电刺激和延迟冷却对宰后牛肉肌原纤维蛋白降解变化的影响

本文研究了电刺激和延迟冷却对中国杂交黄牛宰后肌原纤维蛋白降解变化的影响.分析了宰后成熟0,3,7,10天时肌原纤维小片化指数的变化,并利用蛋白免疫印迹分析了肌间线蛋白Desmin和伴肌动蛋白Nebulin的降解情况.结果表明:电刺激和延迟冷却提高了肌原纤维小片化的程度,促进了Desmin和Nebulin的降解,缩短了成熟时间.     

宰后不同成熟时间对羊肉肌原纤维蛋白降解及钙激活酶活性变化的影响

选择新疆巴什拜羊背最长肌为试验材料,分别在4℃下成熟1、3、7、14、21 d后测定分析其中肌原纤维蛋白、肌间线蛋白(Desmin)、肌钙蛋白-T(Troponin-T)及钙激活酶活性的变化。结果表明,在成熟过程中肌原纤维发生了降解,其中肌间线蛋白、肌钙蛋白-T发生了显著的降解,同时钙激活酶中μ-钙激活酶的活性也逐渐降低。从而证实μ-钙激活酶活性与肌原纤维蛋白降解存在负相关的关系。由此可知钙激活酶在羊肉成熟过程,参与了肌原纤维的降解,促进了羊肉的成熟嫩化。     

CaCl2和木瓜蛋白酶处理对羊肉品质的影响

研究不同浓度的CaCl2注射或不同质量浓度的木瓜蛋白酶浸泡处理对云岭黑山羊股二头肌pH值、肉色、蒸煮损失、肌原纤维小片化指数、胶原蛋白、肌浆蛋白和肌原纤维蛋白降解的影响。结果表明：CaCl2注射或木瓜蛋白酶浸泡处理显著提高了肌原纤维小片化指数;0h和6h时对照组出现极限pH值,分别为6.75和6.15,而CaCl2处理组或木瓜蛋白酶处理组到达极限pH值时间为2h和24h,极限pH值分别为 6.83、6.29和6.90、5.89,SDS-PAGE分析显示CaCl2和木瓜蛋白酶促使羊肉肌肉中的肌动蛋白、肌球蛋白和连接蛋白发生降解。在0.3mol/L CaCl2或0.002g/100mL木瓜蛋白酶处理时,处理组的肌原纤维小片化指数和胶原蛋白的含量达到最大,蒸煮损失最小,表现出最佳的嫩化效果。     

藏羊肉宰后成熟过程中热休克蛋白27对肌原纤维蛋白及细胞凋亡酶的影响

为研究藏羊肉宰后成熟过程中热休克蛋白27(heat shock protein 27,Hsp27)对肌原纤维蛋白降解特性及细胞凋亡酶活力的影响,以经Hsp27抑制剂KRIBB3处理的藏羊肉为研究对象,测定藏羊肉在宰后成熟过程中Caspase-3、Caspase-9活力变化以及肌原纤维蛋白的溶解性和降解特性,并通过体外实验进一步研究Hsp27对μ-钙激活酶和Caspase-3降解肌原纤维蛋白的影响。结果表明:Hsp27能显著抑制Caspase-3、Caspase-9的活力(P0.05),并能够抑制肌原纤维小片化的发生以及肌间线蛋白和肌钙蛋白-T的降解;体外实验发现,Hsp27能够抑制μ-钙激活酶和Caspase-3对肌原纤维蛋白的降解。     

延边黄牛肉干、湿法成熟过程中嫩度的变化

目的：研究干法成熟和湿法成熟过程中牛肉嫩度的变化。方法：以延边黄牛臀肉为材料，测定其在宰后干法成熟与湿法成熟30 d内剪切力值、pH、水分含量、肌原纤维小片化指数、蛋白降解情况及肌纤维组织学特性等指标的变化。结果：随着成熟时间的延长，干法成熟和湿法成熟牛肉样品剪切力值均显著降低，成熟30 d时剪切力值从91.7 N分别降到51.6 N和39.8 N，肌原纤维小片化指数、肌原纤维蛋白降解变化显著；湿法成熟牛肉样品30 d时肌内膜降解，肌纤维排列紧密；牛肉中肌酸激酶和醛缩酶等相对分子质量大的肌浆蛋白分解成为相对分子质量小的蛋白，且种类增多浓度增大。结论：宰后成熟显著提升牛肉嫩度，湿法成熟样品肌原纤维碎裂、肌浆蛋白以及肌细胞中结缔组织降解程度相比于干法成熟更强，对肉嫩度影响较大。     

