Collagen content in farmed Atlantic salmon (Salmo salar,L.) and subsequent changes in solubility during storage on ice

To elucidate whether collagen is an important factor for fish flesh quality, the collagen content and its changes in solubility during storage on ice in muscle of farmed Atlantic salmon (Salmo salar, L.) were measured. The contents of acidsoluble, pepsin-soluble and insoluble collagen in white muscle were determined in fresh fish muscle and at the end of 5, 10 and 15 days storage on ice. Total collagen was found to be 0.66% of fresh weight, with a relative distribution of 6% acidsoluble, 93% pepsin-soluble and 1% insoluble collagen. During storage on ice, a progressive change in solubility of muscle collagen was found. For insoluble collagen, significantly lower values were detected at day 15 compared to day 0. A minor, but even increase in acid-soluble collagen was found from day 0, while no changes were seen in pepsin-soluble collagen during storage. These results show that collagen fibres of farmed Atlantic salmon have a high solubility in acid and salt solutions and contain few cross-links. Some cleavage of intermolecular cross-links seems to occur during storage on ice.     

Pasteurization of Pacific Oysters by Radiation: Post-Mortem Changes in Nucleotides During Storage at 0-2°C

SUMMARY: The concentration of nucleotides was lower in the adductor muscle of the oyster (1.64 μmoles/g) than in the remaining dark tissues of the oyster (2.75 μmoles/g). The concentration was less in the whole oyster meats (2.87 μmoles/g) than is usually found in fish muscle, or other marine invertebrates.
In addition to the adenine nucleotides and inosine monophosphate, uridine triphosphate, guanosine triphosphate, guanosine diphosphate, guanosine monophosphate and guanosine diphosphate-mannose were found in the fresh oysters. Samples collected in summer had greater concentrations of nucleotides than similar winter samples. lnosine monophosphate formed rapidly from adenosine triphosphate during storage at 0°–2°C, while the turnover rate of inosine monophosphate was slow and reflected low 5'-nucleotidase activity. Hypoxanthine, inosine, guanosine, guanine and uracil were formed during ice storage. The nucleotide breakdown in oysters was not changed by 2 mrads of radiation dose. Total nucleosides and free bases increased during storage of both unirradiated and irradiated samples. During the latter part of the storage period the concentrations of nucleosides and free bases were considerably greater in the irradiated samples. This difference probably is due to the utilization of these compounds by bacteria in the un-irradiated samples. After 15 days of storage bacteria had increased to more than 10'organisms per g, while the counts for irradiated samples were very low (less than 103 per g).     

CHANGES IN FRESHNESS OF CHILIPEPPER ROCKFISH (SEBASTES GOODEI) DURING STORAGE AS MEASURED BY CHEMICAL SENSORS AND BIOSENSORS

ABSTRACT Trimethylamine (TMA) and ammonia contents in Chilipepper rockfishes were determined by ion specific electrodes, as a late indicator of fish freshness. After 9 days of storage in ice, TMA contents significantly increased, indicating that bacterial spoilage was in progress. The pattern of changes in ammonia contents was similar to that of TMA. Determination of ATP degradation products in Chilipepper rockfish by HPLC showed that AMP and hypoxanthine levels were low and did not change much during storage. The concentration of IMP initially increased and then continuously decreased as inosine accumulated. Only trace amounts of hypoxanthine were detected in rockfish tissues. Chilipepper rockfish appears to differ from other Sebastes species in that ATP degradation results in inosine accumulation rather than hypoxanthine.     

EFFECT OF DIETARY COMPONENTS AND SURFACE APPLICATION OF OLEORESIN ROSEMARY ON LIPID STABILITY OF RAINBOW TROUT (Oncorhynchus mykiss) MUSCLE DURING REFRIGERATED AND FROZEN STORAGE

The effect of dietary supplementation with α-tocopherol and surface application of oleoresin rosemary on the lipid stability of rainbow trout ( Oncorhynchus mykiss ) muscle during refrigerated storage (4C) and frozen storage (−20C) for 8 days and 6 months, respectively, was investigated. A significant (p < 0.05) reduction in α-tocopherol concentrations in muscle tissue during frozen storage was observed. The 2-thiobarbituric acid-reactive substances (TBARS) values of muscle samples, with or without surface application of oleoresin rosemary, increased during storage for all dietary groups. A significant (P < 0.05) interaction between dietary treatments and the time of storage was observed. Muscle tissue from fish receiving a dietary supplement of 500 mg/kg diet had the lowest TBARS values. The oxidative stability of the lipids was further improved by applying oleoresin rosemary to the surface of the fish fillets.     

