria, yeast, and mold, followed by AcEW and 100-ppm NaClO treatments. All treatments dip washed for 5 min significantly reduced the numbers of total bacteria, yeast, and mold on cabbage. With increasing dipping temperature from 1 to 50 degrees C, the reductions of total bacteria, yeast, and mold were significantly increased from 0.19 to 1.12 log CFU/g in the DIW wash treatment (P < 0.05). Combined 1% CA with AlEW treatment at 50 degrees C showed the reduction of around 3.98 and 3.45 log CFU/g on the total count, and yeast and mold, effective reduction of L. monocytogenes (3.99 log CFU/g), and E. coli O157 : H7 (4.19 log CFU/g) on cabbage. The results suggest that combining AlEW with CA could be a possible method to control foodborne pathogens and spoilage bacteria effectively on produce.
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M: Food Microbiology
& Safety
JFS M: Food Microbiology and Safety
Combined Effects of Alkaline Electrolyzed
Water and Citric Acid with Mild Heat
to Control Microorganisms on Cabbage
S.M.E. RAHMAN,YONG-GUO JIN,AND DEOG-HWAN OH
ABSTRACT: Effects of alkaline electrolyzed water (AlEW), acidic electrolyzed water (AcEW), 100 ppm sodium
hypochlorite (NaClO), deionized water (DIW), 1% citric acid (CA) alone, and combinations of AlEW with 1% CA
(AlEW +CA), in reducing the populations of spoilage bacteria and foodborne pathogens on cabbage were inves-
tigated at various dipping times (3, 5, and 10 min) with different dipping temperatures (1, 20, 40, and 50 C). In-
hibitory effect of the selected optimal treatment against Listeria monocytogenes and Escherichia coli O157:H7on
cabbage were also evaluated. Compared to the untreated control, AlEW treatment most effectively reduced the num-
bers of total bacteria, yeast, and mold, followed by AcEW and 100-ppm NaClO treatments. All treatments dip washed
for 5 min significantly reduced the numbers of total bacteria, yeast, and mold on cabbage. With increasing dipping
temperature from 1 to 50 C, the reductions of total bacteria, yeast, and mold were significantly increased from 0.19
to 1.12 log CFU/g in the DIW wash treatment (P<0.05). Combined 1% CA with AlEW treatment at 50 Cshowed
the reduction of around 3.98 and 3.45 log CFU/g on the total count, and yeast and mold, effective reduction of
L. monocytogenes (3.99 log CFU/g), and E. coli O157 : H7 (4.19 log CFU/g) on cabbage. The results suggest that com-
bining AlEW with CA could be a possible method to control foodborne pathogens and spoilage bacteria effectively
on produce.
Keywords: alkaline electrolyzed water, background flora, cabbage, citric acid, combination treatment,
pathogens
Introduction
Eating patterns among people have changed toward vegetables
and well-being food products due to the increased concerns
on their health (Ahn and Shin 1999). Vegetables such as broccoli,
kale, and cabbage have a high content of vitamins, minerals, and
functional ingredients. The sulfuric chemicals with unique flavor
prevent cancer in our body and inhibit a mutation. Thus, these sul-
furic chemicals have been popular among the convenient and fresh
vegetables (Stoewasand 1995). However, since minimally manufac-
tured agricultural products are usually ingested as an uncooked
form without additional heat treatment, it becomes more likely to
be susceptible to foodborne pathogens and spoilage bacteria, lead-
ing to food poisoning. The United States, Canada, and other North-
American countries reported that cabbage salad could be a main
reservoir of food-poisoning bacteria (Wehr 1987).
