Elisabeth J. Huff-Lonergan

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.

Influence of early postmortem protein oxidation on beef quality

The objective of this study was to examine the effect of early postmortem protein oxidation on the color and tenderness of beef steaks. To obtain a range of oxidation levels, the longissimus lumborum muscles (LM) from both strip loins of 20 steers fed either a finishing diet with vitamin E (1,000 IU per steer daily, minimum of 126 d [VITE]; n = 10 steers) or fed the same finishing diet without vitamin E (CON; n = 10 steers) were used. Within 24 h after slaughter, the LM muscle from each carcass was cut into 2.54-cm-thick steaks and individually vacuum packaged. Steaks from each steer were assigned to a nonirradiated group or an irradiated group. Steaks were irradiated within 26 h postmortem, and were aged at 4 degrees C for 0, 1, 3, 7, and 14 d after irradiation. Steaks from each diet/irradiation/aging time treatment were used to determine color, shear force, and degree of protein oxidation (carbonyl content). Steaks from steers fed the VITE diet had higher (P < 0.01) alpha-tocopherol contents than steaks from steers fed the CON diet. Immediately following irradiation, steaks that had been irradiated had lower (P < 0.05) L* values regardless of diet. Irradiated steaks, regardless of diet, had lower a* (P < 0.05) and b* (P < 0.01) values than nonirradiated steaks at all aging times. Carbonyl concentration was higher (P < 0.05) in proteins from irradiated steaks compared to nonirradiated steaks at 0, 1, 3, and 7 d postirradiation. Immunoblot analysis showed that vitamin E supplementation decreased the number and extent of oxidized sarcoplasmic proteins. Protein carbonyl content was positively correlated with Warner-Bratzler shear force values. These results indicate that increased oxidation of muscle proteins early postmortem could have negative effects on fresh meat color and tenderness.

Progress in reducing the pale, soft and exudative (PSE) problem in pork and poultry meat.

Research in the area of the pale, soft and exudative (PSE) pork and poultry meat is reviewed in this article with an emphasis on genetic, biochemical and metabolic factors contributing to the problem. Over the past five decades, there has been much more work in the pork meat area where a few genetic markers have been identified, and are currently used to remove susceptible animals from the herd. Some of the markers are linked to aberrant calcium regulation in the early postmortem muscle. The poultry industry is still not at the point of using genetic marker(s); however, some recent work has revealed several potential markers. The review also discusses environmental factors such as antemortem stress and early postmortem processing practices (e.g. chilling rate) that can influence the development and severity of the PSE phenomenon. Some of these factors are known to cause protein denaturation at the early stage of postmortem and directly contribute to poor water-holding capacity and inferior texture in fresh meat and later in processed products. A newer hypothesis suggesting that variation in protein oxidation, in response to antemortem stress and early postmortem tissue environment, can contribute to development of PSE pork is also discussed. Finally, a few recommendations for future work are proposed.

High-oxygen modified atmosphere packaging system induces lipid and myoglobin oxidation and protein polymerization.

Beef steaks from longissimus lumborum, semimembranosus, and adductor muscles (n=10; respectively) were cut at 24h postmortem, randomly assigned to either high-oxygen modified atmosphere packaging (HiOx-MAP; 80% O(2), 20% CO(2)) or vacuum (VAC), and displayed for 9days at 1 degrees C. HiOx-MAP packaged beef steaks had a rapid increase in lipid oxidation and a decrease in color stability during display. The steaks in HiOx-MAP had significantly lower tenderness and juiciness scores, and higher off-flavor scores compared to steaks in VAC. HiOx-MAP condition did not affect the postmortem degradation of troponin-T or desmin. Furthermore, autolysis of micro-calpain was not influenced by packaging. SDS-PAGE, immunoblotting, and diagonal-PAGE revealed oxidative cross-linking of myosin heavy chain in meat packaged in HiOx-MAP. These results suggest that the HiOx-MAP system may negatively affect meat quality characteristics by inducing lipid and myoglobin oxidation and cross-linking/aggregation of myosin by protein oxidation.

