Effect of lactic or acetic acid on degradation of myofibrillar proteins in post‐mortem goose (Anser anser) breast muscle

The effects of lactic acid (LA) and acetic acid (AA) on changes in myofibrillar proteins of post‐mortem goose breast muscle marinated for 24 h at 5 °C were studied. Purified myofibrils were prepared from 0.1 M LA or AA samples and controls (non‐marinated samples) after 0, 1, 3, 7 or 14 days of storage at 5 °C. The changes in myofibrillar proteins of goose muscle were examined by SDS‐PAGE. Goose breast muscle marinated in LA and AA exhibited degradation of myosin heavy chains. The appearance of ∼95 and ∼27 kDa components and the disappearance of titin and nebulin were also more rapid than for control muscle. These results suggest that acid marination enhanced the post‐mortem proteolysis of goose breast muscle. © 2000 Society of Chemical Industry     

Cryoprotective additives and cryostabilisation effects on muscle fillets of the freshwater teleost fish Rohu carp (Labeo rohita) during prolonged frozen storage

The effects of various cryoprotective additives separately and in combination were studied on the myofibrillar protein integrity, biochemical enzyme activity levels and muscle ultrastructure in the freshwater teleost fish Rohu carp (Labeo rohita). Fish muscle samples were divided into eight groups and immersed in different mixtures of cryoprotective additives (S1–S8), then frozen at ? 20 or ? 30 °C for 24 months. Electrophoretic studies revealed early (within 6 months) alteration of the myofibrillar proteins myosin light chain, α‐actinin and tropomyosin. Reduction of the storage temperature from ? 20 to ? 30 °C slowed down the degradative processes. Sodium dodecyl sulfate polyacrylamide gel electrophoresis indicated that fish muscle treated with cryoprotective mixture S8 (40 g L^?1 sorbitol/3 g L^?1 sodium tripolyphosphate/4 g L^?1 sodium alginate) showed minimal post mortem changes in myofibrillar proteins. Ultrastructural results also revealed post mortem damage to the muscle, seen earliest (within 6 months) in the sample frozen‐stored without additives (S2), as compared with the normal, unfrozen muscle (S1). The influence of cryoprotectants alone and in combination on fish muscle structural proteins, myosin and actin filaments (A and I bands), during prolonged frozen storage was investigated. After 12 months, samples frozen‐stored with various cryoprotective additives (S2‐S7), except S8, showed signs of myofibrillar disintegration. Beyond that time the degradative processes started showing up in all samples, with minimal muscle ultrastructural damage in sample S8. Again, reducing the storage temperature from ? 20 to ? 30 °C slowed down the degradative processes. Ultrastructural results correlated well with levels of biochemical enzymes (Ca²⁺ myofibrillar ATPase and succinic dehydrogenase) during frozen storage. This is the first report of the cryoprotective effects of these additives on this popular edible fish species. Of the various combinations of additives tested, cryoprotective mixture S8 was found to preserve the muscle structure longest under frozen storage conditions. However, even this mixture was only effective for 18 months at ? 30 °C. Beyond that time the myofibrillar degradative processes were apparent with correlative electrophoretic, biochemical and ultrastructural studies. Copyright © 2006 Society of Chemical Industry     

Lipid and protein deterioration during the chilled storage of minced sea salmon (Pseudopercis semifasciata)

Sea salmon is a very appreciated seafood. The aim of this work was to analyze changes in lipid and protein fractions of minced muscle during chilled storage (1 ± 1 °C). Lipid oxidation was important during the first 6 days of storage according to 2‐thiobarbituric acid (TBA) values determined, decreasing mainly ω3 22:6 fatty acid content. Lipid hydrolysis was evident after 9 days of storage. Interaction compounds between oxidation products and other cellular components were analyzed by fluorescence measurements. The results obtained showed evidence of the formation of interaction products involved, mainly polar components such as proteins. Decreases in myosin and actin thermal stability and myosin denaturation were recorded by differential scanning calorimetry. Solubility of total and myofibrillar proteins decreased after 6 days of storage. The electrophoretic profile of soluble fractions showed an increase in the intensity of bands corresponding to low‐molecular‐weight polypeptides and a decrease in high‐molecular‐weight species. Available lysine content did not change during chilled storage. Copyright © 2007 Society of Chemical Industry     