NaClO has been extensively used as a sanitizing agent in man-
ufacturing companies to improve the microbiological safety, ex-
tend the shelf life of the fresh products by reducing the number
of the initial microorganisms, and inhibit the enzyme activity
when the vegetables are washed. However, it can cause corrosion in
the equipments due to its chemical characteristic of chlorine’s own
stink and oxidation (Yukari and others 1990). Recently, electrolyzed
water (EW) has been found to have an effective bactericidal activ-
ity and leaves no residuals. It, as a washing bactericidal agent, does
MS 20090833 Submitted 8/28/2009, Accepted 12/7/2009. Authors Rahman
and Oh are with Dept. of Food Science and Biotechnology and Inst. of Bio-
science and Biotechnology, Kangwon Natl. Univ., Chuncheon, Gangwon
200-701, Republic of Korea. Author Jin is with College of Food Science and
Technology, Huazhong Agricultural Univ., Wuhan 430070, China. Direct
inquiries to author Oh (E-mail: deoghwa@kangwon.ac.kr).
not produce the secondary pollutions and has been extensively
used (Noriaki and others 2000). EW has been used for a long since
to inactivate pathogens on fresh produce (Izumi 1999; Koseki and
others 2001; Pangloli and others 2009). Huang and others (2008)
reported that electrolyzed oxidizing water (EOW) have the fol-
lowing advantages over other traditional cleaning agents: effective
disinfection, easy operation, relatively inexpensive, and environ-
mentally friendly. The main advantage of EW is its safety that it is
not corrosive to skin, mucous membrane, or organic material like
others chemical compounds. EW has been tested and used as a
disinfectant in the food industry and other applications (Al-Haq
and others 2005). Combinations of EW and other measures are also
possible. There are reports that Clin the EW is in dissolved form
as an ion (Huang and others 2008). The bactericidal ability of this
water is due to the oxidation by chlorine gas. Also, it has been re-
ported that the mechanism of bactericidal ability is mainly due
to its oxidation–reduction potential (ORP) and the amount of dis-
solved oxygen in water, but the correct mechanism has not been
identified yet (Sakai 1995). Also, it has been reported that the or-
ganic acids such as propionic acid, acetic acid, citric acid (CA), tar-
taric acid, lactic acid, malic acid have been used as a single agent or
in combination with other agents to reduce the possible occurrence
of the food poisoning (Ita and Hutkins 1991; Tamblyn and Conner
1997). The combined use of several disinfectant agents has been
widely report in the last few years (Uyttendaele and others 2004;
Beltr´
an and others 2005; Ukuku and others 2005). Combinations of
lactic acid, chlorinated water, thyme essential oil solution, sodium
lactate, CA, hydrogen peroxide, ozone, and peroxyacetic acid were
already tested. In general, combinations of chemical disinfectants
maintain better both sensory and microbial quality of the product.
C
2010 Institute of Food Technologists RVol. 75, Nr. 2, 2010JOURNAL OF FOOD SCIENCE M111
doi: 10.1111/j.1750-3841.2009.01507.x
Further reproduction without permission is prohibited
M: Food Microbiology
& Safety
Combination treatment on cut cabbage . . .
A novel produce-washing procedure using a combination of alka-
line electrolyzed water (AlEW), acidic electrolyzed water (AcEW),
and mild heat demonstrated significant bactericidal effect com-
pared with the treatment with ambient temperature (Koseki and
Isobe 2007). Thus, this study was conducted to determine the con-
ditions for reducing the microorganisms on cabbage by individ-
ual or combination process of organic acid and the EW with mild
heat. The inhibitory effect of combined AlEW and CA against Lis-
teria monocytogenes and Escherichia coli O157:H7inoculatedon
cabbage was also investigated at the selected treatment condition
of sanitizing agents.
Materials and Methods
Bacterial cultures
The 2 strains of L. monocytogenes (ATCC 19116 and ATCC 19111)
and E. coli O157 : H7 (932 and 933) used in this experiment were
obtained from Korean Natl. Inst. of Health (Seoul, Korea) and Dept.
of Food Science, Univ. of Georgia (Griffin, Ga., U.S.A.), respectively.
Stock cultures were maintained on tryptic soy agar (BD Diagnos-
tics, Franklin Lakes, N.J., U.S.A.) slants at 4 C. Prior to use, all
strains were separately grown in tryptic soy broth (TSB; BD Diag-
nostics) at 37 C with 2 consecutive transfers after 24-h periods for
a total of 48 h of incubation. All working cultures (approximately
109CFU/mL) grown in TSB were separately centrifuged at 4000 ×g
for 15 min at 4 C and the supernatants were discarded. The cell
pellets were washed twice with 0.1% peptone water (pH 7.1) and re-
suspended in 10 mL of the same solution to obtain a final cell con-
centration of 109CFU/mL. After this, 2 strains of E. coli O157 : H7
and L. monocytogenes were combined in a cocktail with approxi-
mately equal numbers in the final population.
