Sequential window acquisition of all theoretical mass spectra (SWATH-MS) as an emerging proteomics approach for the discovery of dark-cutting beef biomarkers.

Recent advances in "omics" technologies have enabled the identification of new beef quality biomarkers and have also allowed for the early detection of quality defects such as dark-cutting beef, also known as DFD (dark, firm, and dry) beef. However, most of the studies conducted were carried out on a small number of animals and mostly applied gel-based proteomics. The present study proposes for the first time a Sequential Window Acquisition of All Theoretical Mass Spectra (SWATH-MS) proteomics approach to characterize and comprehensively quantify the post-mortem muscle proteome of DFD (pH24 ≥ 6.2) and CONTROL (5.4 ≤ pH24 ≤ 5.6) beef samples within the largest database of DFD/CONTROL beef samples to date (26 pairs of the Longissimus thoracis muscle samples of young bulls from Asturiana de los Valles breed, n = 52). The pairwise comparison yielded 35 proteins that significantly differed in their abundances between the DFD and CONTROL samples. Chemometrics methods using both PLS-DA and OPLS-DA revealed 31 and 36 proteins with VIP > 2.0, respectively. The combination of different statistical methods these being Volcano plot, PLS-DA and OPLS-DA allowed us to propose 16 proteins as good candidate biomarkers of DFD beef. These proteins are associated with interconnected biochemical pathways related to energy metabolism (DHRS7B and CYB5R3), binding and signaling (RABGGTA, MIA3, BPIFA2B, CAP2, APOBEC2, UBE2V1, KIR2DL1), muscle contraction, structure and associated proteins (DMD, PFN2), proteolysis, hydrolases, and activity regulation (AGT, C4A, GLB1, CAND2), and calcium homeostasis (ANXA6). These results evidenced the potential of SWATH-MS and chemometrics to accurately identify novel biomarkers for meat quality defects, providing a deeper understanding of the molecular mechanisms underlying dark-cutting beef condition.

An analysis of the influence of preslaughter management factors on welfare and meat quality outcomes in fed beef cattle in the United States.

During the preslaughter phase, cattle are transported from their place of origin to a slaughter facility, experiencing transportation, lairage, environmental factors, and novel environments. Although research exists that has focused how the preslaughter phase impacts cattle welfare and meat quality, some significant preslaughter management factors and subsequent welfare and meat quality outcomes have not been thoroughly explored. The objective of this study was to assess the effects of preslaughter management factors on welfare and meat quality outcomes in fed beef cattle in the United States. Transportation factors, environmental characteristics, lairage factors, cattle characteristics, and several meat quality variables were collected from 5 federally inspected commercial processing facilities in the United States. After excluding slaughter lots that included <75% complete datasets, a total of 619 slaughter lots representing 84,508 head of cattle were used for further analysis. Predictor variables of interest included processing plant, cattle breed, sex class, operation shift at the plant, distance traveled to the plant, truck waiting time to unload at the plant, lairage duration and space allowance, temperature humidity index, and windspeed. Outcome variables of interest included cattle mobility, carcass bruising, dark cutting (DC), quality grades, and hot carcass weights. Logistic and linear regressions were used to analyze the associations between the predictor and outcome variables of interest. Increased distance traveled and truck waiting time were associated with higher odds of mobility impairment (P = 0.0009 and P = 0.007, respectively), with each 10 km increase in distance traveled having an odds ratio (OR) of 1.001 (95% confidence interval [CI]: 1.000 to 1.001) and each 1-min increase in waiting time having an OR of 1.003 (CI: 1.001 to 1.004). Conversely, a 10-km increase in distance traveled decreased the odds of carcass bruising (OR: 0.997, CI: 0.996 to 0.998; P < 0.0001). Longer lairage was associated with increased odds of DC (P = 0.0415), with each 60-min increase in duration having an OR of 1.034 (CI: 1.001 to 1.068). The results demonstrate the importance of truck arrival management (i.e., scheduling, prioritizing unloading) on mobility. Focusing on lairage management (i.e., density and time) may provide some opportunities to improve meat quality.

In-depth exploration of the high and normal pH beef proteome: First insights emphasizing the dynamic protein changes in Longissimus thoracis muscle from pasture-finished Nellore bulls over different postmortem times.

