LABORATORY REPORT

FOOD PACKAGING AND STORAGE TECHNOLOGY

FISH DETERIORATION

Written by:
Brigitta Angelina 01034170097
Charlene Octavian 01034170043
Delvin Kho 00000019680
Karenina Caroline 01034170080
Michael Adhimulia 01034170088

FOOD TECHNOLOGY STUDY PROGRAM

FACULTY OF SCIENCE AND TECHNOLOGY
UNIVERSITAS PELITA HARAPAN
TANGERANG
2020
 CHAPTER I
INTRODUCTION

1.1 Introduction
Fish is a perishable food product with limited storage life due to its
biological composition (Sen, 2005). It is particularly known to be susceptible to
spoilage. With slightly alkaline pH of 6.5 to 7.0, high water activity of 0.95 or
more and enzymes present inside, conditions of post-harvests may accelerate
spoilage if stored in higher temperatures. Autolysis and microbial load are the
prime suspects in staleness of fish. While autolysis is caused by enzymes present
in fish, microbial count depends on the environment in which the fish is caught.
Warmer waters carry higher bacterial counts over to fish. Usual techniques to
preserve fish include temperature control and reducing water activity by
refrigerating or freezing. Fish flesh are inherently sterile, hence proper handling
and storage are of question if excessive spoilage does occur (Lie, 2008).
The quality of fish can be evaluated by several methods, which include
both organoleptic, chemical and microbial. Organoleptic tests include texture and
smell, while chemical evaluations include pH and Total Volatile Base Nitrogen
and Trimethylamine content of fish (Obemeata and Christopher, 2012; Sen 2005).

1.2 Objectives
The objective of this experiment was to determine the effects of storage
temperature on chemical, microbiological and organoleptic qualities of fish.

1
 CHAPTER II
LITERATURE REVIEW

2.1 Red Tilapia Fish

Nile tilapia fish (​Oreochromis niloticus​), is a type of fish that can live in
freshwater to brackish water. It grows best at 25-30C, with pH 7-8 and oxygen
concentration of 3-5 ppm (Saparinto, 2012). It is a pelagic fish, which means it
commonly lives in the upper layers of water. In particular, this fish lives, at a
height of 0-1000 m dpl. (Evans, 2009). Containing 4% fat, it is categorized as lean
fish (Suloma ​et al​., 2008). Its distinguishable traits include distinctive stips that
extend from its upper body part to its lower body part. Commonly known tilapia
fish include black, red, and gift tilapia fish.
Red tilapia fish, also known as ‘Nila Merah’, is a hybrid fish that is a
​ ith nile tilapia or ​nila.​ Red
result of a third crossing from tilapia fish or mujair, w
tilapia fish commonly have red body, but some are found to have blackish or
mixed color. It can also have a red colored back and reddish white abdomen. It
has large eyes with bluish green color on its edges (Saparinto, 2012; Kordi, 2010).

2.2 Fresh fish standards according to SNI

According to BSN (2013), fresh fish is defined as fish that has not
undergone preservation, except chilling. Common characterization includes
appearance, odor, and texture. Fish according to the standard has bright and clear
eyes, scent of fresh fish and is elastic, solid, and compact, in terms of texture.
Organoleptic evaluations evaluated include eye, gills, body surface slime or
mucus, meat, smell and texture, all of which are evaluated with a score range of 1
to 9. Fresh fish must have a minimum score of 7 in organoleptic properties. In
terms of microbial properties, it should not exceed total plate count of ​5.0 x 10​5
colonies/g.

