Metmyoglobin Formation in Beef Stored in Carbon Dioxide Enriched and Oxygen Depleted Atmospheres

SUMMARY: A spectroreflectometric technique was used to determine the relative percentages of three myoglobin pigments, reduced myoglobin, oxymyoglobin and metmyoglobin at the surface of fresh beef. It was shown that, at constant humidity, the formation of metmyoglobin in beef was maximal at 6 + 3 mm Hg of oxygen at 0°C and 7.5 ± 3 mm Hg at 7°C for semitendinosus muscles. Carbon dioxide concentrations of 10% and higher had negligible effect on the formation of metmyoglobin, provided the oxygen pressure was above about 5%. At high partial pressures of carbon dioxide, absorption of carbon dioxide increased and the pH of the surface decreased. In air, the formation of metmyoglobin varied widely from muscle to muscle.     

The rate of metmyoglobin formation in beef, pork, and turkey meat as influenced by pH, sodium chloride, and sodium tripolyphosphate

This study investigated the effects of pH (5·5 to 7·0), sodium chloride concentration (0·0-3·0%), and sodium tripolyphosphate concentration (0·0 and 0·5%) on the rate of metmyoglobin formation in ground beef, pork and turkey meat during refrigerated storage. Increasing the sodium chloride concentration produced a progressive increase in the rate of metmyoglobin formation in ground beef. Increasing the pH between pH 5·5 and 6·5 had no effect on the rate of metmyoglobin formation with ground beef and turkey meat, but produced a marked decrease between pH 6·5 and 7·0. In contrast pH had no consistent effect on the rate of metmyoglobin formation of ground pork and the rate of formation remained low at all pH levels. When ground beef contained 0·5% sodium tripolyphosphate, the effect of pH was reversed and the rate of metmyoglobin formation was lowest at pH 5·5, increased as the pH increased to 6·5 and then plateaued.     

Effect of oxygen- and carbon dioxide-enriched atmospheres on shelf-life extension of refrigerated ground pork

The oxidation of the lipids and myoglobin of ground pork meat stored in oxygen- and carbon dioxide-enriched atmospheres at 1 °C has been studied. Elevated oxygen levels (80-100%) depressed myoglobin oxidation, increasing the time to 50% metmyoglobin formation from four to about thirteen days. Twenty per cent carbon dioxide greatly reduced the rate of lipid oxidation, extending the time to reach a TBA number of 5 from five to about twelve days. Tocopherol and ascorbic acids were efficient inhibitors of lipid oxidation but citric acid was not.     

Autoxidation of purified myoglobin from two bovine muscles

To elucidate the behavioural differences between beef muscles from the viewpoint of colour stability, oxymyoglobin was extracted at 2 h post mortem, purified from two different muscles (longissimus lumborum (LL), stable and psoas major (PM), unstable) and the autoxidation rate was measured. Oxymyoglobin was isolated after separation from metmyoglobin by chromatography on DEAE-Sepharose and TSK SW 2000 columns, and its purity was controlled by electrophoresis and IEF. Over a wide range of pH values (5-9), temperatures (20-50°C) and ionic strength (0-500 mM), no difference was noted between autoxidation rates of LL and PM oxymyoglobin extracted at 2 h post mortem. Conversely, when myoglobin was extracted at 192 h post mortem, the autoxidation rate of PM oxymyoglobin was higher than LL myoglobin, particularly at elevated temperatures. These differences in autoxidation rates after extraction of myoglobin at 2h and 192 h post mortem were not associated with differences in Ea (approximately 23 kcal/mol).     