This study aimed to evaluate for the first time the temporal dynamic changes in early postmortem proteome of normal and high ultimate pH (pHu) beef samples from the same cattle using a shotgun proteomics approach. Ten selected carcasses classified as normal (pHu < 5.8; n = 5) or high (pHu ≥ 6.2; n = 5) pHu beef from pasture-finished Nellore (Bos taurus indicus) bulls were sampled from Longissimus thoracis muscle at 30 min, 9 h and 44 h postmortem for proteome comparison. The temporal proteomics profiling quantified 863 proteins, from which 251 were differentially abundant (DAPs) between high and normal pHu at 30 min (n = 33), 9 h (n = 181) and 44 h (n = 37). Among the myriad interconnected pathways regulating pH decline during postmortem metabolism, this study revealed the pivotal role of energy metabolism, cellular response to stress, oxidoreductase activity and muscle system process pathways throughout the early postmortem. Twenty-three proteins overlap among postmortem times and may be suggested as candidate biomarkers to the dark-cutting condition development. The study further evidenced for the first time the central role of ribosomal proteins and histones in the first minutes after animal bleeding. Moreover, this study revealed the disparity in the mechanisms underpinning the development of dark-cutting beef condition among postmortem times, emphasizing multiple dynamic changes in the muscle proteome. Therefore, this study revealed important insights regarding the temporal dynamic changes that occur in early postmortem of high and normal muscle pHu beef, proposing specific pathways to determine the biological mechanisms behind dark-cutting determination.

Preslaughter factors affecting mobility, blood parameters, bruising, and muscle pH of finished beef cattle in the United States.

Decades of work have focused on reducing fear, stress, and discomfort in cattle during the preslaughter phase by improving and promoting animal handling, transportation, and management processes. Even still, there is limited information about the effects of preslaughter factors on animal welfare and meat quality outcomes in finished cattle in the United States. This study aimed to track individual animals through the slaughter process to identify preslaughter factors associated with key welfare and quality outcomes. A total of 454 cattle from one commercial slaughter facility were studied. Preslaughter factors assessed included distance traveled, truck waiting time, lairage density, lairage duration, and season. Animal characteristics, i.e., body weight, breed, and sex, were also recorded. One trained observer scored the mobility of all cattle using the North American Meat Institute's 1-4 scale (i.e., normal to extremely reluctant to move). Exsanguination blood was collected and analyzed for cortisol, creatine kinase, and lactate. Carcass bruising was scored using a modified version of the National Beef Quality Audit's bruise-scoring methodology (i.e., no bruise, one bruise ≤ the size of a deck of cards, one bruise > than the size of a deck of cards, and multiple bruises). Ultimate muscle pH was measured 32 to 36 h postmortem. Multi-predictor models were selected for each outcome variable using Akaike Information Criterion. Continuous outcome variables were analyzed using linear mixed-effect models and categorical outcome variables with mixed-effect logistic regression models. Longer truck waiting times were associated with increased cortisol (P = 0.04) and lactate (P = 0.02) concentrations. Similarly, increased lairage duration was associated with increased creatine kinase concentrations (P = 0.05) and the odds of cattle being bruised (P = 0.03). Less space allowance per animal in lairage was associated with increased odds of cattle having impaired mobility (P = 0.01). There was a seasonal effect for many of the measured outcomes; the summer season was associated with greater lactate concentrations (P < 0.0001), increased odds of impaired mobility (P < 0.0001), and increased odds of carcass bruising (P = 0.003). The findings of this study indicate that many of the preslaughter factors assessed influence critical welfare and meat quality outcomes of finished beef cattle, warranting future research and consideration.

Comparing the effects of packaging normal-pH and atypical dark-cutting beef in modified atmosphere conditions on surface color.

