Biochemical and Physicochemical Properties of Actomyosin from Pre- and Post-spawned Hake (Merluccius hubbsi) Stored on Ice

Biochemical and physicochemical properties of natural actomyosin (NAM) from pre- and post-spawned hake stored on ice were studied. At the beginning of storage both reduced viscosity and ATPases activities of NAM from post-spawned hake were about 3 times higher than those of the pre-spawned ones, except for the Mg²⁺-(Ca²⁺)-ATPase. Higher loss of functional components during pre-spawned hake NAM purification was found compared to NAM from post-spawned fish. Reduced viscosity and Mg²⁺-(EGTA)-ATPase activity of NAM from both pre- and post-spawned hake decreased following ice storage. SDS-PAGE patterns of NAM from pre-spawned hake showed an absence of the myosin heavy chain and the presence of a 160 kDa component at zero time of storage. No proteolysis occurred in ice-stored fish caught in both gonadal stages.     

Biochemical and Functional Properties of Myofibrils from Preand Post-Spawned Hake (Merluccius hubbsi Marini) Stored on Ice

Enzymatic activities assayed at beginning of storage in myofibrils from post-spawned hake were 3 X those in myofibrils from pre-spawned hake. Ca²⁺ sensitivity of myofibrils from pre-spawned hake was 40% less than that of myofibrils from post-spawned hake. The profiles of SDS-PAGE gels of pre-spawned myofibrils at beginning of storage showed a partially denatured myosin heavy chain, and polypeptide bands under myosin heavy chain. They probably represent proteolytic fragments produced by degradation of MHC in vivo. No proteolysis was detected in myofibrils during storage. These results help predict functional properties of fish proteins and changes during storage.     

Biochemical and physicochemical properties of actomyosin from pre- and post-spawned flounder (Paralichtys patagonicus) stored on ice

The biochemical and physicochemical properties of actomyosin from pre- and post-spawned flounder stored on ice were studied. Irrespective of the gonadal condition of the fish, a gradual decrease in reduced viscosity, Mg²⁺ATPase and Mg²⁺Ca²⁺ATPase activities of actomyosin was observed. A similar decrease in the Mg²⁺ EGTA ATPase activity of actomyosin from post-spawned flounder was also observed. The decrease in enzymatic activities was accompanied by an increase in surface hydrophobicity of the protein. No signs of proteolysis of the major components of the actomyosin complex were detected. The relative percentage of myosin decreased and that of actin increased in actomyosin from pre- and post-spawned flounder stored on ice. These changes occur earlier in actomyosin from prespawned flounder. The results of this paper indicate that actomyosin from flounder denatured during fish storage on ice and suggest that this denaturation is due to structural and conformational changes in myosin which led to aggregation of this protein.     

Influence of Gonadal Stage of Hake (Merluccius Hubbsi Marini) on Biochemical Properties of Myofibrils Stored at 2 to 4 °C

ABSTRACT: The influence of the gonadal stage of hake on the biochemical properties of myofibrils stored at 2 to 4 °C was studied. At 0 time and during storage, both Mg²⁺-Ca²⁺-ATPase activity and Ca²⁺ sensitivity of myofibrils from post-spawned hake were significantly higher (p < 0.01) than those of pre-spawned fish. The profiles of SDS-PAGE gels of unstored and stored myofibrils from pre-spawned hake showed a partially denatured myosin heavy chain. The actin-myosin ratio in myofibrils from pre-spawned hake was significantly lower (p < 0.01) than the ratio in post-spawned hake. Irrespective of the gonadal condition of fish, no changes in the myosin-actin ratio of stored myofibrils were observed.     

Proteolytic Activity of Muscle in Pre- and Post-Spawning Hake (Merluccius hubbsi Marini) After Frozen Storage

The proteolytic activity of fillets from fresh fish and after frozen storage of hake muscle from pre- and post-spawned fish was investigated. The autolytic activity of fresh muscle extracts from pre-spawned muscle extracts was significantly higher than that of post-spawned, except for pH 3.4 and incubation temperature of 60 °C where no significant difference was found. In both gonadal conditions, the freezing of hake fillets affected the proteolytic activity of crude muscle extracts. After 50 d of frozen storage, the proteolytic activity values of crude muscle extracts remained higher than the fresh values, except for pre-spawned extracts incubated at 37 °C. At longer storage period (110 d), the proteolytic activity continued with similar or higher values compared with those observed for fresh muscle extracts. Densitometric analysis of crude muscle extracts indicated degradation of MHC in fresh and frozen storage at both gonadal conditions under incubation conditions of pH 6.2 and temperatures 37 and 60 °C, pH 8.5 and temperatures 37 and 60 °C, pH 3.4 and temperature 4 °C.     

