Water distribution and mobility in meat during the conversion of muscle to meat and ageing and the impacts on fresh meat quality attributes--a review

This paper reviews current knowledge on the distribution and mobility of water in muscle (myowater) ante- and post mortem and factors affecting these in relation to fresh meat quality parameters; water-holding capacity (WHC), tenderness and juiciness. NMR transverse relaxometry (T(2)) using bench-top Low-Field Nuclear Magnetic Resonance (LF-NMR) has characterised myowater distribution and mobility as well as structural features in meat which directly affect WHC. The current literature demonstrates that WHC is correlated to the water located outside the myofibrillar network (extra-myofibrillar). This review identifies the critical stages which affect the translocation of water into the extra-myofibrillar space and thus the potential for decreased WHC during proteolysis (the conversion of muscle to meat). This review discusses how the intrinsic properties of the water held within the meat could contribute to juiciness and tenderness. Tenderness has been shown to correlate to T(2), however breed and species differences made it difficult to draw firm conclusions. Further understanding of the inherent water properties of fresh meat and the factors affecting water distribution and mobility using NMR technologies will increase the understanding of WHC and tenderisation of fresh meat.     

The effect of ageing on the water-holding capacity of pork: role of cytoskeletal proteins

The water-holding capacity (WHC) of pork decreases post-mortem but has been shown to increase during subsequent ageing. In order to test a hypothesis that water-holding capacity increases during ageing due to degradation of the cytoskeleton, WHC was followed 10 days post-mortem and related to the extent of proteolysis of cytoskeletal proteins. A fast method for measuring WHC in small meat samples was developed by the use of centrifugation. The WHC of fresh pork decreases in the first part of post-mortem storage after which it increases to the level of 1 day PM. No changes in total water content of the meat were observed which could explain changes in WHC during ageing. Vinculin and desmin degrade gradually during ageing while talin degrades rapidly. These observations are consistent with the hypothesis that degradation of the cytoskeleton slowly removes the linkage between lateral shrinkage of myofibrils and shrinkage of entire muscle fibres, so removing the force that causes flow into the extracellular space. Inflow of previously expelled water is then possible, so increasing WHC as observed in later periods of storage.     

Relationship between water mobility and distribution and sensory attributes in pork slaughtered at an age between 90 and 180 days

Water mobility and distribution were measured in M. longissimus dorsi from 41 pigs slaughtered at an age of 90, 140, 161 or 182 days using low-field proton NMR relaxometry, and in order to investigate the impact on sensory attributes, a sensory evaluation of the pork was performed in parallel. The sensory evaluation demonstrated a significant effect of slaughter age on juiciness of the meat, and a final juiciness score of 8.4, 7.7, 7.2 and 7.8 was obtained for meat from 90, 140, 161 and 182-day old pigs, respectively. The NMR measurements revealed that the higher juiciness in meat slaughtered at an age of 90 days could be ascribed to a longer relaxation time of the extramyofibrillar water, corresponding to more mobile water, in the fresh meat of 90-day old pigs compared with the older pigs. In the cooked meat the higher juiciness of meat from 90-day old animals could be ascribed to a more homogenous distribution of the myofibrillar water compared with meat from older pigs. In contrast, the NMR measurements showed no effects that could explain the higher juiciness in meat from pigs slaughtered at an age of 182 days compared with meat from pigs slaughtered at an age of 140 or 161 days. Possibly the increase in juiciness when the age at slaughter is increased from 161 to 182 days should be ascribed to an increase in intramuscular fat content, which was not evident in the NMR measurements.     

Any link between integrin degradation and water-holding capacity in pork?

