Inactivation of food by pulsed xenon flash light

Transcription

Inactivation of food by pulsed xenon flash light
Food Control 33 (2013) 15e19
Contents lists available at SciVerse ScienceDirect
Food Control
journal homepage: www.elsevier.com/locate/foodcont
Inactivation of food-related microorganisms in liquid environment by
pulsed xenon flash light treatment system
Hirokazu Ogihara a, *, Kenji Morimura a, Hikaru Uruga a, Takuya Miyamae b,
Masayuki Kogure b, Soichi Furukawa a
a
b
Department of Food Bioscience and Biotechnology, College of Bioresource Sciences, Nihon University, Fujisawa-shi, Kanagawa 252-0880, Japan
Iwasaki Electric, Co., Ltd. Ichiriyama 1-1, Gyoda, Saitama-shi 361-8505, Japan
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 19 September 2012
Received in revised form
29 January 2013
Accepted 4 February 2013
Effect of pulsed xenon flash light (PLS) treatment on the inactivation of 13 food-related microorganisms
including food-poisoning bacteria in liquid environment was investigated. Inactivation ratio was proportional to the irradiation energy and irradiation flush number. Higher inactivation was achieved in
transparent environment such as water, phosphate buffer and physiological saline. On the other hand,
pulsed xenon flash light treatment could not inactivate microorganisms in Tryptic Soy Broth. Irradiation
energy 500 J and 1 time pulse treatment brought about above 5 order inactivation in all used strains, and
it was indicated that PLS treatment was effective for inactivating food-related microorganisms including
food-poisoning bacteria.
Ó 2013 Elsevier Ltd. All rights reserved.
Keywords:
Pulsed light
Inactivation
Food-related microorganisms
1. Introduction
Prevention of the outbreak induced by food-poisoning bacteria
is a most important point in food hygiene. In food industry, heat
sterilization is one of the most important process, however, heat
treatment sometimes brings about degeneration of the fragrance
and the functional ingredient components of foods. Therefore,
development of some non-thermal sterilization processes has been
studied and developed.
Representative non-thermal sterilization processes were high
hydrostatic pressure treatment (Hoover, Metrick, Papineau, Farkas,
& Knorr, 1989; Meyer, Cooper, Knorr, & Lelieveld, 2000; Zhang &
Mittal, 2008), pulsed high electric field treatment (Evrendilek,
Zhang, & Richter, 1999; Knorr, Geulen, Grahl, & Sitzman, 1994; Qin
et al., 1995; Rodrigo, Martinez, Harte, Barbosa-Canovas, & Rodrigo,
2001), high pressure carbon dioxide treatment (Yuk, Geveke, &
Zhang, 2010; Zhang et al., 2006), and light treatment usually using
UV light (Bank, John, Schmehl, & Dratch, 1990; Chang et al., 1985;
Kuo, Carey, & Ricke, 1997) etc. Out of above treatments, UV light
treatment is most effective treatment for surface sterilization, and it
has been already used in food and medical fields (Bank et al., 1990;
Chang et al., 1985; Kuo, Carey & Ricke; 1997).
In general, light treatment needs long period for inactivating microorganisms, on the other hand, high energy short period light
* Corresponding author. Tel.: þ81 0466 84 3972.
E-mail address: ogihara@brs.nihon-u.ac.jp (H. Ogihara).
0956-7135/$ e see front matter Ó 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.foodcont.2013.02.012
treatment has been developed. Usually xenon lamp is used in this type
of high energy short period light treatment, named pulsed xenon flash
light treatment (PLS), and it use wide range wave length light (200e
1100 nm) including far UV (200e300 nm), near UV (300e380 nm)
and infrared (780e1100 nm) for inactivating microorganisms.