猪宰后正常肉与PSE肉中肌联蛋白和伴肌动蛋白变化

运用SDS-PAGE电泳技术对猪宰后正常肉与PSE肉的肌联蛋白和伴肌动蛋白的变化进行测定,并分析了它们与持水性的关系。结果显示肌联蛋白在宰后逐步降解。正常肉样中,宰后24 h肌联蛋白开始迅速降解(p0.05);PSE肉样的肌联蛋白在宰后2 d开始迅速降解(p0.05)。伴肌动蛋白在宰后逐步降解。正常肉样中,伴肌动蛋白在宰后2 d出现明显降解(p0.05);PSE样肉中,伴肌动蛋白在宰后24 h出现明显降解(p0.05)。汁液流失率与宰后肌联蛋白和伴肌动蛋白的完整度有正相关性。宰后正常肉样与PSE肉样的细胞间隙面积都随贮藏时间的延长而增大,PSE组的细胞间隙面积始终显著大于正常组(p0.05);汁液流失率与细胞间隙面积呈显著正相关(p0.05);宰后肌联蛋白和伴肌动蛋白完整度与细胞间隙面积呈正相关。结果表明宰后肌联蛋白和伴肌动蛋白降解越多,细胞间隙面积越小,汁液流失率越小,持水力越强。     

宰后成熟期间结构蛋白对秦川牛肉嫩度的影响

为探究宰后成熟期间结构蛋白对秦川牛背最长肌嫩度的影响,测定不同贮藏期（0、2、4、6、8 d）内秦川牛背最长肌剪切力、肌原纤维小片化指数（myofibrillar fragmentation index,MFI）和蛋白含量等,并利用4D-非标记定量（4D-label free quantification,4D-LFQ）蛋白质组学技术分析蛋白质组学变化。结果表明：在贮藏期0～8 d内,剪切力总体呈先升后降的趋势（P＜0.01）,上升的幅度小于下降的幅度;秦川牛背最长肌MFI呈极显著上升趋势（P＜0.01）,总体增长了250.81%;总可溶性蛋白含量呈显著下降趋势（P＜0.05）,总体下降了34.60%;通过宰后肌肉组织代谢变化,肌肉组织结构蛋白发生降解,可能会影响到嫩度的形成,肌原纤维蛋白含量呈显著下降趋势（P＜0.05）,前期下降速度较快,后期下降速度减缓,总体下降了50.56%。在贮藏期0～4 d内,通过骨骼肌组织发育过程调控钙离子结合和细胞骨架蛋白结合途径,4 种蛋白（α-肌动蛋白-1、牛肌球蛋白重链9、牛肌球蛋白轻链2、肌球蛋白调节轻链12B）丰富度发生变化;在贮藏期0～8 d内,通过肌肉器官发育和横纹肌组织发育过程调控钙离子结合途径,8 种蛋白（肌球蛋白调节轻链2、肌球蛋白重链6、α-肌动蛋白-1、心肌肌动蛋白α1、牛肌球蛋白轻链2、肌钙蛋白I 1型、肌钙蛋白I 2型、肌球蛋白重链15）丰富度发生变化,通过肌球蛋白结合、钙离子结合、细胞骨架蛋白结合的肌原纤维组装、骨骼肌组织发育、肌肉器官发育、横纹肌组织发育过程等途径调控细胞的生理状态,结构蛋白降解造成肌原纤维小片化升高,进而促使嫩度提升。     

高浓度CaCl2溶液对某些肌原纤维蛋白的作用研究

用0、100、200、300mmol/LCaCl2溶液处理肌球蛋白、肌动蛋白和肌原纤维,通过SDS-PAGE和免疫印迹电泳研究100、200、300mmol/L高浓度CaCl2溶液处理下肌球蛋白、肌动蛋白、α-肌动素和肌钙蛋白-T的蛋白降解情况。SDS-PAGE结果显示,肌球蛋白、肌动蛋白在100、200、300mmol/LCaCl2溶液处理下1、7d时的电泳结果与对照组相比没有显著差异。免疫电泳结果显示,100、200、300mmol/L高浓度CaCl2溶液处理后,α-肌动素和肌钙蛋白-T与对照组相比发生显著降解,随着浓度升高,降解程度加大,在300mmol/L浓度时,α-肌动素条带密度显著降低,肌钙蛋白-T也裂解为30kD以下的更小片段。     