Limited studies have determined the effects of modified atmospheric packaging (MAP) on atypical dark-cutting beef surface color. The objective was to compare the impacts of using vacuum packaging, carbon monoxide (CO-MAP), and HiOx-MAP (high­oxygen) on the retail color of normal-pH and atypical dark-cutting beef aged 14 d. Atypical dark-cutting beef (pH 5.63) had numerically greater (P > 0.05) pH than normal-pH beef (pH 5.56). Atypical dark-cutting steaks were darker in color (lower L* values; P < 0.05) than normal-pH steaks. Atypical dark-cutting steaks had greater (P < 0.05) oxygen consumption, lower (P < 0.05) relative oxygenation, and less inter-muscle bundle space (P < 0.05) than normal-pH steaks. There were no differences (P > 0.05) in redness between normal-pH and atypical dark-cutting steaks when packaged in HiOx-MAP. Although a minimal increase in pH was observed in atypical dark-cutting beef, steaks in CO-MAP had lower redness than normal-pH steaks.

Sensory quality of beef with different ultimate pH values - A Brazilian perspective.

This study evaluated Brazilian consumer perceptions of beef with different pH values at 48 h post-mortem (pHu) through sensory analysis. A total of 138 consumers evaluated raw and grilled steaks. The steaks were divided according to their pHu (normal pH < 5.8, atypical darkness, firmness, and dryness [DFD] > 5.8 pH < 6, and typical DFD pH ≥ 6). There was no difference in the visual evaluation of raw steaks or purchase intention. Evaluation of the grilled steaks showed that consumers preferred typical DFD steaks in terms of tenderness, and there was a tendency for the same behavior in terms of juiciness. No differences were observed in other evaluated parameters. Cluster analysis identified three consumer segments for visual evaluation, indicating a preference for steaks with higher pHu in terms of freshness. Additionally, four segments were identified for evaluation, with some groups expressing a preference for higher pHu meat in terms of freshness appearance, tenderness liking, and overall liking. The developed regression models for overall liking and purchase intention exhibited favorable adjustment indices, with r2 values of 0.86 and 0.57, respectively, for raw steaks and 0.90 for grilled steaks for in overall liking. Regression models indicated a strong influence of color and freshness appearance, in addition to tenderness liking and juiciness liking, on the overall liking scores of consumers. These results indicate that Brazilian consumers do not dislike dark cutting and, despite differentiating their greater tenderness, do not show a preference between the different pHu values.

High-pressure processing and modified atmosphere packaging combinations for the improvement of dark, firm, and dry beef quality and shelf-life.

This study investigated the effects of high-pressure processing (HPP) and modified atmosphere packaging (MAP) on 'dark, firm, and dry' (DFD) beef. To optimize the HPP, beef steaks (n = 180) were first processed at different pressures (0.1, 200, 300, 400, 500 MPa). It was found that 400 MPa enhanced DFD beef color and shelf-life. This optimized HPP (400 MPa) was combined with 3 MAP formulations, in a second study (40, 60, or 80% O2-MAP), to determine their effect on DFD beef steaks. HPP (400 MPa) combined with MAP improved DFD beef L* and a*, color scores, and delayed discoloration (P < 0.01). Total plate counts for DFD beef held under 60% O2-MAP was ≤6 log10 CFU/g, even after 14 d of chilled storage. These same samples had shear force and TBARS values significantly lower than observed for DFD beef held under 80% O2-MAP. HPP (400 MPa) combined with 60% O2-MAP is recommended to improve DFD beef quality and shelf-life.

Integrative proteomics and metabolomics profiling to understand the biochemical basis of beef muscle darkening at a slightly elevated pH.

Previous studies investigated the biochemical basis of dark-cutting conditions at elevated muscle pH (above 6), but the molecular basis at slightly above normal pH (between 5.6 and 5.8) is still unclear. The objective was to determine protein and metabolite profiles to elucidate postmortem muscle darkening at slightly elevated pH. Loins were selected based on the criteria established in our laboratory before sample collections, such as pH less than 5.8, L* values (muscle lightness) less than 38, and not discounted by the grader (high-pH beef with dark color are discounted and not sold in retail stores). Six bright red loins (longissimus lumborum) at normal-pH (average pH = 5.57) and six dark-colored strip loins at slightly elevated pH (average pH = 5.70) from A maturity carcasses were obtained within 72-h postmortem from a commercial beef purveyor. Surface color, oxygen consumption, metmyoglobin reducing activity, protein, and metabolite profiles were determined on normal-pH and dark-colored steaks at slightly elevated pH. Enzymes related to glycogen metabolism and glycolytic pathways were more differently abundant than metabolites associated with these pathways. The results indicated that oxygen consumption and metmyoglobin reducing activity were greater (P < 0.05) in darker steaks than normal-pH steaks. Enzymes involved with glycogen catabolic pathways and glycogen storage disease showed lower abundance in dark beef. The tricarboxylic acid metabolite, aconitic acid, was overabundant in darker-colored beef than normal-pH beef, but glucose derivative metabolites were less abundant. The majority of glycogenolytic proteins and metabolites reported as overabundant in the previous dark-cutting studies at high pH (>6.4) also did not show significant differences in the current study. Therefore, our data suggest enzymes involved in glycogen metabolism, in part, create a threshold for muscle darkening than metabolites.