In order to investigate a possible relationship between integrin degradation and water-holding capacity (WHC), integrin was quantified using western blot, and water mobility and distribution was measured by proton NMR T(2) relaxometry at 24h postmortem in pork (n=30) with a large variation in WHC (drip loss varying from 2.8% to 11.3%). Regression analyses revealed correlation coefficients of r=-0.32 (P=0.08) and r=0.40 (P=0.03) for the correlations between the content of integrin determined by western blot analysis and WHC determined as either drip loss or by NMR, respectively. Water mobility and distribution was also measured in 18 meat samples upon 7 days of aging, which revealed a correlation (r=0.54) between integrin content determined by western blot analysis 24h postmortem and the mobility of the myofibrillar water (T(21) relaxation time) at day 7. In contrast, no correlation could be established between integrin content 24h postmortem and WHC at day 7 determined by NMR as the amount of extramyofibrillar water (T(22) population) (r=-0.01). In conclusion, both visualisation by CLSM and quantification of integrin by western blot analyses of suggested that a strong link between integrin degradation and WHC in pork is questionable, whereas integrin degradation seems to have impact on the succeeding development in the mobility of the myofibrillar water.     

NMR relaxometry and differential scanning calorimetry during meat cooking

By combining simultaneous nuclear magnetic resonance (NMR) T₂ relaxometry and differential scanning calorimetry (DSC) on pork samples heated to nine temperature levels between 25 and 75 °C, the present study investigates the relationship between thermal denaturation of meat proteins and heat-induced changes in water characteristics. Principal component analysis (PCA) on the distributed ¹H NMR T₂ relaxation data revealed that the major changes in water characteristics during heating occur between 40 and 50 °C. This is probably initiated by denaturation of myosin heads, which however, could not be detected in the DSC thermograms obtained directly on the meat. In contrast, the DSC thermograms revealed endothermic transitions at 54, 65 and 77 °C, probably reflecting the denaturation of myosin (rods and light chain), sarcoplasmic proteins together with collagen and actin, respectively. Simultaneous modelling of DSC and NMR data by partial least squares regression (PLSR) revealed a correlation between denaturation of myosin rods and light chains at 53–58 °C and heat-induced changes in myofibrillar water (T₂ relaxation time 10–60 ms) as well as between actin denaturation at 80–82 °C and expulsion of water from the meat. Accordingly, the present study demonstrates a direct relationship between thermal denaturation of specific proteins/protein structures and heat-induced changes in water mobility during heating of pork.     

Effect of freezing temperature, thawing and cooking rate on water distribution in two pork qualities

The effects of freezing temperature (-20 versus -80°C) in combination with long-term freezer storage (-20°C) on water mobility and distribution in pork of two qualities (normal and high ultimate pH) were explored using low-field NMR T(2) relaxometry. A marked effect of freezing temperature on the characteristics of intra- and extramyofibrillar water (T(2) relaxation times) in the thawed pork was demonstrated, implying that the freezing temperature in combination with prolonged freezer storage affects the distribution and chemical-physical state of water in the thawed meat. Determination of technological properties (thawing and cooking loss) revealed that the observed T(2) variations related to water distribution and water properties, which were found to be consistent with the degree of structural damage assessed by light microscopy, also were reflected in the technological quality of the meat. Low freezing temperature in combination with prolonged freezer storage was associated with increased thawing and cooking loss. In addition, pH in the fresh meat had a pronounced effect on the distribution of myofibrillar water, as a more homogenous pore size distribution was evident in meat with high pH compared with normal pH. A clear effect of cooking rate on the T(2) relaxation characteristics in the cooked pork was also demonstrated, probably reflecting a cooking rate-induced effect on the myofibrillar structures. The effect of cooking rate on water distribution resulted in a significantly lower cooking loss upon a slow cooking rate (0.5°C/min from 25 to 65°C and 0.3°C/min from 65 to 80°C) compared with a fast cooking rate (1°C/min).     

