Relationship between wooden breast severity in broiler chicken, antioxidant enzyme activity and markers of energy metabolism.

This study aims to provide new insight on the association between the development of wooden breast myopathy and mitochondrial and glycolytic activity under oxidative stress. Myopathic muscle had higher oxidative stress together with altered glycolytic metabolism and tricarboxylic acid (TCA) cycle. This was evidenced by significantly elevated antioxidant enzyme activities (catalase, superoxide dismutase, and glutathione peroxidase), decreased citrate synthase activity and postmortem glycolytic potential with increasing wooden breast severity. In addition, affected muscles also exhibited higher initial and ultimate pH values as well as reduced total glucose and lactate contents. Citrate synthase activity was negatively correlated to antioxidant enzyme activities. Taken together, we propose that the development of the wooden breast lesion is a chronic process that may be related to the failure of muscle fibers to defend against the excessively generated oxidative products promoted by mitochondrial damage accompanied by impaired TCA cycle. Furthermore, there was a positive correlation between citrate synthase activity and glycolytic potential, which suggests that the wooden breast condition is linked to the overall altered energy metabolism of the muscle, including the oxidative phosphorylation and glycolytic pathways.

Effects of Wooden Breast Syndrome in Broiler Chicken on Sarcoplasmic, Myofibrillar, and Connective Tissue Proteins and Their Association with Muscle Fiber Area.

This study was conducted on chicken pectoralis major muscle with different wooden breast severity in combination with different sampling locations to investigate the effects of wooden breast syndrome on protein traits and total myofiber area, and their associations. Contents of sarcoplasmic, salt-soluble myofibrillar and salt-insoluble protein and proportion of total myofiber area significantly declined with increasing severity in the superficial part of muscle, whereas the amount of heat-soluble/insoluble collagen and protein denaturation as well as the area of degenerated myofibers, connective tissue and cellular infiltrates increased. Myofibril protein content indicators showed strong positive correlations to total myofiber area. Moreover, PCA results indicated that severe wooden breast is positively linked to muscle collagen content and to protein denaturation. Our results suggest that decrease in sarcoplasmic and myofibrillar proteins is associated with reduction of myofiber area. In turn, the muscle fibers are replaced by connective tissue, accompanied by excessive myofibrillar and sarcoplasmic protein denaturation.

Changes of Raw Texture, Intramuscular Connective Tissue Properties and Collagen Profiles in Broiler Wooden Breast during Early Storage.

A recently identified broiler myopathy known as wooden breast (WB) is predominantly found in the pectoralis major muscle of fast-growing broiler hybrids and is causing significant losses to the poultry industry. The aim of this study was to investigate the effects of WB syndrome on raw meat texture, purge loss and thermal properties of intramuscular connective tissue of pectoralis major muscle in the early postmortem period (1-3 days). Results showed that the presence of the WB muscles condition at 1 day postmortem was associated with significantly increased stiffness (27.0 N vs. 23.1 N) and significantly increased purge loss (1.8% vs. 1.0%) compared to normal breast (NB). However, on 3 days postmortem, these parameters did not differ between WB and NB groups. Insoluble and total collagen content was significantly higher in WB muscles compared to NB muscles, and the extractability of intramuscular connective tissue (IMCT) of WB was also higher (0.42% vs. 0.37%) compared to NB and remained stable in the early postmortem period. There was significantly lower protein content in the sarcoplasmic protein fraction and myofibrillar protein fraction of WB muscles compared to NB muscles (p < 0.05). The IMCT of these two groups showed different thermal properties, as the enthalpy of denaturation (ΔH) was significantly lower in WB muscles compared to NB muscles. The WB syndrome had a great effect on the texture and connective tissue properties of the meat compared to normal muscle, with a tendency for having a lower purge loss and higher raw meat hardness.

Relationship between pre-rigor temperature of pork longissimus muscle, myofibril-bound calpain activity and protein degradation.

The effect of pre-rigor temperature incubation on the activity and distribution in sarcoplasmic and myofibrillar fractions of calpains, and meat quality attributes was investigated. Porcine longissimus thoracis muscles were incubated pre-rigor at 14, 22, 30 and 38 °C to 6 h postmortem, followed by another 2 h incubation at 14 °C. Thereafter, muscles were stored at 2 °C for 1 or 4 days. With higher pre-rigor temperature, sarcoplasmic Ca2+ concentration, purge loss and myofibril-bound calpain-1 content increased, while shear force declined. Water-holding capacity of isolated myofibrils was lower after pre-rigor incubation at 38 °C. Desmin and troponin T degradation, and myofibril fragmentation was greater upon incubation of isolated myofibrils with added Ca2+ in the order 800 µM Ca2+ > 40 µM Ca2+ > no Ca2+, suggesting that calpain-1 and calpain-2 were associated to myofibrils and proteolytically active with sufficient Ca2+. Activity of myofibril-bound calpain-1 in muscle incubated pre-rigor at 22 and 30 °C were higher than when incubated at 14 and 38 °C. These results indicate that calpains translocate from the sarcoplasm onto myofibrils with higher pre-rigor temperature to 30 °C and the proteolytic potential of myofibril-associated calpains is thereby increased.