2
2.3 Fish spoilage
2.3.1 Factors affecting changes in fish properties
Fish is a highly perishable food commodity, therefore fish can easily
undergo a deterioration process. According to Rahman (2007), there are a few
factors that can contribute to the deterioration process of fish. Those factors such
as the mechanical handling damage, environmental factors, biotic factor, lipid
oxidation and hydrolysis are very crucial in keeping the quality of fish. The
nutritional content of fish also plays a great role in which fatty fish that contains a
high amount of polyunsaturated fatty acids will undergo oxidative rancidity since
they are chemically reactive. In the case of lean fish, the spoilage is mainly
microbial spoilage. Carbohydrates content in fish muscle are very low, this results
in bacteria using nitrogenous materials which in turn will produce unwanted
off-odours and off-flavours (Adams and Moss, 2007).
For chilled fish, spoilage mainly occurs due to psychotropic Gram
negative bacteria such as ​Shewanella putrefaciens a​ nd ​Pseudomonas spp.
Bacterial catabolism of these amino acids will produce compounds such as TMA,
skatole and other amines (Adams and Moss, 2007).
2.3.2 Fish spoilage mechanism
Fish can experience spoilage because they undergo several
microbiological and chemical changes immediately after they are captured. When
the fish are improperly handled, fish will undergo spoilage like color, texture and
odor changes, biochemical changes and nutritional content changes. Based on
Rahman (2007), fish undergo 4 phases during spoilage when stored in ice. The
first phase is when the fish is stored for less than a week. Fish at the first phase
will show no sign of spoilage. When the fish is stored for 7 to 10 days, the fish
still shows no sign of odor production. However, after 11 to 14 days of storage,
the fish will produce fruity odors. After storage of more than 2 weeks, there will
be some sulfide compounds production like hydrogen sulfite and fecal ammonia
odours.

3
 Fish will also show some color and texture changes like in the gills,
surface, flesh and eyes. Fish gills of early storage will be red or orange colour, and
as storage time increases, the color will fade to pale gray or blue. For the surface
of fish, increase of storage time will result in thick yellow slime on the body of the
fish. While for the texture, fish that is kept for more than 2 weeks will retain
finger indentation (Rahman, 2007).

2.4 Fish storage

Fish products are considered to have relatively short shelf life influenced
by their quality and storage conditions especially fish’s temperature. To obtain the
best shelf life possible, fish must be stored at near 0​°C to prevent the fish from
being frozen and minimize the microorganism and enzymatic activities in the fish
product. When the storage temperature rises above ​0​°C, the bacterial and
enzymatic activities also rise significantly causing lower shelf life. The most
acceptable storage temperature for fish is between -23°C to -29°C. If possible,
crushed ice might be possible to maintain fish quality. The microorganism can be
washed by the melted water from the ice that prolongs the shelf life. Little to no
packaging is needed for fish.
Rapid freezing in fish is necessary to be implemented in fish storage due to
the smaller ice crystal that is less damaging the flesh structure in fish products by
minimizing destructive concentration of cellular constituents. Rapid freezing
should be done when thin processors, prechilled fish packages, high capacity low
temperature freezers are used (Caballero ​et al​. 2003).

2.5 Principle of Analysis

2.5.1 Drip loss
Drip loss is a parameter that depends on the species of the product,
particularly in shrimps. The initial quality of the fish as well as the rate of freezing
and the time period. Fish that were slowly frozen will have greater drip loss than
quick-frozen fish (Sen, 2005). The water-holding capacity of the muscles will

