Food Control 70 (2016) 242e248

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Food Control
journal homepage: www.elsevier.com/locate/foodcont

High pressure processing extend the shelf life of fresh salmon, cod and
mackerel
Tone Mari Rode*, Maria Befring Hovda
Noma, Norwegian Institute of Food, Fisheries and Aquaculture Research, Richard Johnsens Gate 4, N-4021 Stavanger, Norway

a r t i c l e i n f o a b s t r a c t

Article history: High pressure processing (HPP) is used on several types of seafood, though its potential for extending the
Received 15 March 2016 shelf life of fresh sh has not been fully exploited. This study compared the effect of HPP (200 and
Received in revised form 500 MPa, 120 s) on salmon, cod and mackerel. Immediately after processing, and during storage up to 26
25 May 2016
days different analysis were carried out to evaluate the microbiological shelf life, oxidation, acid phos-
Accepted 26 May 2016
phatase level and pH of the different sh species during the storage period. For cod and mackerel, HPP at
Available online 27 May 2016
500 MPa restrained the bacterial ora and it did not reach spoilage level, opposite to the salmon samples
exposed to 500 MPa. Analysis of TBARS is often used as a measure of lipid oxidation. The TBARS-level was
Keywords:
Microbiology
differently affected by pressure in salmon and cod. HPP at 200 MPa did not have any effect on the
TBARS oxidation level in salmon during the storage period while this was observed for cod. The TBARS level in
pH mackerel was high, independent of pressure treatment or not. Phosphatase activity in sh has been
Acid phosphatase suggested to be related to freshness. The acid phosphatase (ACP) levels in control samples showed
HPP signicant differences between salmon and cod. For salmon both control and samples treated at 200 MPa
Shelf life showed a decrease in ACP level at day 11 compared with day 0, whereas the HPP500 values were even
throughout the study. This latter observation was also the case for cod treated at 500 MPa. This study
have shown that HPP induce different levels of changes in various sh species.
2016 Published by Elsevier Ltd.

1. Introduction 2000; Ramirez-Suarez & Morrissey, 2006; Yagiz et al., 2009).

High pressure processing (HPP) has been increasingly imple-
mented in the food industry for extending the microbiological shelf
life and food quality of different foods. The effect of HPP on sh and
seafood has not been as intensively studied, and the industrial use
of this technology has been limited compared with other types of
food like juice, cooked meat and dry fermented sausages. The shelf
life of fresh sh and seafood is short, and processing methods to
inhibit spoilage and thereby increase shelf life would be an
advantage. Some challenges need to be addressed to ensure a
proper inhibition of spoilage and pathogenic bacteria, avoid lipid
oxidation and maintaining sensory characteristics.
There are some studies on HPP of sh (Campus et al., 2010;
Cheret, Chapleau, Delbarre-Ladrat, Verrez-Bagnis, & De Lamball-
erie, 2005; Go mez-Estaca, Lo  pez-Caballero, Go mez-Guille
n, Lo
 pez
de Lacey, & Montero, 2009; Matser, Stegeman, Kals, & Bartels,
* Corresponding author. Noma, P.O. Box 8034, N-4021 Stavanger, Norway.
E-mail addresses: tone.mari.rode@noma.no (T.M. Rode), mariabhovda@
hotmail.com (M.B. Hovda).
However, results from different studies using high pressure pro-
cessing can be difcult to compare, as different equipment, pres-
sure level, holding time, temperature, storage time and analysis are
used. Furthermore, different sh species have different biological
composition, shelf life, and are affected by processing in various
ways. The quality of fresh sh is rapidly reduced due to various
breakdown processes. Lipid oxidation and increased microbial load
during storage are main factors for the spoilage of sh.
The shelf life of fresh sh depends on the initial sh quality and
storage conditions (temperature, atmosphere and packaging). To
extend the shelf life of sh, thermal processes can be used. How-
ever, the quality of the sh can be severely reduced if the heating
process is designed for a shelf life of more than 21 days at chilled
conditions. Use of high pressure processing can be an interesting
application to extend the shelf life of sh without using heat.
Enzymatic time-temperature integrators (TTI) have been used
as intrinsic biological markers for heat induced quality changes or
bacterial inactivation during thermal processes. There have also
been some attempts trying to make TTIs that are relevant for HPP
(review by Grauwet et al. (2012)). Acid phosphatase (ACP) has been