Effects of Modified Atmosphere Packaging with Varied CO2 and O2 Concentrations on the Texture, Protein, and Odor Characteristics of Salmon during Cold Storage.

The effect of gas ratio on the growth of bacteria has been well demonstrated, but some adverse effects of modified atmosphere packaging (MAP) on seafoods have also been found. To provide a better understanding of the effects of CO2 and O2 concentrations (CO2 from 40% to 100% and O2 from 0% to 30%) in MAP on the texture and protein contents and odor characteristics of salmon during cold storage, the physiochemical, microbial, and odor indicators were compared with those without treatment (CK). Generally, MAP treatments hindered the increase of microbial counts, total volatile basic nitrogen, and TCA-soluble peptides, and decreased the water-holding capacity, hardness, springiness, and sarcoplasmic and myofibrillar protein contents. The results also indicated that 60%CO2/10%O2/30%N2 was optimal and decreased the total mesophilic bacterial counts by 2.8 log cfu/g in comparison with CK on day 12. In agreement, the concentration of CO2 of 60% showed the lowest myofibrillar protein degradation, and less subsequent loss of hardness. The electronic nose characteristics analysis indicated that 60%CO2/20%O2/20%N2 and 60%CO2/10%O2/30%N2 had the best effect to maintain the original odor profiles of salmon. The correlation analysis demonstrated that microbial growth had a strong relationship with myofibrillar and sarcoplasmic protein content. It can be concluded that 60%CO2/10%O2/30%N2 displayed the best effect to achieve the goal of preventing protein degradation and odor changes in salmon fillets.

Utilization of fermented and enzymatically hydrolyzed soy press cake as ingredient for meat analogues.

The aim of the present study was to improve the properties of soy press cake to be utilized as an ingredient of meat analogues. Soy press cakes were fermented with lactobacillus strains, and separately hydrolyzed by cellulase/xylanase mixture and α-amylase. Meat analogues were produced with 10% fermented or hydrolyzed soy press cakes. The effect of applied processes on protein oxidation, physical and functional properties of soy press cakes were analyzed, as well as sensory and textural properties of meat analogues. The results indicated that soy press cake was a suitable source of fibre and energy with low content of saturated fatty acids, and provided plant-based proteins and essential amino acids. The study demonstrated the potential of lactic acid fermentation, and enzymatic hydrolysis to improve water- and oil-holding capacity and reduce protein oxidation in soy press cakes. L. acidophilus 336 and cellulase/xylanase mixture were recommended for fermentation and hydrolysis of soy press cakes, respectively, regarding reduction of protein oxidation. Fermentation of soy press cakes with L. plantarum P1 improved the texture of meat analogues. Press cakes fermentation reduced bitterness, increased juiciness, and balanced the taste of meat analogues. Fermented soy press cake was recommended for the production of meat analogues.

Role of freezing-induced myofibrillar protein denaturation in the generation of thaw loss: A review.

Formation of thaw loss cannot generally be avoided when meat is frozen and then thawed. Explanations have mainly focused on the damage to muscle fibers resulting from ice crystallization and the freezing-induced denaturation of myofibrillar proteins, the latter of which has, however, not received much research focus. This review discusses the relationship between myofibrillar protein denaturation and water-holding capacity of meat in freezing-thawing with the aim to improve the understanding the relative importance of protein denaturation in the formation of thaw loss. The contribution of decreased pH and high ionic strength in the unfrozen water in freezing is emphasized and we hypothesize that these two factors are causing protein denaturation and conformational changes within muscle fibers, and consequently loss of water-holding capacity. Slow freezing produces more thaw loss than fast freezing, and this is discussed here in relation to the impacts on myofibrillar protein denaturation induced by the freezing rate.

Sarcoplasmic and myofibril-bound calpains during storage of pork longissimus muscle: New insights on protein degradation.

