Kinetics analysis of the reactions responsible for myoglobin chemical state in meat using an advanced reaction-diffusion model.

Here we developed an advanced reaction-diffusion model to predict the evolution of the myoglobin state in beef meat using numerous reactions with rate constants of different orders of magnitude. The initial scheme included 44 reactions from the literature. Sensitivity analysis proved that this initial scheme was equivalent to a simple 22-reaction scheme. Results calculated with this scheme were compared against the spatial distributions of oxymyoglobin (MbO2), metmyoglobin (MMb) and deoxymyoglobin (DMb) measured in meat cuts stored at 20°C under air-permeable packaging. We found global agreement between measured and calculated distributions when adequate rate constant values were used, particularly for the formation of MbO2 from DMb. The model was used to calculate evolutions in MbO2 and MMb distributions under different situations (modified-atmosphere packaging, Fenton chemistry with or without water-soluble antioxidants, increased mitochondrial oxygen consumption). Results were used to discuss the underlying kinetics reaction mechanisms and the performances and limits of the model.

Comparison of proteolysis in native, heat-treated and aged proteins from turkey meat.

1. The present study compared the ability of native, heat-treated and aged turkey breast muscle proteins to undergo proteolysis by digestive tract proteases. 2. Domestic turkey toms were slaughtered under laboratory conditions. Breast muscles were excised immediately post mortem; one was placed under conditions to develop exudative meat by maintaining the muscle at 40 degrees C for at least 30 min and the other was refrigerated under commercial conditions. 3. Meat was collected and stored for 7 d at 4 degrees C. Breast samples removed at d 0 and d 7 were frozen and stored at -80 degrees C until used for determination of solubility, protein surface hydrophobicity and protein oxidation through carbonyl content. Measurements of pepsin and trypsin/chymotrypsin activities were performed in vitro on myofibrillar proteins. 4. Storage increased carbonyl content in control samples while the oxidation increase was not significant in heat-treated myofibrillar protein. Hydrophobicity was not affected by storage time or treatment or protein solubility. 5. Storage significantly increased trypsin + chymotrypsin activity only in the control group. The activities of pepsin and trypsin + chymotrypsin were negatively correlated with protein surface hydrophobicity.

Review: divergent selection for residual feed intake in the growing pig.

This review summarizes the results from the INRA (Institut National de la Recherche Agronomique) divergent selection experiment on residual feed intake (RFI) in growing Large White pigs during nine generations of selection. It discusses the remaining challenges and perspectives for the improvement of feed efficiency in growing pigs. The impacts on growing pigs raised under standard conditions and in alternative situations such as heat stress, inflammatory challenges or lactation have been studied. After nine generations of selection, the divergent selection for RFI led to highly significant (P<0.001) line differences for RFI (-165 g/day in the low RFI (LRFI) line compared with high RFI line) and daily feed intake (-270 g/day). Low responses were observed on growth rate (-12.8 g/day, P<0.05) and body composition (+0.9 mm backfat thickness, P=0.57; -2.64% lean meat content, P<0.001) with a marked response on feed conversion ratio (-0.32 kg feed/kg gain, P<0.001). Reduced ultimate pH and increased lightness of the meat (P<0.001) were observed in LRFI pigs with minor impact on the sensory quality of the meat. These changes in meat quality were associated with changes of the muscular energy metabolism. Reduced maintenance energy requirements (-10% after five generations of selection) and activity (-21% of time standing after six generations of selection) of LRFI pigs greatly contributed to the gain in energy efficiency. However, the impact of selection for RFI on the protein metabolism of the pig remains unclear. Digestibility of energy and nutrients was not affected by selection, neither for pigs fed conventional diets nor for pigs fed high-fibre diets. A significant improvement of digestive efficiency could likely be achieved by selecting pigs on fibre diets. No convincing genetic or blood biomarker has been identified for explaining the differences in RFI, suggesting that pigs have various ways to achieve an efficient use of feed. No deleterious impact of the selection on the sow reproduction performance was observed. The resource allocation theory states that low RFI may reduce the ability to cope with stressors, via the reduction of a buffer compartment dedicated to responses to stress. None of the experiments focussed on the response of pigs to stress or challenges could confirm this theory. Understanding the relationships between RFI and responses to stress and energy demanding processes, as such immunity and lactation, remains a major challenge for a better understanding of the underlying biological mechanisms of the trait and to reconcile the experimental results with the resource allocation theory.

