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Effect of Chilling and Freezing on Fish Muscle



Muscle samples of Mystus seenghala were stored at two different low temperatures, i.e at 4±1 o C (chilled) and at -12±2 o C (frozen) for 21 days. Weekly analysis was conducted to measure protein, lipid, ash, moisture, free fatty acid, pH and total plate count. The result clearly reveals that during storage, both the samples showed a highly significant (p<0.01) decreasing trend in protein, lipid, ash and moisture content. After 21 days, the percentage decrease was 54.35% and 22.70% for protein and 82.64% and 56.68% for lipid, 10.96% and 4.89% for moisture and 38.92% and 37.88% for ash content in chilled and frozen sample respectively. However, the free fatty acid and pH showed highly significant (p<0.01) increasing trend in both the samples. Similarly, the bacteriological studies revealed that the total plate count (TPC) in chilled and frozen samples also showed an increasing trend. It was found within acceptable limits (TPC=6.04±0.11 log cfu/g) in chilled sample upto 10 th day and up to 14th day (TPC=5.78±0.2 log cfu/g) in frozen sample. Thereafter, the microbial quality further deteriorates and became inedible for human consumption.
IOSR Journal of Pharmacy and Biological Sciences (IOSRJPBS)
ISSN: 2278-3008 Volume 2, Issue 5 (Sep-Oct. 2012), PP 05-09 5 | Page
Effect of Chilling and Freezing on Fish Muscle
Roopma Gandotra, Shalini Sharma*, Meenakshi Koul and Sweta Gupta
Department of Zoology, University of Jammu, Jammu And Kashmir
Abstract: Muscle samples of Mystus seenghala were stored at two different low temperatures, i.e at 4±1oC
(chilled) and at -12±2oC (frozen) for 21 days. Weekly analysis was conducted to measure protein, lipid, ash,
moisture, free fatty acid, pH and total plate count. The result clearly reveals that during storage, both the
samples showed a highly significant (p<0.01) decreasing trend in protein, lipid, ash and moisture content. After
21 days, the percentage decrease was 54.35% and 22.70% for protein and 82.64% and 56.68% for lipid,
10.96% and 4.89% for moisture and 38.92% and 37.88% for ash content in chilled and frozen sample
respectively. However, the free fatty acid and pH showed highly significant (p<0.01) increasing trend in both
the samples. Similarly, the bacteriological studies revealed that the total plate count (TPC) in chilled and frozen
samples also showed an increasing trend. It was found within acceptable limits (TPC=6.04±0.11 log cfu/g) in
chilled sample upto 10th day and up to 14th day (TPC=5.78±0.2 log cfu/g) in frozen sample. Thereafter, the
microbial quality further deteriorates and became inedible for human consumption.
(Keywords: chiller, freezer, spoilage, acceptable limit, inedible.)
I. Introduction
Fish has been playing an important role in addressing nutritional and livelihood security of people in
the developing countries. Besides, it is good source of polyunsaturated fatty acids (PUFA’s), protein, minerals
and vitamins which are vital to our health. Although fish is highly nutritious, yet it is one of the most rapid
perishable foods because of its short shelf life. The extension of shelf life can be achieved by freezing, chilling,
salting, smoking, glazing etc. Consumers usually buy fish in bulk and store in refrigerator. Deterioration of fish
quality in refrigerator storage have great impact on the nutritious value of fish and the health of consumers.
Considering the importance from consumer view point, this study was designed to study the effect of two
different low temperatures (4±1oC and-12±2oC) on fish quality.
II. Materials And Methods
Fresh samples of Mystus seenghala were purchased from local market. They were immediately brought
to the lab in polythene bags along with crushed ice. The viscera of fish were removed; the fish was washed with
water and cut into pieces. These pieces were washed and immediately wrapped in aluminium foil, kept in air
tight plastic container and stored at 4±1°C (chilled storage) and at-12±2°C (frozen storage). Analytical
procedures for biochemical and microbiological changes were done on 0, 7th, 14th and 21st day of storage.