Effect of salt on oxidative changes in pre- and post-rigor ground beef

Pre- and post-rigor beef was ground and salt was added to give 0·0, 0·5, 2·0 and 4·0% NaCl (w/w). Samples were removed after 0, 24, 48, 72 and 96 h at 4°C and analyzed for pH, TBA numbers and percentages of reduced myoglobin (Mb), metmyoglobin (MMb) and oxymyoglobin (MbO(2)). After holding for 96 h the samples were cooked in a boiling water bath to an internal temperature of 80°C and held at 4°C for 48 h before TBA analysis. Pre-rigor grinding and salting reduced the post-mortem pH decline and the extent of meat discoloration as shown by the differences in the amount of MMb. The extent of lipid oxidation as measured by TBA numbers was not significantly different for the pre- and post-rigor ground salted meat samples, although salt accelerated oxidation during storage. Results demonstrated that pre-rigor grinding and salting of beef produces a more stable bright red color, which appears to be associated with a lower percentage of MMb and a higher ultimate pH in the pre-rigor salted meat.     

Metmyoglobin and inorganic metals as pro-oxidants in raw and cooked muscle systems

The pro-oxidant activities of metmyoglobin (Mb) and metal ions on the induction of lipid oxidation in raw and heated water-washed muscle systems from fish, turkey, chicken, pork, beef and lamb and during storage of these systems at 4°C, were investigated. Lipid oxidation was invariably faster in heated than in raw systems. In raw Mb-catalyzed systems, oxidation was slow over a 5-day period, except in fish, where significant (P < 0·05) increases in TBA values occurred; in contrast, significant (P < 0·05) increases in TBA values occurred in cooked fish, turkey, chicken and pork after 3 days of storage. Cooked beef and lamb, however, showed significant lipid oxidation only after 5 days of storage. Fe(2+) was found to be highly catalytic in cooked muscle. Cu(2+) and Co(2+) were less effective catalysts than Fe(2+); the overall pro-oxidant activity was in the order Fe(2+) > Cu(2+) > Co(2+) > Mb, and the susceptibility to lipid oxidation of the muscle systems was in the order: fish > turkey > chicken > pork > beef > lamb, probably reflecting the degree of unsaturation of the constituent triglyceride fractions.     

Post-slaughter influences on the formation of metyyoglobin in beef muscles

By the use of electrical stimulation and/or hot-boning and/or water baths adjusted to temperatures in the range 1-41°C, beef muscles (Semimembranosus, Longissimus dorsi and Psoas major) were exposed to different pH/temperature/time regimes following slaughter and the formation of metmyoglobin (metMb) at their surfaces was monitored during subsequent aerobic storage at 1±1°C. The time course of formation was complex but, in general, consisted of an initial, rapid phase, lasting a few days, followed by a slower, or equilibrium, phase which ultimately yielded to a second rapid phase. Exposure to high temperature and low pH led to increased rates of metMb formation in the Longissimus dorsi and Semimembranosus muscles sliced 48 h after death but not in Psoas major muscles sliced 23 or 48 h post slaughter. However, ageing the Psoas major muscles, in vacuo, at 1°C caused the high temperature treated samples to be less colour stable than those held at 1°C. When the air-stored samples were transferred to an anaerobic environment at 23·5°C reduction of the metmyoglobin occurred, the rate of reduction being inversely proportional to the concentration of metMb developed during aerobic storage. It is suggested that the rate of metmyoglobin formation at the surface of beef muscle slices is dependent on at least two factors-the oxygen consumption rate (OCR) and the activity of an enzymic reducing system. In most practical circumstances the activity of the reducing system is the most important factor and this can be affected by the time, temperature and pH history of the muscle.     

Biochemical factors influencing metmyoglobin formation on beef from muscles of differing colour stability

Biochemical parameters, such as oxygen consumption rate by muscle post mortem (OCR), depth of oxygen penetration into meat, rates of myoglobin oxygenation and deoxygenation and myoglobin content and succinic dehydrogenase activity, were determined for muscles of differing colour stability. Metmyoglobin reduction, in anoxia following oxidation with ferricyanide (MRA) and aerobically following oxidation with low pO(2) (ARA), were also determined. M. psoas major (poor colour stability) has higher enzymic activity than M. longissimus dorsi (good colour stability). This difference, together with the low myoglobin content in M. psoas major, results in relatively high OCR with consequent low oxygen penetration and rapid conversion of oxymyoglobin to myoglobin in M. psoas major, disposing it to rapid formation of metmyoglobin. Metmyoglobin reduction occurs both under anaerobic and aerobic conditions but no significant correlation is found between actual metmyoglobin reduction and rate of discoloration of different muscles. The most significant factor affecting colour stability of beef muscles appears to be their enzymic activity which determines the rate of myoglobin oxidation.     

