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Volume 64, Issue 1,
, Pages 69-75
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https://doi.org/10.1016/S0309-1740(02)00163-8Get rights and content
Changes in myofibrillar protein content and centrifugal drip proteins of psoas major and minor (PM) and semitendinosus (ST) muscles of calves, heifers and cows taken from carcasses on the 1st, 6th and 12th day of post-mortem cold storage were estimated. Washed myofibrils and centrifugal drip from muscles were analysed using SDS-PAGE 10 and 12% polyacrylamide gels. No significant changes were observed in content of contractile proteins, α-actinin and regulatory proteins (except for TN-T). There were no significant differences between muscles from investigated groups and between muscles aged in chilled conditions. The levels of titin T1 during ageing varied slightly. The 30 kDa-fraction appearance was fastest in calf, slower in heifer and slowest in cow muscles. More pronounced differences in the level of protein degradation with regards to muscle type, age of animals and time of storage were found in the centrifugal drip of meat. In the drip, the level of high molecular weight proteins was higher in muscles from young animals and in the muscles stored longer. The opposite was observed in case of 26–28 kDa proteins. Their amount in muscle drip decreased with increased storage time. The rate of proteolysis and release of cytoskeletal proteins during cold storage of muscles were related to change in shear values of roasted meat. The highest rate of protein degradation was observed in PM calf muscle and the lowest rate in ST cow muscle. The fastest tenderization process was registered in calves muscles and the slowest tenderization in cows.
Structural changes occur in bovine muscles during post-mortem storage of meat in the refrigerated conditions (Wegner et al., 2000, Will et al., 1980, Ho et al., 1996). According to Kołczak, Pospiech, Palka, and Lącki (2002) during 12 days of cold storage of meat the length of sarcomeres increases, mainly due to the enlargement of I-bands and also some changes in A-band, which took place especially in PM of veal. These structural changes are more rapid and intense in muscles from younger animals, are faster and have wider range in psoas major and minor (PM) than in semitenidinosus (ST) muscle.
The main contractile proteins of myofibrils, such as myosin and actin, do not appear to undergo degradation post-mortem at 4°C even after 56 days of cold storage (Bandman and Zdanis, 1988, Yates et al., 1983). However changes in structure of muscle fibres are considered to be caused by degradation of cytoskeletal proteins. These proteins are part of costameres binding myofibrils with sarcolemma (Huff-Lonergan et al., 1996, Robson et al., 1997, Taylor et al., 1995) and stabilise the intracellular myofibrillar protein arrangement (Taylor et al., 1995, Chiung-Ying et al., 1996, Huff-Lonergan et al., 1995), especially the Z-discs’ structure (Shimada et al., 1999, Takahashi, 1996). The changes are associated with degradation of two very large proteins: nebulin and titin, however the disappearance of smaller proteins has also been brought to the attention of investigators. Recent studies show that this disappearance and the breaks of structures near the Z-line may be related to the titin degradation at the N2 line (Boyer-Berri & Greaser, 1998) and proteolysis of desmin, which is responsible for transverse stabilisation of muscle fibrils and myofibrils (Purslow, Ertbjerg, Baron, Christensen, & Lawson, 2001).
One of the most common symptoms of proteolytic protein degradation during the cold ageing is the appearance of troponin T (TN-T) degradation products having molecular weight of about 30 kDa (Ho et al., 1994, Geesing et al., 1995, MacBride and Parrish, 1977, Penny and Dransfield, 1979). According to Penny and Dransfield (1979) they can be used as an indicator of the rate of meat ageing.
Despite many studies describing degradation of cytoskeletal proteins and TN-T, there is a lack of information on their changes in muscles, which come from animals of different maturity. The aim of this study was to describe changes in myofibrillar (including cytoskeletal) proteins and centrifugal drip proteins from PM and ST muscles of calves, heifers and cows during 12-days post-mortem cold storage and to relate the results to Warner-Bratzler shear force measurements.