EW generation
The EW was provided using EW generator (A2-1000, Korean E &
S Fist Inc., Seoul, Korea) including a small amount of salt solution
and tap water. AcEW (pH of 2.4 to 2.6, ORP of 1000 mV to 1100 mV,
and an available chlorine concentration of 50 to 60 mg/L) and AlEW
(pH of 11 to 11.2 and ORP of 830 mV to 850 mV) were produced
from EW generator, respectively. The pH, ORP, and available chlo-
rine concentration of the treatment solutions (AcEW and AlEW)
were measured immediately before treatment with a dual scale pH
meter (Accumet model 15, Fisher Scientific Co., Fair Lawn, N.J.,
U.S.A.) bearing pH and ORP electrodes. The residual chlorine con-
centration was determined via a colorimetric method using a dig-
ital chlorine test kit (RC-3F, Kasahara Chemical Instruments Corp.,
Saitama, Japan). The detection limit for this measurement was 0 to
300 mg/L.
Sample preparation and inoculation
The cabbages used in this experiment were purchased from a
local supermarket or big warehouse stores in Chunchon, Korea,
and then quickly transported to laboratory and stored at 4 Cfor
no more than 3 h before testing. Uneatable, wilted, and dam-
aged portions were trimmed. Cabbages were cut into 3 ×3cm
slices using a sterile knife. Before inoculation, each sample weighed
10 ±0.1 g. Each trimmed leaf was placed on sterile aluminum foil
in a biosafety hood. Pieces of sliced cabbage were inoculated by
pipetting 0.1 mL of pathogen cocktail suspensions (approximately
109CFU/mL) onto the surface of the samples. The samples were
then left in a laminar flow hood for 30 min at room temperature
(23 ±2C) to allow for bacterial attachment to the leaf surfaces.
This procedure resulted in initial pathogen levels of approximately
6 to 7 log CFU/g on cabbage.
Sanitizing treatments
Inoculated and uninoculated cabbage samples (10 g) were
placed in sterile containers and immersed in each sanitizing agent,
deionized water (DIW). Unwashed cabbages were used as control.
To evaluate the effect of dipping time on the reduction of microor-
ganisms, each piece (10 g) was dipped for 3, 5, and 10 min and the
samples were immersed at 1, 20, 40, and 50 Ctoevaluatetheef-
fect of temperature. For sanitizing agents (dipping solutions), DIW,
100 ppm NaClO, electrolyzed water (AcEW and AlEW) and 1% CA
(Fisher Scientific Co., Norcross, Ga., U.S.A.) either alone or in com-
bination with AlEW were used. After the dipping of each cabbage,
the moisture on the each piece was drained using sterile gauze. All
experiments were replicated 3 times.
Microbiological analysis
Following treatments, all samples (each of 10 g) were aseptically
excised and immediately placed in a stomacher bag (Nasco Whirl-
Pak, Janesville, Wis., U.S.A.) containing 90 mL of buffered peptone
water (BPW; Difco, Sparks, Md., U.S.A.) and homogenized for 2 min
with a Seward stomacher (400 Circulator, Seward, London, U.K.).
After homogenization, 1-mL aliquots of the sample were serially
diluted in 9 mL of sterile BPW and 0.1 mL of sample or diluent
was spread-plated onto each selective medium (in this case limit
of detection was 100/g, that is, 2 log CFU/g). In the case of 1 mL
placed on the plates, the limit of detection was 1 log CFU/g. Two se-
lective media of Sorbitol MacConkey agar (Difco) and Oxford Agar
Base (Difco) were used for the enumeration of E. coli O157:H7and
L. monocytogenes, respectively. For uninoculated cabbage samples,
the mixture were plated onto plate count agar (Difco) for the total
counts and potato dextrose agar (Difco with 10% tartaric acid solu-
tion) for yeasts and molds. All plates were incubated at 37 Cfor24
h, except for yeasts and molds at 25 Cfor3to5dandexpressedas
log CFU/g.