Gelling Properties of Actomyosin from Pre- and Post-Spawning Hake (Merluccius hubbsi)

The biochemical properties and the characteristics of heat-induced gelation of natural actomyosin (NAM) from pre- and post-spawning hake were studied. Mg²⁺ ATPase activity, reduced viscosity and myosin/actin mole ratio of NAM from post-spawning fish were higher than those of pre-spawning ones. Gelation of both actomyosin at 10 mg mL^?1 of protein concentration was optimal at 60°C and pH 6.0. The highest rigidity was reached at 0.40M and 0.44M KCl with NAM from pre and post-spawning hake, respectively. Irrespective of heating temperature, ionic strength conditions and at pH range 5.5–7.5, rigidity of post-spawning hake NAM gels was higher than those of pre-spawning fish. Scanning electron micrographs of pre- and post-spawning hake NAM showed “actin-type” and “myosin-type” ultrastructures, respectively.     

Physicochemical Changes in Pacific Whiting Muscle Proteins during Iced Storage

No changes in actomyosin Ca²⁺-, Mg²⁺-, or Mg²⁺-Ca²⁺-ATPase activities were observed during iced storage of Pacific whiting fillets, but Mg²⁺-EGTA-ATPase increased with a loss of Ca²⁺-sensitivity. Surface hydrophobicity of actomyosin increased substantially within 2 days, but not total sulfhydryl (SH) content. During longer storage, the SH content decreased gradually, but surface hydrophobicity remained constant. Autolytic degradation products increased in fish muscle with storage time. Myosin heavy chain (MHC) was degraded by 45% within 8 days, but no noticeable difference was observed in actin. Results indicated that autolysis may be the main cause of physicochemical changes in Pacific whiting muscle proteins during iced storage.     

Biochemical Properties of Actomyosin of Cold Stored Striated Adductor Muscles of Aulacomya ater ater (Molina)

Expressible juice showed highest increase within the second day of storage. Actomyosin was partially purified from stored muscles. Both reduced viscosity and Mg²⁺-ATPase activity of actomyosin decreased about 44% in the first day. These changes were due neither to actomyosin dissociation nor fragmentation of major proteins of the complex. Relative percentage of myosin decreased, and actin increased in actomyosin after the second day. These changes are related to a decrease in the myosin heavy chain and could explain the slow decrease in reduced viscosity and Mg²⁺-ATPase of this protein after the second day.     

Conformational Change in Actomyosin from Post-spawned Hake Stored on Ice

The changes that lead to a decrease in the viscosity of actomyosin of post-spawned hake stored on ice were investigated. Surface hydro-phobicity of actomyosin from hake increased significantly in the first 3 days of storage. Compared with that stored 3 days, the viscosity of actomyosin from day 0 was greater and its changes suggested greater hydrophylic surface area. Electron microscopy showed changes in the filamentous structure of actomyosin during ice storage. Actomyosin from hake stored in ice is hypothesized to undergo conformational changes including exposure of hydrophobic groups resulting in drastic changes in hydrodynamic properties and a loss of the characteristic arrowhead structure of fresh acotmysin.     

Properties of Kamaboko Made From Red Hake (Urophycis chuss) Fillets, Mince, or Surimi

Fresh red hake (Urophycis chuss) was randomly assigned to three treatments: whole fillets, minced flesh, and surimi. These treatments were stored at −5°C for 10 wk and tested for salt-extractable protein, actomyosin, and Ca⁺⁺– ATPase activity prior to the manufacture of Kamaboko. Kamaboko quality was determined by the fold test and percent expressible fluid. Salt-extractable protein, actomyosin, and Ca⁺⁺– ATPase activity all decreased throughout the storage period and were found to be good measures of the ultimate quality of Kamaboko. The data suggest red hake is suitable for the manufacture of surimi which can be stored frozen and subsequently manufactured into an acceptable Kamaboko.     

Effect of post-rigor fish storage on ice on physicochemical properties of actomyosin

Actomyosin was purified from post-rigor fish stored for various times on ice and was characterised by Mg²⁺-ATPase activity, ATP-sensitivity and SDS-polyacrylamide gel electrophoretic patterns. It had a similar reduced viscosity to that of the myofibrillar extract and the viscosity of both extracts fell in a similar manner as the time of storage of the whole fish on ice increased. No changes in the extractability, the Mg²⁺-ATPase activity or the electrophoretic pattern of actomyosin were detected when extracted from fish muscle held for up to 11 days in ice.     