Material and methods
PM and ST muscles were taken from left half-carcasses of female calves (3-months old), heifers (18-months old) and cows (around 8 years old) 24 h after slaughter. In each of the examined groups, analyses were carried out on muscles taken from three carcasses. Each of the muscles was divided into 3 parts, which were vacuum-sealed and stored at 4°C for 12 days.
Myofibrils from samples after 1, 6 and 12 days of cold storage were isolated and their proteins were determined using PAGE-SDS (Fritz,
Results and discussion
Electrophoresis of washed myofibrils originating from examined PM and ST muscles showed from 18 to 24 bands of proteins. Fig.1 presents their electrophoretical pattern after 1st, 6th, and 12th days of cold storage of samples taken from heifers muscles. This figure also depicts the separation of proteins from centrifugal drip of these muscles.
No significant changes were observed in the amount of the contractile proteins myosin and actin. The share of MHC (myosin heavy chains) in the fraction of
During post-mortem storage of beef muscles the degradation of myofibrillar and cytoskeletal proteins depends on muscle type and animal age. Changes of titin T1 in the fraction of washed myofibrillar proteins observed on SDS-PAGE are small. More pronounced changes occur in the fraction of 30 kDa, i.e. Tn-T degradation products. Their appearance is slower in cow than in heifer and calf muscles. Clear differences in concentration of protein degradation fractions in centrifugal drip are observed
- L.D. Yates et al.
Effect of temperature and pH on the post-mortem degradation of myofibrillar proteins
- L.D. Yates et al.
Quantitative determination of myosin and actin in rabbit skeletal muscle
Journal of Molecular Biology
- K. Takahashi
Structural weakening of skeletal muscle tissue during post- mortem ageing of meat. The non-enzymatic mechanism of meat tenderization
- H. Negishi et al.
The origin of the 30 kDa component appearing during post-mortem ageing of bovine muscle
- M. Mikami et al.
Degradation of myofibrils from rabbit, chicken and beef by cathepsin L and lysosomal lysates
- C.Y. Ho et al.
Identification of the 30 kDa polypeptide in postmortem skeletal muscle as a degradation product of troponin-T
- J.D. Fritz et al.
Titin content of beef in relation to tenderness
- E. Bandman et al.
An immunological method to assess protein degradation in post-mortem muscle
- P.J. Bechtel et al.
Effects of postmortem storage and temperature on muscle protein degradation: analysis by SDS gel electrophoresis
Journal of Food Science
- J.W.C. Bird et al.
Proteolytic enzymes in striated and non-striated muscle
Effect of postmortem storage on the Z-line region of titin in bovine muscle
Journal of Animal Science
Effect of electrical stimulation on post-mortem titin, nebulin, desmin, and troponin-T degradation and ultrastructural changes in bovine longissimus muscle
Journal of Animal Science
Relationship between proteolytic changes and tenderness in prerigor lactic acid marinated beef
Journal Science of Food Agriculture
Changes in titin and nebulin in postmortem bovine revealed by gel electrophoresis western blotting and immunofluorescence microscopy
Journal of Food Science
Factors affecting polyacrylamide elektrophoresis and electroblotting of high molecular weight myofibrillar protein
Determinants of tenderization in beef longissimus dorsi and triceps brachii muscles
Evaluation of structural changes in cured meat through the determination of myosin in the centrifugal drip
Roczniki Instytutu Przemysłu Miêsnego i Tłuszczowego
Effect of electrical stimulation on postmortem titin, nebulin, desmin, and troponin-T degradation and ultrastructural changes in bovine longissimus muscle
Journal of Animal Science
Preliminary investigation of the use of Raman spectroscopy to predict beef spoilage in different types of packaging
2020, Meat Science
In this study, pH, meat color analysis, microbial counts and Raman spectroscopic data were obtained from beef steaks stored at 4°C for up to 21days using two different packaging methods: vacuum (VP) and modified atmosphere packaging (MAP). Models using partial least square regression (PLSR), indicated that Raman spectroscopy was able to predict total viable counts (TVC) and lactic acid bacteria (LAB) measured at 21d post mortem (TVC in VP: R2cv=0.99, RMSEP=0.61; TVC in MAP: R2cv=0.90, RMSEP=0.38; LAB in VP: R2cv=0.99, RMSEP=0.54; LAB in MAP: R2cv=0.75, RMSEP=0.60). The results of this study demonstrate that Raman spectroscopy may have potential for the rapid determination of meat spoilage.