The binding of N-trifluoroacetyl chito-oligosaccharides to wheat-germ agglutinin: a fluorescence investigation.

We describe the synthesis of N-trifluoroacetyl chito-oligosaccharides and their use as ligands to probe the binding sites of wheat-germ agglutinin, a lectin specific for N-acetylglucosamine. The binding is monitored using intrinsic protein fluorescence, which is due to tryptophan side-chains. We present arguments purporting to show the presence of a fluorophore close to each of the four sites. The binding of chito-oligosaccharides to wheat-germ agglutinin is complex and can only be approximately described by an independent and equivalent sites model. This model applies when the ligand concentration range is restricted to higher values. The possible role of ligand-mediated protein aggregation and of site inequivalence is discussed. We find that the affinity of trifluoroacetylated chito-oligosaccharides for wheat-germ agglutinin is higher than that of the N-acetylated parent compounds, the difference increasing with chain length. Our results are in agreement with a model of the binding site previously proposed by Clegg et al. (Biochemistry 22 (1983) 4797-4804).

Effect of finishing mode (pasture- or mixed-diet) on lipid composition, colour stability and lipid oxidation in meat from Charolais cattle.

Effects of pasture- or mixed-diet finishing mode on colour and lipid stability were measured in meat from Charolais steers, heifers and cows of different ages after refrigerated storage. Meats from pasture- and mixed-diet finished cattle have more or less significant differences in lipid composition which will influence the colour and lipid stability. The mixed-diet finished cattle have about the same proportions of saturated and monounsaturated lipids as pasture-diet finished animals; on the contrary, cattle fed on grass have higher proportions of n-3 PUFA, and to a less extent, of n-6. Finishing diet had an important effect on lipid stability, meat from pasture-finished animals showing significantly lower TBA-RS level than meat from mixed-diet finished animals. Effect of diet was also significant on myoglobin content with a higher content of haeminic iron in mixed-diet finished animals. Effect of diet on colour stability was slight with a beneficial effect of pasture finishing mode, significant only in heifers, and after a meat storage of six days in air.

Influence of dietary fat and vitamin E supplementation on free radical production and on lipid and protein oxidation in turkey muscle extracts.

The objectives of this study were to investigate the effects of dietary fat (6% soy oil or rapeseed oil or tallow) and alpha-tocopheryl acetate supplementation at two levels (30 or 200 ppm) on radical production, measured by ESR spectroscopy, and on lipid and protein oxidation in turkey muscle extracts oxidized by an enzymic system (NADPH, ADP, FeSO(4)/cytochrome P450 reductase). Two muscles were tested: pectoralis major (glycolytic) and sartorius (oxidative) muscles. Radical production measured by ESR was higher in pectoralis major muscle than in sartorius muscle, whereas lipid and protein oxidation was more important in sartorius muscle, showing the importance of the pro-/antioxidant ratio in oxidative processes in muscular cells and of the measurement methodology to appreciate the free radical production. Dietary fat had no effect on the level of ESR signals, whereas feeding of animals with soy oil induced higher oxidation of lipids. Protein oxidation was less sensitive to the nature of the dietary fat than lipid oxidation. Vitamin E supplementation significantly decreased radical production, as measured by ESR spectroscopy. Vitamin E also decreased lipid and protein oxidation, but the effect of vitamin E on protein oxidation was less pronounced than on lipid oxidation.

Influence of dietary fat and vitamin E on antioxidant status of muscles of turkey.