There were some studies on the effect of PLS on the inactivation
of some food related or food-poisoning bacteria on the various
surfaces (mainly agar media surface) (Anderson, Rowan,
MacGregor, Fouracre, & Farish, 2000; Gomez-Lopez, Devlieghere,
Bonduelle, & Debevere, 2005a,b; MacGregor et al., 1998; Rowan
et al., 1999; Takeshita et al., 2002). On the other hand, there were
a few reports on the inactivation of microorganisms in liquid
environment, such as water, apple juice, orange juice and milk
(Haffman, Slifko, Salisbury, & Rose, 2000; Palgan et al., 2011; Sauer
& Moraru, 2009). PLS treatment is a promising method for inactivating food-poisoning bacteria, however there was no enough
studies on the effect of PLS on the food related or food-poisoning
bacteria in liquid environment and its inactivation mechanism.
Therefore, effect of PLS treatment on some food-related and
food-poisoning bacteria in liquid environment was investigated.
2. Materials and methods
2.1. Pulsed xenon flash lamp light (PLS) system
PLS system was used (Iwasaki Electric, Co. Saitama, Japan). This
system is comprised of xenon lamp (tube type), irradiation vessel,
16
H. Ogihara et al. / Food Control 33 (2013) 15e19
cooling water system and power source (Fig. 1). FLASH UV
MONITOR (Iwasaki Electric, Co. Saitama, Japan) was used for
measuring the irradiation intensity of 4e16 cm points from lamp.
2.2. Used bacteria and culture
The following bacteria, Listeria monocytogenes ATCC 49594,
Staphylococcus aureus ATCC 25923, Streptococcus faecalis ATCC
29212, Enterobacter cloacae ATCC 23355, Enterobacter aerogenes ATCC
13047, Escherichia coli ATCC 25922, Providencia alcalifaciens ATCC
51902, Pseudomonas aeruginosa ATCC 27853, Serratia marcescens
ATCC 8100, Salmonella Enteritidis IFO 3313, Aeromonas hydrophila
subsp. hydrophila IFO 13286, Yersinia enterocolitica JCM 1677, and
Salmonella Typhimurium IID 1000 were obtained from the American
Type Culture Collection (ATCC) (Rockville, MD, USA), Institute for
Fermentation Osaka (IFO) (Osaka, Japan), Japan collection of Microorganisms (JCM) (Saitama, Japan) and Institute of Medical Science,
University of Tokyo (IID) (Tokyo, Japan), respectively. These bacteria
were cultured in Tryptic Soy Broth (TSB) (Difco, Detroit, MI, USA) at
37 C for 24 h. Second successive full-growth cultures (around
1089 CFU/ml) were washed two times by phosphate buffer (1/15 M),
and these cell suspensions were used for PLS treatment. After PLS
treatment, inactivation ratios were measured by culturing at 37 C for
48 h on Tryptic Soy Agar (TSA) (Difco, Detroit, MI, USA). Inactivation
ratios were directly described by cell numbers.
2.3. Inactivation of food-related bacteria by PLS
Applicable irradiation widths at 7e16 cm irradiation distance
(from lamp to liquid surface) points by PLS treatment (1 flush,
500 J) were measured by using E. coli (100 ml) on TSA in
polystyrene rectangle Petri dish (235 mm 85 mm 15 mm).
Effects of solutions of cells on the inactivation of bacteria by PLS
treatment (1, 5 and 10 flushes, 500 J, 10 cm distance) were investigated by using E. coli (5 ml) in polystyrene Petri dish (4 48 mm)
without cover. E. coli cultures were centrifuged at 8000 rpm for
10 min, and those were dissolved in 0.1% peptone added physiological saline (PSS), TSB, 1/15 M phosphate buffer (PPB) and refined
water (RFW). Optical density at 660 nm of each suspensions were
measured by BACTOMNITOR$BACT-550 (JIKCO LTD., Tokyo).
Effects of container on the inactivation of bacteria by PLS
treatment (1 flush, 500 J, 10 cm distance) were investigated by
using E. coli (4 ml) in crystal boat (73 mm 17 mm 9 mm; boat
like shape), crystal beaker (4 40 mm 60 mm) and polystyrene
Petri dish (4 48 mm) without cover. E. coli cells were dissolved in
PPB. Irradiation area, suspension depth and thickness of containers
were described in Table 1.