Effect of frozen storage on thermal stability of sarcoplasmic protein and myofibrillar protein from common carp (Cyprinus carpio) muscle

Thermal stability of sarcoplasmic protein and myofibrillar protein extracted from fresh and frozen common carp was comparatively studied. Total sulphydryl content (SH) in sarcoplasmic protein solution from 5‐month frozen carp decreased by 19.43% compared with fresh sample. The SDS‐PAGE patterns showed that all the bands of sarcoplasmic protein from frozen‐stored samples were almost invisible at 80 °C. Myofibrillar protein from fresh sample exhibited lower turbidity and surface hydrophobicity and higher Ca²⁺‐ATPase activity and SH content than frozen‐stored sample when heated from 20 to 80 °C. The Ca²⁺‐ATPase activity from fresh (M0), 2 (M2)‐ and 5 (M5)‐month frozen‐stored carp was completely lost at 48, 46 and 46 °C, respectively. When heated to 80 °C, the SH content of myofibrillar solutions in M0, M2 and M5 decreased by 26%, 60% and 70%, respectively. Sarcoplasmic and myofibrillar proteins from frozen carp were more susceptible to aggregate during heating treatment.     

Myosin heavy chain degradation during post mortem storage of Atlantic cod (Gadus morhua L.)

Post mortem proteolytic degradation of fish fillets leads to textural changes like muscle softening and gaping. In this study proteolytic degradation of myosin heavy chain (MHC) was monitored during storage of muscle and of isolated myofibrils at different temperatures and pH-values by the use of MHC-specific antibodies. The ability of cathepsin D to associate to myofibrillar proteins was also studied. Muscle stored at 6 °C and isolated myofibrils stored at 0 °C, 6 °C and 20 °C were degraded at pH 6.3 or lower. Cathepsin D could be found associated with extensively washed myofibrils. Inhibition of cathepsin D during storage affected the observed MHC-degradation at pH 5.5, but not at pH 6.3. This indicates that cathepsin D to a less extend than formerly believed, is responsible post mortem degradation of MHC.     

Influence of rapid chilling on biochemical parameters governing the water‐holding capacity of pork longissimus dorsi

The aim of this study was to investigate the effects of rapid chilling (RC) of carcasses on the quality of pork longissimus dorsi (LD) compared to conventional chilling (CC). Carcasses subjected to RC were chilled at ?20 °C for 1.5 h and then moved to 4 °C until 24 h post‐mortem, while carcasses that were CC were placed directly in a chill at 4 °C for 24 h post‐mortem. RC accelerated the rates of temperature decline post‐mortem (P < 0.01) and increased pH values at 1.5 and 8 h post‐mortem (P < 0.05). RC could significantly decrease the denaturation of myosin and sarcoplasmic proteins during post‐mortem ageing. Samples of RC had a significant lower purge loss compared to CC group during post‐mortem ageing (P < 0.05). Our study implies that RC could influence the biochemical changes of pork LD which may further regulate water‐holding capacity during post‐mortem ageing.     

Gilthead seabream (Sparus aurata): suitability for freezing and commercial alternatives