Proteomic approach to investigate the impact of different dietary supplementation on lamb meat tenderness
2017, Meat Science
Citation Excerpt :
The highest MLC2 percentage in LS meat suggests that dietary supplementation could affect the postmortem degradation processes and the alteration of the acto-myosin complex integrity. However, previous studies (Kolczack, Pospiech, Palkaa, & Lacki, 2003; Lametsch, Roepstorff, & Bendixen, 2002) reported that MLC2 does not undergo major changes during postmortem storage. Degradation of myofibrillar proteins, including desmin and proteins of troponin complex, could explain the tenderization process of meat (Anderson et al., 2012; Huff-Lonergan et al., 1996; Marino et al., 2015).
The aim of this study was to evaluate the effect of dietary supplementation of linseed and/or quinoa on tenderness and on proteome of lamb meat. Thirty-two Italian Merino lambs were distributed into 4 groups with different diet: control (CO) with no supplemental fat, linseed (LS), quinoa (QS) and QS+LS diets. Meat obtained by lamb fed linseed showed the lowest values of WBSF (P<0.001), hardness (P<0.01), gumminess (P<0.01) and chewiness (P<0.01). Proteomic changes of myofibrillar and sarcoplasmic proteins were estimated with SDS-PAGE, Western Blot and Two-Dimensional Gel Electrophoresis. In linseed group proteomic analysis revealed a degradation of desmin and TnT proteins complex and a major number of spots and phosphorylation isoforms of fast MLC2 patterns. Meat obtained by lamb fed quinoa showed a minor effect on the instrumental evaluation of meat tenderness and a major number of spots ascribed to sarcoplasmic proteins and fMHC. Data suggest that dietary supplementation may act on meat tenderness and on proteolytic pattern of myofibrillar fraction.
Exudate Protein Composition and Meat Tenderness of Broiler Breast Fillets
2016, Poultry Science
The relationship between meat tenderness and the protein composition of muscle exudates collected from broiler breast fillets deboned at different postmortem times was investigated. A total of 85 broilers were processed and breast fillets from each carcass were deboned at either 2 h (early-deboned, EB) or 24 h (control) postmortem. One fillet per carcass was used for 1 d postmortem meat tenderness measurements and the other fillet was stored at 4°C until 6 d postmortem for the collection of exudate prior to tenderness evaluation. Protein content and composition of muscle exudates were determined by a biuret assay and SDS-PAGE. Fillet pH, color, drip loss, and cook loss were also measured. Early-deboned fillets exhibited greater (P < 0.05) Warner-Bratzler shear force (WBSF) than controls at 1 d (7.4 vs. 3.1 kg) and 6 d (4.1 vs. 2.5 kg). Deboning time did not influence pH or color values (L*a*b*). Control fillets exhibited less drip loss after 6 d of storage (P = 0.005) and less cook loss at 1 and 6 d (P < 0.001). Exudate protein concentration was not influenced by deboning time. From the SDS-PAGE profiles of the exudates, the relative abundances of seventeen protein bands were quantified. Electrophoresis analysis revealed that, in general, the protein profiles of exudates from control and EB fillets were not distinct from each other. However, the band corresponding to 225 kDa was more abundant in controls (P = 0.021). Although the protein concentrations and SDS-PAGE profiles of muscle exudates varied widely between breast fillets, variations in exudate protein characteristics were not strongly associated with changes in the tenderness of broiler breast meat due to the combined effects of postmortem deboning time and post-deboning aging.