Statistical analysis
Means of bacterial populations (log10 CFU/g) from each treat-
ment were calculated from 3 replications for each experiment. Data
were expressed as the means ±standard errors. The results were
analyzed using SPSS statistical package (SPSS Inc., Chicago, Ill.,
U.S.A.) and the significance of difference was defined at P<0.05.
Results and Discussion
Effects of dipping times on
inactivation of microorganisms
Effects of dipping times (3, 5, and 10 min) on the reduction of
total counts and yeast and mold on cabbage treated with sanitiz-
ing agents are shown in Figure 1A and 1B, respectively. For all dip-
ping times, DIW treatment reduced the populations by less than
0.5 log CFU/g, while AlEW treatment showed the greatest reduc-
tion of 2.71 and 2.74 log CFU/g for total bacteria and yeast and
mold count, respectively compared to the untreated control. Com-
pared with other sanitizers, the relative influence of inactivation
was AlEW >AcEW >NaClO >DIW. Though, there was no sig-
nificant difference (P>0.05) found in dipping times effect, yet 5
and 10 min dipping time from each treatment showed better effect
than 3 min on the reduction of total counts on cabbage (Figure 1A).
Similar results were observed in the reduction of yeast and mold
on cabbage (Figure 1B). From these results, there was no any re-
markable difference found in microbial reduction between 5 and 10
min dipping. Also 10 min dipping resulted in changing color (visual
estimation) of cut cabbage. According to these results, 5 min ex-
posure would be the most appropriate to minimize the quality
M112 JOURNAL OF FOOD SCIENCEVol. 75, Nr. 2, 2010
M: Food Microbiology
& Safety
Combination treatment on cut cabbage . . .
deterioration (color), thus it was chosen for following treatments.
Yang and others (2003) observed a 2 log CFU/g reduction in
Salmonella typhimurium,E. coli,andL. monocytogenes on the sur-
A
a
a
a
a
a
a
a
a
a
a
a
a
0
1
2
3
4
5
6
AlEWAcEWNaClODIWControl
Treatments
Log CFU/g
3 min
5 min
10 min
B
a
a
a
a
a
a
a
a
a
a
a
a
0
1
2
3
4
5
6
AlEWAcEWNaClODIWControl
Treatments
Log CFU/g
3 min
5 min
10 min
Figure 1 --- Effect of electrolyzed water on inactivation of
total counts (A), and yeast and mold (B) on cabbage by dif-
ferent dipping times. Means with different letters in the
same figure are significantly different (P<0.05). (Con-
trol: not treated; DIW: deionized water; NaClO: 100 mg/L
sodium hypochlorite; AcEW: acidic electrolyzed water;
AlEW: alkaline electrolyzed water.)
faces of romaine lettuce after a 5 min dip in neutral EOW (pH 7; 300
ppm total residual chlorine, TRC). On the other hand, dip (10 min)
treatment of spinach in near-neutral EOW at 100 and 120 ppm TRC
resulted in a 4 to 5 log CFU/mL reduction of bacterial counts for
five organisms (E. coli,Staphylococcus typhimurium,S. aureus,L.
monocytogenes,andEnterococcus faecalis) tested which is obtained
by Guentzel and others (2008).
Carson (1991) found that the bactericidal effect of the EW was
higher than the chlorine water because ORP in the EW was higher
than that of the chlorine water. Some researchers believe that the
antimicrobial activity of AcEW is due to the presence of chlorine
species, while others believe that the low pH is responsible. A few
studies have suggested that this activity is due to its high ORP (Kim
and others 2000; Liao and others 2007). Park and others (2002) re-
ported that Campylobacter jejuni was more sensitive to AcEW than
chlorine water in a culture medium and AlEW has better bacterici-
dal effect against microorganisms such as yeast and mold or total
counts on vegetables than AcEW, thus it reduced the total num-
ber of microorganisms (Park and others 2004a). Koide and oth-
ers (2009) revealed that slightly acidic electrolyzed water (SlAEW)
(pH 6.1, 20 mg/L available chlorine) has an equivalent or high disin-
fectant efficacy for fresh cut cabbage compared to NaClO solution
(pH 9.6, about 150 ppm available chlorine). They found SlAEW re-
duced about by 1.5 log CFU/g for total aerobic bacteria and 1.3 log
CFU/g for yeasts and molds, compared to fresh cut cabbage before
dipping.