Table 1
Irradiation area, suspension depth and thickness of containers.
Treatment container
Irradiation
area (mm2)
Suspension
depth (mm)
Thickness
(mm)
Crystal board
Crystal beaker
Polystyrene Petri dishes
1.24 102
1.25 102
1.81 102
5.5
3.2
2.2
1.4
1.4
1.2
Effects of PLS treatment (1 flush, 200, 300, 400 and 500 J, 10 cm
distance) on the 13 food-related bacteria including 8 pathogenic
bacteria, L. monocytogenes, S. aureus, S. faecalis, A. hydrophila,
E. cloacae, E. aerogenes, E. coli, P. alcalifaciens, P. aeruginosa, S. Enteritidis, S. Typhimurium, S. marcescens, and Y. enterocolitica (each 4 ml),
in crystal boat without cover were investigated. Cells were dissolved
in PPB.
2.4. Statistical analysis
All experiments were performed in three or more replications.
The data presented are the means of at least three replicate experiments. The error bar shows the standard deviation. Significant
differences were determined by Student’s t-test (P < 0.05).
3. Results
3.1. Studies on the properties of PLS treatment on the inactivation of
bacteria
Relationships among irradiation distance, irradiation energy
and irradiation intensity in PLS treatment were described (Table 2),
and it was indicated that irradiation intensities increased in proportion to the increase of irradiation energy. However, there was no
inverse proportional relationship between irradiation intensities
and irradiation distance, and 9e12 cm irradiation distances were
best for irradiation intensity. Effect of the irradiation distances
(7e16 cm) on the applicable irradiation width by PLS treatment
of E. coli on the agar plate was investigated (Fig. 2), and irradiation distances (10e12 cm) showed the narrowest applicable
irradiation width (4e5 cm).
Effects of solutions (TSB, PSS, PPB and RFW) of the cells on the
inactivation of E. coli by PLS treatment was investigated (Fig. 3), and
inactivation ratios increased in order of TSB, PSS, PPB and RFW.
Especially in TSB, increase of flush number brought about no increase in inactivation ratio, on the other hand, 10 flushes PLS
treatment brought about above 7 orders in activation ratios in other
three solutions. Transmission ratios of the PLS in TSB, PSS, PPB and
Fig. 1. Schematic model of pulsed xenon flash lamp system.
H. Ogihara et al. / Food Control 33 (2013) 15e19
Table 2
Relationships among irradiation distance, irradiation energy and irradiation intensity in PLS treatment in PLS treatment.
4 cm
5 cm
6 cm
7 cm
8 cm
9 cm
10 cm
11 cm
12 cm
13 cm
14 cm
15 cm
16 cm
8.28
8.07
8.16
8.88
11.60
13.78
15.54
15.32
14.26
11.90
10.16
7.71
7.62
200 J
300 J
0.30
0.77
0.22
0.25
0.27
0.58
0.66
0.56
0.12
0.89
0.55
0.35
1.04
13.49
13.61
13.41
14.94
19.09
22.91
26.09
25.92
22.21
17.63
14.24
13.04
12.74
400 J
0.32
0.98
1.44
0.82
0.46
0.63
1.15
1.76
0.99
3.30
1.26
1.19
1.39
18.56
17.32
17.38
23.22
25.29
28.39
33.06
32.87
30.32
23.95
21.66
19.89
16.43
500 J
1.12
0.48
1.44
1.67
0.77
1.90
0.67
0.56
0.98
1.65
0.95
1.70
1.45
22.9
20.51
21.04
26.71
30.05
37.25
41.48
41.26
35.71
33.21
30.64
25.04
21.85
1.21
0.93
1.03
1.56
1.84
2.93
2.15
2.57
2.00
2.94
3.30
3.09
2.56
9
log 10 CFU/ml
[mJ/cm2]/(n ¼ 10) (Mean Standard deviation)
10
8
7
6
Number of cell survivors
Irradiation
distance (cm)
5
4
3
2
1
0
TSB
RFW were described in Table 3, and it was indicated that there were
inverse proportional relationships between inactivation ratios and
light transmission ratios.