4
determine the rate of the drip loss, once the water content is larger than the water
holding capacity, water will be lost either immediately or during processing
further on. Therefore the muscle protein content is important to reduce the drip
loss, making the post-harvest handling and marketing a critical step to retain the
water content in the fish properly. Delaying the rigor mortis by rapid death of the
fish will help retain the protein of the muscle and retain the water holding capacity
of the fish (Rahman, 2007).
The rate of weight loss is affected by temperature, the temperature
fluctuation, humidity, heat transfer, air flow of the product, radiation effects of
lighting, the product itself (including the type, shape and size) as well as the
wrapper used (Goncalves, 2012). Weight loss in fish occurs due to the formation
of ice crystals from the water molecules in fish. Cold storage at high temperatures
will cause the ice to form more slowly, forming larger and less uniform ice
crystals. This will also cause the muscle structure of the fish to be damaged more
easily compared to quick freezing. As the muscle fiber in fish shrinks and retracts
due to the dehydration of concentrated proteins and minerals, larger drip loss will
occur as it is common in fish frozen more slowly (Dawson ​et.al, ​2018).
2.5.2 TVBN and TMA
Trimethylamine oxide is an osmoregulatory compound which is present in
considerable amounts in marine fish. Freshwater fish usually lack this compound,
with the exception of Nile perch and tilapia. After death, TMAO is reduced to
TMA by TMAO-reductase present in intestinal enterobacteria or bacteria
introduced from outside such as S. putrefaciens. Along TMA release, off-odor
from breaking down of sulfur-containing amino acids are also produced. Hence,
TMA presence is used to indicate the degree of spoilage in mostly marine fish to
be correlated with sensory result (Venugopal, 2006).
Total Volatile Base Nitrogen (TVBN) is a classic indicator of fish
spoilage, and is a better index of quality as not all fish have TMAO within. TVBN
includes TMA, ammonia and other volatile amines. TMA is also produced only
by microbial spoilage, and hence cannot be used as an indicator of freshness. Like

5
TMA, TVBN also correlates with sensory results. 10-15 mg of TMA and 30-40
mg of TVBN per 100 grams are considered to be the limits, as sensory rejection
starts at the limits aforementioned (Venugopal, 2006).
2.5.3 Total Plate Count
Total plate count is a method used to calculate the amount of aerobic
microorganisms present on a media plate that has been inoculated with a sample.
This method is done by swabbing the sample like air, equipment or food into a
culture medium and letting it incubate for a certain period of time. The samples
are usually diluted several times before being inoculated. Because the desired
microorganisms to be counted are of a specific group, plate count must be done in
a sterile condition to avoid contamination of other unwanted groups of
microorganisms. Total plate count results only show the number of
microorganisms present, not the type of microorganisms that grow on the sample
(Mariott and Robertson, 2012).

6
 CHAPTER III
MATERIALS AND METHODS

3.1 Materials and Equipment

The materials used in this experiment were Red Tilapia fish, ice, sulfuric
acid, boric acid, formaldehyde, TCA, methyl red and bromocresol green,
physiological salt solution and PCA media. The equipment used in this
experiment were burette, Erlenmeyer flasks, beakers, bunsen burners, refrigerator,
test tubes, dilution bottles, mass balance, knives, Ziploc bags, Petri dishes and
micropipettes.

3.2 Procedure
3.2.1 Sample preparation
1. The fish were submerged in an ice bath to shock.
2. The fish were descaled, gutted and washed.
3. The fish samples were packed and weighed
4. The samples were stored in room temperature or chilled storage, according
to the treatments.
5. All analyses were performed at hour 0, 2,5 and 22.
3.2.2 Sensory analysis
1. Sensory analysis was performed on the fish, in accordance with SNI
2729:2013.
3.2.3 Weight loss analysis
1. Weight the fish stored at the specified times
2. Calculate the loss from the difference with the initial weight.
3.2.4 TPC analysis
1. 10 grams of fish was put in a dilution bottle of 90 ml physiological salt
solution.
2. The diluted samples were then diluted further to 10​-4​, 10​-5​ and 10​-6​.

7
 3. The diluted samples were inoculated onto PCA media and incubated at
37°C for 24 hours.
4. The colonies were counted for each dilution levels.
3.2.5 pH check
1. 10​-1​ dilutions of the samples were drawn from the dilution bottle.
2. The pH of samples were checked with a pH meter.
3.2.6 TVBN and TMA analysis
1. 100 grams of the fish samples were blended with 200 ml of TCA.
2. The mixtures were then centrifuged. The clear extracts were drawn.
3. For TVBN determination, 25 mL of the clear extracts and 6 mL of 10%
NaOH were added into a Kjeldahl flask.
4. The samples were distilled and captured in Erlenmeyer flasks containing
15 mL of 4% boric acid.
5. The samples were added with two drops of phenolphthalein indicator and
titrated With 0.03 N H​2​SO​4​.
6. For TMA determination, steps 3-5 were repeated with the addition of 20

mL 35% Formaldehyde.