Effects of dipping temperatures on
inactivation of microorganisms
The effects of dipping temperatures (1, 20, 40, and 50 C) on the
efficacy of sanitizers including DIW, 100 ppm NaClO (pH 7.48), 1%
CA (pH 1.87), AcEW (pH 2.45), AlEW (pH 11.46), and CA +AlEW
(pH 2.32) in the reduction of the total bacteria, yeast, and mold
on cabbage are shown in Table 1 and Figure 2A and 2B. Washing
with AlEW (2.06 log reduction of total counts) and 1% CA (2.10 log
reduction of total counts) at 1 C for 5 min showed significantly
greater (P<0.05) sanitizing effect than DIW treatment (0.20 log re-
duction of total counts) compared to control, while sanitizing effect
with AlEW (2.34 log reduction of total counts) and 1% CA (2.30 log
reduction of total counts) was much more enhanced than DIW
treatment (0.45 log reduction of total counts) at 20 C. As dipping
temperatures increased from 1 to 50 C, DIW treatment showed
Table 1 --- Effect of alkaline electrolyzed water and citric acid, either alone or in combination on inactivation of total
counts, and yeast and mold on cabbage by different dipping temperatures for 5 min.
Treatment Temperature (C) Total count (log CFU/g)aYeast and mold (log CFU/g)
Control – 5.23 ±0.16Aa 4.15 ±0.17Aa
DIW 1 5.03 ±0.18a 3.96 ±0.17a
20 4.78 ±0.21ab 3.77 ±0.21ab
40 4.32 ±0.18b 3.45 ±0.18b
50 4.11 ±0.16Bb 3.32 ±0.19Bb
AlEW 1 3.17 ±0.24b 1.75 ±0.21b
20 2.89 ±0.20bc 1.57 ±0.15b
40 2.40 ±0.16c 1.25 ±0.14bc
50 2.15 ±0.19cd 1.10 ±0.11Cc
CA 1 3.13 ±0.11b 2.02 ±0.18b
20 2.93 ±0.16b 1.85 ±0.10b
40 2.43 ±0.15c 1.52 ±0.21bc
50 2.22 ±0.21Cc 1.37 ±0.20Cc
AlEW +CA 1 2.75 ±0.18b 1.64 ±0.17b
20 2.53 ±0.19b 1.46 ±0.19b
40 2.00 ±0.11c 1.16 ±0.20bc
50 1.25 ±0.13Dd 0.70 ±0.12Dc
aMeans with different letters in the same column are significantly different (
P
<0.05).
Control: not treated; DIW: deionized water; AlEW: alkaline electrolyzed water; CA: 1% citric acid; AlEW +CA: alkaline electrolyzed water +1% citric acid.
Vol. 75, Nr. 2, 2010JOURNAL OF FOOD SCIENCE M113
M: Food Microbiology
& Safety
Combination treatment on cut cabbage . . .
significantly increased the reduction of total counts from 0.19 to
1.12 log CFU/g (P<0.05). Like to AlEW, and 1% CA treatments
at 1 C, DIW treatment at 50 C exhibited a significant (P<0.05)
bactericidal effectiveness. Similar results were observed in yeast
and mold at each treatment. However, there was no significant
difference on reduction among 1% citric, AcEW, and AlEW wash-
ing at 20 and 40 C, but significantly enhanced reduction was ob-
served for all treatments at 50 C compared to 1 C for the total
counts,andyeastandmold(Table1andFigure2Aand2B).In
the meantime, combined 1% CA and AlEW treatment showed
the significant reduction of the total counts and yeast and mold
on cabbage at 50 C compared to each single treatment alone
(Table 1). There have been reports that mild heat treatment im-
proved the efficacy of a sanitizer in killing or removing microor-
ganisms on produce. Shredded lettuce treated with chlorinated
water for 3 min at 47 C reduced aerobic bacterial count by 3 log
CFU/g, while that reduced by approximately 1 log CFU/g at 4 C
(Delaquis and others 1999). Kim and others (2002) reported that
the bactericidal effect was more enhanced in the treatment of 50
mg/L chlorine at 50 C when compared to washing cabbage with
distilled water at 50 C. Park and others (2004a) elucidated that the
combination treatment of the AlEW with organic acid had better
reduction (2 log CFU/g) than AlEW or CA and lactic acid alone.