A bright cherry-red color beef is ideal during meat retail and carcass grading. Any deviation from a bright red color, such as dark red color, at the interface of the 12th and 13th rib-eye area leads to carcass discounts. Various studies have determined protein, metabolite, and mitochondrial profiles to understand the biochemical basis of dark-cutting beef (muscle pH greater than 6); however, limited knowledge is currently available on muscle darkening at a slightly elevated pH. Bright red loins at normal muscle pH and darker color loins at slightly elevated pH (not discounted by a grader) were collected 72-h postmortem from a commercial beef purveyor. Surface color, oxygen consumption, metmyoglobin reducing activity, protein, and metabolite profiles were determined on normal-pH and dark-colored steaks at slightly elevated pH. The results indicated that oxygen consumption and metmyoglobin reducing activity were greater in darker steaks than normal-pH steaks. Furthermore, the protein abundance profiles of enzymes related to glycogen metabolism and glycolytic pathways were more differently abundant than metabolites associated with these pathways. Understanding the factors involved in the occurrence of dark color steaks help to minimize losses due to discount carcasses.

Bushfire exposure is associated with increased pH and dark-cutting in beef longissimus thoracis at grading.

To investigate if bushfire exposure is associated with increased loin pH, this study analysed temporal and geospatial data on fire incidence in South-Eastern Australia together with beef carcase quality and production records for fire affected animals (n = 451,299). Two outcomes were modelled: 1) loin pH at time of grading, and 2) the incidence of "high pH" defects (pH > 5.70). For both models, decreasing "time since closest fire" and "distance of property from closest fire" were associated with increasing loin pH and increased incidence of high pH carcases(p < 0.05 for all); interactions for "distance from the closest fire" with feed type (grain vs grass) and "days of fire exposure" with HGP (hormonal growth promotant) treatment (yes vs no) (p < 0.05 for both) indicate high pH outcomes were exacerbated in grass-fed and HGP treated cattle. It is concluded that exposure to bushfire is associated with increasing pH but the extent and magnitude of these increases are modulated by production factors.

Dark-cutting beef mitochondrial proteomic signatures reveal increased biogenesis proteins and bioenergetics capabilities.

Mitochondria remain active in postmortem muscles and can influence meat color via oxygen consumption. Previous studies have shown that dark-cutting compared with normal-pH beef has greater mitochondrial protein and DNA content per gram of muscle tissue. However, the mechanism regulating mitochondrial content in dark-cutting vs. normal-pH beef is still unknown. Therefore, the objective was to compare mitochondrial proteomes of dark-cutting vs. normal-pH beef using LC-MS/MS-based proteomics and mitochondrial respiratory capacity using a Clark oxygen electrode. Dark-cutting compared with normal-pH beef has up-regulation of proteins involved in mitochondrial biogenesis, oxidative phosphorylation, intracellular protein transport, and cellular calcium ion homeostasis. Mitochondria isolated from dark-cutting phenotypes showed greater mitochondrial complex II respiration and uncoupled oxidative phosphorylation. However, mitochondrial membrane integrity and respiration at complexes I and IV were not different between normal-pH and dark-cutting beef. These results indicate that dark-cutting beef has greater mitochondrial biogenesis proteins than normal-pH beef, increasing mitochondrial content and contributing to dark-cutting beef. SIGNIFICANCE: Defective glycogen metabolism resulting from chronic stress before slaughter coupled with the greater mitochondrial protein and DNA content per gram of muscle tissue promotes muscle darkening in dark-cutting phenotypes in beef. However, the mechanistic basis for this occurrence in dark-cutting phenotypes is still unknown. In this work, we show that dark-cutting beef phenotype is caused, in part, as a consequence of over-proliferation of mitochondria. This is supported by the up-regulation of proteins involved in mitochondrial biogenesis, mitochondrial electron transport, calcium homeostasis, and fatty acid metabolism. Hence, the study of mitochondrial proteome changes provides a set of mitochondrial biogenesis proteins that could be used as potential candidate markers for detecting changes in pre-slaughter developmental events contributing to dark-cutting phenotypes in beef.