http://dx.doi.org/10.1016/j.foodcont.2016.05.045
0956-7135/ 2016 Published by Elsevier Ltd.
 T.M. Rode, M.B. Hovda / Food Control 70 (2016) 242e248
used as an intrinsic TTI in thermal processes in cod (Skipnes,
Johnsen, Skara, & Lekang, 2010), but in minor degree been tested
for its relevance during HPP. In a study of sea bass (Teixeira et al.,
2013), the acid phosphatase activity was lower at 400 MPa
compared with the nontreated sample.
The aim of this study was to investigate the effect of HPP on
three different kinds of sh (salmon, cod and mackerel). The sh
samples were pressurized at 200 MPa or 500 MPa for 2 min, and
stored for up to 26 days at 0.5
C. On selected days the microbial
load, TBARS, pH and acid phosphatase was analysed. Furthermore,
this experiment investigated the potential use of ACP as an intrinsic
biological marker for HPP by studying the correlation between ACP
activity and the quality and microbiological contents of the sh. The
ACP method was also suggested as a method for determining the
thermal load during HPP.
2. Materials and methods
2.1. Raw material
Skinned and lleted back loin from farmed salmon (Salmo salar),
farmed cod (Gadus morhua), and llet of wild-caught mackerel
(Scomber scombrus) was obtained after slaughtering. The salmon
and cod lets were vacuum packed and used 2 and 3 days after
slaughter. During packaging of cod, the package was ushed with
CO2 before vacuum was employed. The mackerel was used 5 days
after catching, though only one day after lleting. The lets were
cut into samples, vacuum packed individually and stored at 0.5
C
on ice before high pressure processing (HPP). Both control and HPP
samples were included in the study. After HPP, the samples were
stored on ice in a cooling room (0.5 0.5
C). On given days, ana-
lyses of microbiology and pH were conducted. Samples for TBARS
and ACP were frozen at 80
C until further analysis. Day 0 was set
as the day of HPP processing, irrelevant of time of slaughtering. The
same was for the control samples, though they were unprocessed.
2.2. High pressure treatment
HPP was carried out in a high hydrostatic pressure machine QFP
2L-700 (Avure Technologies Inc., Columbus, USA). The HPP unit was
cooled to 8e9
C prior to treatments. Samples were pressurized at
either 200 (HPP200) or 500 MPa (HPP500) for 120 s. Come up times
were approx. 50 and 100 s for 200 and 500 MPa, respectively,
whereas the pressure release was immediate. The duration of
treatment did not include the come up time. Prior to and after
treatment, samples were kept on ice. Control samples were non-
pressurized samples (0.1 MPa).
2.3. Microbial analyses
For microbial analyses, 5 samples of each sh (10 g) were diluted
1:10 in peptone water (BactoPeptone, Merck, Darmstadt, Germany)
added 0.85% NaCl, and homogenised for 3 min in a stomacher.
Aerobic quantication of psychrotrophic and heat labile microor-
ganisms (colony forming units [cfu] ml1) was performed by sur-
face plating on Long & Hammer agar plates. The plates were
incubated for 5e7 days at 15
C. A mechanical spiral plater (Eddy
Jet, IUL Instruments, Barcelona, Spain) was mainly used for this
purpose. However, some manual plating was performed for low
dilutions. Aerobic count was also performed on Iron Agar to
determine hydrogen sulphide producing bacteria. A pour plate
technique was used, adding Iron Agar with 0.8% L-cysteine to 1 ml
of sample. Plates were incubated for 3e4 days at 20
C. Both the
total number (aerobic count) and the number of black colonies
were counted (hydrogen sulphide producing bacteria). The
243
detection level on Long & Hammer and Iron Agar was 102 and
10 cfu/g, respectively. For plates with no colonies detected, the level
was set to half of the detection limit. The microbial analysis was
performed on selected days, depending on the sh species. For cod
at day: 0, 7, 11, 14 and 22, salmon: day 0, 7, 11, 15, 18, 21 and 26, and
for mackerel: day 0, 4, 7, 11, 14 and 19. Day 0 was set as the day of
HPP processing.
2.4. pH
pH was measured in room tempered, homogenised samples
(n 5) using a pH-meter instrument PHM210 (Meterlab, Copen-
hagen, Denmark).
2.5. Lipid analyses
The amount of total lipids was analysed in triplicate sh samples
at day 0. The analysis was performed according to Bligh and Dyer
(Bligh & Dyer, 1959).
2.6. Analyses of thiobarbituric acid reactive substances (TBARS)
Triplicate samples (100 g) of sh were frozen at 80
C at
selected days after storage. For all sh species, samples at day 0 and
11 were analysed. In addition, samples of salmon at day 25 and cod
at day 18 were analysed. The TBARS analysis was performed ac-
cording to Srensen and Jrgensen (Sorensen & Jorgensen, 1996).
Shortly described the procedure included: at the day of analysis,
thawed samples (10 g) were homogenised and TBARS was extrac-
ted by homogenisation in trichloroacetic acid, ltrated, and added
TBA. Steam distillation was carried out in a water bath at 100
C for
40 min. After cooling to room temperature the absorbance was
measured at 532 nm, and the TBARS values were calculated. Cali-
bration was performed towards a standard curve of TEP (1,1,3,3-
tetraethoxypropane). The level of TBARS were presented as mg
malondialdehyde (MDA)/kg of sample.
2.7. Analyses of acid phosphatase (ACP)
The acid phosphatase (ACP) was determined using the method
of Kuda, Matsumoto, and Yano (2002), and analysis was performed
on samples (n 5) from selected days. The procedure for ACP an-
alyses and calculations was performed as in Skipnes et al. (2010).
Briey, muscle sample (1 g) was added 1% Triton X-100. After
centrifugation, the supernatant was used in an enzymatic assay and
added a substrate solution. The ACP activity was measured in a
spectrophotometer as the absorbance of p-nitrophenol (405 nm)
released from the substrate.
2.8. Statistics
General linear modelling (GLM) was performed to determine
statistically signicant effects of the different parameters (Minitab
Statistical Software v15, Minitab Ltd., Coventry, UK) using Tukeys
HSD test at level P < 0.05 (95%). ANOVA was applied to log trans-
formations of the microbiological counts.
3. Results and discussion
3.1. Effect of HPP on the microbial inactivation
Aerobic bacteria. The number of background aerobic bacteria
varied between the sh species (Fig. 1). The salmon samples
showed no detectable growth at day 0, while the untreated cod and
mackerel samples showed initial bacterial numbers of 3.8 0.3 and
244 T.M. Rode, M.B. Hovda / Food Control 70 (2016) 242e248