INTEGRATED STUDIES ON THE FRESHNESS OF RAINBOW TROUT (ONCORHYNCHUS MYKISS WALBAUM) POSTMORTEM DURING CHILLED AND FROZEN STORAGE

Rainbow trout were killed by two methods, asphyxiation and clubbing. The concentration of ATP in specimens of skeletal muscle taken immediately after death was significantly (P<0.01) higher in clubbed (4.41 ± 0.86 μmol/g) than in asphyxiated (2.00 ± 0.69 μmol/g) fish. The shear force (Warner-Bratzler) required to cut the muscle was higher (P<0.05) in clubbed (8.33 ± 0.61 N) than in asphyxiated (6.85 ± 0.98 N) fish. Changes in the concentration of adenine nucleotides and in shear force were measured at intervals during storage at 3C and - 30C. The K value was calculated and was found to be correlated inversely with changes in shear force, Torrymeter readings and sensory assessment. There were no significant differences in the concentrations of ATP and metabolites between muscle sites. There were no differences in shear force measurements between the locations sampled nor between muscle taken from the right and left sides of the fish.     

ISOLATION OF INTACT TYPE V COLLAGEN FROM FISH INTRAMUSCULAR CONNECTIVE TISSUE

A purification method for intact type V collagen from fish intramuscular connective tissue was developed. Collagens were extracted with 0.5 M acetic acid from fish intramuscular connective tissue. Intact type V collagen could not be isolated by conventional salt fractionation which has been used for isolation of pepsinized collagens. Isolation was achieved by ion-exchange chromatography in the presence of 5 M urea. This is the first report on isolation of intact type V collagen from muscle tissue.     

Changes in Free Amino Acids and Creatine Contents in Yellowtail (Seriola quinqueradiata) Muscle during Ice Storage

Free amino acids (FAA) and creatine contents of white and dark muscles of yellowtail (Seriola quinqueradiata) were measured and compared during ice storage. White muscle contained a large quantity of free histidine (1,160 mg/100g), while an equally large amount of taurine (1,150 mg/100g) was present in the dark muscle. Little change in the levels of most FAA occurred in the white muscle during ice storage for over 40 days but in the dark muscle the levels of almost all FAA except taurine increased significantly over a period of 33 days. Creatine was present in higher concentration in the white than in the dark muscle (510 vs 170 mg/100g), while its level did not change in either muscle during the entire storage period.     

IDENTIFICATION OF TWO MATRIX METALLOPROTEINASES IN THE SKELETAL MUSCLE OF PACIFIC ROCKFISH (SEBASTES SP.)

Two heat-stable metalloproteinases with collagenase activity have been identified in the skeletal muscle of Pacific rockfish (Sebastes sp.) and partially characterized. The 47 kDa and 95 kDa enzymes appear to be similar to mammalian matrix metalloproteinases (MMPs), with respect to molecular weight, pH-activity profile, substrate specificity, dependence on calcium for activity, and inhibition-activation profiles. The fish metalloproteinases readily hydrolyzed collagen and gelatin, but not β- or K-casein, in SDS-PAGE substrate zymograms. They also hydrolyzed collagen and gelatin in solution, but showed very low activity in solubilizing native fibrillar collagen. They did not hydrolyze solutions of β- or K-casein, azocasein, hide powder azure or synthetic substrates for trypsin, chymotrypsin or collagenase. To our knowledge, this is the first report where fish muscle proteinases have been identified as possible members of the family of MMPs. The rockfish muscle MMPs appear to be different from other currently identified fish muscle proteinases in regard to molecular weight, substrate specificity and activation-inhibition profiles. These two enzymes could be partly responsible for the degradation of collagen and other extracellular matrix proteins in fish muscle and for the texture softening of seafood products.     