Biomarkers and biosensors for the diagnosis of noncompliant pH, dark cutting beef predisposition, and welfare in cattle.

Meat quality can be affected by stress, exhaustion, feed composition, and other physical and environmental conditions. These stressors can alter the pH in postmortem muscle, leading to high pH and low-quality dark cutting (DC) beef, resulting in considerable economic loss. Moreover, the dark cutting prediction may equally provide a measure for animal welfare since it is directly related to animal stress. There are two needs to advance on-site detection of dark cutters: (1) a clear indication that biomarker (signature compounds) levels in cattle correlate with stress and DC outcome; and (2) measuring these biomarkers rapidly and accurately on-farm or the abattoir, depending on the objectives. This critical review assesses which small molecules and proteins have been identified as potential biomarkers of stress and dark cutting in cattle. We discuss the potential of promising small molecule biomarkers, including catecholamine/cortisol metabolites, lactate, succinate, inosine, glucose, and ß-hydroxybutyrate, and we identify a clear research gap for proteomic biomarker discovery in live cattle. We also explore the potential of chemical-sensing and biosensing technologies, including direct electrochemical detection improved through nanotechnology (e.g., carbon and gold nanostructures), surface-enhanced Raman spectroscopy in combination with chemometrics, and commercial hand-held devices for small molecule detection. No current strategy exists to rapidly detect predictive meat quality biomarkers due to the need to further validate biomarkers and the fact that different biosensor types are needed to optimally detect different molecules. Nonetheless, several biomarker/biosensor combinations reported herein show excellent potential to enable the measurement of DC potential in live cattle.

Influence of oxygen concentration on the fresh and internal cooked color of modified atmosphere packaged dark-cutting beef stored under chilled and superchilled conditions.

The current research evaluated the combined effect of oxygen concentration (20%, 50% and 80% O2) and storage temperature (4°C or -1.5°C) on the fresh and internal cooked color of dark-cutting (DC) beef. Steaks were packaged in modified atmosphere packaging (MAP) over a 12-day storage period. Although the metmyoglobin reducing activity (MRA) of DC steaks decreased with increasing O2 levels, lower oxygen consumption (OC) and deeper oxygen penetration depth (OPD) improved the surface color of 50%O2-MAP and 80%O2-MAP DC steaks. 80%O2-MAP resulted in the highest (P < 0.05) percentage myoglobin denaturation and alleviated the phenomenon of persistent pink (PP) observed in DC beef. Compared with normal chilling, superchilling significantly increased the redness values of DC steaks through inhibition of lipid oxidation and enhancement of MRA, while it had no effects on PP due to the limited OPD. The results suggest that 80%O2-MAP combined with superchilling can be used to improve fresh color and minimize PP of DC beef.

Sarcoplasmic and myofibrillar phosphoproteins profile of beef M. longissimus thoracis with different pHu at different days postmortem.

BACKGROUND: The abnormal ultimate pH (pHu ) in postmortem muscles affect the meat quality and results in substantial economic losses. Dark, firm, and dry (DFD) meat linked with the higher postmortem pHu values and exhibited many quality issues such as dark color, tough texture and shorter shelf life. This research aimed to investigate the effect of protein phosphorylation on variations in beef pHu in order to explore the possible mechanisms underlying DFD meat formation. RESULTS: Glycogen and lactate contents were higher, while L* and a* were lower in high pHu beef. Shear force was higher in intermediate pHu group. Global phosphorylation of sarcoplasmic proteins was higher in low pHu samples on day 1 and of myofibrillar proteins was higher in intermediate pHu meat on days 1 and 2 postmortem. Sarcoplasmic protein bands with different phosphorylation levels were identified as containing some glycometabolism and stress response proteins and phosphorylated myofibrillar protein bands were identified sarcomeric and metabolic proteins. CONCLUSIONS: Phosphorylation of multiple proteins of glycolytic pathway and contractile machinery may play critical roles in development of DFD beef. © 2021 Society of Chemical Industry.