Changes in meat quality traits and sarcoplasmic proteins during aging in three different cattle breeds
2014, Meat Science
Citation Excerpt :
Several studies (Revilla & Vivar-Quintana, 2006; Ruiz de Huidobro, Miguel, Onega, & Blazquez, 2003) reported that these parameters are dependent on the rate and extent of post mortem aging. A number of factors contribute to post mortem tenderization of meat, in particular, many authors (Huff-Lonergan, Zhang, & Lonergan, 2010; Kolczak, Pospiech, Palka, & Lacki, 2003; Koohmaraie, 1996; Taylor, Geesink, Thompson, Koohmaraie, & Goll, 1995) indicate that proteolytic degradation of myofibrillar proteins plays an important role in tenderization showing ultrastructural changes in skeletal muscle. Besides myofibrillar proteins, sarcoplasmic fraction is the other compound of muscle proteins that accounts for 30–35% of total proteins and is primarily represented by glycolytic enzymes and myoglobin.
The effects of breed and aging time (1, 7, 14, 21d) were evaluated on physical meat properties and on sarcoplasmic protein changes in 24 young bulls from Romagnola×Podolian, Podolian and Friesian breeds. Aging affects lightness showing an increase in all breeds while changes in redness varied according to the breed. Podolian breed showed meat with the darkest and the reddest color and the lowest drip loss compared to the other breeds. Extending aging to 21d reduced drip loss from meat. SDS-PAGE and 2DE showed that many changes in the sarcoplasmic proteins occurred among breeds and during aging. During post-mortem some sarcoplasmic proteins decline in intensity after 21d highlighting that they were susceptible to aging. Protein identification and western blotting showed the presence of myosin light chains, Troponin T and tropomyosin proteins during aging, suggesting a degradation of myofibers and a more intense proteolysis especially in the Podolian breed.
Effect of pH and postmortem aging on protein extraction from broiler breast muscle
2014, Poultry Science
Citation Excerpt :
Alterations to the protein contents and profiles of the extracts in this study could have been partially due to aging and pH induced protein shifts between the 2 fractions upon extraction. Several reports on beef have suggested that proteins of myofibrillar origin accumulate in the sarcoplasmic fraction or drip extract with postmortem aging (Xiong and Anglemier, 1989; Kolczak et al., 2003). These protein bands were thought to be aging-related degradation products of high molecular weight myofibrillar proteins that were more water-soluble than the intact proteins.
This study determined the effects of extraction buffer pH and postmortem aging on the extraction of salt-soluble and water-soluble proteins from broiler pectoralis muscle. Deboned broiler breast fillets were collected at 4 h postmortem, packaged, and then stored at 4°C until 1, 5, or 8 d postmortem. After the designated aging period, salt-soluble and water-soluble protein extractions were performed using buffers at 7 different pH levels (pH 5.4, 6.4, 6.9, 7.2, 7.5, 8.0, 9.0). Protein concentrations of the extracts were measured and SDS-PAGE analysis was performed. Salt-soluble protein concentration increased (P < 0.0001) as buffer pH increased from pH 5.4 to 6.9 and then remained unchanged from pH 6.9 to 9.0. Water-soluble protein concentration increased (P < 0.0001) as buffer pH increased from pH 5.4 to 7.2 and then remained unchanged from pH 7.2 to 9.0. There was not a significant extraction buffer pH by aging treatment interaction for the total protein concentration of either the salt-soluble or water-soluble protein extracts. The protein concentrations of salt-soluble extracts were similar at both 1 and 8 d postmortem but lower (P < 0.0001) at 5 d postmortem. The protein concentrations of water-soluble extracts were similar at both 1 and 5 d postmortem, but higher (P < 0.0001) at 8 d. Both extraction buffer pH and postmortem aging influenced the SDS-PAGE protein profiles of salt-soluble and water-soluble protein extracts from breast muscles. Data demonstrate that postmortem aging and extraction buffer pH influence both the total amount and the composition of the myofibrillar and sarcoplasmic proteins that can be extracted from broiler breast fillets.