9 1A 9 1B
8 8
7 7
6 6

Log cfu/g
Log cfu/g 5
5
4 4
3 3
2 2
1 1
0 0
0 7 11 14 18 22 26 0 7 11 14 18 22 26
Storage at 0.5 C (days)
Storage at 0.5 C (days)

9 9
2A 2B
8 8
7 7
6 6
Log cfu/g

Log cfu/g
5 5
4 4
3 3
2 2
1 1
0 0
0 7 11 15 18 21 26 0 7 11 15 18 21 26
Storage at 0.5 C (days) Storage at 0.5 C (days)

9 3A 9 3B
8 8
7 7
6 6
Log cfu/g

Log cfu/g

5 5
4 4
3 3
2 2
1 1
0 0
0 4 7 11 14 19 21 26 0 4 7 11 14 19 21 26
Storage at 0.5 C (days) Storage at 0.5 C (days)

Fig. 1. Aerobic count on Long & Hammer (A) and Iron agar (B) plates of spoilage ora in salmon (1), cod (2) and mackerel (3) after storage at 0.5
C. The solid lines are control
samples not exposed to HPP. The dashed and dotted lines are sh samples exposed 200 and 500 MPa, respectively. The detection level on Long & Hammer and Iron agar was 1.7 and
0.7 log cfu/g, respectively. For plates with no detected colonies, the level was set to half of the detection limit.