The subcellular distribution of calpain-1 and -2 and the proteolytical activity of myofibril-bound calpains in pork were investigated during 12 days cold storage. The content of sarcoplasmic calpain-1 decreased during storage while myofibril-bound calpain-1 content first increased (P < 0.05) to 17% of that of 12 h-sarcoplasmic calpain-1 on day 6 followed by a gradual decrease with subsequent storage, suggesting that calpain-1 gradually translocated from sarcoplasm to myofibrils during the initial 6 days of postmortem storage. Intact desmin decreased (P < 0.05) after incubation of myofibrils with 0.05 mM Ca2+, and this was more pronounced with 5 mM Ca2+ (P < 0.05). Ca2+ titration curves of day 6 myofibrils showed two distinct proteolytic activities becoming activated in the range 0.03 to 0.06 mM and 0.4 to 0.8 mM Ca2+, respectively. The results suggest that both calpain-1 and calpain-2 binds to myofibrils during storage and subsequently degrade structural proteins including desmin.

Freezing of meat and aquatic food: Underlying mechanisms and implications on protein oxidation.

Over the recent decades,protein oxidation in muscle foods has gained increasing research interests as it is known that protein oxidation can affect eating quality and nutritional value of meat and aquatic products. Protein oxidation occurs during freezing/thawing and frozen storage of muscle foods, leading to irreversible physicochemical changes and impaired quality traits. Controlling oxidative damage to muscle foods during such technological processes requires a deeper understanding of the mechanisms of freezing-induced protein oxidation. This review focus on key physicochemical factors in freezing/thawing and frozen storage of muscle foods, such as formation of ice crystals, freeze concentrating and macromolecular crowding effect, instability of proteins at the ice-water interface, freezer burn, lipid oxidation, and so on. Possible relationships between these physicochemical factors and protein oxidation are thoroughly discussed. In addition, the occurrence of protein oxidation, the impact on eating quality and nutrition, and controlling methods are also briefly reviewed. This review will shed light on the complicated mechanism of protein oxidation in frozen muscle foods.

Near-Infrared Reflectance Spectroscopy for Predicting the Phospholipid Fraction and the Total Fatty Acid Composition of Freeze-Dried Beef.

Research on fatty acids (FA) is important because their intake is related to human health. NIRS can be a useful tool to estimate the FA of beef but due to the high moisture and the high absorbance of water makes it difficult to calibrate the analyses. This work evaluated near-infrared reflectance spectroscopy as a tool to assess the total fatty acid composition and the phospholipid fraction of fatty acids of beef using freeze-dried meat. An average of 22 unrelated pure breed young bulls from 15 European breeds were reared on a common concentrate-based diet. A total of 332 longissimus thoracis steaks were analysed for fatty acid composition and a freeze-dried sample was subjected to near-infrared spectral analysis. 220 samples (67%) were used as a calibration set with the remaining 110 (33%) being used for validation of the models obtained. There was a large variation in the total FA concentration across the animals giving a good data set for the analysis and whilst the coefficient of variation was nearly 68% for the monounsaturated FA it was only 27% for the polyunsaturated fatty acids (PUFA). PLS method was used to develop the prediction models. The models for the phospholipid fraction had a low R2p and high standard error, while models for neutral lipid had the best performance, in general. It was not possible to obtain a good prediction of many individual PUFA concentrations being present at low concentrations and less variable than other FA. The best models were developed for Total FA, saturated FA, 9c18:1 and 16:1 with R2p greater than 0.76. This study indicates that NIRS is a feasible and useful tool for screening purposes and it has the potential to predict most of the FA of freeze-dried beef.

Myofibrillar protein characteristics of fast or slow frozen pork during subsequent storage at -3 °C.

This study aimed to investigate the effect of storage at -3 °C on myofibrillar protein in fast or slow frozen pork. Five pork loins at 48 h post-mortem were subjected to either fast (cold metal plate/-80 °C) or slow freezing (still air/-20 °C) followed by storage at -3 °C for 0, 1, 3, and 7 days before thawing. Freezing rate significantly influenced myofibrillar proteins within 3 days at -3 °C, evidenced by higher thaw loss, higher surface hydrophobicity and reduced water-holding of myofibrils, and accelerated appearance of a myosin-4 fragment (160 kDa) in slow freezing. However, these observed differences disappeared after 7 days of storage at -3 °C. The meat pH after thawing did not differ between fast and slow freezing rate. However, the pH values after thawing in both groups decreased with extended storage at -3 °C. Our results suggest that the beneficial effects of fast freezing are gradually lost by holding at -3 °C due to more extensive protein denaturation.

Effect of LTLT heat treatment on cathepsin B and L activities and denaturation of myofibrillar proteins of pork.