1.1. Biochemical composition:-
Moisture content was measured by using hot air oven and aluminium moisture dishes.
Ash content was measured by using Muffle furnace,
Total lipid content was estimated by the method of Folch et al. (1957).
Total protein content was estimated by following the method of Lowry et al. (1951).
pH was measured by the method of Keller et al. (1974) using digital pH meter.
Free fatty acid was determined by method of US Army laboratories (Natick) described by Koniecko (1979).
1.2. Bacteriological Profile
Total plate count (TPC) in the fish muscle was determined by method described by APHA (1984).
Readymade media (Hi-media) were used for the analysis. Serial ten-fold dilutions were made for inoculation.
The sample preparation was done near flame under laminar flow.
1.3. Statistical Analysis
Means and standard errors were calculated for different parameters, the data analysis was performed
using SPSS software. Differences between treatments were analysed by using independent measures one way
ANOVA. Post-hoc comparisons were conducted using Ducan’s test. The values were expressed as mean ±SE. p
value <0.05 were considered as significant and p value <0.001 were considered as highly significant.
Effect Of Chilling And Freezing On Fish Muscle 6 | Page
III. Result and Discussion:
3.1. Total Protein Content
At 4±10C, the initial value of total protein content of muscle of Mystus seenghala was found to be
18.01±0.06% and on 21st day this value was found to be least i.e. 8.22±0.2%. At -12±20C, it was found to be
18.81±0.06 % on the first day of frozen storage and on the last day i.e., 21st day of experimentation the value
was further decreased to 14.54±0.04%. The overall decreased was found to be highly significant (p≤0.01). In
present investigation, decreasing trend was observed. In chilled sample, there was 54.35% decrease, whereas in
frozen samples, there was 22.70% decrease after 21st day of storage. The present results were found to be in line
with those of Kandeepan and Biswas (2007). They found 23% decrease in total protein content of buffalo meat
at 4±10C (Chilled Storage) and 3.5% decrease at -10±10C (Frozen Storage) after 7 days. According to them, the
lower protein content of chilled meat at 4±10C was due to increased microbial growth resulted from higher
water activity (aw) and enzymatic autolysis and on frozen storage the protein content was decreased due to
protein denaturation and proteolysis induced by enzymatic activities of psychrotrophic microbial growth.
Siddique and Ali (1979) attributed the decrease in protein content of prawn during ice storage to the leaching
effect of the amino acids and water soluble proteins leaching out with melting ice. Similarly, Kolodjiejska et al.
(1987) while working on biochemical changes in fish muscle during low temperature storage found a
remarkable rate of denaturation and autolysis of fish protein. Zamir et al. (1998) calculated 21.79% decrease in
total protein content after 7 days of storage of crab meat at 7±20C. Arannilewa et al. (2005) calculated 27.94%
decrease in protein content of Tilapia (Sarotherodun galiaenus) after 60 day of frozen storage. Siddique et al.
(2011) while assessing the effect of freezing time on nutritional value of Puntius sophore, P. sarana and P.
gonionotus registered similar trends during the frozen storage at -50C of 20 days. They observed 9.44% decrease
in P. sophore, 9.94% decrease in P. sarana and 5.34% in P. gonionotus in protein content during frozen storage
of 20 days.