5.3 0.2 log cfu/g, respectively. For both cod and mackerel, pro-
cessing at 500 MPa lowered the bacterial number to undetectable
levels right after processing. Salmon control samples were spoiled
after about 18e22 days of storage at 0.5
C. For unprocessed cod the
shelf life of was between 11 and 15 days, whereas mackerel be-
tween day 0 and 4. Bacterial numbers above log 6 cfu/g on Iron agar
(IA) and log 7 cfu/g on Long & Hammer (L&H) is often used to
indicate spoilage (ICMSF, 1986; Senturk & Alpas, 2012).
The sample period for each sh was longer then the shelf life
recommended. According to the producer of the salmon and cod
used in these experiments, they had a best-before-date 10 days
after slaughter. This would be equivalent to day 8 and 9 for salmon
and cod, respectively, in this study. After 11 days, growth was at an
undetectable or below log 2 cfu/g on L&H and IA, respectively, for
the salmon HPP500 samples, Fig. 1A and B. HPP200 extended the
time to around 7 days before the bacteria started growing on L&H
compared to around 11 days for HPP500. After 18 days of storage
the HPP salmon was still acceptable for consummation, while the
control was spoiled, showing bacterial numbers >106 on IA. A study
by Briones, Reyes, Tabilo-Munizaga, and Perez-Won (2010) showed
that aerobic count in salmon increased to upper acceptable limit
after 17 4 days, and for salmon processed at 200 MPa this value
was reached after 21 days. This is the same as we observed for the
HPP200 samples. Our ndings correspond to the ndings another
study which showed that HP treatment signicantly reduced the
bacterial growth in farmed coho salmon (Aubourg, Tabilo-
Munizaga, Reyes, Rodriguez, & Perez-Won, 2010). Furthermore
HPP (>300 MPa) of rainbow trout resulted in a 6 log reduction in
bacteria, and no growth after 6 days of storage (Yagiz, Kristinsson,
Balaban, & Marshall, 2007). Based on the bacterial numbers from
IA plates, the salmon processed at 200 and 500 MPa was spoiled
when analyzing after 22 days. Even though HPP increased the lag
phase and the bacterial numbers were just above the detection
level up to day 11, there might be indications, based on the results
of this study that the bacterial growth escalated when the bacteria
rst started to grow in the salmon samples. This (same) pattern was
not observed for neither cod nor mackerel.
For cod, the control samples were spoiled between day 11e15,
Figs. 2A and B. The cod HPP500 samples had a lag phase of around 7
days, and did not reach the level of spoilage during the 26 days of
storage. However, the HPP200 treated cod reached the spoilage
level at the end of the experiment around day 21. The shelf life after
slaughtering of mackerel, given by the supplier, was 8 days. After 4
storage days the control sh was spoiled, which was 9 days after
slaughtering. HPP200 mackerel was spoiled after 7 storage days,
hence 12 days after slaughtering. However, the HPP500 mackerel
samples showed bacterial levels below the spoilage level during the
19 days the storage experiment lasted, Figs. 3A and B. Senturk and
Alpas (2012) showed that HPP at 200 and 400 MPa extended the
shelf life of Atlantic mackerel and resulted in a shelf life of 17 and 19
days, respectively. Compared with our study, the other study was
performed on thawed-frozen samples, but that cannot explain the
difference in shelf life after HPP. Other studies have shown a pos-
itive effect in extending the shelf life of sh by the use of HPP
(Chevalier, Le Bail, & Ghoul, 2001; Erkan, Uretener, & Alpas, 2010;
Erkan, retener, & Alpas, 2010; Gomez-Estaca, Montero, Gimenez,
 T.M. Rode, M.B. Hovda / Food Control 70 (2016) 242e248 245

6 6
A B
5 5

4 4

Log cfu/g

Log cfu/g
3 3

2 2

1 1

0 0
0 7 11 15 18 21 26 0 4 7 11 14 19 21 26
Storage at 0.5 C (days) Storage at 0.5 C (days)

Fig. 2. Hydrogen producing bacteria in cod (A) and mackerel (B) after storage at 0.5
C on Iron agar plates. The solid lines are control samples not exposed to HPP. The dashed and
dotted lines are sh samples exposed 200 and 500 MPa, respectively. The detection level was 10 cfu/g. For plates with no colonies detected, the level was set to half of the detection
limit.