The aim was to study biophysical and chemical changes during low-temperature long-time (LTLT) heat treatment of pork by measuring cathepsin B+L activity, surface hydrophobicity of myofibrils, particle size of myofibrils and effect on meat toughness as indicated by Allo-Kramer shear force. Longissimus thoracis et lumborum muscles were divided into large pieces, vacuum packaged and cooked in water baths at 53, 58, 63, 68 and 73 °C for 1, 8 and 24 h. The results showed that the meat toughness was markedly lower at temperatures of 53 °C and 58 °C and decreased with increasing holding time. Myofibrillar surface hydrophobicity increased with temperature, but not with time, indicating aggregation and/or gelation phenomena took place. Treatments with the lowest shear force values generally had smaller particles and were associated with high cathepsin B+L activity. A mechanism by which these cathepsins might affect the aggregation dynamics and change the mechanical properties of meat is proposed.

Mimicking myofibrillar protein denaturation in frozen-thawed meat: Effect of pH at high ionic strength.

This study aims at providing new insight on protein denaturation in freezing-thawing. Freezing-thawing minced pork reduced water-holding of myofibrils and increased surface hydrophobicity. One additional freezing-thawing cycle at slow freezing rate caused appearance of a 160 kDa myosin-4 fragment in SDS-PAGE, further decreased water-holding of myofibrils and increased surface hydrophobicity. Fresh minced pork was exposed to either high salt (2 M KCl) only or high salt with lower pH to mimic conditions in freezing. Exposure to high salt only increased water-holding of myofibrils and hence did not reproduce myofibrillar protein changes in freezing. Exposure to combinations of lower pHs and high salt decreased water-holding and increased surface hydrophobicity, suggesting myofibrillar protein denaturation occurred by a comparable mechanism as in freezing-thawing. We propose that exposure to decreased pH combined with high solute concentrations in the unfrozen water of frozen meat is the primary cause of myofibrillar protein denaturation in frozen-thawed meat.

Ca2+-induced binding of calpain-2 to myofibrils: Preliminary results in pork longissimus thoracis muscle supporting a role on myofibrillar protein degradation.

The aim of this study was to investigate the role of Ca2+ in the process of calpain-2 becoming associated to myofibrils and the potential of myofibril-bound calpain to degrade myofibrillar proteins. Different Ca2+ concentrations were applied to myofibrils mixed with partially purified calpain-2. Ca2+ induced binding of calpain to myofibrils in a concentration-dependent manner. The half-maximal Ca2+ requirements for binding of calpain-2 to myofibrils and for calpain-2 proteolysis of myofibrils were 0.60 mM and 0.29 mM, respectively. To investigate the proteolytic activity of myofibril-bound calpain, a mixture of myofibrils and calpain-2 was briefly incubated with Ca2+. Unbound calpain was removed by washing with a Ca2+-free buffer. The myofibril-bound calpain maintained proteolytic activity and degraded desmin when re-activated with Ca2+. In conclusion, the results suggest that an increase in Ca2+ will activate and induce binding of calpain to myofibrils. Subsequently, calpain is relatively tightly bound and proteolytically active.

MEATabolomics: Muscle and Meat Metabolomics in Domestic Animals.

In the past decades, metabolomics has been used to comprehensively understand a variety of food materials for improvement and assessment of food quality. Farm animal skeletal muscles and meat are one of the major targets of metabolomics for the characterization of meat and the exploration of biomarkers in the production system. For identification of potential biomarkers to control meat quality, studies of animal muscles and meat with metabolomics (MEATabolomics) has been conducted in combination with analyses of meat quality traits, focusing on specific factors associated with animal genetic background and sensory scores, or conditions in feeding system and treatments of meat in the processes such as postmortem storage, processing, and hygiene control. Currently, most of MEATabolomics approaches combine separation techniques (gas or liquid chromatography, and capillary electrophoresis)-mass spectrometry (MS) or nuclear magnetic resonance (NMR) approaches with the downstream multivariate analyses, depending on the polarity and/or hydrophobicity of the targeted metabolites. Studies employing these approaches provide useful information to monitor meat quality traits efficiently and to understand the genetic background and production system of animals behind the meat quality. MEATabolomics is expected to improve the knowledge and methodologies in animal breeding and feeding, meat storage and processing, and prediction of meat quality.

Protein degradation of black carp (Mylopharyngodon piceus) muscle during cold storage.