2014, Encyclopedia of Meat Sciences
Modeling is an emerging technique to qualitatively understand the complex interactions among phenomena that affect meat quality and to make quantitative predictions of how these phenomena affect the many quality attributes that are important to consumers. This article reviews both causal and statistical modeling approaches, their advantages and disadvantages, and thus their applicability to different contexts and applications. Examples of each approach are provided, along with indications of the insights that can be gained from each.
Enzyme immunoassay and proteomic characterization of troponin I as a marker of mammalian muscle compounds in raw meat and some meat products
Meat Science, Volume 105, 2015, pp. 46-52
The skeletal muscle protein troponin I (TnI) has been characterized as a potential thermally stable and species-specific biomarker of mammalian muscle tissues in raw meat and meat products. This study proposed a technique for the quantification of TnI comprising protein extraction and sandwich enzyme-linked immunosorbent assay (ELISA). The technique is characterized by a TnI detection limit of 4.8ng/ml with quantifiable concentrations ranging from 8.7 to 52ng/ml. The method was shown to be suitable for detection of TnI in mammalian (beef, pork, lamb, and horse) meat but not in poultry (chicken, turkey, and duck) meat. In particular, the TnI content in beef was 0.403±0.058mg/g of wet tissue. The TnI estimations obtained for the pork and beef samples using ELISA were comparable to the proteomic analysis results. Thus, the quantitative study of TnI can be a convenient way to assess the mammalian muscle tissue content of various meat products.
Proteome basis of pale, soft, and exudative-like (PSE-like) broiler breast (Pectoralis major) meat
Poultry Science, Volume 95, Issue 11, 2016, pp. 2696-2706
Pale, Soft, and Exudative (PSE) broiler breast meat has poor protein functionality, which leads to quality problems and economic loss in the poultry industry. Proteomics has been applied to characterize the biochemical mechanisms governing tenderness, color, and water-holding capacity in meat. However, the proteome basis of PSE has not yet been characterized for broiler breast meat. Therefore, this study was conducted to determine the differences in meat quality (cooking loss and shear force), descriptive sensory characteristics, consumer acceptance, and whole muscle proteome between normal and PSE-like broiler breast meat. Male Hubbard × Cobb 500 birds (n = 1,050) were raised in commercial houses. Prior to harvest, a sample of the broilers (n = 900) were subjected to short-term stress (38°C for 2 h), and the remaining broilers (n = 150) were maintained at control conditions (21°C for 2 h). Broiler breast (Pectoralis major) meat was collected and characterized by pH24 and L*24 as normal (pH24 5.8 to 6.2, L*24 45 to 55) or PSE-like (pH24 5.4 to 5.7, L*24 55 to 65) samples. Normal broiler breast meat had lower shear force values than PSE-like meat (P < 0.05). Based on sensory descriptive analysis, normal cooked chicken breast meat was more tender and juicier than PSE-like breast meat (P < 0.05). Consumer sensory analysis results indicated that 81% of consumer panelists liked normal breast meat whereas 62% of the panelists liked PSE-like breast meat. Whole muscle proteome profiling identified fifteen differentially abundant proteins in normal and PSE-like broiler breast samples. Actin alpha, myosin heavy chain, phosphoglycerate kinase, creatine kinase M type, beta-enolase, carbonic anhydrase 2, proteasome subunit alpha, pyruvate kinase, and malate dehydrogenase were over-abundant (P < 0.05) in PSE-like broiler breast whereas phosphoglycerate mutase-1, alpha-enolase, ATP-dependent 6-phosphofructokinase, and fructose 1,6-bisphosphatase were over-abundant (P < 0.05) in normal meat. Thus, results indicated that differences in proteome abundance could be related to the meat quality differences between normal and PSE-like broiler breast meat.