Similar results were observed in our experiments that combina-
tions of AlEW and 1% CA had greater inhibitory effect than each
1°C20°C40°C50°C
A
aa
a
a
ab
ab
b
ab
b
ab
bc
b
bc
b
c
b
0
1
2
3
4
5
6
AlEWAcEWNaClODIWControl
Treatments
Log CFU/g
1°C20°C40°C50°C
B
a
a
a
a
a
ab
b
ab
ab
b
b
b
b
b
b
b
0
1
2
3
4
5
6
AlEWAcEWNaClODIWControl
Treatments
Log CFU/g
Figure 2 --- Effect of electrolyzed water on inactivation of
total counts (A), and yeast and mold (B) on cabbage by
different dipping temperatures for 5 min. Means with dif-
ferent letters in the same figure are significantly different
(P<0.05). (Control: not treated; DIW: deionized water;
NaClO: 100 mg/L sodium hypochlorite; AcEW: acidic elec-
trolyzed water; AlEW: alkaline electrolyzed water.)
single treatment. Koseki and others (2004) observed that the ap-
pearance of the mildly heated (50 C) lettuce was not deteriorated
after the treatment. They also have illustrated the efficacious ap-
plication of AlEW as a prewash agent and the effective combined
use of AlEW and AcEW. Mild heat treatment of fresh produce is re-
ported to enhance the bactericidal effect of sanitizers and the phys-
iological and sensory quality of the produce. A significantly more
enhanced antibrowning effect was observed (Jin and others 2009)
when the lettuce leaves were subjected to both 1% CA and AlEW
treatment at temperatures greater than 40 C.
Effect of combined treatment on the
reduction of foodborne pathogens
It would be expected that combinations of sanitizers and/or
other intervention methods would have additive, synergistic, or an-
tagonistic interactions (Parish and Davidson 1993; Rajkowski and
Baldwin 2003). According to our results, combination of AlEW with
1% CA for 5 min at 50 C had a strongest sanitizing effect on the
reduction of indigenous total counts or yeast and mold on cab-
bage. Thus, this optimal condition was chosen for the reduction
of L. monocytogenes and E. coli O157:H7inoculatedoncabbage
(Figure 3A and 3B). The dipping cabbage leaves in DIW for 5 min
at 50 CforL. monocytogenes reduced populations only by 0.34
CFU/g, while 100 ppm NaClO, AlEW, and 1% CA treatment alone
decreased populations by 2.31, 2.69, and 2.83 log CFU/g, respec-
tively (Figure 3A). Combined treatment of AlEW and 1% CA sig-
nificantly (P<0.05) decreased populations of L. monocytogenes
by 3.99 log CFU/g compared with each treatment alone. More en-
hanced reductions were observed in E. coli O157 : H7 inoculated
on cabbage at the same condition (Figure 3B). The DIW, NaClO,
A
c
b
b
b
a
a
0
1
2
3
4
5
6
7
8
AlEW+CACAAlEWNaClODIWControl
Treatments
Log CFU/g
B
c
b
b
b
a
a
0
1
2
3
4
5
6
7
8
AlEW+CACAAlEWNaClODIWControl
Treatments
Log CFU/g
Figure 3 --- Effect of alkaline electrolyzed water and cit-
ric acid, either alone or in combination on inactivation of
L.monocytogenes (A) and E.coli O157:H7 (B) on cabbage
at 50 C for 5 min. Means with different letters in the same
figure are significantly different (P<0.05). (Control: not
treated; DIW: deionized water; NaClO: 100 mg/L sodium
hypochlorite; AlEW: alkaline electrolyzed water; CA: 1%
citric acid; AlEW +CA: alkaline electrolyzed water +1%
citric acid.)
M114 JOURNAL OF FOOD SCIENCEVol. 75, Nr. 2, 2010
M: Food Microbiology
& Safety
Combination treatment on cut cabbage . . .