Mechanisms of water-holding capacity of meat: The role of postmortem biochemical and structural changes

Unacceptable water-holding capacity costs the meat industry millions of dollars annually. However, limited progress has been made toward understanding the mechanisms that underlie the development of drip or purge. It is clear that early postmortem events including rate and extent of pH decline, proteolysis and even protein oxidation are key in influencing the ability of meat to retain moisture. Much of the water in the muscle is entrapped in structures of the cell, including the intra- and extramyofibrillar spaces; therefore, key changes in the intracellular architecture of the cell influence the ability of muscle cells to retain water. As rigor progresses, the space for water to be held in the myofibrils is reduced and fluid can be forced into the extramyofibrillar spaces where it is more easily lost as drip. Lateral shrinkage of the myofibrils occurring during rigor can be transmitted to the entire cell if proteins that link myofibrils together and myofibrils to the cell membrane (such as desmin) are not degraded. Limited degradation of cytoskeletal proteins may result in increased shrinking of the overall muscle cell, which is ultimately translated into drip loss. Recent evidence suggests that degradation of key cytoskeletal proteins by calpain proteinases has a role to play in determining water-holding capacity. This review will focus on key events in muscle that influence structural changes that are associated with water-holding capacity.     

Functionality of myofibrillar proteins as affected by pH, ionic strength and heat treatment - a low-field NMR study

Myofibrills were extracted from porcine muscle, and their water properties were characterized using low-field nuclear magnetic resonance (NMR) T(2) relaxometry. A T(2) relaxation pattern very similar to the pattern observed in intact meat and water contents comparable to the water content in meat were observed, implying that the myofibrillar structures are responsible for retaining the majority of water in meat. The effect of pH and ionic strength in the samples was investigated as pH was adjusted to 5.4, 6.2, and 7.0 and ionic strength to 0.29, 0.46 and 0.71 M, respectively. Even though there were interactions between pH and ionic strength, the water content in the samples increased significantly with increasing pH and ionic strength. Moreover, mean T(2) relaxation times likewise increased with increasing pH and ionic strength, which reveals that the increased water retention could be ascribed to a swelling of the myofibrils and thereby increased spacing between filaments. The present study demonstrates that NMR T(2) relaxometry is a potential tool to explore how processing factors such as pH and ionic strength affect the microstructure of meat.     

Effect of salt addition on the thermal behavior of proteins of bovine meat from Argentina

Research was undertaken to investigate how the addition of sodium chloride (NaCl) and/or sodium tripolyphosphate (TPP) to sous vide cooked meat pieces produces an increase in water holding capacity (WHC). Semitendinosus muscles were injected to obtain tissue final concentrations of 0.70% NaCl, 0.25% TPP, 0.70% NaCl+0.25% TPP, and 1.20% NaCl+0.25% TPP. SDS-PAGE analysis showed increased protein solubilization in those treatments which included NaCl. Thermal analysis of whole muscles and isolated myofibrils showed the destabilizing effect of NaCl and a global stabilizing effect of TPP. Both salts together induced a destabilizing global effect, where TPP assisted NaCl in breaking the meat structure. It is suggested that the WHC increments are related to conformational changes in myofibrillar proteins and to the weakening of myofibrillar structure by the removal of myofibrillar proteins.     

The relationship between water-holding capacity and cooked meat tenderness in some beef muscles as influenced by acidic conditions below the ultimate pH

Previous studies have indicated the beneficial influence of higher than normal ultimate pH (pH(u)) on the tenderness of cooked meat. Such benefits have been indirectly linked to the influence of increased pH on the water-holding capacity (WHC) of meat above the iso-electric point (IEP) of the myofibrillar proteins. In the present study, relationships between WHC and the tenderness of some beef muscles were investigated under pH conditions within and below the IEP of the myofibrillar proteins, such that the maximum range of meat swelling was achieved. It was found that increased WHC, as measured by swelling ratio in both raw and cooked meat, markedly influenced cooked meat tenderness, irrespective of the connective tissue content of the muscles. The results fitted a series of exponential decay equations relating swelling ratio to cooked meat toughness. Additionally, strong linear decreases in toughness were apparent over the pH range 4·6 to 4·1 for the three muscle types studied.     