The quality of portion‐size farmed gilthead seabream (Sparus aurata) during frozen storage and the influence of post‐mortem treatments were studied in order to find new ways of marketing this species. Portion‐sized gilthead seabream, fasted for 48 h prior to slaughter, were frozen and stored at ?20 °C for up to one year. Whole fish were frozen immediately after rigor mortis; gutted fish were frozen immediately after rigor and after 5 days of storage in ice. All lots were stored at ?20 °C for up to one year. The myofibrillar protein of this species was very stable and a slight decrease of solubility in salt solutions was found only after one year of frozen storage. A slight decrease in water‐holding capacity and a slight increase in shear strength were observed, but these were lower than in other species. These changes were reflected as increased toughness and reduced juiciness in sensory analysis of the cooked fillets after one year. The main changes in the cooked fillets were observed in odour and flavour. No significant detrimental effect due to the guts was detected during frozen storage. Storage in ice prior to freezing was reflected in sensory assessment of the raw fish, mainly in terms of initial higher demerit points for fishy odour, gills and eyes; however, no effect was observed in the cooked fillets. Copyright © 2004 Society of Chemical Industry     

Myofibril-Bound Serine Proteinase (MBP) and its Degradation of Myofibrillar Proteins

Proteolysis of a myofibril-bound serine proteinase (MBP) from carp Cyprinus carpio on myofibrillar proteins and their gel formation ability were investigated. MBP readily decomposed myosin heavy chain as indicated by SDS-PAGE. In the preparation of kamaboko, the gel formation ability was diminished by addition of MBP. The optimum degradation temperatures of MBP to myosin heavy chain in myofibril and kamaboko gel were 55°C and 60°C, respectively. The degradation effects of MBP on actin, α-actinin and tropomyosin were studied by the immunoblotting method. Because of its myofibril-bound and myofibrillar protein degradation characteristics, MBP was regarded as the proteinase most probably involved in the modori effect.     

Separation of Cod (Gadus morhua) Fillet Proteins by Electrophoresis and HPLC after Various Frozen Storage Treatments

Myofibrillar, sarcoplasmic (SAR) and total extractable proteins (TEP) were isolated from the same lot of frozen cod (Gadus morhua) fillets stored at -30°C, -22°C, -15°C, and -12°C and a simulated industrial fluctuating temperature (SIFT) program. HPLC and SDS-PAGE showed that myofibrillar and SAR of the fillets stored at -12°C and -15°C exhibited low and high molecular weight changes similar to the SIFT. A new indicator of myofibrillar frozen storage quality was developed using SDS-PAGE. TEP profiles by HPLC were time and temperature dependent. These observations by SDS-PAGE and HPLC were further evidence that changes in frozen stored cod muscle were partially due to covalent S-S crosslinking.     

Autolytic degradation of belly tissue in anchovy (Engraulis encrasicholus)

Tissue degradation in anchovy was studied during storage under different conditions. Belly bursting seems to be caused by leakage of proteolytic enzymes from the pyloric caeca and intestine to the ventral muscle. These proteases have optimum pH in the alkaline range and they are very active in the neutral range. At pH 6.5, which is the post mortem pH of the tissue, the basic proteases play an important role in the degradation of myofibrillar protein and in the solubilization of connective tissue. The rate of tissue degradation during storage was considerably reduced if the fish was chilled by refrigerated or iced sea water to about 0°C, and the pH of the medium was reduced to 5 by addition of lactic or acetic acid.     

In vitro proteolysis of myofibrillar and sarcoplasmic proteins of European sea bass (Dicentrarchus Labrax L) by an endogenous m‐calpain

The effects of m‐calpain isolated from the skeletal muscle of sea bass on sarcoplasmic and myofibrillar proteins isolated from the same tissue were examined in vitro. Incubation of sarcoplasmic proteins with m‐calpain resulted in only a slight decrease (0.7 kDa) in the molecular weight (MW) of a 26.5 kDa protein. Degradation of myofibrils, monitored by quantification of TCA‐soluble peptides generated, resulted in the maximum amount of peptides being generated after 1 h of incubation at 25 °C. Noticeable modifications in the SDS‐PAGE profile of digested myofibrils were observed, including partial denaturation of myosin heavy chain and the release of tropomyosin, ～69 and ～27 kDa doublet bands and a few polypeptides of MW lower than 20 kDa in the soluble fraction. Examination of the degradation patterns of myofibrillar proteins using Western blotting showed that α‐actinin was partially degraded, with release of native α‐actinin and its fragments from myofibrils, whereas desmin was highly degraded after 2 h of digestion. © 2002 Society of Chemical Industry     