Contribution of postmortem changes of integrin, desmin and μ-calpain to variation in water holding capacity of pork

The purpose of this study was to examine the relationship between integrin, desmin, μ-calpain and water holding capacity in fresh pork. High levels of intact integrin at one day postmortem were negatively correlated with day 1 (P<0.05) and days 1-5 (cumulative) drip loss (P<0.05). High levels of intact integrin at five days postmortem were negatively correlated with days 1-7 (cumulative) purge loss (P<0.05). Intensity of intact desmin at one day postmortem was positively correlated with days 1-7 purge loss (P<0.01). There were positive correlations between intensity of intact desmin at day 7 and day 1 (P<0.01), days 1-5 drip loss (P<0.01) and days 1-7 purge loss (P<0.05). Autolysis of μ-calpain was associated with the degradation of desmin and drip or purge loss postmortem. Our results indicate that low levels of degradation of integrin and high levels of desmin degradation were associated with low drip loss values in fresh pork.

Rate and extent of pH decline affect proteolysis of cytoskeletal proteins and water-holding capacity in pork

The objective of this study was to determine the extent to which early postmortem (PM) pH decline influences proteolysis of the intermediate filament protein desmin, the costameric proteins vinculin and talin and autolysis of μ-calpain in the longissimus muscle (LM) of pigs from two genetic lines. Based on the LM 3h pH (H=3h pH of LM>6.0; L=3h pH of LM pH<5.7) PM, 10 carcasses per line and pH group were selected. The average 3h pH within pH group was 6.23 (H) and 5.44 (L). The LM samples were collected 24, 48, 72, and 120h PM and percent drip loss was measured after 1, 2, and 4d of storage. Samples collected at 24, 48, 72, and 120h PM were used to monitor desmin, vinculin, and talin degradation and samples collected at 24h PM were used to determine the extent of μ-calpain autolysis by immunoblotting. Higher (P<0.01) pH values at 45min, 6h, and 24h PM and lower (P<0.01) drip losses after 1, 2, and 4d of storage were recorded in the H-compared to the L-group. Abundance of the 76kDa μ-calpain autolysis product was greater (P<0.01), proteolysis of talin at all measured time points and proteolysis of desmin after 24 and 48h PM was greater (P⩽0.03) in the H-group than in the L-group. The current findings indicate activation rate of μ-calpain may be associated with proteolysis of desmin and talin and could play a role in the development of drip loss. The rate of early PM pH decline can partly explain the variation of desmin and talin degradation by affecting the activation of μ-calpain.

Identification of genetic markers associated with residual feed intake and meat quality traits in the pig

Residual feed intake (RFI) has become increasingly important and is being considered as a more reasonable approach to evaluate feed efficiency in livestock. However, the cost and technical difficulties in measuring this trait restrict the extensive adoption of RFI selection, and this makes marker assisted selection (MAS) a feasible tool. In addition, the effects on meat quality caused by low RFI selection have yet to be clarified. In this study, 11 SNPs from eight candidate genes were evaluated in a Yorkshire pig experimental population (n=169) consisting of a low RFI selection line and a randomly selected control line. Associations of these SNPs with RFI, growth rate, carcass composition, and meat quality measures including water holding capacity, pH at 2d postmortem, meat color and sensory traits were analyzed. The SNPs FTO p.Ala198Ala and TCF7L2 c.646+514A>G showed significant (P<0.05) and suggestively significant (P<0.1) associations with RFI, respectively. The MC4R SNP p.Asp298Asn was associated with backfat but it was not with ADG and meat quality attributes. Both SNPs within HNF1A were associated with intramuscular lipid content and sensory juiciness. The SNPs ACC1 c(*)384C>T and TCF7L2 c.646+514A>G were significantly (P<0.05) associated with ADG. The SNPs CTSZ p.Arg64Lys and TCF7L2 c.646+514A>G were associated with both visual scoring of meat color and the objective L-value measure of meat color. This study has identified potential genetic markers suitable for MAS in improving RFI, ADG, and meat color traits, but these associations need to be validated in other larger populations.