Effects of container on the inactivation of E. coli by PLS treatment
(1 flush, 500 J, 10 cm distance) were investigated in polystyrene
Petri dish, crystal boat and crystal beaker without cover, and
inactivation ratios increased in order of polystyrene Petri dish,
crystal beaker and crystal boat (Table 4). These results indicated
that crystal containers were better for its high light permeability,
and difference between crystal boat and crystal beaker was
considered to be dependent on their shapes.
3.2. Inactivation of food-related bacteria by PLS
Effects of PLS treatment (1 flush, 200, 300, 400 and 500 J, 10 cm
distance) on the 13 food-related bacteria including 8 pathogen,
L. monocytogenes, S. aureus, S. faecalis, A. hydrophila, E. cloacae,
E. aerogenes, E. coli, P. alcalifaciens, P. aeruginosa, S. Enteritidis, S.
Typhimurium, S. marcescens, and Y. enterocolitica in crystal boat
were investigated (Figs. 4 and 5). Inactivation ratio was proportional to the irradiation energy. It was indicated that except for
E. aerogenes and S. Typhimurium, all strains were inactivated more
than 6-log-orders by 500 J, 1 flush PLS treatment, and all strains
were inactivated completely with the increase of the flush number
(data not shown). In many strains, treatment above 400 J showed
significantly higher inactivation. Interestingly, gram-positive bacteria showed higher sensitivity to PLS treatment.
Non
n tretment
PSS
1flashes
1fl
f ashes
PPB
RFW
5 flashes
f ashes
fl
500J
10 flashes
Fig. 3. Effects of solutions of the cells on the inactivation of E. coli by PLS treatment.
Used solutions were tryptic soy broth (TSB), peptone added physiological saline (PSS),
Potassium phosphate buffer (PPB) and refined water (RFW).
4. Discussion
There were some reports on the effect of PLS treatment on the
inactivation of microorganisms including pathogens on surface of
food stuff, such as fish, vegetables, fruit etc (Dunn, 1997; Dunn, Ott,
& Clark., 1995; Gomez-Lopez et al., 2005a,b; Kaack & Lyager, 2007;
Keklik, Demirci, Patterson, & Puri, 2010; Lagunas-solar, Pina,
MacDonald, & Bolkan, 2006; Ozer & Demirci, 2006), and it was
indicated that PLS was effective for inactivating microorganisms on
food surface and also for extending their shelf-life. On the other
hand, there were a few reports on the inactivation of microorganisms in liquid environment (Huffman et al., 2000; Palgan et al.,
2011; Sauer & Moraru, 2009). Therefore, effect of PLS treatment
on some food-related and food-poisoning bacteria in liquid environment should be investigated, and its inactivation mechanism is
also necessary to be discussed.
Relationships among irradiation distance, irradiation energy,
irradiation intensity and applicable irradiation width in PLS treatment was investigated (Table 2 and Fig. 2), and it was indicated that
irradiation intensities increased in independent on the increase of
irradiation energy, and it was also indicated that there was
Table 3
Transmission ratios of the PLS in TSB, PSS, PPB and RFW.
12
applicable irradiation width [cm]
17
10
8
Replacement solution
Transmission rate (%)
Tryptic soy broth (TSB)
0.1% peptone added physiological saline (PSS)
1/15 M phosphate buffer (PPB)
Refined water (RFW)
0.57
20.00
98.77
98.93
0.05
0.41
0.54
0.54
6
Table 4
Effects of container on the inactivation of E. coli by PLS treatment (1 flush, 500 J,
10 cm distance).
4
2
Treatment container
Pulsed light treatment
CFU/ml
0
7cm
8cm
9cm
10cm
11cm
12cm
13cm
Irradiation distance [cm]
14cm
15cm
16cm
(n=3)
Fig. 2. Effect of the irradiation distances on the applicable irradiation width by PLS
treatment (1 flash, 500 J) of E. coli.