3.2. Total Lipid Content
At 4±10C, the total lipid content was found to be decreased from initial value of 4.84±0.015% (0 day)
to the final value of 0.84±0.04% (21st day). At -12±20C, it was estimated to be 4.94±0.03% (o day zero) and
reaches up to 2.14±0.06 % (21st day). There was 82.64% decrease in total lipid content when stored at 4±10C
and when stored at -12±20C, there was 56.68% decrease in total lipid content after 21st day. The overall decrease
found to be highly significant (p ≤0.01). These results get strong support from the findings of Kandeepan and
Biswas (2007) who conducted a similar experiment on buffalo meat. They found 47.94% decrease in chiller
(4±10C) and 17.80% decrease in freezer in total lipid content during the storage period of 7 days. They
attributed this marked decrease in chiller to the exposure of strong light, as in display cabinets, which
accelerated oxidation of fats causing discoloration. In freezer they extended the storage up to 75 days. During
this prolonged storage, the lipid oxidation occurred mainly due to losses in triglyceride fraction. Agnihotri
(1988) reported deterioration in meat lipids took place due to intermediary activities of endogenous meat
enzymes leading to hydrolysis of fat. Zamir et al. (1998) attributed the loss in lipid of crab meat stored at
refrigerator temperature 7±20C for one week due to the oxidative rancidity. Arannilewa et al. (2005) calculated
25.92% decrease in total lipid content in Tilapia after storing it in freezer compartment of the refrigerator for 60
days and associated the changes in fat content during frozen storage with the oxidation of fat. Siddique et al.
(2011) found that total lipid content decreased during frozen storage of three species of Puntius. They calculated
35.34%, 19.24% and 21.78% decrease in total lipid content in P. sophore, P. sarana and P. gonionotus
3.3. Moisture Content
At 4±10C, the initial moisture content in raw fish muscle was 88.64±0.1%. On the final day of storage,
it was observed to be 78.92±0.02%. At -12±20C, the initial moisture content in raw muscle stored in freezer was
88.04±0.04% and on 21st day of the storage, the value decreased to 83.03±0.02%. The overall decrease found to
be highly significant (p ≤0.01). Here, the percent decrease in moisture content of raw fish stored at 4±10C was
calculated to be 10.96% decrease on 21st day, while it was calculated to be 4.89% decrease at -12±20C.These
results are in agreement with those observed by Kandeepan and Biswas (2007), when they stored buffalo meat
in chiller and freezer compartment of refrigerator. Here, the percent decrease in moisture loss was found to be
5.11% in chiller and 1.57% in freezer after seven days of storage. They advocated that the more decrease in
moisture content was due to evaporation of moisture from meat in chiller, whereas the decrease in moisture
content was due to sublimation of surface water of the meat in the freezer. In freezer, the storage was extended
up to 75 days, the significant moisture losses in later storage periods was due to the myofibrillar distortion
resulting in the poor water detention ability of the meat. According to Kirschnik et al. (2006), moisture content
was constant for 14 days in samples of tail meat of the giant river prawn, (Macrobrachium rosenbergii) stored
without direct contact in ice, while in samples stored in direct contact with ice it increased approximately 6%,
Effect Of Chilling And Freezing On Fish Muscle 7 | Page
revealing absorption of water through exposed surface of meat and decrease in solid content (crude protein)
during early stages of storage in ice.
3.4. Ash Content
Fish muscle at 0 day of storage at 4±10C, the ash content was found to be 1.49±0.09% and the final
value on 21st day was found to be 0.91±0.03%. At -12±20C, it was about 1.61±0.07% on day zero and on 21st
day it was estimated about 1±0.02%.
The overall decrease found to be highly significant (p ≤0.01). Okeyo et. al. (2009) observed that the ash
context of the frozen raw Nile perch decreases with storage time. They calculated 12.69% decrease after 22 days
of ice storage. However, Kandeepan and Biswas (2007) registered 14.87% decrease in chiller and 20.66%
decrease in freezer after 7 days of storage. According to Arannilewa et al., 2005, the ash content remains almost
the same throughout the sixty days frozen storage of Tilapia. It changed from 26.13±2.20 (recorded on 0 day) to
26.80±1.44 (recorded on 60th day).
3.5. Free Fatty Acid (FFA)
At 4±10C in present studies, it was observed to be 0.5±0.01% on day 0 and 12.27±0.01% on 21st day.