Liberation of actin from actomyosin in meats heated to 65°C

This study investigated whether actin liberation from myofibrils occurs during the heating of various muscles, as well as squid mantle muscle at temperatures, such as 60°C, employed for vacuum cooking of meats. Actin liberation was demonstrated in scallop striated adductor muscle, but not in beef, pork, or chicken, using the detection method previously employed with squid muscle, in which liberated actin was detected with SDS-PAGE, in the supernatant obtained by centrifugation of the homogenate of heated muscle in 0.2M KCl at a neutral pH. However, actin liberation was demonstrated in beef, pork and chicken by a new detection method, in which heated muscle was homogenized in 0.6M KCl or NaCl at a slightly alkaline pH and maintained at 4°C for 16h with stirring, after which the homogenate was diluted three times with water and centrifuged to obtain the supernatant containing the liberated actin. This new method indicated that actin liberation in beef, pork, and chicken was marked by heating at 65°C, but scarcely induced at 80°C. Thus, the liberation of actin from myofibrils may contribute to the greater tenderness of vacuum-cooked meat (meat heated at a low temperature for long time), as compared with meat prepared by cooking at a higher temperature.     

STRUCTURAL CHANGES BETWEEN BEEF OXY- AND METMYOGLOBIN AS REVEALED BY MONOCLONAL ANTIBODIES

Oxy- and metmyoglobin were purified from beef muscle using ion-exchange chromatography on Mono-Q. Their reactivity against six monoclonal antibodies (MAbs) to beef myoglobin was compared in a competitive ELISA using biotinylated beef oxymyoglobin. Four MAbs reacted with higher affinity for oxymyoglobin than for metmyoglobin. A similar higher reactivity of oxymyoglobin versus metmyoglobin was obtained with a sandwich ELISA using peroxidase-labelled rabbit anti-serum against beef myoglobin. Such a result has never been reported for myoglobins of any species. These MAbs should be valuable tools for analyzing conformational changes of oxymyoglobin solutions when subjected to chemical or physical treatments.     

Low-temperature osmotic dehydration improves the quality of intermediate moisture meats

Dehydration of meat at low temperature (4°C) with a new dehydrating sheet to obtain intermediate moisture meat produced more desirable results than did more rapid dehydration at 25°C, because the native state of meat proteins was better retained at the lower temperature. At low temperature with replacement of the dehydrating sheet once per day, the water content decreased to 57% after 150 h. Water activity (a(w)) dropped to 0·965. Myofibrils could be prepared readily from meat after 150 h at 4°C, and the myofibrils were induced to contract by addition of Mg(2+)-ATP solution and therefore retained biological activity. Electron microscopic observation showed that intermediate moisture meat prepared at 4°C retained the original muscle structure to a large extent. Myosin could be extracted easily from intermediate moisture meat dehydrated at 4°C after 10 days of storage at 25°C. No marked difference in the effect of dehydration on muscle was observed among species (beef, pork, chicken and rabbit). Results show that dehydration at low temperature (4°C) with use of the new dehydrating sheet enables intermediate moisture meat of good quality to be manufactured.     

Color Characteristics of Irradiated Vacuum-Packaged Pork, Beef, and Turkey

Changes in color of irradiated meat were observed to be species-dependent. Irradiated pork and turkey became redder due to irradiation but irradiated beef a* values decreased and yellowness increased with dose and storage time. The extent of color change was irradiation dose-dependent and was not related to myoglobin concentration. Visual evaluation indicated pork and turkey increased in red ness whereas beef decreased in redness as dose levels increased. Reflectance spectra showed that irradiation induced an oxymyoglobin-like pigment in pork and that both oxymyoglobin and metmyoglobin developed in beef as a result of irradiation.     