高强度超声对鹅胸肉嫩度及品质的影响

为了解超声技术对鹅胸肉品质的影响,采用高强度超声（超声功率800 W,超声总时间42 min,工作时间 2 s、停歇时间3 s）对鹅胸肉进行处理。测定未超声和超声处理的鹅胸肉于4 ℃下放置不同时间（0、6、12、24、 36、48 h）的pH值、肌原纤维小片化指数、蒸煮损失率、剪切力、表面微观结构、肌原纤维蛋白降解情况、肉品 的热力学性质等指标。结果显示：与未经超声处理的鹅胸肉相比,超声处理后随着放置时间的延长,肉品pH值升 高,肌原纤维小片化指数显著增大,蒸煮损失率和剪切力显著减小（P＜0.05）;表面微观结构发生明显变化,肌 动蛋白和肌球蛋白热敏性增大,导致更多肌原纤维蛋白降解,游离肌动蛋白含量显著增加（P＜0.05）,从而显著 改善鹅胸肉的嫩度。因此,超声处理可破坏鹅胸肉肌原纤维的完整结构,缩短宰后肉品成熟所需时间,进一步提高 鹅胸肉的品质。     

宰后僵直及成熟过程中羊背最长肌理化性质的变化

动物宰后从肌肉到肉品经过僵直、成熟等一系列复杂的生理生化反应。僵直及成熟过程的研究可为肉品质改善及肉制品加工提供理论依据。选取杂交公羊(小尾寒羊×北京本地羊)双侧背最长肌在4℃成熟0.5、2.0、6.0、12.0、24.0、48.0、72.0、120.0、168.0 h,测定不同时间的pH值、剪切力、肌节长度、ATP含量、肌原纤维小片化指数(myofibril fragmentation index,MFI)、钙蛋白酶活力。结果表明:宰后成熟过程中,pH值先下降后逐渐趋于稳定,从第24小时后变化不显著(P0.05);剪切力先上升后下降,在第24小时达到最大值;肌节长度在宰后先缩短后逐渐变长,且在第48小时缩至最短;ATP含量先上升后下降,48 h后趋于稳定;μ-钙蛋白酶80 kD大亚基在宰后24 h基本降解完全,其降解的78 kD大亚基在第48小时降解完全;肌间线蛋白和肌钙蛋白T作为μ-钙蛋白酶的降解底物,在成熟过程中发生降解,第168小时几乎观察不到完整的蛋白条带;随着宰后时间的延长,羊背最长肌从第2小时开始僵直,到第24小时程度达到最大,从第48小时开始解僵,解僵后肉的嫩度逐渐改善。     

Effect of meat ultimate pH on rate of titin and nebulin degradation

The processes involved in the tenderisation of meat were studied on muscles with a range of ultimate pH values (5.4–7.0), produced by subcutaneous injection of various doses of adrenaline and exercise. The m. longissimus thoracicum et lumborum (LD) was removed from carcasses stored at 10 °C and held for 1, 3 or 6 days after slaughter, then frozen until tenderness assessment. The tenderness of meat cooked from the frozen state was determined as the force to shear samples of 10 mm × 10 mm cross-section using a MIRINZ tenderometer. The maximum toughness of 15 kgF occurred at an ultimate pH (pH_u) of about 6.0, resulting in a curvilinear relationship between tenderness and pH_u at 1 day post-slaughter. By 6 days post-slaughter, all meat had reached the same low shear value of approximately 3 kgF. SDS-PAGE patterns obtained from samples at 12, 24 and 48 hr post-slaughter showed increasing titin and nebulin degradation over time, with the slowest rate of degradation occurring at pH_u values 6.0–6.3. Titin and nebulin are known to play an important role in the stabilisation of myofibril structure, and it is suggested that the curvilinear relationship results when pH-dependent titin and nebulin degradation occurs.     

The relationship between meat tenderization, myofibril fragmentation and autolysis of calpain 3 during post-mortem aging

The objective was to study the potential role of calpain 3 in postmortem myofibril breakdown and meat tenderization. We determined the temporal changes in calpain 3 protein in the ovine m. longissimus thoracis et lumborum (LTL, n=4) during post-mortem storage. Concurrently, we also determined the kinetics of tenderization level, changes in MFI, degradation of nebulin and desmin, and autolysis of calpain 1. The autolysis of calpains 1 and 3 were strongly correlated with the kinetics of tenderization and changes in MFI. The best correlation was between the appearance of the autolyzed calpains 1 and 3 and nebulin degradation. Taken together, the results indicated that calpains 1 and/ or 3 might be playing a key role in post-mortem tenderization of LTL via the proteolysis of specific muscle structural proteins such as nebulin. This is the first report that relates calpain 3 to myofibrillar protein degradation in post-mortem skeletal muscle.     