9 3.2. pH
0.1 200 500
8
In the unprocessed control samples the pH varied from 5.96 to
7
6.46 for the different types of sh at day 0, Table 1. For salmon there
mol MDA/kg

6 was a statistically signicant difference in pH between control and

5 HPP500 samples during the storage period (P < 0.05). The unpro-
4 cessed cod samples generally had a pH ranging between 6.30 and
3 6.48 during the storage period, having the highest pH compared
2 with salmon (6.16e6.27) and mackerel (5.96e6.34). Generally,
1 there were relatively small differences in pH comparing unpro-
cessed sh and the HPP sh. However, there was a tendency of pH
0
increasing up to day 4 or 7 for all samples. Then there was a
0 11 25 0 11 18 0 11
decrease resulting in pH levels lower than the initial pH. This was
Salmon Cod Mackerel the case for all samples stored longer than 18 days. The same results
Fish and days of storage of increasing pH after some days, and then decreasing pH levels to
almost the initial level has been reported in another HPP study of
Fig. 3. Changes in lipid oxidation as TBARS (mg MDA/kg) for salmon, cod and mack-
tuna (Ramirez-Suarez & Morrissey, 2006).
erel. Samples exposed to 0.1, 200 and 500 MPa followed by storage at 0.5
C.
In a study by Angsupanich and Ledward (1998) on cod they also
found an increase in pH with increasing pressure level used. It was
suggested in that study that the increased pH was associated with
& Gomez-Guillen, 2007; Karim et al., 2011; Ramirez-Suarez & the denaturation of some protein fractions. After storage for 7 days,
Morrissey, 2006; Yagiz et al., 2007). Several of these studies have the pH increased most for the control samples, and gave the highest
studied pressure levels below 300 MPa. One reason for this might values, even higher than the samples exposed to 800 MPa.
be the in many cases detrimental both textual and sensory effect of
using pressure levels above 300 MPa. 3.3. TBARS
Hydrogen sulphide producing bacteria. Bacteria producing
hydrogen sulphide (H2S) appear as black colonies on IA. For salmon The amount of lipids varies for different types of sh. The
only one black colony was detected for two individual samples, amount of fat was 12.37 2.72, 1.03 0.06 and 3.53 1.42% in
during the entire storage period. On salmon HPP500 samples there salmon, cod and mackerel, respectively. The TBARS analysis was
were no H2S producing bacteria detected. The latter was also the employed as a measure of lipid oxidation, and TBARS have been
case for cod and mackerel, Fig. 2. During the experiment none of the shown to correlate well with the development of rancid odor and
samples, not even the unprocessed controls, reached the spoilage avor (Mielnik, Herstad, Lea, Nordal, & Nilsson, 2002). The result of
level, often set to log 8 cfu/g (Jorgensen, Gibson, & Huss, 1988). the TBARS analysis is shown in Fig. 3. There was a major difference
Spoilage of H2S producing bacteria is mainly caused by Shewanella in TBARS level of the different sh types tested. Both unpressurised
putrefaciens and other Vibrionaceae which are known to be present and 200 MPa samples of salmon and cod showed low levels of
only in low numbers in farmed sh like cod and halibut (Hovda, oxidation during the whole period tested (25 days for salmon and
Lunestad, Sivertsvik, & Rosnes, 2007; Hovda, Sivertsvik, Lunestad, 18 days for cod). Pressurizing at 500 compared to 200 MPa showed
Lorentzen, & Rosnes, 2007; Sivertsvik, 2007). a signicant difference (P < 0.05) in TBARS level for all sampling
Amanatidou et al. (2000) had in their study presence of S. days, except day 0 for both salmon and cod.
putrefaciens in vacuum packed untreated salmon samples. After One difference between salmon and cod was that the TBARS
storage for 8 days at 5
C, and the total counts was >7 log. For level in the former was low and stable, below 0.12 mg MDA/kg, for
samples exposed to 150 MPa for 10 min, the equivalent samples unprocessed and 200 MPa samples also during storage. The low
contained 4.63 log cfu S. putrefaciens. Even though the storage level of TBARS in salmon, processed around 200 MPa, is also
temperature was different, the large amount of bacteria at day conrmed by others (Amanatidou et al., 2000; Yagiz et al., 2007).
0 was probably the reason for the diverging results in this and the The results of Yagiz et al. (2007) also support the increasing level of
other paper. oxidation during storage in salmon processed for >450 MPa. A
study of cod showed no increase in TBA values after 7 days of
storage for cod processed at 200 MPa, compared with day
246 T.M. Rode, M.B. Hovda / Food Control 70 (2016) 242e248