Peptides in rainbow trout (Oncorhynchus mykiss) muscle subjected to ice storage and cooking

Although proteolysis during post mortem storage is an important factor affecting fish texture, little is known about degradation end-products. This study was performed to investigate the occurrence of low molecular weight peptides (<5 kDa) in post mortem rainbow trout muscle, during ice storage, and to evaluate their stability during cooking. It combined quantitative (amino acid analysis) and qualitative approaches (mass spectrometry). The results showed that muscle of trout was poor in peptides. These were mainly anserine and glutathione. Their concentration was almost unaffected by the seven days of ice storage and vacuum cooking for 5 min at 70 °C. MS analysis revealed a limited but highly reproducible appearance of small peptides in trout muscle during the ice storage and after cooking. The compounds detected by MS analysis remain to be characterised.     

Changes in Electrophoretic Patterns of Gadoid and Non-gadoid Fish Muscle during Frozen Storage

Sodium dodecyl sulfate polyacrylamidc gel electrophoresis of fish muscle during frozen storage showed that a crosslinked protein of 280,000 daltons formed with gadoids, whereas this band did not occur with non-gadoid fish species. For cod, 20 days at -7°C are needed for the crosslink to appear, whereas for whiting only 3 days arc needed. Formaldehyde (FA) may be responsible for the crosslinking. Adding ascorbic acid to cod mince prior to freezing accelerated dimcthylamine and FA formation during frozen storage and accelerated the formation of the crosslinked protein, while leaving cod fillets on ice for 10 days prior to freezing, reduced the Instron hardness and the crosslinked protein did not appear with 50 days of frozen storage at -7°C.     

STRUCTURE AND SOLUBILITY OF COLLAGEN AND GLYCOSAMINOGLYCANS IN TWO BOVINE MUSCLES WITH DIFFERENT TEXTURAL PROPERTIES

Meat samples were collected from two bovine muscles , psoas major and semitendinosus, exhibiting different textural properties as measured by Warner Bratzler Shear Force. The structure and composition of the intramuscular connective tissue was compared by transmission electron microscopy, fibril diameter measurements, statistical analysis, collagen and glycosaminoglycan solubility studies in acetic acid and acetic acid added pepsin. The mean fibril diameter was significantly larger in the tougher muscle and the fibrils were tightly bundled into fibers. The tender muscle exhibited mainly randomly oriented fibrils with more space in between. The biochemical study confirmed the microscopy, showing a higher hexuronic acid/hydroxyproline ratio in the tender muscle, mainly in pepsin digested samples. The content of hydroxyproline after defatting in acetone was 30% higher in ST. No major differences were seen in the proportion of acid labile or stable cross-links in the collagen of the tough and tender muscle.     

Relationship between lysyl oxidase activity,pyridinoline content and muscle texture during ice storage of jumbo squid (Dosidicus gigas)

Collagen fibres, stabilised by lysyl oxidase (LOX), play an important role in jumbo squid because they are responsible for the union between various cells; therefore, a close interdependence between their functions and muscle firmness during ice storage has been suggested. In this study, the relationship between LOX activity, pyridinoline (Pyr) content and muscle texture (SF) during ice storage of jumbo squid mantle was evaluated. LOX activity was confirmed within the range of 4.1–7.1 × 10^?3 U g^?1 of protein, leading to an increase in Pyr content, detected in the range of 0.85–1.32 mmol mol^?1 of collagen after 5–20 days. The SF of the muscle became harder during the ice storage time, increasing from 21.08 to 37.95 N. It was therefore possible to establish the relationship between LOX activity, collagen cross‐links (Pyr content) and texture patterns during ice storage of jumbo squid muscle, which increased after 20 days.     