At -12±20C, it was 0.57±0.02% on day zero and 5.61±0.05% on 21st day of storage. Here, more FFA content
(12.27±0.01%) was observed in chilled muscle as compared to FFA content (5.61±0.05%) in frozen muscle
after 21 days. At 4±10C, the raw fish sample was found to be near the acceptable limit (5%) on 7th day
(4.89±0.04%) and at -12±20C, the sample crossed the acceptable limit on 21st day (5.61±0.05%). Rodriquez et
al. (2007) observed increasing FFA during frozen storage farmed coho salmon (Oncorhynchus kisutch).
According to Pacheo-Aguillar et al. (2000) during post mortem period, lipid (glycerol-fatty acids esters) present
in the fish muscle undergo hydrolysis, resulting in the release of fatty acids. This is also supported by Okeyo et
al. (2009), who stated that the accumulation of FFA could be due to lipases and phospho-lipase activity in
digestive organs in muscle of Nile perch.
Table: 1 Changes in raw muscle of Mystus seenghala stored under chilled condition at 4±10 C.
Table: 2 Changes in raw muscle of Mystus seenghala stored under frozen condition at -12±20 C.
-Mean±SD with different superscript in a row differs significantly (P<0.05).
3.6. pH
In present studies, fish muscle that was stored in chiller (4±1o C) showed rapid increase in pH, while
when in freezer (-12±2o C), there was comparatively slow increase in pH. It increased from 6.8 to 7.4 in chilled
Lipid (%)
Ash (%)
Protein (%)
Lipid (%)
Moisture (%)
Ash (%)
Effect Of Chilling And Freezing On Fish Muscle 8 | Page
sample and from 6.8 to 7.1 in frozen sample during 21 days. These results are in line with those of Kandeepan
and Biswas (2007) while conducting experiment on chilled and frozen buffalo meat. Erkan and Ozden (2008)
stated that the increase was due to an increase in volatile bases from the decomposition of nitrogenous
compounds by endogenous or microbial enzymes. Obemeata et al. (2011) observed that the increase in pH was
higher in the 4oC stored sample of fish-Tilapia, than in the -18 oC stored samples, indicating that biochemical
and microbial changes are occurring faster in the fish of 4 oC stored fish.
3.7. Total Plate Count (TPC)
As recommended by International Commission on Microbiological Specification for Food,
ICMSF, (1986), an increase of total plate count (TPC) up to levels exceeding the value of 6 log CFU/g is
regarded as microbial spoiled fish muscle not fit for human consumption. In present investigation at 4±10C, the
bacterial load in raw fish muscle for period of 21 days showed considerable increase. The TPC in raw fish
muscle on day zero was rather low i.e. 2.74±0.2 log cfu/g and on 21st day of storage, it was increased further to
the final value of 9.90±0.02 log cfu/g. At -12±20C, the TPC in fish muscle on day 0 was found to be low in
2.70±0.09 log cfu/g and at the end of storage i.e. 21st day, it was found to be 7.77±0.2 log cfu/g. In present
investigation, it was clearly depicted that the microbial growth was more rapid with increasing storage
temperature. It has been reported that at -12±20C, the TPC (5.78 log cfu/g) was found to be within the
permissible limit on 14th day of storage whereas at 4±10C, the TPC (6.04 log cfu/g) crossed the permissible
limit on 7th day of storage. Lawire (1998) attributed the microbial growth to the growth promoting effect of
moisture on microbes in meat stored in chiller. Bao et al. (2007) reported a faster microbial growth in chilled
than in superchilled samples of Arctic Charr fillet under the effect of dry ice and superchilling. Liu et al. (2010)
reported increase in mesophilic bacterial load was found to be 85% in tray packed tilapia fillets stored at 00C.