Table 1
pH in the different sh (salmon, cod and mackerel) in control (0.1) and HPP samples (200 and 500 MPa) during storage (day 0e26). pH values with standard deviation is shown.

Fish Pressure (MPa) Time of storage (day)

0 4 7 11 14 18/19 21/22 26

Salmon 0.1 6.22 0.04 nd 6.27 0.03 6.19 0.03 6.17 0.04 nd 6.16 0.02 nd
200 6.28 0.03 nd 6.3 0.02 6.26 0.04 6.22 0.06 nd 6.23 0.07 nd
500 6.34 0.03 nd 6.36 0.02 6.35 0.04 6.32 0.02 nd 6.31 0.06 nd

Cod 0.1 6.36 0.07 nd 6.48 0.10 6.37 0.09 6.36 0.05 6.37 0.12 6.30 0.10 nd
200 6.42 0.06 nd 6.48 0.07 6.45 0.04 6.45 0.08 6.42 0.05 6.36 0.07 6.42 0.08
500 6.46 0.02 nd 6.51 0.07 6.47 0.08 6.47 0.06 6.44 0.07 6.41 0.09 6.43 0.05

Mackerel 0.1 5.96 0.05 6.22 0.06 6.34 0.07 6.34 0.09 nd nd nd nd
200 6.09 0.13 6.04 0.11 6.29 0.17 6.31 0.03 nd nd nd nd
500 6.08 0.12 6.18 0.09 6.15 0.07 6.08 0.06 6.13 0.07 6.06 0.04 nd nd

nd: not determined.