The significance of cooling rate on water dynamics in porcine muscle from heterozygote carriers and non-carriers of the halothane gene-a low-field NMR relaxation study

The post mortem changes in the chemical/physical state distribution of water were followed in pig muscle (M. longissimus dorsi) from heterozygote (n=12) and non-carriers (n=12) of the halothane gene exposed to two different cooling profiles using continuous low-field NMR relaxation measurements. T₂ relaxation data were analyzed using distributed exponential fitting analysis. Independent of genotype post mortem changes were observed in the two water populations characterizing water within the myofibrillar space (T₂₁) and the extra-myofibrillar space (T₂₂), respectively, as a function of chilling regime. The effect was most pronounced in samples from heterozygote carriers of the halothane gene. The obtained results strongly suggest that improved water-holding capacity of muscles upon fast chilling can be ascribed to a reduced accumulation of extra-myofibrillar water in the meat post mortem, and it is hypothesized that differences in the accumulation of extra-myofibrillar water post mortem can be ascribed largely to the time at which disruption of cell membrane integrity takes place.     

A structural approach to understanding the interactions between colour,water-holding capacity and tenderness

The colour, water-holding capacity (WHC) and tenderness of meat are primary determinants of visual and sensory appeal. Although there are many factors which influence these quality traits, the end-results of their influence is often through key changes to the structure of muscle proteins and their spatial arrangement. Water acts as a plasticiser of muscle proteins and water is lost from the myofibrillar lattice structure as a result of protein denaturation and consequent reductions in the muscle fibre volume with increasing cooking temperature. Changes in the myofilament lattice arrangement also impact the light scattering properties and the perceived paleness of the meat. Causes of variation in the quality traits of raw meat do not generally correspond to variations in cooked meat and the differences observed between the raw muscle and cooked or further processed meat are discussed. The review will also identify the gaps in our knowledge and where further investigation would beneficial.     

Physical changes of significance for early post mortem water distribution in porcine M. longissimus

The post mortem changes in water mobility and distribution were followed in porcine muscle (M. longissimus dorsi) samples using continuous low-field NMR relaxation measurements and simultaneous measurement of changes in muscle impedance as an indirect measure of membrane integrity as well as muscle contraction measurements using a rigormeter instrument. Distributed exponential fitting analysis of NMR T₂ relaxation data revealed the presence of three distinct water populations (T₂₀, T₂₁, T₂₂) within the muscle during its conversion to meat. Comparison of T₂ relaxation patterns and contraction data indicates that rigor development affects the attributes of the T₂₁ water population and thereby contributes to myofibrillar water characteristics post mortem, as the T₂₁ water population is believed to reflect inter/intra-myofibrillar water. The volume of the water population believed to reflect extra-cellular water (T₂₂) in the living muscle. Early post mortem T₂₂ decreased slightly within the first 2–3 h post mortem followed by an increase and a change in its characteristic time constant. This was ascribed to an initial muscle cell swelling followed by water being expelled from the cellular space into the extra-myofibrillar space. Comparison of changes in the T₂₂ water population and impedance characteristics within the muscle during its conversion to meat revealed close relationship between progresses in the two attributes. Obtained data strongly support that the post mortem reorganization of water is closely associated with membrane properties, which moreover was found to affect the final water-holding capacity of the meat. Finally, a model for early post mortem events leading to changes in the distribution of water within muscles is proposed.     

Pelvic suspension and fast post-mortem chilling: Effects on technological and sensory quality of pork - A combined NMR and sensory study

The effects of pre-rigor excision, which results in a fast post-mortem chilling; pelvic suspension, which results in a muscular stretching; control treatment of m. longissimus dorsi on water distribution measured by (1)H NMR relaxometry and on sensory properties were investigated in two sub-studies including a total of 16 pigs. Determination of sarcomere length revealed significant effects of the treatments on the degree of contraction, as pre-rigor excision and pelvic suspension resulted in shorter and longer sarcomere length, respectively. In addition, an effect of treatment on pH measured at 24h post-mortem was found, as pre-rigor excision was associated with a higher ultimate pH. NMR measurements revealed that water distribution in the meat was affected to a minor degree by the various treatments. However, an interaction between treatment and ageing period was observed, as the data demonstrated an effect of pre-rigor excision on water distribution in the cooked samples when meat was only aged for 2 days, but this effect was eliminated when meat was aged for 5 days. The effect of pre-rigor excision on water distribution in cooked meat after 2 days of ageing is suggested to reflect structural constraints in the meat that are eliminated during ageing. Sensory analyses demonstrated strong interactions between treatment and position on the muscle and between treatment and ageing period. However, in general pre-rigor-excised meat was sensed as significantly juicier compared with the other two treatments. Accordingly, the study demonstrates that under the present conditions the ultimate pH is more important for juiciness than the sarcomere length.     