Analysis of meat pigments with tissue spectrophotometer TS-200

The colour of meat and rice flour pastes containing known amounts of myoglobin and the colour of intact beef and pork samples were analyzed with tissue spectrophotometer TS-200. The differences in the spectra of myoglobin among three types of derivatives were successfully distinguished with this instrument. In addition, there is a close relationship between I_HB value, a parameter for estimating the content of pigments in animal tissues, and myoglobin content in model systems (rice flour paste and meat paste). Especially, the I_HB value is proportional to myoglobin content in intact beef and pork meat whose myoglobin is mostly in the state of oxymyoglobin and/or deoxymyoglobin: Y = 208·26 x + 6·72, where y is the I_HB value, x is the myoglobin content (%) and R = 0·94.     

Physico-chemical and processing quality of porcine M. longissimus dorsi frozen at different temperatures

Longissimus dorsi muscle from six pigs (24 h post-mortem) was cut into portions of similar size and shape (c. 700 g) and vacuum-packed in polyfilm. The muscle specimens were divided into three samples, one frozen at -20°C, another at -80°C and the third served as the control (not frozen). The meat sample frozen at -80°C was transferred to the -20°C freezer. After one month, both frozen pork samples were thawed at -2°C and drip loss (%) was measured. Hunter colour, metmyoglobin (MetMb) formation (%), water-holding capacity (WHC), TBA value, transmission value (TM) and myofibril fragmentation were also determined. There was no significant difference in drip loss for the two frozen samples. No MetMb formation could be detected and Hunter values were basically the same for all three samples. WHC, TBA value and TM were essentially the same for all three samples. TBA value was quite low for each frozen sample, indicating that lipid oxidation did not occur during freezing. Histological examination of both frozen samples indicated inter- and intracellular ice crystal formation at -20°C, and intracellular ice at -80°C, the extent being less than at -20°C. At -20°C, ice crystals were larger and muscle fibre diameter smaller than for the control or -80°C sample. Myofibril fragmentation in both frozen samples was significantly higher than in the control. Pork sausage was prepared from all three samples by adding 2% NaCl and 100 ppm NaNO(2). Cooking loss and colour forming ratios were essentially the same. The sausage sample made from the -20°C frozen meat was harder than that of the other two samples according to rheological measurement.     

Effects of salt reduction on the rheological and gelation properties of beef, pork and poultry meat batters

The gelation and rheological properties of beef, pork and poultry meat batters as affected by salt reduction (2·50, 1·25 and 0·00%) were studied by using a Haake rotational viscometer and a thermal scanning rigidity monitor. Beef batters showed a decrease in shear stress with the decrease in salt levels at both high and low shear rates. Pork batter showed a mixed behavior (no definite trend in shear stress versus shear rate) and the poultry meat batters showed a Bingham pseudoplastic behavior, except for the no-salt treatment. During heating the beef batters showed the highest G values followed by the pork and the poultry meat batters. The rigidity modulus profiles exhibited two major transition temperatures at 47-53°C and at 64-76°C. Beef batter with 2·50% salt developed the highest average G value (16·6 kPa) and the poultry batter with 2·50% salt the lowest (7·3 kPa).     

Tenderising in M. longissimus dorsi of beef, veal, rabbit, lamb and pork

The decrease in toughness of M. longissimus dorsi with storage time at 1°C was effectively described by an exponential decay equation. The average rate constant for beef, veal and rabbit was 0·17 whilst that for lamb was 0·21 and that for pork, 0·40 days(-1). However, the rate constants were not significantly different due to variations both within muscles and between animals. On average, 50% of the tenderising occurred in 2 days for pork and in 4·2 days for beef, veal and rabbit, and 80% in 4·9 and 9·5 days, respectively. At the completion of tenderising, beef and rabbit were the toughest and pork the most tender, whilst the greatest tenderising occurred in beef and lamb.     