AlEW, and 1% CA treatment alone decreased populations by 0.21,
2.39, 2.69, and 2.85 log CFU/g, respectively, but the combined treat-
ment of AlEW and 1% CA showed the strongest inactivation of
4.19 log CFU/g. Many combinations of physical and chemical treat-
ments have been tested in recent years to enhance the antimicro-
bial action of different disinfectant agents. Effective combined use
of AlEW and AcEW with mild heat has been reported by Koseki and
others (2004). On the other hand, AcEW has been combined with
ozone by using sequential washes (Wang and others 2004). A newer
tendency has been reported by Bari and others (2005), they investi-
gated the efficacy of nisin and pediocin treatments in combination
with EDTA, CA, sodium lactate, potassium sorbate, and phytic acid
in reducing L. monocytogenes on fresh-cut produce. Yuk and oth-
ers (2007) showed that ozone treatment gave less than 1 and 0.5 log
count reductions on E. coli O157 : H7 and L. monocytogenes,respec-
tively on enoki mushroom. Efficacy was improved with combined
3 ppm ozone and 1% CA treatment, resulting in 2.26 and 1.32 log
count reductions, respectively.
The antimicrobial effect of EW for killing bacteria on cabbage
might be dependent on microbial species and strain, exposure
time, pH and temperature, and delivery method (agitation, soak-
ing, spraying, or bubbling). Park and others (2004b) demonstrated
that EOW is very effective for inhibiting E. coli O157 : H7 and L.
monocytogenes in a wide pH range (between 2.6 and 7), if suffi-
cient residual chlorine is present. The residual chlorine concentra-
tion of EOW will only need to be maintained at a very low level for
practical applications, because EOW containing only 1 to 2 mg/L of
residual chlorine showed very strong bactericidal activity against E.
coli O157:H7andL. monocytogenes. Shigenobu and others (2004)
found that AlEW treatment on the E. coli O157 : H7 inoculated on
cabbage at 50 C for 5 min reduced the populations by 2.79 log
CFU/g, giving similar results in our experiments. Jung and oth-
ers (1996) revealed that complete bactericidal effect of EW against
Bacillus spp. and Staphylococcus in broth condition was observed,
but its reduction effect was extensively decreased in the food due to
the interaction of complex food matrix.
In conclusion, the most significant finding in this study was
the synergistic effect of AlEW and CA on the inactivation of to-
tal aerobic bacteria, yeast and mold, L.monocytogenes,andE.coli
O157 : H7. The use of the combined treatment could greatly ben-
efit the vegetable industry as post harvest intervention method
that would improve the microbiological safety of vegetables. There-
fore, the results provide practical information on the applica-
tion of the combined EW with organic acid, as effective surface
sanitizer, on controlling foodborne pathogens and spoilage
bacteria on vegetables. Therefore, further studies should be eluci-
dated to study both sensory and microbial quality of the product as
well to determine the synergistic effects of combining technologies.
Acknowledgments
This work is financially supported by the Ministry of Education,
Science and Technology (MEST) and the Ministry of Knowledge
Economy (MKE), and the Ministry of Labor (MOLAB) through the
fostering project of the Industrial-Academic Cooperation Centered
Univ.
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This study was conducted to determine the effects of alkaline electrolyzed water (AIEW), acidic electrolyzed water (AcEW), 1% citric acid, and 100 ppm sodium hypochlorite, either alone or in combination with citric acid, in reducing the populations of spoilage bacteria and foodborne pathogens (Listeria monocytogenes and Escherichia coli O157:H7) on lettuce at various exposure times (3, 5, and 10 min) with different dipping temperatures (1, 20, 40, and 50°C). In addition, the inhibitory effect of alkaline electrolyzed water combined with citric acid on the browning reaction during storage at 4°C for 15 days was investigated. Compared to the untreated control, electrolyzed water more effectively reduced the number of total bacteria, mold, and yeast than 100 ppm sodium hypochlorite under the same treatment conditions. All treatments exposed for 5 min significantly reduced the numbers of total bacteria, yeast, and mold on head lettuce. The inactivation effect of each treatment on head lettuce was enhanced as the dipping temperature increased from 1 to 50°C, but there was no significantly difference at temperatures greater than 40°C (p<0.05). The total counts of yeast and mold in head lettuce were completely eliminated when a combination of 1% citric acid and AlEW treatment was used at temperatures greater than 40°C. However, decreased reduction in L. monocytogenes (2.81 log CFU/g), and E. coli O157:H7 (2.93 log CFU/g) on head lettuce was observed under these treatment conditions. In addition, enhanced antibrowning effect was observed when the samples were subjected to both 1% citric acid and AlEW treatment at temperatures greater than 40oC compared to when single treatments alone were used. Thus, this combined treatment might be considered a potentially beneficial sanitizing method for improving the quality and safety of head lettuce.