Disulfide bond within µ-calpain active site inhibits activity and autolysis

Oxidative processes have the ability to influence mu-calpain activity. In the present study the influence of oxidation on activity and autolysis of mu-calpain was examined. Furthermore, LC-MS/MS analysis was employed to identify and characterize protein modifications caused by oxidation. The results revealed that the activity of mu-calpain is diminished by oxidation with H2O2 in a reversible manner involving cysteine and that the rate of autolysis of mu-calpain concomitantly slowed. The LC-MS/MS analysis of the oxidized mu-calpain revealed that the amino acid residues 105-133 contained a disulfide bond between Cys(108) and Cys(115). The finding that the active site cysteine in mu-calpain is able to form a disulfide bond has, to our knowledge, not been reported before. This could be part of a unique oxidation mechanism for mu-calpain. The results also showed that the formation of the disulfide bond is limited in the control (no oxidant added), and further limited in a concentration-dependent manner when beta-mercaptoethanol is added. However, the disulfide bond is still present to some extent in all conditions indicating that the active site cysteine is potentially highly susceptible to the formation of this intramolecular disulfide bond.

Effects of selection for decreased residual feed intake on composition and quality of fresh pork

The objectives of this study were to determine the extent to which selection for decreased residual feed intake (RFI) affects pork composition and quality. Pigs from the fifth generation of selection for decreased RFI (select) and a randomly selected line (control) were utilized. Two experiments were conducted. In Exp. 1, barrows (22.6 ± 3.9 kg) from select and control lines were paired based on age and BW. The test was conducted in 8 replicates of pairs for the test period of 6 wk. Calpastatin activity and myosin isoforms profile were determined on samples from the LM. Control barrows were heavier (59.1 vs. 55.0 kg; P < 0.01) at the end of the test period. Calpastatin activity was greater (P < 0.01) in LM of select barrows than control barrows. In Exp. 2, composition and quality of gilts (114 kg) from control and select lines were determined. The model included fixed effects of line, slaughter date, melanocortin-4 receptor (MC4R) genotype, barn group, line × slaughter date, genotype × line interactions, a covariate of off-test BW, and sire, pen, and litter fitted as random effects. The select line (n = 80) had 0.043 kg less (P < 0.05) RFI per day than the control line (n = 89). Loin quality and composition were determined at 2 d postmortem. Desmin degradation was measured at 2 and 7 d postmortem. Purge, cook loss, sensory traits, and star probe texture were measured at 7 to 10 d postmortem on cooked chops. Residual correlations between RFI and composition and quality traits were calculated. Compared with the control line, carcasses from the select line tended to have less (P = 0.09) backfat, greater (P < 0.05) loin depth, and greater (P < 0.05) fat free lean. Loin chops from the select line had less (P < 0.01) intramuscular lipid content than loin chops from control line. Significant residual correlations between RFI and both tenderness (r = 0.24, P < 0.01) and star probe (r = -0.26, P < 0.01) were identified. Selection for decreased RFI has the potential to improve carcass composition with few effects on pH and water-holding capacity. However, decreased RFI could negatively affect tenderness and texture because of decreased lipid content and decreased postmortem protein degradation.

Divergent genetic selection for residual feed intake impacts mitochondria reactive oxygen species production in pigs

The objective of this study was to determine the extent to which genetic selection for residual feed intake (RFI) impacts electron leakage and reactive oxygen species (ROS) production in mitochondria from muscle and liver tissue. Understanding how genetic selection for RFI impacts animal physiology and growth efficiency is of the utmost importance as the world population increases. Production efficiency is tied directly to energy use. Mitochondria were used in this study because they produce 90% of the ATP in the body and use a large majority of dietary energy. Mitochondria were isolated from both muscle and liver tissue from pigs genetically selected for RFI (n = 8 per RFI line; 34 ± 4 kg). A 2,7-dichlorofluorscein diacetate assay was used to detect differences in hydrogen peroxide production between the more efficient low RFI line and the less efficient high RFI line. Our hypothesis was that greater efficiency would be linked to less ROS production from the mitochondria. There was less ROS production in mitochondria from the white portion of the semitendinosus in the low RFI line compared with the high RFI line, when both NADH and Flavin Adenine Dinucleotide (FADH2) energy substrates were used (glutamate and succinate, respectively). Additionally, mitochondria from the red portion of the semitendinosus in the low RFI line had less ROS production when succinate was used as an energy substrate (P < 0.05). A positive correlation was observed between RFI and ROS in mitochondria from the LM. These data indicate genetic selection for RFI may influence mitochondrial ROS production and efficiency of pork production.