8
 CHAPTER IV
RESULTS AND DISCUSSION

4.1 Effect of temperature on fish drip loss

The fish that were stored in both room temperature and refrigerated room
were weighed before and after their treatment after a period of time in order to see
how storage temperature affects the weight of the fish.
Table 4.1 Drip loss of fish
Treatment Hour Drip loss (%)

2 1.07

Room temperature 5 0.29

22 -1.45

2 0.63

Refrigeration 5 0.38

22 1.27

Figure 4.1 Weight loss of fish stored in room temperature vs refrigerated

As seen in figure 4.1, the weight of the refrigerated fish will decrease with
time while the room temperature fish increases their weight with time. While the

9
refrigerated fish is as predicted, the room temperature stored fish does not match
the literature review. According to Goncalves ​et.al (2012) and Dawson ​et. al.,
(2018), both room temperature and refrigerated stored fish should have a decrease
in weight loss as time goes by due to the muscle proteins of the fish decreasing
and the WHC falling under the water content of the fish, causing water to escape.
The increase in weight loss may be due to human error as both fish and the water
excess were weighed together.

4.2 Effect of temperature on fish TVBN and TMA

TVBN and TMA values were determined at hour 0, 2, 5 and 22 for all the
treatments. This was done to observe the effects of temperature on the degree of
spoilage of fish.

Figure 4.2 TVBN analysis of fish stored in room temperature vs refrigerated

10
 Figure 4.3 TMA analysis of fish stored in room temperature vs refrigerated

As seen in the 2 figures above (Figure 4.2 and 4.3), the data collected
showed that protein breakdown in all the samples were minimal up to the fifth
hour. Normally, TVBN values would rise along with TMA values, as the TMAO
breakdown was included in TVBN along with other volatile amines. According to
Venugopal (2006), TMA rose due to the increase in breakdown rate of TMAO
that is caused by proliferation of bacteria in fish. This appears to be true as the
results at hour 22 showed that TMA content rose while remaining below TVBN
content in all treatments. At hour 22, refrigeration treatments showed to be
effective at reducing the degree of spoilage as TVBN and TMA values were
below half of the values resulting from room temperature treatments. This is in
accordance with a literature by Caballero et al​. (2003) which stated that chilled
storage minimizes microbial growth. 10-15 mg of TMA and 30-40 mg of TVBN
per 100 grams are considered to be the limits before sensory rejection occurs
(Venugopal, 2006). From the graph, it can be said that all the fish under room
temperature and refrigerated conditions remained acceptable throughout the
experiment, as the values are far below the limits aforementioned.

11
4.3 ​Effect of temperature on fish pH
The fish pH that was stored in room temperature and refrigerated room
were measured after their treatment to determine the influence of storage
temperature to the pH of the fish.

Figure 4.4 pH of fish stored in room temperature vs refrigerated

Fish in optimum condition usually have alkaline pH of 6.5 to 7 (Lie,

2008). The graph shows that the fluctuation of pH of fish stored in room
temperature drops faster compared with the refrigerated fish. The fish stored at
room temperature drops significantly in the second hour however the refrigerated
fish is stable until the fifth hour.
According to literature, one of the changes of pH of fish flesh in the post
mortem phase is caused by a shift in bacteria (Sivertsvik, 2003). The
microorganisms present in freshwater fish are usually ​Aeromonas, Alcaligenes,
Brevibacterium, Lactobacillus, and Streptococcus.​ The decrease of pH indicates
the spoilage of fish (Erkmen ​et al​. 2016). The acidification of fish tissue is caused
by the presence of lactic acid (Sen, 2005). The increase of pH in fish is caused by
ammonia and trimethylamine produced by endogenous enzymes and bacterial
spoilage (Sharifian ​et al​. 2011). The pH increase also indicates the decrease of
carbohydrate in fish (Hall, 2012).