0 (Angsupanich & Ledward, 1998). For cod processed at 400 MPa
the TBARS level was more than doubled from day 0 to day 7
(0.74e1.67), which is higher levels then observed in this study. Here
we detected an increase in lipid oxidation during storage with
highest values seen at the last sampling point (after 18 days), 0.60
and 1.22 mg MDA/kg, respectively for HPP200 and HPP500. The
mackerel samples in this study showed very high levels of lipid
oxidation even at day 0 for both unprocessed and processed sam-
ples, where the level was above 4 mg MDA/kg. It has been stated
that TBARS values in salmon above 1.9 mg MDA/kg are character-
ized as a rancid taste (Amanatidou et al., 2000), and a level between
1 and 2 mg MDA/kg has been denoted by others as the limit of
acceptability (Goulas & Kontominas, 2005). So these high levels will
probably be noticed by the consumer as a rancid taste.
A study by Go mez-Estaca et al. (2009) showed that the oxidative
values of fresh salmon were lower compared to tuna in spite of the
high fat content, and stated that this was due to the high carotenoid
content of salmon. Our salmon results conrm this nding, as the
TBARS level was very low in the unprocessed control samples. The
high levels of astaxanthin in salmon is responsible for the red
colour of the sh meat. This carotenoid has high antioxidant ac-
tivity, and may be the reason why low numbers of oxidation was
observed for salmon during storage. For cod exposed to treatment
at 200 MPa, there was a steady increase in the oxidative values of
the processed samples, and the level after 18 days were higher
compared with salmon stored for 25 days (P < 0.05). Astaxanthin
probably also gave a protective effect for salmon when processed at
200 MPa, but when processed at 500 MPa, the antioxidant effect
was not adequate to prevent oxidation. There are also human
studies showing the positive connection between astaxanthin and
oxidation (Iwamoto et al., 2000; Park, Chyun, Kim, Line, & Chew,
2010). For mackerel the TBARS levels at day 0 were 30 times
higher in both processed and unprocessed samples compared with
both salmon and cod samples. And at the last sampling points, day
25 and 18 for salmon and cod, respectively, the TBARS levels were
much lower than the start level of mackerel. The amount of fat in
mackerel (3.53%) was not so much higher compared with cod
(1.03%). However, transition metals are major pro-oxidants, and the
amount of iron, haemoglobin and myoglobin in dark muscle of
mackerel is very high. Studies have also indicated that the dark
muscles are rich in oxidative bers that are more susceptible to
oxidative reactions than white muscles (Alasnier, David-Briand, &
Gandemer, 2000; Undeland, Ekstrand, & Lingnert, 1998).
3.4. ACP
Phosphatase activity in sh has been suggested to be related to
freshness. The ACP absorbance in HPP treated salmon showed small
change in enzyme activity during 25 days storage, although some
differences were observed (Fig. 4A). HPP200 treatment did not alter
the enzyme activity, compared to untreated samples, whereas
HPP500 treatment gave lower enzyme activity (P < 0.001) right
after processing. Statistically signicant changes were observed for
the control and HPP200 comparing the day 0 and 11 absorbance
values, whereas the HPP500 values were even throughout the
study.
HPP of cod resulted in a decrease in ACP activity after process-
ing, and both 200 and 500 MPa were signicantly different from the
control (P < 0.05) (Fig. 4B). HPP200 and 500 were signicantly
different at day 11 (P < 0.05), however this difference disappeared
after storing the samples for 25 days. During storage the amount of
ACP in the HPP200 samples showed a statistically signicant
decrease from day 0 to day 25 for both salmon and cod (P < 0.05).
For the HPP500 samples there were no signicant changes. In
mackerel there were no signicant difference between the control
and HPP samples during storage. The only exception was that the
HPP500 was statistically signicant form the control at day 11
(P < 0.05) (Fig. 4C). During the storage period, there were no sta-
tistically signicant changes for the different mackerel samples.
The observed effects on ACP can be related to HPP treatment, as
the adiabatic temperature during treatment will be below 30
C
and too low to affect the ACP. Previously Kuda et al. (2002) have
shown differences in the ACP content of different sh species. The
ACP content of the untreated controls in this study revealed sta-
tistically signicant differences between the salmon, and cod
samples at day 0 (P 0.007). A study of ACP in sea bass showed
lower activity for samples treated by 400 MPa compared with
nontreated controls. The decrease in ACP has been suggested to be a
result of enzyme inactivation caused by HPP (Teixeira et al., 2013).
This can explain the results for salmon and cod in this study. At day
0 there were signicant differences, but these differences were
smaller during the storage period. For mackerel it was the opposite
as the changes between the samples were increasing during
storage.
This study have shown that HPP induce different levels of
chemical and microbiological changes in various sh species. HPP
can extend the shelf life of sh by inhibiting bacterial growth. The
microbiological lag phase in salmon was extended by treatment at
500 MPa, but then showed an exponential growth phase to spoilage
level. For cod and mackerel HPP500 samples, the bacterial growth
was within acceptable levels during the storage period. Pressur-
izing at 500 MPa might have a negative effect on different quality
parameters, whereas HPP at 200 MPa does not have the same effect
of changing the physical and chemical properties of sh. If pressure
levels around 200 MPa were used on sh this could have a positive
effect on increasing the shelf life. For all sh there were an extended
microbial shelf life by using HPP, and also at 200 MPa. The results of
this study have shown that different pressure levels ought to be
 T.M. Rode, M.B. Hovda / Food Control 70 (2016) 242e248 247

3.0 Acknowledgements
A
The authors thank Line Bach Christensen, Leena Shinde, Laila
2.5
a ab Budal and Karin Trany for skilled work in the laboratory. This work
bc c was supported by The Norconserv Foundation.
2.0
cd
de
e
e References
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1.5
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