Preservation of iced refrigerated sea bream (Sparus aurata) by irradiation: microbiological, chemical and sensory attributes

Quality and shelf life of non-irradiated and irradiated (2.5 and 5 kGy) sea bream in ice conditions and stored at +4 °C were investigated by measurement of microbiological, chemical and sensory analysis. Microbial counts for non-irradiated sea bream samples were higher than respective irradiated fish. Total volatile base nitrogen (TVB-N) values increased value of 38.64 mg/100 g for non-irradiated, sea bream during iced storage whereas for irradiated fish lower values of 13.48 and 12.06 mg/100 g were recorded at 2.5 and 5 kGy, respectively (day 19). Trimethylamine (TMA-N) values and thiobarbituric acid (TBA) values for irradiated samples were lower than non-irradiated samples. Acceptability scores for odour, taste and texture of cooked decreased with storage time. The sensory scores of sea bream stored in control and 2.5–5 kGy at +4 °C were 13 and 15 days, respectively. The results obtained from this study showed that the shelf life of sea bream stored in ice, as determined by overall acceptability all data, is 13 days for non-irradiated sea bream and 15 days for 2.5 kGy irradiated and 17 days for 5 kGy irradiated sea bream.     

DETERMINATION OF COLLAGEN CROSSLINKS IN ROCKFISH SKELETAL MUSCLE

The low collagen content in fish muscle makes the quantitative analysis of collagen crosslinks difficult. We have developed a method for the quantitative determination of reducible difunctional collagen crosslinks in samples containing as little as 0.5 picomoles of crosslink per microgram of hydroxyproline. The low collagen-containing tissue is first enriched by removing most of the non-collagenous protein with cold sodium hydroxide solution. The lyophilized alkali-insoluble extract is then reduced with NaB³H₄ and acid-hydrolyzed. Initial fractionation of hydrolysates on a Bio-Gel P-2 gel filtration column provides partial separation of components with a sharp peak of radioactivity containing more than 90% of the difunctional crosslinks. This peak is ultimately analyzed by HPLC for the quantitative determination of specific difunctional crosslinks. This method has been used to show that the difunctional crosslink composition of muscle collagen from two species of Pacific rockfish differs quantitatively from each other and from that of rat tail tendon.     

The effect of storage on ice and various freezing treatments on enzyme leakage in muscle tissue of rainbow trout (Oncorhynchus mykiss)

In order to study the biochemical changes in fish muscle during ice storage and freezing-thawing processes, the activities of certain marker enzymes in the cell interstitial fluid from muscle tissue of rainbow trout (Oncorhynchus mykiss) were measured. The enzymes analysed were: lysosomal α-glucosidase (E.C. 3.2.1.20), β-N-glucosaminidase (E.C. 3.2.1.30) and acid phosphatase (E.C. 3.1.3.2). The activity in centrifuged tissue fluid (CTF) was compared with the activity in total homogenate. When ice storage was varied between 3 and 14 days, it did not affect enzyme leakage into the CTF significantly. However, there was a distinct difference between fresh fish and fish iced even for only 1 day, which gave increased leakage of marker enzymes. When the ice-stored samples were subject to a freezing-thawing cycle they showed a marked increase in enzyme activity in the press juice. When the freezing process was varied so as to achieve different freezing rates, the slowest freezing rate caused the highest enzyme leakage.     

Histamine Formation and Bacterial Spoilage of Albacore Harvested off the U.S. Northwest Coast

Iced and previously frozen albacore were monitored for histamine formation and bacterial growth during storage at 0–37°C. The optimum temperature for histamine formation in albacore tuna (Thunnus alalunga) was 25°C, and whole fish were more susceptible to histamine formation than dressed fish at that temperature. Storage at 25°C resulted in the highest histamine level, 60.4 mg/100g in whole fish stored for 7 days. When albacore were frozen prior to storage, reduced amount of histamine was found at 7.14 mg/100 g after 7 day storage at 25°C, only after decomposition became obvious. No histamine was found in any of the albacore samples stored in ice for 18 days.     