12
4.4 Effect of temperature on fish microbial aspect
Total bacterial count was done on media inoculated with fish sample
stored in room temperature and refrigerator. The result was shown in Table 4.1 to
see the effect of storage temperature on microbiological count of fish samples.

Table 4.2 Total colony count of room temperature and refrigerated stored fish

Treatment Hour Total Plate Count (CFU/​2g​)

Fresh 0 4.5 x 10​5

2 <2.5 x 10​5 ​ (1.4 x 10​5​)

Room temperature 5 <2.5 x 10​5 ​ (4.5 x 10​4​)

22 8.6 x 10​7

2 <2.5 x 10​5 ​ (4.5 x 10​4​)

Refrigeration
5 <2.5 x 10​5 ​ (1.4 x 10​5​)

22 1.4 x 10​8

As seen in Table 4.1, fresh fish has an initial microbial count of 4.5 x 10​5​.
While the bacteria count for room temperature stored fish increases rapidly as
storage time increases. From ​1.4 x 10​5 for the second hour to ​8.6 x 10​7 in the 22nd
hour. While the microbial count for refrigerator stored fish increases slowly as the
storage time increases. From ​4.5 x 10​4 ​for a second hour increasing to ​1.4 x 10​8 ​for
22nd hour. From this result, it can be inferred that storage temperature has a
significant effect on reducing the microbial count of fish. This result matches the
literature mentioned above, which states that refrigeration of fish samples can help
decrease microbial count ​(Caballero ​et al.​ 2003).
The growth of microorganisms in fish is greatly affected by the
temperature of storage. According to Rahman (2007), signs of microbial spoilage
can even be detected at 0 to -4℃, but spoilage can be prevented by -10℃ storage

13
temperature. While the species of bacteria that may be present in fish are
Pseudomonas spp.,​ Vibrio parahaemolyticus​, and ​Listeria monocytogenes which
can grow at chill temperature. Bacteria in cold water fish is mostly Gram-negative
while bacteria in tropical fish is mostly Gram-positive (Rahman, 2007).

4.5 Effect of temperature on fish organoleptic properties

The organoleptic properties of fish with different storage temperatures
were evaluated according to the evaluation parameters in SNI 2729:2013, which
include eyes, body surface slime, meat, smell and texture by giving a score from 1
to 9. The summary of organoleptic scores were then tabulated.
Table 4.3 Organoleptic evaluation of fish
Hour Temperature Eyes Body Meat Smell Texture
Surface
Slime

0 Fresh 9 9 9 9 9

2 Room temperature 8 8 9 9 9

Refrigerated 7 8 9 9 9
temperature

5 Room temperature 6 9 7 6 8

Refrigerated 6 9 7 6 7
temperature

22 Room temperature 5 6 3 1 5

Refrigerated 9 9 7 8 9
temperature

Two graphs, each for both storage temperatures, were then created based
on the results obtained to allow more effective analysis of the effect of
temperature on the organoleptic properties of fish.

14
 Figure 4.5 Organoleptic properties of fish stored in room temperature

Figure 4.6 Organoleptic properties of fish stored in refrigeration

Based on the graphs above, it can be seen that all of the organoleptic
properties of fish stored in room temperature experience a decline, except for
body surface which experienced a slight increase in score, but later experience

15
decline again. Meanwhile, the organoleptic properties of fish stored in refrigerated
storage temperature experience a slight decline before later inclining back either
to initial state or slightly lower. This is except for the meat of fish which undergo
decline, however, the decline of the meat condition was not as steep as those of
the fish stored at room temperature. This is in accordance with ​Caballero ​et al.​
(2003), which stated that storing the fish in lower temperature, which can be done
by refrigeration, could ​minimize the microorganism and enzymatic activities in
the fish product. Lower microbial activities and enzymatic activities can slow
down deterioration of organoleptic properties.