Relationship between hardness and myowater properties in Wooden Breast affected chicken meat: A nuclear magnetic resonance study
LWT, Volume 86, 2017, pp. 20-24
The role of myowater-holding on the development of the hardness of Wooden Breast (WB) affected chicken breasts was investigated. Transverse (T2) relaxation times and proportions of myowater populations (T2B, T21 and T22) were assessed using low-field nuclear magnetic resonance (NMR) relaxometry and integrated with meat compression measurements. Two muscle conditions (M: Normal (N) vs WB), four sampling locations (L), four sampling times (T) and interactions (M x L and M x T) were considered. Compared to N, WB was harder, the extramyofibrillar myowater population (T22) was increased and the relaxation time of the water trapped into the myofibrillar matrix (T21) was also increased. A link between the T21 relaxation time of water trapped into the myofibrillar matrix and hardness was suggested for the WB muscles. During storage, a redistribution of water occurred over time, as revealed by an increasing trend of the T21 population, but a concomitant texture evolution did not reflect this change. The cranial/superficial part of the breasts exhibited the highest amount of the extramyofibrillar water population (T22), and the texture of this muscle part was harder than the deep layers. However, the role of myowater on muscle hardness was not fully clarified by this study.
Microstructure and physical changes in the Mexican cooked lamb meat barbacoa made with chilled and frozen meat
Meat Science, Volume 118, 2016, pp. 122-128
Longissimus dorci (LD) samples of different origin (imported and domestic) with pre-treatments (imported meat stored at −18°C for 6months, domestic meat stored at −18°C for 10days, and domestic meat stored at 4°C for 24h) were cooked as barbacoa and frozen using two treatments (air blast and liquid immersion) and then evaluated after 30days of storage. The results showed that the origin and pre-treatment of meat affected L*, a*, instrumental texture and microstructure; that the storage time affected pH, aw, b* and microstructure; and that the freezing treatments did not affect the meat. Overall, the frozen cooked lamb dish barbacoa could present some problems at the conservation stage due to an increase in pH, aw and changes in microstructure; however, the physical traits (color and texture) remained mostly unchanged and depended more on the quality of the raw meat.
Regulation of calpain-1 activity and protein proteolysis by protein nitrosylation in postmortem beef
Meat Science, Volume 141, 2018, pp. 44-49
The effects of nitric oxide (NO) and its induced protein nitrosylation on calpain-1 activation and protein proteolysis in beef during postmortem aging were investigated. Five semimembranosus muscles were removed from beef cattle carcass. Beef samples were incubated with one of following treatments for 24 h at 4 °C: control (normal saline), NO donor (100, 200 and 400 μM S-nitrosoglutathione (GSNO)) or nitric oxide synthase (NOS) inhibitor (0.05, 0.1 and 0.15 M Nω-Nitro-l-arginine methyl ester hydrochloride (L-NAME)). After incubation, the beef samples were vacuum-packaged and aged at 4 °C for 1, 4, and 7 days. Results showed that GSNO decreased and L-NAME increased the extent of calpain-1 autolysis at d 1. Degradation of desmin and troponin-T was increased by L-NAME while decreased by GSNO. These results suggest that NO could regulate calpain-1 autolysis and its proteolysis activity during postmortem aging in beef SM muscle.
Cleavage of the calpain inhibitor, calpastatin, during postmortem ageing of beef skeletal muscle
Food Chemistry, Volume 148, 2014, pp. 1-6
The objective of this study was to investigate the contribution of caspase and calpain, on the proteolysis of calpastatin in postmortem beef muscle, by examining the influences of calpain inhibitor MDL-28170 and caspase-3 inhibitor DEVD-CHO on calpastatin degradation and the in vitro proteolysis of calpastatin by caspase-3, -6 and μ-calpain. In this study, both calpain- and caspase-3-inhibitors suppressed postmortem degradation of calpastatin. In vitro treatment of calpastatin with μ-calpain resulted in degradation products similar in size to those occurring naturally in aged beef muscle. With addition of caspase-3, only the 100kDa degradation fragment was present during the early phase of ageing, and subsequently, was likely to have been inactivated by calpain or other factors. Therefore, calpain was the major contributor to the proteolysis of calpastatin in postmortem beef muscle. While caspase-3 was involved in calpastatin degradation during the early postmortem period, calpastatin maybe plays an important role in bridging the gap between caspase and calpain systems.
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