They calculate initial load of 3-4 log CFU/G/g, reaching 7.4 log CFU/G/g on day 13.Obemeata et al. (2011)
showed an increase in bacterial count from 7.9×103 to 7.6×107 cfu/g when Tilapia fish was stored at 40C and
from 7.9×103 to 5.4×101cfu/g when Tilapia was stored at -180C. They stated that freezing of fish at -180C
created an unfavourable environmental condition for the growth and the survival of the micro-organisms, while
freezing at 40C allows the rapid proliferation of the micro-organisms.
IV. Conclusion
Freezing of fish muscle at -12±2o C causes comparatively lesser spoilage than chilling at 4±1o C. This
study reveals that as the storage period increases, there occurs degrading changes in biochemical and
bacteriological composition that has direct effect on shelf life and market value of the fish. Freezing of fish
creates unfavourable environmental conditions which slow down the bacterial growth and biochemical
decomposition of fish muscle, thereby increasing the shelf life; while chilling at 4±1oC allows the comparatively
rapid proliferation of bacteria, protein denaturation, lipid hydrolysis and oxidation; thereby reducing the shelf
life. Hence, for storage purposes freezing is recommended, since frozen fish muscle will have the enhanced
shelf life.
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... The pH standard of fresh fish ranged from 6.8-7.0. Gandotra et al., 2012, mentioned that cooling storage at temperature 4 °C caused the increased of pH 6.8 to pH 7.4 for 21 days. The increase of pH was associated with the increased TVB by enzymes and bacteria decomposition. ...
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As a good source of protein and other nutrition that useful for human growth, fish is categorized as perishable food that decaying quickly without good handling. In order to prevent the fish decay and to maintain the quality and freshness of the fish as long as possible, it is need good handling and storage sanitation. Chilling and freezing are the most common technology applied to prolong the shelf life of fresh fish. Chilling is one method of handling that most widely used because it can be practiced easily and quickly. This study aimed to determine the changes in the quality of fresh skipjack (Katsuwonus pelamis) during chilled storage. Parameters as quality indicator measured were total plate count (TPC), total volatile bases (TVB), and pH. Chilled storage were 0, 2and 4days. TPC of fresh skipjack during chilled storage at 0, 2 and 4 days were 1.08 x 102, 8.11 x 102 and 1.06 x 103 cfu/g, respectively. TVB of fresh skipjack during chilled storage at 0, 2 and 4 days were 9.33, 16.00 and 20.00 mgN/100g, respectively. The pH of fresh skipjack during chilled storage at 0, 2 and 4 days was 4.7, 4.8 and 5.4, respectively
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Quantitative and qualitative flesh production in the Silurus glanis species was comparatively studied between two fish groups: one from aquaculture (AG) and the other from a natural environment, the Prut River (RG). Morphometry was carried out on the fish, and then biometric and conformational indices were calculated. Better values were found in the aquaculture catfish. The Fulton coefficient was 0.82 in the Prut River fish and 0.91% in the farmed ones. The fleshy index reached 19.58% in the AG fish and 20.79% in the RG fish, suggesting better productive capabilities in the AG fish. Postslaughter, the flesh yield and its quality were assessed at different moments throughout the refrigeration period (0–15 days), and chemical compound loss occurred. In the AG samples, the water content decreased by 8.87%, proteins by 27.66%, and lipids by 29.58%. For the RG samples, the loss reached 8.59% in water, 25.16% in proteins, and 29%in lipids. By studying the fatty acids profile and sanogenic indices, good levels of PUFA (31–35%) were found, and the atherogenic index reached 0.35–0.41 while the thrombogenic index ranged between 0.22 and 0.27. Consequently, it can be stated that fish origin and especially the refrigeration period influence the flesh proximate composition and nutritional value of European catfish.