Post-mortem changes in myofibrillar proteins of breast and leg muscles from broilers,spent hens and Taiwanese Country Chickens

Post mortem ageing at 4°C was studied in the breast and leg muscles from broilers, White Leghorn spent hens and Taiwanese Country Chickens (TCC). Purified myofibrils were prepared from muscles after 0, 1, 3, 5, and 7 days of post mortem storage at 4°C. SDS-PAGE was used to examine the changes in myofibrillar proteins of the muscles. Results showed that 30 kDa components appealed earlier in the breast muscles than in the leg muscles and in the order: broiler, TCC, and spent hen. The intensity of 30 kDa band increased with post mortem time. In the breast muscles, a decrease in the intensity of the α-actinin band could be observed at 1 day post mortem in broilers and TCC and at 3 days post mortem in spent hens. This decrease, however, could not be found in the leg samples until 5 or 7 day post mortem. Titin 1 band disappeared within 3 day post mortem in the breast samples but within 5 days in the leg samples. Similar results could be observed in the degradation of nebulin although traces of nebulin remained in the teg muscles of TCC after 7 days post mortem.     

Electrophoretic patterns of microwaved and γ‐irradiated beef liver proteins

The effects of γ‐irradiation treatments (2.5, 5 and 10 kGy) and microwaves generated from an oven at low and defrost power settings for 0.5, 1 and 2 min on the total proteins and protein patterns of beef liver immediately after treatment and during frozen storage (?18 °C) for different periods were studied. Chemical analyses indicated that the protein content of beef liver was reduced after exposure to γ‐radiation or microwaves and also during frozen storage. Sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS‐PAGE) was used to illustrate the changes in protein bands of different molecular weights and their percentages before and after exposure to gamma and microwave radiation. The main effect of γ‐radiation on the protein patterns of beef liver was the disappearance of some high‐molecular‐weight protein bands and the development of other bands characterised by moderate and low molecular weights. This finding indicates the degradation of beef liver proteins by γ‐irradiation. In contrast, microwave treatment caused an increase in the levels of high‐molecular‐weight protein bands with a concomitant decrease in low‐molecular‐weight protein bands. This phenomenon demonstrates the polymerisation of low‐molecular‐weight proteins under the influence of microwaves. © 2001 Society of Chemical Industry     

Relationship between proteolytic changes and tenderness in prerigor lactic acid marinated beef

A meat tenderising procedure involving injection of a lactic acid solution into prerigor muscle was investigated using beef M pectoralis profundus. The distribution of lysosomal enzymes in subcellular fractions, densities of myofibrillar protein bands after SDS‐PAGE and shear force were measured in non‐injected, 0.5 M and 1.0 M lactic‐acid‐injected samples during a 21 days ageing period. The activities of cathepsin B + L and β‐glucuronidase in the soluble fraction increased with level of lactic acid and with time post‐mortem (P < 0.001). Lactic acid and storage decreased densities of SDS‐PAGE bands migrating at the position of myosin heavy chain (MHC) and α‐actinin and increased densities of a 150 kDa band (P < 0.01). SDS‐PAGE of isolated perimysium cleaved with CNBr showed proteolytic cleavage of collagen after prolonged storage. Lactic acid injection significantly reduced shear force (P < 0.001). The cathepsin B + L activity in the soluble fraction correlated to shear force (r = −0.8), the degradation of MHC and α‐actinin (r = −0.88 and −0.90) and the generation of the 150 kDa fragment (r = 0.90) but not to the generation of a 31 kDa fragment (r = 0.05). A major part of the tenderness improvement after lactic acid injection was complete at 24 h post‐mortem, and was therefore due to a rapid process, eg pH‐induced swelling of the muscle structure. The data on enzyme activities and protein degradation, however, suggested that the action of lysosomal cathepsins also contributed to textural changes. © 1999 Society of Chemical Industry     

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.     