THE DEGRADATION OF MYOFIBRILLAR PROTEINS IN BEEF AND LAMB USING DENATURING ELECTROPHORESIS - AN OVERVIEW

Degradation of specific myofibrillar proteins has been followed in many studies using SDS-PAGE (denaturing) electrophoresis and these have shown that proteins such as titin, nebulin, troponin-T, desmin, filamin and vinculin are degraded at different rates during postmortem storage of meat. Although informative, electrophoresis is usually restricted to qualitative interpretation of the gels and there have been few studies that have quantitatively linked the degradation of specific proteins to changes in toughness. Improved quantitative methods (including scanning densitometry, image analysis, and modeling approaches) will be necessary to determine the protein and/or protein alterations that cause postmortem tenderization. This approach is complementary to other measurements of proteolysis where the objective is to more adequately explain the variation in toughness.     

Combination of low voltage electrical stimulation and early postmortem temperature conditioning on degradation of myofibrillar proteins in Korean native cattle (Hanwoo)

The combination effect of low voltage electrical stimulation (LVES) and early postmortem (PM) temperature conditioning (2, 16, and 30°C until 3 h PM) on degradation of myofibrillar proteins were determined from Korean native cattle (Hanwoo). Myofibrils were removed at 1, 2, 3, 7, and 14 days of PM storage (2°C) and analyzed for titin, nebulin, desmin, and troponin-T by SDS-PAGE and by Western blot analysis. Degradation rate of myofibrillar proteins was affected by the combination of LVES and temperature conditioning. LVES-30°C treatment resulted in faster degradation of titin, nebulin, desmin, and troponin-T during PM storage than the other treatments. Degradation of titin took place more slowly than nebulin, desmin or troponin-T.     

不同品种牛和部位肌肉在成熟过程中品质差异比较研究

为探究肌肉部位和成熟时间对鲁西黄杂交牛和安秦杂交牛牛肉食用品质的影响,分别选取2种牛宰后的背最长肌、腰大肌和半膜肌3个部位肉,在成熟5、7、14、21?d时测定其嫩度、肌节长度、肉色和蛋白降解程度等指标.结果表明:随着成熟时间的延长,2种牛腰大肌剪切力值较低,肌节长度较长,肉色更好,而背最长肌和半膜肌剪切力更高,肌节长...     

Preliminary study on the effect of caspase-6 and calpain inhibitors on postmortem proteolysis of myofibrillar proteins in chicken breast muscle

The objective was to determine the effect of three different protease inhibitors, caspase-6 specific inhibitor VEID-CHO (N-Acetyl-Val-Glu-Ile-Asp-al), calpain inhibitor leupeptin or calpain inhibitor EGTA on protein degradation, ultrastructure of myofibrils and calpain activity during postmortem (PM) aging of chicken muscle. Results showed that proteolysis of nebulin, troponin-T and desmin during 14-days postmortem storage were inhibited significantly by leupeptin. Inhibitive effects of VEID-CHO and EGTA on these protein degradations were significant only during 1-day postmortem storage. The activities of calpains were inhibited noticeably by leupeptin and EGTA, but not by VEID-CHO. Samples treated with VEID-CHO, leupeptin and EGTA retarded structural disruption of chicken muscle fibers. These results demonstrate that calpain is a major contributor to PM tenderization; while caspase-6 plays, if any, a minimal role in the conversion of chicken muscle to meat.     

Influence of caspase3 selective inhibitor on proteolysis of chicken skeletal muscle proteins during post mortem aging

The objective of this study was to investigate the potential contribution of apoptosis related downstream executioner caspase3 to post mortem skeletal muscle proteolysis by use of caspase3 selective inhibitor DEVD–CHO (N-acetyl–Asp–Glu–Val–Asp–CHO). After slaughter, four chicken breast muscles were removed and cut into small pieces, then marinated in treatment solution containing DEVD–CHO, or in control solution, and stored at 4 °C for 1, 3 or 7 d. Meat samples were obtained and used for detecting muscle protein degradation or calpain activity. Results showed that DEVD–CHO had inhibited the degradation of muscle skeletal proteins (titin, nebulin, desmin and troponin-T) significantly, whereas the activity of calpains had not been influenced. Therefore, the degradation of muscle proteins should not been exclusively attributed to the calpain system, and the effector caspase3 may be a new protease involved in meat post mortem tenderization.     