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The study was designed to investigate the effects of freezing periods and the combination of packaging with or without turmeric on the proximate composition of whole and sliced Labeo bata. The samples were subjected to 28 days of frozen (-20 °C) storage periods and analyzed for proximate composition such as moisture, protein, lipid and ash at intervals of 0, 14 and 28 days. Data obtained was subjected to analysis of variance (ANOVA) at 95% significant level. Different preservation periods and methods significantly affected the nutritional composition of fresh fish. Moisture, protein, lipid and ash content decreased with increasing storage periods when compared to the fresh fish as control. The highest percentage of protein was found in turmeric-treated whole fish preserved in polythene, C4, after the 14 th (15.70±0.14) and 28 th (15.27±0.21) day. On the other hand, the lowest percentage (14.01±0.10 and 13.22±0.18 for the 14 th and 28 th day, respectively) was found in sliced fish preserved without polythene, C5. In conclusion, the turmeric-treated whole fish preserved in polythene retained fish nutrients and ensured its good quality and composition longer than other treatments, thereby extending the shelf life during frozen storage.
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The influence of different cooling techniques (dry ice/ice packs) and storage temperature (-2°C/3°C) to prolong the shelf life of Arctic charr (Salvelinus alpinus) fillets were evaluated by sensory analysis, physical methods, chemical and microbial analysis. The effects of storage temperature were stronger than of different cooling agents. Superchilling (-2°C) of fillets packed with dry ice resulted in 6 days extension of shelf life compared to chilling (3°C). The use of dry ice parallel to superchilling prolonged shelf life for 1 day compared to fillets stored with ice packs. No negative effects on quality of the fillets where detected that could be linked to cell destruction caused by partial freezing or to sour taste, caused by absorption of CO2 gas in fish flesh.
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The objective of this study was to evaluate the shelf-life of peeled giant river prawn Macrobrachium rosenbergii stored directly in contact with ice (DCI), and without direct contact with ice (WCI). The prawns from DCI treatment showed an intense leaching of non-protein nitrogen (NPN) and total volatile bases nitrogen (TVB-N), thus suggesting that NPN or TVB-N should not be used as freshness indicators of peeled tails stored directly in contact with ice. Loss of flavor and a quick texture tactile decrease with time occurred in both treatments. The shelf-life of peeled tails prepared from M. rosenbergii was 7 days for DCI and 10 days for WCI.
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Buffalo meat is the only future remedy for nutritional security in India. If the quality gets deteriorated, the meat preserved in refrigerator would impact greatly on the health of consumers. Hence meat samples from five year old sixteen buffalo bulls were analyzed in the fresh state (0 day) and after 4 and 7 days in chiller (4±1°C) and 4, 7, 14, 30, 60 and 75 days in freezer (-10±1°C) in a domestic refrigerator. The values of ERV, WHC and proximate composition decreased with increasing storage period. Whereas pH, TBA no., tyrosine value, chilling loss and drip loss showed an increasing trend. The chiller storage increased but freezer decreased the microbial counts (SPC, PC and Coliforms). The values of odour and flavour scores decreased with increasing storage period. Whereas, texture, tenderness and juiciness scores showed an increasing trend. Thus it was concluded that a storage period upto 4 days in chiller and 30 days in freezer could satisfactorily maintain the buffalo meat quality.
Drying characteristics were evaluated for summer sausages (50% beef, 50% pork) prepared with three meat particle sizes obtained through grinding variations. Increases of chemical components (protein, fat, ash, salt, lactic acid) during 45 days of drying were dependent on the rate of moisture removal from sausages. Summer sausage produced with a 9 mm grinder plate for the pork and a 6 mm plate for beef (9-6 grinding combination) had a 34% shrinkage at 45 days, whereas sausages of a 3-6 and a 6-6 grind combination had shrinkages of 37% and 40%, respectively. The rate of moisture removal for an all beef summer sausage was lower for larger diameter sausage when 52, 62, and 73 mm sizes were compared. Moisture content of the outer one third radius portion of the sausages was 5 to 7% lower than the moisture content of the inner two thirds radius portion from 5 days through 45 days of drying. Both types of summer sausages (beef pork and all beef) having greater than 1.2 kg/cm 2 of shearing force were generally of poor eating and slicing quality because of the dried fibrous condition of the meat.