Extra-Cold Storage of Hake and Mackerel Fillets and Mince

Extra-cold storage (?30 and ?40°C) of mackerel (Scomber scombras) mince and fillets showed lower free fatty acid formation. Extra-cold storage (-30°C) of white hake (Urophycti tenuis) fillets produced fish with better quality based on sensory and chemical indices. The colder the storage temperature, the less firm the hake mince and fillets. Ascorbic acid accelerated cohesiveness development of mackerel mince and fillets. Over time, the quality of the hake and mackerel decreased according to sensory and chemical indices. They became tougher and generally more cohesive.     

Total volatile acids content as a quality index of irradiated hake (Merluccius merluccius hubbsii) fillets

Irradiation of hake (Merluccius merluccius hubbsii) fillets with 500 krad (5 k Gy) of Co⁶⁰ gamma rays caused a delay in the formation of volatile acids during storage under refrigeration at 4–5°C. Loss in organoleptic acceptability could be correlated with the increase in volatile acids both in the unirradiated and irradiated fish stored at 4–5°C, suggesting that the volatile acids number (VAN) can be a useful tool for the quality evaluation of hake fillets.     

Comparison of gilthead sea bream (Sparus aurata) and hake (Merluccius merluccius) muscle proteins during iced and frozen storage

Gilthead sea bream (Sparus aurata) and hake (Merluccius merluccius) muscle behave differently during storage, whether in ice or deep frozen. Rapid changes have been observed in the texture of hake muscle during frozen storage, while gilthead sea bream has proved to be more stable. In order to ascertain the role of muscle proteins in the changes observed during storage, parameters related to protein functionality and the properties of extracted natural actomyosin (NAM) were studied initially and during storage in ice or at ?20 °C. Initially, the parameters related to functionality had higher values in hake muscle and extracted NAM than in gilthead sea bream. At the end of iced storage (22 days), less myosin heavy chain (MHC) and actin were extracted from hake, but there was practically no change in gilthead sea bream. This decrease was not accompanied by lower Ca²⁺‐ATPase activity. Freezing produced no drastic changes, with lower values for gilthead sea bream. However, this species was more stable after 1 year, except for the Ca²⁺‐ATPase activity of NAM. This suggests that the changes that hake proteins underwent during storage particularly affected properties related to aggregation, whereas in gilthead sea bream the changes hardly affected the formation of soluble or insoluble aggregates but did affect the active sites of myosin. © 2002 Society of Chemical Industry     

Biochemical Variations in Salt Soluble Fractions of Ultrasonicated Actomyosin Isolated from Broiler Breast-Muscle

Solubility and quality of actomyosin play crucial roles during storage and processing of a particular muscle type. We report here the effect of low intensity (20 kHz) ultrasonication on the solubility and some other biochemical properties of the actomyosin isolated from chicken breast muscle. Although there was an overall enhancement in the solubility of actomyosin during sonication, the major increase of ~61.55% occurred at 0.2M NaCl after 10–12 min of exposure. The distinctive feature of sodium dodecyl sulphate-polyacrylamide gel electrophoretic (SDS-PAGE) profiles of this fraction was the presence of free or non-interacting actin in considerable amount. Even at higher salt concentrations (0.4 and 0.5M NaCl), the protein solubility was almost twice as high as that of the corresponding controls. At low salt concentrations (<0.3 M NaCl or KCl) where the protein solubility was high, Ca²⁺-, K⁺(EDTA)- and Mg²⁺-ATPase activities were low. However, a remarkable increase in each of the enzymatic activities occurred at the salt concentrations higher than 0.3 M. More importantly, the high level of Mg²⁺-ATPase strongly indicated that actomyosin was reconstituted. The ultraviolet spectra also supported the reconstitution of actomyosin at high salt concentrations. These data suggested that ultrasonication caused conformational changes which modified properties of myosin and actin affecting the intermolecular relationships within actomyosin complex. The shift in the solubility and functional modifications of sonicated actomyosin provide some basis to explain the processing behavior of sonicated poultry meat. In addition, the low frequency sonication may be suggested as a non-destructive method to compare actomyosin extracted from various sources.     

Protein Concentration, pH and Ionic Strength Affect Apparent Viscosity of Actomyosin

Response surface methodology was used to compare effects of NaCl protein concentration, and pH on apparent viscosity (ζapp) of natural actomyosin from hake, pork and chicken muscle. Models for the three species had R² of 0.921, 0.915 and 0.814 (P<0.01) for chicken, pork and hake, respectively. Protein concentration and pH increased qapp, whereas higher ionic strength decreased it and had less influence. Interactions occurred between protein concentration and pH. Models did not reveal inter-species differences (P<0.05). However, analysis of variance of regression coefficients for each variable showed some differences due to pH.     