The aim of this study was to better understand the effects of more or less unsaturated fat source (tallow/soy oil/rapeseed oil) and/or vitamin E dietary supplementation (200 ppm) on the antioxidant status (at day 1 post-mortem) of turkey muscles [pectoralis major (Pm) and sartorius (S)]. More particularly, when turkeys were fed tallow, supplementation was sufficient to improve significantly the vitamin E status. Feeding rapeseed oil increased the antioxidant enzyme (AOE) activities (catalase, superoxide dismutase, glutathion reductase), glutathione concentration, and value from the benzoic acid test. Dietary soy oil increased glutathione peroxidase activity, compared to other dietary fat sources. With tallow, most of AOE activities were lower than with rapeseed or soy oil. Whatever the feeding mode, vitamin E supplementation did not affect the AOE activities, glutathione concentration, or values from the benzoic acid test. AOE activities were always higher in the oxidative S muscle than in the glycolytic Pm muscle. After feeding tallow, 9 days of storage increased TBA-RS and carbonyl contents, whereas the activity of many antioxidant enzymes and the total antioxidant activity (TEAC test and benzoic acid test) decreased.

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 diet finishing mode (pasture or mixed diet) on antioxidant status of Charolais bovine meat.

The aim of this study was to better understand the effect of diet finishing mode (pasture or mixed diet) on the antioxidant status of bovine meat (M. Longissimus dorsi). Effects of sex (cow, heifer and steer) and age (only in cows) were also studied. Vitamin E content of muscle and activity of antioxidant enzymes, such as superoxide dismutase (SOD), catalase (Cat) and glutathione peroxidase (GPx), were measured. Total antioxidant status was also estimated by measuring Trolox equivalent antioxidant capacity (TEAC) and benzoate hydroxylation. Vitamin E content was higher in meat from pasture-finished animals than from mixed diet-finished animals. Diet had also an important effect on antioxidant enzyme activity. Pasture finishing mode increased SOD activity and decreased GPx activity in muscle. Effect of diet on catalase activity was less pronounced. Total antioxidant status measured by benzoate hydroxylation was higher in the mixed diet-group but no effect of diet was observed on TEAC measurement. Effect of sex was observed on SOD and catalase activity and also in benzoate hydroxylation. Effect of age was only noticed on benzoate hydroxylation.

Lipid and protein oxidation in vitro, and antioxidant potential in meat from Charolais cows finished on pasture or mixed diet.

In this work, the effect of finishing diet (pasture- or mixed-diet) on lipid and protein oxidation in beef homogenates was evaluated. Oxidation was chemically induced by ferrous iron and hydrogen peroxide. Lipid and protein oxidation were respectively measured by determining TBA reactive substances (TBA-RS) and protein carbonyl groups (DNPH coupling method). Evaluation of the antioxidant status of meat was also performed by measuring vitamin E concentration and antioxidant enzyme activities: superoxide dismutase (SOD), glutathione peroxidase (GPx) and catalase. Pasture-finishing mode of animals significantly protected lipids in meat from oxidation but diet mode did not affect protein oxidation. Concentration of vitamin E was higher in meat from pasture-fed animals. Different diets had opposite effects on SOD and GPx activities, pasture-finishing mode of animals increasing SOD activity but decreasing GPx activity. No significant effect of diet was noted on catalase activity.

Antioxidant enzyme activities in beef in relation to oxidation of lipid and myoglobin.

Lipid- and oxy-free radical generation has been implicated in oxidative processes which occur during meat maturation but the importance of the antioxidant enzyme (AOE) activity in these processes is not known. It was shown that metmyoglobin (MetMb) % and lipofuscin content were higher in colour-unstable muscles such as psoas major (PM) and diaphragma (D) compared to longissimus lumborum (LL) and tensor fasciae latae (TFL). Although Superoxide dismutase (SOD) activity is higher post mortem in PM and D muscles than in LL and TFL muscles, catalase and glutathione peroxidase (GPx) activities were higher only in D muscle. The higher AOE activity in colour-unstable muscles such as PM and D was not sufficient to prevent increased formation of MetMb and lipofuscin in these muscles compared to LL and TFL muscles.

A rapid method of oxymyoglobin purification.