The quality deterioration of tray-packed tilapia (genetically improved farmed tilapia strain of Oreochromis niloticus) fillets stored at 0°C were studied by integrated evaluations of sensory, microbiological, biochemical and physical analysis, in order to expound the mechanism of fish spoilage and develop the most reliable indicators for quality assessment. The results showed that four quality index as Pseudomonas counts, total volatile basic nitrogen (TVBN), cadaverine (CAD) and K value were highly correlated (r > 0.90) with storage time and sensory acceptability. Protein degradation was visible on SDS-PAGE when microbiological load exceeded 6 log cfu/g. Thiobarbituric acid reactive substances (TBARS) value remained at a very low level throughout the storage, suggesting low lipid oxidation in muscle. Hardness decrease tested by texture machine was consistent with texture softening of fillets in the sensory evaluation. Considering fish freshness and microbiological safety, the shelf life of tilapia fillets stored at 0°C was approximately 10 - 12 days.
The quality and shelf life of whole ungutted and gutted sardines (Sardina pilchardus) stored in ice were studied. The changes in the fish were investigated by sensory assessments, chemical analyses and microbiological analyses. The sensory scores of uneviscerated and gutted sardines stored in ice at +4 °C were 7 days. The chemical indicators of spoilage, total volatile basic nitrogen and trimethylamine values of gutted sardine increased very slowly, whereas for whole ungutted samples higher values were obtained reaching a final value of 15.03–29.23 mg per 100 g and 2.36–4.16 mg per 100 g, respectively (day 9). Peroxide and thiobarbituric acid values remained lower for whole ungutted sardine samples until day 9 of storage, whereas for gutted fish were higher. The level of histamine exceeded the legal limit in whole ungutted sardine after 7 days of storage in ice, during which sardines were rejected by the sensory panel. Mesophilic aerobic bacteria count, H2S-producing bacteria, sulphide reducing anaerobe Clostridias, Enterobacteriaceae count of whole ungutted sardine samples are higher than gutted sardine samples during the storage. Psychrotrophic bacteria counts of the two groups were not different. The limits of microbiological data were not exceeded throughout the storage in both the groups’ samples.
Postmortem changes of sardine muscle during 15 d storage at 0 °C were studied to evaluate its quality and functionality. No microbial deterioration was detected since trimethylamine and histamine concentration remained low with final values of ≤ 1.62 mg/100g and 0.00018 ppm, respectively. A final proteolytic activity ≤ 20 μg Tyrosine/min/g protein was detected. Lipid oxidation from moderate to advanced was detected after day 5 with values of 31.8 to 33.9 meq/kg and 26 mg/kg for peroxide value and thiobarbituric acid value respectively. Muscle protein showed no gel-forming ability. Extraction of myofibrillar protein decreased 45% and 81% at day 5 and 15 respectively. Overall results indicated that good quality was maintained during the storage period with a final K value of 50.7% when proper handling practices were implemented.
Since 1922 when Wu proposed the use of the Folin phenol reagent for the measurement of proteins (l), a number of modified analytical pro- cedures ut.ilizing this reagent have been reported for the determination of proteins in serum (2-G), in antigen-antibody precipitates (7-9), and in insulin (10). Although the reagent would seem to be recommended by its great sen- sitivity and the simplicity of procedure possible with its use, it has not found great favor for general biochemical purposes. In the belief that this reagent, nevertheless, has considerable merit for certain application, but that its peculiarities and limitations need to be understood for its fullest exploitation, it has been studied with regard t.o effects of variations in pH, time of reaction, and concentration of react- ants, permissible levels of reagents commonly used in handling proteins, and interfering subst.ances. Procedures are described for measuring pro- tein in solution or after precipitation wit,h acids or other agents, and for the determination of as little as 0.2 y of protein.