Quantitative changes in bacteria, amino acids and biogenic amines in sardine (Sardina pilchardus) stored at ambient temperature (25–28°C) and in ice

Freshly caught sardines contained high levels of bacteria located mainly on the skin and the gills. These bacteria invaded and grew rapidly in sardine muscle, reaching 5x10⁸ c.f.u. g^-1 and 6x10⁸ c.f.u. g^-1 respectively after 24h at ambient temperature and 8 days in ice.
Histidine, arginine, lysine, tyrosine and methionine levels decreased during storage. The other amino acids, except proline and taurine, accumulated in the fish muscle, indicating an extensive proteolysis.
Histamine, cadaverine and putrescine accumulated to levels of 2350ppm, 1050ppm and 300ppm respectively, after 24h storage at ambient temperature. Histamine and cadaverine reached similar levels after 8 days storage in ice, whereas putrescine formation was insignificant. Spermidine and spermine levels increased slightly under ambient conditions.
Salting the fish at 8% delayed bacterial and chemical changes but only in iced sardines.
The high content of free histidine found in sardines and the susceptibility of its muscle to histamine and cadaverine formation could explain its increasing implication in incidents of histamine poisoning.     

Properties of actomyosin isolated from cod (Gadus morhua) after various periods of storage in ice

Natural actomyosin was isolated from cod (Gadus morhua L) stored in ice for up to 28 days. The gelling properties, apparent viscosity, Ca^2--ATPase activity and component protein composition by sodium dodecyl sulphate (SDS) electrophoresis were determined for each preparation of natural actomyosin. The apparent viscosity, protease activity, trimethylamine (TMA) content and pH of the fish muscle were also determined. The results showed that the apparent viscosity and Ca^2--ATPase activity tended to decrease slightly during ageing of the fish in ice, whereas some of the gelling properties showed a maximum between 3 and 6 days of storage. However, there was no change in the apparent viscosity of the muscle as a whole even after the fish were considered to be stale according to the TMA values. The ratio of myosin heavy chain to actin in the actomyosin changed with the time of storage of the fish, being highest at 3 days when gelling properties were maximal and decreasing progressively thereafter.     

The impact of collagen on softening of grass carp (Ctenopharyngodon idella) fillets stored under superchilled and ice storage

The objective of this work was to study the impact of collagen on softening of grass carp (Ctenopharyngodon idella) fillets during chilled storage. The fillets were stored under superchilling (?1.5 ± 0.2 °C) and with ice (0.2 ± 0.1 °C) for 21 days, and texture properties, collagen and the related enzyme activities were measured. Results showed that firmness and collagen content were strongly influenced by storage temperature and time. Fillet firmness decreased by 32.3% (superchilling) and 49.6% (ice stored) of the initial values after 3 days of storage, respectively. Total collagen content decreased with time, but different collagen fractions showed variations. Collagen degraded to different extents depending on storage conditions as indicated by SDS‐PAGE and amino acid analysis. In addition, collagenase activity declined significantly during the first 3 days, followed by a slow increase. This study demonstrated that collagen degradation was involved in grass carp fillet softening and provided useful information for fillet quality improvement.     

BACTERIAL FORMATION OF HISTAMINE IN JACK MACKEREL (TRACHURUS SYMMETRICUS)

Peak histamine concentrations of 0.023, 0.031 and 0.027 g histamine/100 g muscle and maximal bacteria concentrations of 1.75, 1.59 and 0.423 g dry cells/100 g muscle were observed in muscles of jack mackerel stored at 25, 15 and 5C, respectively. Incubated fish homogenates suggest rate and transport limitations in histamine formation in muscle. The Mulchandani model predicted bacterial growth in muscle. The Luedeking and Piret expression fitted histamine formation in muscle; α values were 3.0 × 10⁻³, 1.23 × 10⁻² and 4.17 × 10⁻² g histamine/g dry cells, while β-values were 4.5 × 104, 8.0 × 10⁻⁵ and 0 g histamine/g dry cells × h at 25, 15, and 5C, respectively. The model predicts that jack mackerel could be stored from 4.5 to 5.5 days in ice, from 1 to 2 days at 15C and from 17 h to 2 days at 25C before fishmeal quality might be affected.