Oxymyoglobin was isolated from bovine Longissimus lumborum muscle by precipitation with ammonium sulfate and purification in rapid conditions with only one chromatographic step on Mono-Q HR column with a HPLC system. Purity of oxymyoglobin was controlled by SDS-polyacrylamide gel electrophoresis and isoelectrofocusing.

Lipid and protein oxidation in microsomal fraction from turkeys: influence of dietary fat and vitamin E supplementation.

The objectives of this study were to investigate the effects of dietary fats (6% soya oil or rapeseed oil or tallow) and α-tocopheryl acetate supplementation (30 ppm for control and 200 ppm for supplemented animals), on lipid and protein oxidation, induced by Fe(3+)/ascorbate, of microsomal fraction in turkey muscles (M. pectoralis major and M. sartorius). Supplementation of turkeys with α-tocopheryl acetate increased the vitamin E content of microsomal membranes. Vitamin E supplementation strongly decreased lipid oxidation in membranal fractions when animals were fed rapeseed oil or tallow; this effect was less pronounced in animals fed soya oil. Vitamin E supplementation induced a slight decrease in protein carbonyl content, especially in animals fed soya oil. Level of protein free thiols was considerably enhanced in diet enriched with soya oil. Vitamin E supplementation had a stabilizing effect on glucose-6-phosphatase activity of microsomes when oxidized by Fe(3+)/ascorbate. No muscle effect was detected on the level of lipid and protein oxidation in membranal fractions even if M. sartorius is known to be more oxidative than M. pectoralis major.

Effect of a dietary vitamin E supplementation on colour stability and lipid oxidation of air- and modified atmosphere-packaged beef.

The effects of dietary vitamin E supplementation on tissue α-tocopherol level and on the susceptibility of fresh and modified atmosphere-packaged beef on myoglobin and lipid oxidation were investigated. Charolais cattle, aged 32-44 months, were fed diets containing 75 (control, n=8) or 1000 mg (supplemented, n=8) α-tocopheryl acetate/kg feed/day for 111 days prior to slaughter. Following vacuum packaging, M. Longissimus lumborum and M. triceps brachii were aerobically packaged and held under refrigerated display (3°C) for 9 days or packaged under modified atmosphere (MAP; 20% CO(2): 80%O(2)) and held under refrigerated display (8°C) for 13 days under fluorescent light. α-tocopherol concentrations were significantly higher (P<0.05) in meat from the supplemented group than from the basal one. Whatever the measured colour characteristics (a*, R(630)-R(580),% MetMb), the vitamin E supplementation had a positive but non-significant effect on the rate of discoloration. But by visual assessment, essentially with MAP, a significant and positive effect of vitamin E supplementation was noted to lower discoloration (P<0.05). TBARS values were significantly lowered (essentially at the end of storage time for the two packaged modes) after an α-tocopheryl acetate-supplementation.

Effect of dietary fat and vitamin E on colour stability and on lipid and protein oxidation in Turkey meat during storage.

The objectives of the study were to investigate the effects of dietary fat (6% soya oil or rapeseed oil or tallow), together with tocopheryl acetate at either a basal (30 ppm) or a supplemented (400 ppm) level for 16 weeks on lipid and protein oxidation, including myoglobin, during refrigerated storage of turkey muscles. When turkeys were fed tallow in particular, vitamin E supplementation improved the vitamin E status of the muscles. Vitamin E supplementation significantly delayed lipid oxidation measured by TBARS, whatever the dietary fat. TBARS were highest in meat from animals fed soya oil. Vitamin E supplementation had no positive effect on colour stability of meat during refrigerated storage. Feeding soya oil induced significantly higher oxidation of proteins (carbonyl content) than rapeseed oil or tallow and vitamin E supplementation induced a slight decrease in carbonyl content at day 9 of storage for M. sartorius. SH content was significantly higher in vitamin E supplemented M. sartorius and M. pectoralis than in controls.

NMR relaxation of water protons in normal and malignant hyperthermia-susceptible pig muscle.