Ageing as a method to increase bloom depth and improve retail colour in beef graded AUS-MEAT colour 4.

Meat colour is an important attribute for consumer acceptance but there is an interval between colour grading and retail display. This experiment investigates the effect of time post-slaughter (5, 14 and 40 days) on colour and bloom depth (after 1 and 24 h) for beef carcasses graded AUS-MEAT colour 4 (MC 4). Sixteen carcasses were selected at grading, 8 carcasses were graded MC 4 and 8 as compliant colour (AUS-MEAT score 2 or 3). At 5 days post-slaughter, compliant loins had greater overall bloom depth and were more red than MC 4 loins. Bloom depth (24 h) increased with ageing time but reached maximum bloom at 14 days for both treatments. After ageing for 14 and 40 days, the bloom depth of MC 4 loin was no different to the 5 day aged compliant loins after 24 h on display. Colour at grading may not be a reliable measure of retail colour considering the changes post slaughter.

Investigation of the physicochemical, bacteriological, and sensory quality of beef steaks held under modified atmosphere packaging and representative of different ultimate pH values.

The objective was to evaluate the effect of packaging conditions on bacteriological and biochemical changes in beef steaks classified here, as normal ultimate pH (pHu; 5.40-5.79), intermediate pHu (5.80-6.09) or high pHu (≥ 6.10). Steaks were packaged in vacuum packaging (VP) and modified atmosphere packaging with high oxygen (HiO2-MAP) or carbon monoxide (CO-MAP). Headspace, bacterial counts, total volatile basic nitrogen, pH, purge loss, surface color, lipid oxidation, and sensory attributes were monitored over a 14-day storage period at 2 °C. HiO2-MAP and CO-MAP of high pHu steaks resulted in a bright red color even when observed to be spoilt. Reduced purge loss and lipid oxidation, as well as increased sensory acceptability were observed for steaks with elevated pHu. Higher pHu was also accompanied by an acceleration of microbial proliferation. Of all packaging types investigated, CO-MAP exerted the best preservation, regardless of pHu and based on its antibacterial and antioxidant properties.

Preliminary study on the characterization of Longissimus lumborum dark cutting meat in Angus × Nellore crossbreed cattle using NMR-based metabolomics.

The aim of this preliminary study was to evaluate meat quality properties, muscle metabolite profile and metabolic pathways associated with the occurrence of dark cutting meat in Angus x Nellore crossbreed cattle. After 14 days' ageing, dark cutting meat presented a higher pH, lower cooking loss and colour parameters, and greater tenderness compared with normal meat. Dark cutting meat had a higher ATP level and lower concentrations of glucose-6-phosphate, lactate, glucose, serine, threonine, creatine phosphate, inosine, leucine, methionine, succinate and glucose-1-phosphate compared to normal meat. In dark cutting samples, the ultimate pH was positively correlated with carnitine and negatively correlated with glucose-6-phosphate. However, in normal meat, the ultimate pH presented a positive correlation with arginine, leucine, methionine, proline, threonine, tyrosine and valine. Pathway analysis showed that differentiation of the groups was linked to energetic pathways such as starch and sucrose metabolism, the pentose phosphate pathway, amino sugar, nucleotide sugar metabolism, and glycolysis or gluconeogenesis. In conclusion, the occurrence of dark cutting meat has a notable impact on meat quality attributes and concentrations of post-mortem glycolytic metabolites, appears to be correlated with mitochondrial activity and affects energetic metabolic pathways.

Changes in glycolytic and mitochondrial protein profiles regulates postmortem muscle acidification and oxygen consumption in dark-cutting beef.