The function of endogenous cathepsin in quality deterioration of grass carp (Ctenopharyngodon idella) fillets stored in chilling conditions

Changes of textural properties and cathepsin activity of grass carp fillets during storage in superchilling (?1.5 ± 0.2 °C) and ice (0.2 ± 0.1 °C) was investigated, and the function of cathepsin in quality deterioration of grass carp fillets was discussed. Results showed that shear force of superchilled and ice‐stored fillets decreased by 50% and 55% after 6 days of storage, respectively. The cathepsin activies in different fractions changed significantly during storage at both conditions. Cathepsin B, B+L activities in sarcoplasma, myofibrillar and heavy mitochondrial fraction significantly increased during the early 3 days postmortem, accompanied by remarkable reduction of corresponding activity in lysosome. Cathepsin D activity in sarcoplasma and myofibrillar significantly increased during 6 days of storage with corresponding decrease in lysosome and heavy mitochondria. Correlation analysis showed that changes of shear force of grass carp fillets were significantly correlated with integration of cathepsin B, D and B+L throughout storage time (r² > 0.94). As derived from the first‐order exponential decay model, the enzyme efficiencies (k_a) of cathepsin B and B+L were more than twofold higher than those of cathepsin D, suggesting the major role of cathepsin B and B+L in textural deterioration of grass carp fillets in chilling storage.     

CHARACTERISTICS OF MUSCLE FROM TWO SPECIES OF BIGEYE SNAPPER, PRIACANTHUS TAYENUS AND PRIACANTHUS MACRACANTHUS

Composition and some properties of muscle from two species of bigeye snapper, P. tayenus and P. macracanthus, were investigated. Both species had a similar composition with the same myofibrillar protein content. However, muscle proteins from P. tayenus had higher thermal stability than those from P. macracanthus, as indicated by the higher enthalpy for transitions as well as the lower inactivation rate constant (K_D). Upon 15 days of iced storage, natural actomyosin Ca^2*‐ATP ase and Mg²⁺‐Ca²⁺‐ATPase activities decreased, whereas Mg²⁺‐EGTA‐ATPase activity increased, suggesting the denaturation of myosin, actomyosin and troponin/tropomyosin complexes, respectively. Increased surface hydrophobicity and decreased sulfhydryl groups indicated the denaturation possibly occurred via hydrophobic interaction and disulfide formation. Heading and eviscerating offish retarded the denaturation and physicochemical changes of proteins during iced storage. The results indicated that a rapid and proper post harvest handling was of importance to maintain the muscle quality of bigeye snapper.     

Physico-chemical and functional properties of myofibrillar proteins from different species of molluscs

The physico-chemical and functional properties of myofibrillar proteins from scallop (Zygochlamys patagonica), bivalve molluscs (Aulacomya ater ater), and squid (Illex argentinus) were investigated. Actomyosins were studied by SDS-PAGE, Mg²⁺-(Ca²⁺-dependent) ATPase activity, reduced viscosity, and surface hydrophobicity determinations. The emulsion capacity (EC) of both soluble protein extracts and actomyosins was also determined. Densitometric analysis of the SDS-PAGE 10% gels showed that in comparison with the actomyosin from striated muscle of scallop, the actomyosins of Aulacomya, squid, and smooth plus striated muscle of scallop have a significant higher relative percentage of paramyosin and a lower relative percentage of myosin. The reduced viscosity of actomyosin from both striated and smooth plus striated adductor muscle of scallop was significantly lower than that of squid and Aulacomya. No significant differences were observed in the enzymatic activity of the actomyosins analysed. Surface hydrophobicity and EC of actomyosin from striated muscle of Aulacomya were lower than those corresponding to actomyosin of the other species. The highest values of EC were observed in extracts of squid mantle. In addition, the EC of the extracts was significantly higher than the EC of their respective actomyosins.     

Oxidation desensitizes actomyosin to magnesium pyrophosphate-induced dissociation

This study aimed to establish the influence of protein oxidation on the ability of magnesium pyrophosphate (PP) to dissociate actomyosin. Actomyosin isolated from pork muscle then suspended in 0.1 M NaCl at pH 6.2 was oxidatively stressed with 10 μM FeCl₃/0.1 mM ascorbate/1 mM H₂O₂ for 6 or 12 h at 4 °C. Protein oxidation was evidenced by the loss of myosin and actin, the concomitant formation of disulphide-cross-linked polymers, and elevated myosin ATPase activity. The intrinsic viscosity of oxidized actomyosin had a weaker response to PP-Mg²⁺ than that of non-oxidized actomyosin, indicating the suppression of actomyosin dissociation. Moreover, oxidized actomyosin solutions were devoid of small particles (<10 nm) and the stressed actomyosin exhibited weaker binding of PP-Mg²⁺ than non-oxidized, which further suggested a reduced myosin–PP interaction and subsequent dissociation of the actomyosin complexes.