(1)H-NMR has been used to study the evolution of water proton transversal relaxation times in ageing skeletal muscles of normal and halothane-positive Pietrain pigs. Malignant hyperthermia was confirmed by the caffeine contracture test. Lactic acid, creatine phosphate, and ATP levels in muscle biopsies were measured by biochemical analysis. The NMR dynamic results revealed malignant hyperthermia, but knowledge of animal age and muscle type improved significantly the detection ability. The NMR results revealed large differences between muscles. Halothane sensitivity detection seems to be less affected by animal age, than by muscle effect but discrimination was more efficient in the older animal group. It is concluded that (1) H-NMR is a suitable method for diagnosing halothane sensitivity on a well identified muscle biopsy and that water dynamics might be related to acidosis in muscle fibres.

Application to proteomics to understand and modify meat quality.

The use of proteomics in the field of meat science has gained in robustness and accuracy. This is consistent with the genomic and bioinformatic tools. Its application to sensorial and technological meat quality traits is discussed as well as the emergence of sanitary and nutritional issue which will grow in a next future.

Kinetics of protein physicochemical changes induced by heating in meat using mimetic models: (1) relative effects of heat and oxidants.

Optimizing the nutritional quality of cooked meat needs a better understanding of the mechanisms responsible for protein changes induced by heating. The relative contributions of chemical and thermal effects on protein physicochemical changes were studied using meat models. Two models were tested: a basic model made of an aqueous suspension of myofibrillar proteins, and a complex model, in which oxidants were added in physiological concentrations. Various heating time-temperature combinations were applied to both models in the ranges 45-90 °C and 5-120 min. Protein oxidation was evaluated by carbonyl and free thiol contents. Conformational changes of proteins were assessed by measurements of surface hydrophobicity and aggregation. Carbonyl formation was weakly affected by the thermal process alone but exacerbated by oxidants. A synergistic effect of oxidants and heat treatments on protein oxidation was noted. Changes in protein hydrophobicity and aggregation were dominated by the thermal process.

Kinetics of protein physicochemical changes induced by heating in meat using mimetic models: (2) effects of fibre type, peroxides and antioxidants.

Heating-induced changes in meat proteins were investigated using models made of aqueous suspensions of myofibrils according to muscle fibre types and cellular compounds (oxidants and antioxidants). These changes were evaluated by measurements of carbonyl groups and protein surface hydrophobicity. Model results were compared to trial results obtained on pork meat (M. Longissimus dorsi) heated under the same conditions (45 and 75°C, from 5 to 120 min). Myofibrillar proteins from α-white fibres were more sensitive to oxidation and thermal denaturation than those from ß-red fibres. At 45°C, there were negligible differences due to peroxide or antioxidant types. At 75°C, organic peroxides (ROOH) were less oxidative than hydrogen peroxide (H2O2), and antioxidant enzymes were less efficient than vitamin E and carnosine at protecting proteins against oxidation. Protein oxidation observed in meat is lower than in the mimetic models and the increase in hydrophobicity remained limited in meat.

Characterization of Longissimus thoracis, Semitendinosus and Masseter muscles and relationships with technological quality in pigs. 1. Microscopic analysis of muscles.

Three porcine muscles (Longissimus thoracis, Semitendinosus, Masseter), known to have large differences in biochemical and histological traits, were fully characterized and the link between muscle structure and quality evaluated. The oxidative Masseter had more pigment, higher content of metmyoglobin, haem iron, protein and collagen, and was redder with higher fibre numbers, fibre circularity, pH and water holding capacity than the glycolytic Longissimus. Fibre type distribution showed predominance of type IIB in Longissimus and Semitendinosus white, type I in Semitendinosus red and IIA in Masseter. Type I fibres were larger than type IIB and IIA in Semitendinosus and Masseter, respectively, but not in the Longissimus, indicating that fibre size is muscle dependent. Muscle redness was positively correlated with type I fibre traits, haem iron and metmyoglobin, and negatively associated with type II fibre characteristics, non-haem iron and oxymyoglobin. Expressible juice had positive correlation with fibre size and negative with fibre number and connective tissue.