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To study the effect of citric, acetic, lactic, and hydrochloric acids on Listeria monocytogenes Scott A, growth, survival, and intracellular pH (pHin) values were determined during growth in a pH-controlled fermentation vessel. Under the experimental conditions, L. monocytogenes Scott A grown in tryptic soy (plus yeast extract) broth survived even when the pH was reduced to 3.5. For most acids, L. monocytogenes maintained a pH gradient (intracellular pH-external pH) of about 1.0 pH unit and a pHin near 5.0. When the citric and lactic acid-treated cells at pH values 3.5, 4.0, and 4.5 were incubated for a longer time (24 h), both the pH gradient and the pHin values decreased. Although citric and lactic acids were more effective in lowering the pHin, acetic acid had the greatest effect on cell survival. A greater than 4-log reduction in cell number occurred when L. monocytogenes was held in acetic acid-treated broth for 24 h at pH 3.5 even though the pHin was 5.0. The results suggest that inhibition of L. monocytogenes by acids is caused not by a decrease in the intracellular pH, per se, but rather by specific effects of undissociated acid species on metabolic or other physiological activities.
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The influence of electrolyzed acid-water (oxidation-reduction potential (ORP): above 1,150 mV, pH : 2.5) on the survival of some microorganisms was investigated. It was observed that the ORP of electrolyzed acid-water was kept at the level of above 1,000 mV for 15 days at room temperature. Escherichia coli. Salmonella typhi, Staphylococcus aureus and Saccharomyces cerevisiae were not detected after 10 to 40 min in electrolyzed acid-water. However, Bacillus cereus showed higher tolerance to electrolyzed acid-water than other test microorganisms. After 60 min of inoculation, only 0.4% of initial population remained. The investigation of surface sterilization effect on some vegetables was carried out too. Total count of cabbage, Chinese cabbage and kale were reduced to below 3% of initial count, and no coliform was detected after 20 to 60 min of immersion in 5 volumes of electrolyzed acid-water. In the lettuce, total and coliform counts were reduced to 90% and 2% of initial population. This study shows that the electrolyzed acid-water has a potential for the sterilization of food products such as vegetables and fruits which cannot be thermally sterilized.
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Chemicals containing SH-groups as sulfites and chlorine-based agents are commonly employed in the fresh-cut process of vegetables such as potatoes to prevent browning and to sanitize produce. However, there is a concern over the application of these compounds in fresh-cut commodities as they might affect human and environmental safety and this has created the need to investigate alternatives. In the present work, the effectiveness of different traditional and non-traditional sanitizers on the sensory and microbial quality of fresh-cut potatoes stored under passive modified atmosphere packaging (MAP) and vacuum packaging was investigated. Six different washing treatments consisting of water, sodium sulfite, sodium hypochlorite, Tsunami, ozone and the combination of ozone–Tsunami were evaluated. Browning and growth of aerobic mesophilic bacteria, psychrotrophic bacteria, coliforms, lactic acid bacteria (LAB), anaerobic bacteria, moulds and yeasts were studied. In general, vacuum packaging preserved the appearance better than MAP. Under MAP only sodium sulfite prevented browning although it conferred off-odors. After 14 days of storage, there was no evidence of browning in fresh-cut potatoes dipped in ozonated water or ozone–Tsunami and stored under vacuum and these treatments maintained initial texture and aroma. However, the use of ozonated water alone was not effective in reducing total microbial populations. Ozone–Tsunami resulted in the most effective treatment to control microbial growth achieving 3.3, 3.0 and 1.2 log-reductions for LAB, coliforms and anaerobic bacteria, respectively. Therefore, although microbial growth was not slowed down by ozone alone, the combination of ozone–Tsunami resulted an efficient and promising treatment for controlling microbial growth and maintaining sensory quality of potato strips under vacuum.