Evidence against the non-enzymatic calcium theory of tenderization

The objective of the present study was to determine whether variation in the tenderization of lamb longissimus could be attributed to variations in the rise in free calcium postmortem and sarcomere lengthening post rigor. The longissimus muscle of 10 crossbred lambs (Romney×Coopworth) was sampled at 1 and 7 days postmortem for determination of MIRINZ shear force, myofibrillar fragmentation index (MFI), sarcomere length, free calcium, and proteolysis of troponin-T. Despite considerable variation in tenderness and tenderization of the muscles, sarcomere lengthening was not observed. The concentration of free calcium at 7 days postmortem correlated significantly with the MFI (r=0.640; P<0.05) and tended to correlate with the shear force (r=−0.596; P<0.1) and degradation of troponin-T (r=0.625; P<0.1). Degradation of troponin-T was significantly correlated with tenderization (r=0.664; P<0.05). Troponin-T is a calpain substrate, but reportedly is not degraded through a direct effect from calcium. The present results, therefore, suggest that the variation in free calcium in postmortem muscle affects tenderization through an effect on the calpain system and not through a direct effect of calcium on myofibrillar proteins. Consequently, the results of this study do not support the (calcium) theory that calcium directly affects tenderization.     

Postmortem role of calpains in Pekin duck skeletal muscles

BACKGROUD: It is generally agreed that calpains are involved in postmortem proteolysis of skeletal muscle and improve meat tenderness. However, little information regarding the postmortem role of calpains in duck skeletal muscle is known. Therefore, the purpose of this study was to examine the role of calpains in Pekin duck postmortem breast muscles (BM) and leg and thigh muscles (LM) muscles at 5 °C. RESULTS: The postmortem pH was lower (P < 0.05) in BM than in LM. Western blots indicated that postmortem desmin degradation and the 30/32 kDa troponin-T degradation product accumulation were more rapid in BM than in LM. Casein zymograms showed that at-death μ-calpain activity was higher in BM than in LM. As time post mortem increased, μ-calpain was activated and autolyzed more rapidly and extensively in BM than in LM, but μ/m-calpain was activated at a relative slower rate compared with μ-calpain. Correlation results showed that μ-calpain activity, rather than μ/m-calpain activity, in BM samples was highly correlated with the abundance of desmin and the 30/32 kDa troponin-T degradation components across the postmortem period. However, no such correlations were found with LM μ- and μ/m-calpains. CONCLUSION: Therefore, our results suggest that BM μ-calpain with a faster and more extensive activation and autolysis would play a relatively dominant role in dictating degradation of desmin and troponin-T in postmortem duck muscle.     

Biochemistry of postmortem muscle — Lessons on mechanisms of meat tenderization

It is certain that meat tenderness is a highly valued consumer trait and thus definition of the multiple processes that influence meat tenderness will provide clues toward improving meat quality and value. The natural process by which meat becomes tender is complex. Tenderness development is dependent on the architecture and the integrity of the skeletal muscle cell and on events that modify those proteins and their interaction. Specifically protein degradation and protein oxidation have been identified as processes that modify proteins as well as the tenderness of meat. The intracellular environment is a major factor that controls these events. Ultimately, the interplay between these events determines the rate and extent of tenderization. Given the intricacy of the structure of the muscle cell, coupled with the complexity of the regulation of protein modification and the ever-changing intracellular environment it is not surprising that this area of research is a very dynamic field. Just as the overall integrity and function of muscle cells does not depend on a single protein, but rather on the coordinated interaction of several proteins, the structural weakening of muscle cells during postmortem aging also must not depend on the degradation of a single myofibrillar or other cytoskeletal protein. The proteins mentioned in this review are located in different regions of the muscle cell, and most have been implicated in some manner as being important in maintaining the structure and function of the muscle cell. Oxidation of myosin heavy chain, a predominant protein in the myofibril, is known to promote aggregation and toughening of meat. Degradation of proteins such as desmin, filamin, dystrophin, and talin (all located at the periphery of the Z-line) may disrupt the lateral register and integrity of the myofibril themselves as well as the attachments of the peripheral layer of myofibril to the sarcolemma. Degradation of the proteins within the myofibril that are associated with the thick and thin filaments may allow lateral movement or breaks to occur within the sarcomeres of postmortem aged samples. Titin, nebulin, and troponin-T, by their ability to directly interact with, or modulate the interaction between, major proteins of the thick and thin filaments and (or) the Z-line, play key roles in muscle cell integrity. Disruption of these proteins, especially titin and nebulin, could initiate further physicochemical and structural changes that result in myofibril fragmentation and loss of muscle cell integrity, and ultimately in tenderization of the muscle. In order to make real progress in this area, the scientific community must have a global appreciation of how both the structural proteins and the key proteases are influenced by the vast changes that occur during the conversion of muscle to meat.