This study investigated the effects of cold storage at different temperatures (4, -0.5, -3, and -20 °C) on protein degradation and its relationship to structural changes of black carp muscle. At -0.5 and 4 °C, major structural changes occurred, including the formation of gaps between myofibers and myofibrils, breakage of myofibrils and myofibers, and degradation of sarcoplasmic reticulum. Gel-based proteomic analysis showed that these structural changes were accompanied by degradation of a series of myofibrillar proteins, including titin, nebulin, troponin, myosin, myomesin, myosin-binding protein, and α-actinin. Loss of extractable gelatinolytic and caseinolytic protease activities was also observed. At -3 and -20 °C, formation of ice crystals was the most noticeable change. The major proteins were degraded at different locations in the black carp muscle, and gelatinolytic and caseinolytic proteases appear to contribute to the degradation of those proteins.

Metabolite profile based on 1H NMR of broiler chicken breasts affected by wooden breast myodegeneration.

The objective was to characterize the effect of wooden breast (WB) myodegeneration on the metabolite profile of chicken meat by 1H NMR and multivariate data analysis. The results displayed that the metabonome of chicken breast consisted predominantly of 30 metabolites, including amino acids, organic acids, carbohydrates, alkaloids, nucleosides and their derivatives. WB-affected samples showed higher leucine, valine, alanine, glutamate, lysine, lactate, succinate, taurine, glucose, and 5'-IMP levels, but lower histidine, ß-alanine, acetate, creatine, creatinine, anserine and nicotinamide adenine dinucleotide levels compared to normal fillets (p < 0.05). In conclusion, results indicated that WB-affected fillets possessed a unique biochemical signature. This unique profile could identify candidate biomarkers for diagnostic utilization and provide mechanistic insight into biochemical processes leading to WB myopathy in commercial broiler chickens.

On the origin of thaw loss: Relationship between freezing rate and protein denaturation.

The role of protein denaturation in formation of thaw loss is currently not well understood. This study investigated denaturation of myofibrillar and sarcoplasmic proteins of pork loins caused by freezing-thawing in relation to freezing rate. Compared to fast freezing, slow freezing caused 28% larger thaw loss, decreased water-holding capacity of myofibrils and increased surface hydrophobicity, indicating more pronounced denaturation of myofibrillar proteins. We here propose a model: In slow freezing protons are concentrated in the unfrozen water resulting in reduced pH in proximity of structural proteins causing protein denaturation. In parallel, large ice crystals are formed outside of muscle fibers resulting in transversal shrinkage. In fast freezing small ice crystals trap protons and cause less severe protein denaturation and reduced thaw loss. Differential scanning calorimetry and tryptophan fluorescence spectra indicated sarcoplasmic protein denaturation in drip due to freezing-thawing. However, sarcoplasmic protein denaturation was independent of freezing rate.

Effects of protein oxidation on the texture and water-holding of meat: a review.

Protein oxidation readily occurs in postmortem muscle during storage and processing. Over the past decade new analytical methods have been developed and new aspects of protein oxidation in meat have been studied, such as the reaction mechanism, and impacts on eating quality and nutritional value. It is now evident that amino acid side chains in myofibrillar proteins undergoes modifications due to oxidative stress. In turn this will lead to formation of new protein-protein cross-links in structural proteins, however, also the overall level of fixed-charge groups attached to the peptide backbones is modified. Meat texture and water-holding are important quality attributes and they are affected by the oxidation of structural proteins. Different mechanisms have been suggested to explain the oxidation-induced quality changes, focusing mainly on reduced proteolysis and formation of cross-links. This review explores the current understanding of protein oxidation in fresh meat in relation to texture and water-holding. The consequences of protein oxidation at molecular level in relation to oxidation-induced cross-linking and changes in net charges of myofibrillar proteins, and the impacts on texture and water-holding are discussed.

Effects of frozen-then-chilled storage on proteolytic enzyme activity and water-holding capacity of pork loin.

This study aimed to determine the effect of frozen-then-chilled storage on free Ca2+, proteolytic enzyme activity of calpains and the proteasome, water-holding capacity and shear force of porcine longissimus thoracis et lumborum muscle. Pork loins were subjected to either chilled storage at 2 ±â€¯1 °C for 1, 2, 4, 6 and 9 days, or frozen-then chilled storage (-20 ±â€¯1 °C for 1 week followed by thawing overnight). Free Ca2+ increased with chilled storage in the non-frozen group. Frozen-then-chilled storage increased free Ca2+ concentration, followed by a faster decrease of calpain-1 activity and activation of around 50% of calpain-2. Proteasome activity was reduced by around 40% following freezing-thawing. Purge loss increased and water-holding capacity of myofibrils decreased in the frozen-thawed group, suggesting considerable denaturation of myofibrillar proteins. Shear force was not affected by freezing-thawing, and we speculate that the tenderizing effect of calpain activation was counteracted by loss of proteasome activity and substantial exudate loss.