中性氧化电解水对冷鲜草鱼肉品质及质构的影响

为探讨中性氧化电解水对冷鲜草鱼肉的保鲜效果,对冷藏条件下(4℃)冷鲜草鱼肉的品质指标及质构进行检测,并对比贮藏开始和结束后草鱼的肌原纤维组织结构的变化。结果表明:中性氧化电解水能够降低冷鲜鱼肉贮藏过程中菌落总数、假单胞菌数、挥发性盐基氮及硫代巴比妥酸值等品质指标的变化,可将货架期延长4 d左右;与对照组及减菌剂Ⅰ组相比,电解水处理可以减缓冷藏过程中鱼肉肌原纤维的分解,进而减少冷鲜草鱼的硬度、弹性和回复性等质构的变化。在贮藏至第6天,中性氧化电解水处理的冷鲜鱼肉硬度、弹性及回复性分别为对照组的1.23、2.13、1.83倍,且电解水组的肌原纤维相对较长且排布较规整。利用中性氧化电解水可以有效减轻冷鲜草鱼肉贮藏过程中品质及质构的变化,可为淡水鱼冷鲜鱼肉产品的贮藏保鲜技术开发提供参考。     

Sarcoplasmic proteins influence water-holding capacity of pork myofibrils

Water-holding capacity (WHC) is one of the main pork quality characteristics. The objective of this study was to determine the influence of (denaturation of) sarcoplasmic proteins on WHC. Myofibrils extracted from red, firm, non-exudative (‘normal’) and PSE (pale, soft, exudative) pork longissimus muscle were combined with sarcoplasmic extracts (with or without proteins) from PSE and normal pork longissimus samples. Weight increase of myofibrils (mg increase mg⁻¹ myofibrillar protein) was used as a measure of WHC. When combined with protein-containing sarcoplasmic extract from normal pork, WHC of myofibrils from PSE (2.6 mg mg⁻¹) and normal (2.8 mg mg⁻¹) pork was higher (P < 0.05) than when combined with sarcoplasmic extract from PSE meat (1.3 mg mg⁻¹ for PSE and 1.9 mg mg⁻¹ for normal myofibrils). Protein-free sarcoplasmic extracts were prepared by heating the extracts for 30 min at 80 °C. WHC of myofibrils combined with protein-free sarcoplasmic extract from PSE and normal pork was not significantly different. WHC of myofibrils combined with protein-free extract was lower than WHC of myofibrils combined with protein-containing extract. Ionic strength or pH could not explain the observed differences. It was concluded that sarcoplasmic proteins do influence WHC. The mechanism of this influence still needs to be determined. © 1999 Society of Chemical Industry     

FIBRE FRACTURE OF BOVINE M. PECTORALIS PROFUNDUS MUSCLE COOKED PRE- AND POSTRIGOR

The ultimate tensile strength and extensibility in the direction of the muscle fibres of cooked meat decreased exponentially during storage of the raw meat for up to 3 days at 15C. The mode of fracture was examined by light microscopy of longitudinal sections taken after testing to break. In meat cooked soon after stunning, 'brittle' fracture had been induced across the fibres and the high extensibility was proposed to be due to sequential fibre fracture. At 1 day postmortem, the fractures crossed several fibres occurring in a repetitive fashion every 30 μm along the fibres. This was thought to be due to a stiff fibre embedded in an extensible surface matrix connecting the fibres. In aged meat, discontinuous fractures were prominent within each fibre and which stopped at the surface of each fibre showing further weakening of the fibre and the composite matrix. All the myofibrils remained in register. These changes are consistent with a gradual reduction in cohesion within the meat and indicates that the weakening of the focal adhesions is primarily responsible for postmortem tenderisation.     