16
 CHAPTER V
CONCLUSION

From this experiment, it is found that the Red Tilapia fish will gradually
lose weight through drip loss as times goes by. TVBN and and TMA values will
increase as time passes, however refrigeration storage helps keep both values to a
minimum due to colder storage minimizing the microbial growth and minimizes
the production of both components. This is also evident in the total plate count
where in the refrigerated stored fish, the CFU count was much smaller compared
to the room temperature stored fish. Refrigerated fish also helps retain the
organoleptic properties compared to the room temperature stored fish where all of
them experience a larger decline as the colder temperature can help minimize the
microorganism and enzymatic activities in the fish which slows down the
deterioration of organoleptic properties.

17
 BIBLIOGRAPHY

Adams, M. R. and M. O. Moss. ​Food Microbiology​. New Delhi: New Age

International, 2007.

Badan Standardisasi Nasional (BSN). “SNI 2729:2013 Ikan Segar”. Jakarta:

Badan Standardisasi Nasional, 2013.

Caballero, Benjamin, Luiz C. Trugo, and Paul M. Finglas. ​Encyclopedia of food

sciences and nutrition.​ Academic, 2003.

Dawson, Paul. Al-Jeddawi, Wesam. Remington, Nanne. “Effect of Freezing on

the Shelf Life of Salmon.” ​International Journal of Food science 3​ 40 (2018):
1-12.

Egna, Hillary S. and Claude E. Boyd. ​Dynamics of Pond Aquaculture​. Boca

Raton: CRC Press, Inc., 2017.

Evans, Judith A. ​Frozen Food Science and Technology.​ New Delhi: John WIley &
Sons. 2009.

Goncalves, Alex Augusto. Nielsen, Jette. Jessen, Fleming. “Quality of Frozen

Fish.” In the ​Handbook of Meat, Poultry and Seafood Quality. Edited by Leo
M. L. Neollet. John Wiley & Sons, 2012.

Hall, George M. ​Fish processing technology​. Springer Science & Business Media,
2012.

Kordi, M. Ghufran H. ​Budi Daya Ikan Nila di Kolam Terpal: Lebih Mudah, Lebih
Murah, Lebih Untung.​ Yogyakarta: Lily Publisher, 2010.

Mariott, G. Norman and Gill Robertson. ​Essentials of Food Sanitation.​ New

York: Springer Science & Business Media, 2012.

Obemeata, Oriakpono and Ndome Christopher. “Organoleptic Assessment and

Proximate Analysis of Stored Tilapia guineensis​.” ​Annual Review &
Research in Biology​ 2, no.2 (2012): 48-52.

​ oca Raton: CRC Press,

Øyvind, Lie. ​Improved Farmed Fish Quality and Safety. B
2006.

Rahman, M. Shafiur. Handbook of Food Preservation.​ New York: CRC Press,

2007.
Saparinto, Cahyo. ​Budi Daya Ikan di Kolam Terpal.​ Jakarta: Niaga Swadaya,
2012.

Sen, D.P. ​Advances in Fish Processing Technology. Mumbai: Allied Publishers

PVT. LTD., 2005.

Sharifian, Salim, Ebrahim Alizadeh, Mohammad Seddiq Mortazavi, and Mohsen

Shahriari Moghadam. "Effects of refrigerated storage on the microstructure
and quality of Grouper (Epinephelus coioides) fillets." ​Journal of food
science and technology​ 51, no. 5 (2014): 929-935.

Sivertsvik, M. "Active packaging in practice: Fish." ​Novel food packaging

techniques​ (2003): 384-400.

Suloma, A., H.Y. Ogata, E.S. Garibay, D.R. Chavez, and E. R. El-Haroun. “Fatty
Acid Composition of Nile Tilapia ​Oreochromis niloticus ​Muscles: A
Comparative Study with Commercially Important Tropical Freshwater Fish in
Philippines.” ​8th International Symposium on Tilapia in Aquaculture
(January 2008): 921-932.