Effect of high pressure treatment on the color of fresh and processed meats: A review

High pressure (HP) treatment often results in discoloration of beef, lamb, pork, and poultry. The degree of color changes depends on the physical and chemical state of the meat, especially myoglobin, and the atmospheric conditions during and after pressurization. A decreased redness is attributed to a large degree to the oxidation of the bright red oxymyoglobin or the purplish deoxymyoglobin into the brownish metmyoglobin, as well as to the denaturation of myoglobin. Surely, the high myoglobin content makes beef more exposed to this discoloration compared to the white chicken meat. In addition, HP treatment causes denaturation of myofibrillar proteins followed by aggregation, consequently, changing the surface reflectance and increasing lightness. Other intrinsic and extrinsic factors may affect the pressure-induced color changes positively or negatively. In this review, the pressure-induced color changes in meat are discussed in relation to modification of the myoglobin molecule, changes in the meat microstructure, and the impact of the presence of different chemical compounds and physical conditions during processing.     

Determination of the myoglobin states in ground beef using non-invasive reflectance spectrometry and multivariate regression analysis

Seventy-two samples of ground beef from M. semimembranosus of two 5 and two 1.5 year old animals were prepared. Two types of fat tissues from either beef or pork were added to the ground beef. The samples were prepared to contain predominantly deoxymyoglobin (DMb), oxymyoglobin (OMb) and metmyoglobin (MMb) states on surfaces using selected methods based on chemical treatment (for MMb) and oxygen pressure packaging to induce the two other states. Reflectance spectra were measured on ground beef after three storage times. Partial least regression analysis was used to make calibration models of the desired myoglobin states. Validated models using leave-one-sample out cross validation gave, after correction and normalization, prediction errors of about 5%. Long term storage of ground beef was unsuitable for preparing pure MMb states due to gradual reduction of the pigment to DMb, presumably by bacteria.     

Effects of pyruvate on lipid oxidation and ground beef color

Our overall objective was to better understand the effects of added pyruvate on enhanced beef color stability. The 2 possible mechanisms assessed were the role of pyruvate in lipid oxidation and direct interaction between pyruvate and beef myoglobin. Microsomes were incubated with pyruvate at pH 5.6, 25 °C, and lipid oxidation was measured hourly for 3 h. Bovine oxymyoglobin at pH 5.6 was incubated with pyruvate and used to quantify both redox stability (metmyoglobin formation) and pyruvate-myoglobin adduction using mass spectrometry analysis. Surface color and lipid oxidation were measured on ground beef patties stored for 6 d in polyvinyl chloride over-wrap (PVC) or high oxygen. Addition of pyruvate to microsomes decreased lipid oxidation compared with controls (P < 0.05). Conversely, no effect on myoglobin was observed (no changes in redox stability and no peaks corresponding to pyruvate were observed; P > 0.05). However, pyruvate increased color stability and decreased lipid oxidation of ground beef patties packaged in PVC and high oxygen. Pyruvate decreased nitric oxide metmyoglobin-reducing capacity and oxygen consumption of patties compared with controls (P < 0.05). This research suggests that pyruvate may improve beef color stability primarily through its antioxidant effect on lipids. Practical Application: Discoloration of meat often results in significant revenue loss. This study suggests that pyruvate can improve the color stability of patties packaged in high oxygen and PVC primarily through its antioxidant effect on lipids.     

Differentiation between fresh and thawed meat by an enzyme profile test

Exudates from fresh (stored at +4°C) and thawed pork and beef (frozen and stored below -20°C) was assayed by the rapid test kit API-ZYM(TM) to determine enzyme profiles. The test kit consists of 20 wells for different enzyme substrates. Of altogether 1040 results, only few reactions of the enzymes differed in their intensity between frozen and thawed pork or beef. Fresh pork showed a more intensive β-galactosidase and N-acetyl-β-glucosaminidase reaction while with fresh beef a more intensive reaction could only be detected for N-acetyl-β-glucosaminidase. Only N-acetyl-β-glucosaminidase showed significant differences between fresh and frozen meat in both species (α = 0·01). Considering the indistinct results of the test kit differentiation between frozen and thawed meat, the API-ZYM(TM) test kit is considered not suitable for distinguishing frozen from thawed pork or beef.