牛肉排酸成熟过程中的品质变化

为探究排酸成熟过程中肉类品质的变化,以牛肉为研究对象,测定排酸1～7 d肉样的硬度、保水性、pH值、肌原纤维小片化指数（myofibril fragmentation index,MFI）、菌落总数、大肠菌群数以及排酸1、3、5d时肌原纤维的超微结构。结果表明：牛肉排酸3 d时出现最大僵直期,硬度达到最高,保水性达到最低,pH值降到最低值5.56;随着解僵及成熟的进行,牛肉嫩度、保水性回升,排酸7 d时pH值回升到5.84,达到排酸成熟肉标准（5.8～6.2）;排酸1 d时,肌原纤维的Z线、粗细丝清晰可见,随着肌原纤维小片化程度的不断加深,排酸5d时,Z线变得模糊,几乎消失,成为晕染状,几何构型发生明显变化。上述结果表明,牛肉在0～4 ℃条件下排酸成熟需要7d,前3 d是排酸的关键时期。     

Ultrastructural changes in pre- and post-rigor beef muscle caused by conventional and microwave cookery

Examination of cooked pre-rigor muscle by TEM (transmission electron microscopy) and SEM (scanning electron microscopy) revealed that all cookery methods resulted in the development of supercontraction bands alternating with areas showing tissue fragmentation and tearing. Microwave cookery produced smaller and less dense supercontraction nodes in pre-rigor muscle with less tearing and fragmentation but more fibre separation. Although cold-shortened pre-rigor muscle cooked by all methods also exhibited supercontraction bands with some tearing and fragmentation in adjacent sarcomeres, the samples cooked by microwaves showed a more uniform repeating pattern of small stretched areas alternating with dense contracted areas. Cooking of muscle in full rigor resulted in myofibrillar protein coagulation and shrinkage but supercontraction nodes were absent. Cold-shortened bicarbonate treated muscle was relatively intact after cooking, exhibiting fusion of the myofibrils and an absence of intermyofibrillar spaces. Results are discussed in relation to possible effects upon tenderness.     

The role of the endomysium in the salt-induced swelling of muscle fibres

Variable uptake of water is a persistent problem in meat processing. To try to understand the changes that occur during water uptake, and thereby explain this variability, we have measured the swelling of salt-treated single muscle fibres from rabbit longissimus dorsi. By dissolving out the myofibrillar proteins with SDS we have been able to tell whether or not a dissected fibre has an endomysial connective sheath. We find that some fibres do lack a sheath and that these stripped fibres swell on average approximately five times more than intact fibres in 0·25 m KI. This shows that the endomysium limits the swelling of the fibre in salt, as is also shown by the greater swelling of those parts of fibres from which the endomysium has been deliberately removed. On prolonged immersion in 0·25 m KI the stripped parts of fibres shrink after swelling whereas the smaller swelling of unstripped parts is maintained. With careful, slow dissection, the proportion of stripped fibres is low at all times post mortem, but if a quicker method is used, the proportion of stripped fibres increases with time post mortem. As ageing proceeds, an increasing incidence of transverse fractures reduces the productivity of the quicker method, so the slow method has to be used. Regardless of the method of dissection the swelling of stripped and unstripped fibres does not depend on time post mortem. Thus it is the changing proportion of stripped fibres that underlies the earlier report (Wilding et al. Meat Sci., 1986 18, 55) of a dependence of swelling on time post mortem. The ease of stripping of endomysium from muscle fibres may be important in meat processing. Thus whether or not fibres produced during comminution and mixing of meat are stripped directly affects water uptake. The extraction of myofibrillar proteins may also be affected, which would be important for the adhesion of cooked reformed meat products.