Venugopal, V. ​Seafood Processing: Adding Value Through Quick Freezing,

Retortable Packaging, and Cook-Chilling. ​Boca Raton: CRC Press, 2006.
 APPENDIX

A. TVBN and TMA analysis

Table A.1 TVBN analysis of fish
Hour Treatment Weight Blank Volume of TVBN (mg
of fish (mL) H​2​SO​4 N/100 g)
(g) (mL)

73.19 0.3 0
0 Fresh
80 0.3 0

Room
92.24 0.4 0.2 0
temperature
2
0.3
Refrigeration 97.12 0.2 0.3 0

Room
100.06 0.3 0.3 0
temperature
5

Refrigeration 100.43 0.2 0.3 0

Room
68.3 10.7 10.7 1.393
temperature
22 0.4

Refrigeration 94.18 3.5 3.2 0.289

Refrigeration - Hour 2
TVBN (mgN/100 g)
(V 1−V 0) x 14N g/mol x (0.055 mol) x 100
= 8.333 x W
(0.3−0.3) x 14N g/mol x (0.055 mol) x 100
= 8.333 x 97.12 g

= 0 mg N/100 g of fish
Table A.2 TMA analysis of fish
Hour Treatment Weight Blank Volume of TMA (mg
of fish (mL) H​2​SO​4 N/100 g)
(g) (mL)

73.19 0.1
0 Fresh 0.012
80 0.1

Room
92.24 0.2 0.2 0.020
temperature
2
0
Refrigeration 97.12 0.2 0.2 0.019

Room
100.06 0.2 0.2 0.018
temperature
5

Refrigeration 100.43 0.2 0.2 0.018

Room
68.3 4.9 5.2 0.616
temperature
22 0.5

Refrigeration 94.18 2.1 2.5 0.177

Refrigeration - Hour 2
TMA (mgN/100 g)
(V 1−V 0) x 14N g/mol x (0.055 mol) x 100
= 8.333 x W
(0.2−0) x 14N g/mol x (0.055 mol) x 100
= 8.333 x 97.12 g

= 0.019029 mg N/100 g of fish

B. Total plate count analysis
Table B.1 Total plate count analysis of fish
Hour Treatment Dilution CFU Colony count (CFU/g)

10​-4 0/0

0 Fresh 10​-5 1/0 <2.5 x 10​6 ​(4.5 x 10​5​)

10​-6 0/0

10​-4 2/1

Room temperature 10​-5 0/0 <2.5 x 10​5 ​ (1.4 x 10​5​)

2
10​-6 0/0

10​-4 1/0

Refrigeration 10​-5 0/0 <2.5 x 10​5 ​ (4.5 x 10​4​)

10​-6 0/0

10​-4 1/0

Room temperature 10​-5 0/0 <2.5 x 10​5 ​ (4.5 x 10​4​)

5
10​-6 0/0

10​-4 1/2

Refrigeration 10​-5 0/0 <2.5 x 10​5 ​ (1.4 x 10​5​)

10​-6 0/0

10​-4 TNTC/TNTC

Room temperature 10​-5 72/54 8.6 x 10​7

22
10​-6 41/44

10​-4 TNTC/TNTC

Refrigeration 10​-5 128/98 1.4 x 10​8

10​-6 28/77

Refrigeration - Hour 2
ΣC 90 gram
= [(1x n1 )+(0.1x n1 ) x d
x 10 mL

1 90 gram
= x
[(1x1) + (0.1x0)] x 10−5 10 mL

= ​<2.5 x 10​5 ​ (4.5 x 10​4​)

C. Drip loss analysis
Table C.1 Drip loss analysis of fish
Hour Treatment Initial weight (g) Weight after Drip loss (%)
treatment (g)

2 Room temperature 237.56 235.01 1.07

Refrigeration 256.01 254.40 0.63

5 Room temperature 278.19 277.39 0.29

Refrigeration 245.49 244.55 0.38

22 Room temperature 235.94 239.36 -1.45

Refrigeration 225.21 222.36 1.27

Refrigeration - Hour 2
Initial weight (g) − W eight af ter treatment (g)
= Initial weight (g)
x 100
278.19 g − 277.39 g
= 278.19 g
x 100

= 0.29 %