湿热地区不同贮藏温度对热鲜牛肉品质变化的影响

本文研究了湿热地区不同温度热鲜牛肉贮藏过程中食用品质、新鲜度、肌糖元含量和肌原纤维小片化指数的变化,旨为热鲜牛肉的食用提供参考。选取20月龄左右锦江黄牛6头,宰后立即取背最长肌作为原料,分别贮藏在温度为5℃、15℃、25℃和35℃和湿度为80%恒温培养箱。测定不同贮藏温度处理对热鲜牛肉贮藏过程中pH、感官指标(色泽、风味、弹性、组织状态和总体可接受性)、菌落总数、肌糖元含量和肌原纤维蛋白降解的变化规律。结果表明;随着贮藏时间长,不同贮藏温度热鲜牛肉pH呈先降低后升高,肌糖元含量和感官评分降低,菌落总数和MFI增加。综合分析;贮藏温度对热鲜牛肉的pH值、感官评分、菌落总数、肌糖元含量以及MFI变化具有一定影响。贮藏温度越高,失水率越高,微生物生长繁殖越快,肌原纤维蛋白降解程度越大。     

Broad‐spectrum inhibition of proteolytic enzymes by allicin and application in mitigating textural deterioration of ice‐stored grass carp (Ctenopharyngodon idella) fillets

The inhibitory effect of allicin on proteolytic enzymes and textural deterioration of ice‐stored grass carp (Ctenopharyngodon idella) fillets was investigated. The results of in vitro study showed that allicin inhibited the activity of cathepsin B, L and D, calpain and collagenase in crude extract of grass carp muscle. Among endogenous enzymes, cathepsin B, L and collagenase were the most susceptible to allicin. Proteolysis of myofibrillar proteins by either crude enzyme or cathepsin B and L was almost prevented by allicin when employed at a concentration higher than 100 mm . After storage of 21 days, shear force of fillets treated with allicin at 10–100 mm was 39–51% higher than that of control. Myofibrillar proteins of fillets during storage were well protected against degradation when allicin concentration increased to 100 mm , as evidenced by SDS‐PAGE. Therefore, allicin could be a potential broad‐spectrum inhibitor to retard softening of fish fillets via mitigating myofibrillar proteolysis by endogenous enzymes especially cathepsin B and L during ice‐storage.     

Glass Transition Values of Muscle Tissue

Reports of glass transition (Tg') values for frozen muscle tissue are not common and reported values are mostly much lower than would be expected. Tg’ values for muscle tissue and isolated proteins were studied using differential scanning calorimetry. Apparent Tg's of mackerel, cod and beef were similar (ca ?11 to ?13°C) and substantially higher than most published values (?15 to ?77°C for tuna and beef), but in accord with expectations for substances of high molecular weight. Dialyzed insoluble and soluble protein fractions from mackerel yielded apparent Tg’ values (ca ?7°C) that were similar, with both being higher than those for whole muscle. Apparent Tg’ values of ca ?7°C were determined for aqueous samples of gelatin and collagen, but none was detected for zein.     

Effect of heating temperature and duration on the texture and protein composition of Bighead Carp (Aristichthys nobilis) muscle

The effect of heating temperature and duration on the texture and protein composition of bighead carp (Aristichthys nobilis) muscle was investigated. Samples were heated at 90°C, 100°C, 110°C, and 120°C for different times. The results showed that cooking loss increased significantly with increasing temperature and heating time, and a fractional conversion model was used to describe the increase. Hardness, chewiness, and gumminess exhibited a similar four-phase change with two peaks at 90°C, 100°C, and 110°C, and the trend was not observed at 120°C. Analysis of sodium dodecyl sulfate polyacrylamide gel electropheresis (SDS-PAGE) revealed that contents of myosin heavy chain decreased entirely to disappear and contents of actin significantly decreased. Increasing the intensity of the heavy molecular weight bands indicated the aggregation of myosin. Content of water-soluble proteins presented an increasing trend, which reflected the protein degradation and the release of low-molecular-weight compounds.