Non treatment
Crystal board
Crystal beaker
Polystyrene Petri dishes
9.08
5.21
7.23
8.05
0.05
0.19
0.12
0.05
18
H. Ogihara et al. / Food Control 33 (2013) 15e19
10
Number of cell survivors
log 10 CFU/ml
9
8
7
6
5
4
3
2
1
0
Non tretment
ment
200J
300J
400J
500J
Fig. 4. Effects of PLS treatment (1 flush, 200, 300, 400 and 500 J, 10 cm distance) on the
5 food-related bacteria, S. faecalis, E. cloacae, E. aerogenes, E. coli, and S. marcescens, in
crystal boat.
optimum irradiation distance at around 10e11 cm for inactivating
microorganisms.
And then, effects of solutions (TSB, PSS, PPB and RFW) of the
cells on the inactivation of E. coli by PLS treatment (Fig. 3), and it
was indicated that there were inverse proportional relationships
between inactivation ratios and light transmission ratios (Table 3).
In TSB, there were some components that can absorb light
including UV, and those components would inhibit the transmission of light necessary for inactivating bacteria. PLS treatment
also could not inactivate microorganisms enough in milk and juice
environment for their high UV absorbancy.
Inactivation ratios increased in order of polystyrene Petri dish,
crystal beaker and crystal boat (Table 4). These results indicated
that crystal containers were better for its high light permeability.
Distance was measured between lamp and surface of cell suspensions and suspension depths were inversely proportional to the
inactivation ratios. We considered that reflected light from the
bottom of the crystal boat also would significantly affect the
viability of the cells. In addition boat like shape of the crystal boat
would also contribute to the low depth of cell suspensions.
From above results, it was indicated that direct irradiation of
light including UV to microbial cells would be most important point
in inactivating bacteria. This hypothesis was also supported by the
results that using crystal boat was most effective for inactivating
bacteria by PLS treatment.
Mechanisms of the inactivation of microorganisms by PLS
treatment would mainly depend on the UV, and inactivation was
mainly induced by formation of thymine dimer in DNA (Mitchell,
Jen, & Cleaver, 1992). However, PLS treatment brought about
higher inactivation ratio more than expected from UV treatment,
and it was indicated that higher inactivation ratios would be
induced by broad wave length and momentary high-energy flush.
Thermal energy would support the inactivation. However the
temperatures after 500 J and 10 time pulse have just increased
(0.9 C) crystal boat, (1.2 C) crystal beaker and (0.7 C) polystyrene
Petri dish (4 48 mm), respectively. From these results, it was
concluded that thermal energy did not support the inactivation.
Effects of PLS treatment on the 13 food-related bacteria
including 8 pathogen in crystal boat were investigated (Figs. 4 and
5), and PLS treatment (500 J, 1 flush) inactivated most used bacteria
more than 6-log-orders except for E. aerogenes and S. Typhimurium.
All strains were inactivated completely with the increase of the
flush number. E. aerogenes aggregated in its preparation, and that
would be one of the reasons of their resistance. There were many
studies on the biofilm formation of S. Typhimurium, and S. Typhimurium also would form slight aggregates. Used E. aerogenes and S.
Typhimurium would have higher UV resistance than other used
strains. Their genes related to UV resistance would have higher
activity or they would produce some extracellular substances that
can absorb UV.
From these results, PLS treatment was promising method to
inactivate food-related bacteria including pathogen in high light
permeable liquid environment. The precise mechanism of the
inactivation of microorganisms by PLS treatment should be
investigated.
Number of cell survivors
log 10 CFU/ml
10
9
8
7
6
5
4
3
2
1
0
Non tretment
200J
300J
400J
500J
Fig. 5. Effects of PLS treatment (1 flush, 200, 300, 400 and 500 J, 10 cm distance) on the 8 food-related pathogenic bacteria, L. monocytogenes, S. aureus, A. hydrophila, P. alcalifaciens,
P. aeruginosa, S. Enteritidis, S. Typhimurium, and Y. enterocolitica in crystal boat were investigated.
H. Ogihara et al. / Food Control 33 (2013) 15e19
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