Dark-cutting beef is a condition in which beef fails to have a characteristic bright-red color when the cut surface is exposed to oxygen. However, the mechanistic basis for this occurrence is not clear. Protein expression profiles were compared between dark-cutting and normal-pH beef using LC-MS/MS-based proteomics. Mass spectrometry analysis identified 1162 proteins in the proteomes of dark-cutting and normal-pH beef. Of these, 92 proteins had significant changes in protein abundance between dark-cutting versus normal-pH beef. In dark-cutting beef, 25 proteins were down-regulated, including enzymes related to glycogen metabolism, glucose homeostasis, denovo synthesis of adenosine monophosphate (AMP), and glycogen phosphorylase activity. In comparison, 27 proteins were up-regulated in dark-cutting beef related to oxidation-reduction processes, muscle contraction, and oxidative phosphorylation. Down-regulation of glycogenolytic proteins suggests decreased glycogen mobilization and utilization, while the up-regulation of mitochondrial transport chain proteins indicates a greater capacity to support mitochondrial respiration in dark-cutting beef. These results showed that changes in proteins involved in glycogenolysis and mitochondrial electron transport would promote the development of high-pH and greater oxygen consumption, respectively; thus limiting myoglobin oxygenation in dark-cutting beef. SIGNIFICANCE: The current understanding indicates that defective glycolysis causes less carbon flow, leading to less postmortem lactic acid formation and elevated muscle pH in dark-cutting beef. However, to the best of our knowledge, limited research has evaluated how changes in glycolytic and mitochondrial protein abundance regulate postmortem muscle acidification and oxygen consumption in dark-cutting beef. We utilized a shotgun proteomics approach to elucidate potential differences in protein profiles between dark-cutting versus normal-pH beef that may influence differences in postmortem metabolism and muscle surface color characteristics. Our study shows that down-regulation of glycolgenolytic and IMP/AMP biosynthetic proteins results in elevated postmortem muscle pH in dark-cutting beef. In addition, the up-regulation of mitochondrial protein content coupled with the higher muscle pH are conducive factors for enhanced oxygen consumption and less myoglobin oxygenation, contributing to a dark meat color typically associated with dark-cutting beef.

Determination of a pH threshold for dark cutting beef based on visual evaluation by Asian consumers.

This study defined the pH threshold for dark cutting beef of Asian consumers using a web-based survey and examined their preferences for beef appearance. Ninety individual images of beef steaks with different pH values were randomly selected to be scored by 4322 respondents. Gender, age and beef eating habits of respondents were found to impact on color preferences. With respondent's visual score of 4 set as the acceptance threshold, linear regression analysis between visual scores and beef instrumental colorimetrics, showed that the acceptance thresholds of instrumental color values were L* ≥ 31.4, a* ≥ 16.4, b* ≥ 6.5, chroma ≥17.4 and hue ≥22.5. These corresponded to pH threshold values of 6.11, 6.20, 6.28, 6.25 and 6.29, respectively. This pH benchmark (6.11 to 6.29) for dark cutters is higher than pH thresholds applied in most countries, indicating that Asian consumers prefer darker meat than expected, however, the likely tenderness of beef with pH's at this level merits consideration.

Extent of dark-cutting in beef carcasses graded Canada B4.

Recovering value from dark cutting carcasses (Canada B4) was investigated by examining twelve muscles in the loin, fore- and hindquarters of atypical (AT, pH < 5.8), borderline (BD, 5.8 < pH < 6.0) and classic (CL, pH > 6.0) dark cutting carcasses. Subjective and objective colour, purge loss, and colour stability were measured over 4 days of retail display. Forequarter muscles from all dark cutting carcasses were not different from those of normal Canada AA carcasses, suggesting that forequarter muscles may be preferentially harvested for sale through normal retail outlets. None of the adductor, biceps femoris, gluteus medius, and semitendinosus muscles in the AT carcasses were dark and all had retail display colour stability comparable to that of normal steaks, indicating that these muscles in these carcasses are undervalued. Sorting of dark cutting carcasses by longissimus thoracis pH or a* and b* values will allow for value to be recovered from atypical dark cutting carcasses.

Investigation of muscle-specific beef color stability at different ultimate pHs.

OBJECTIVE: This study was aimed to investigate the muscle-specific beef color stability at normal and high ultimate pHs. METHODS: The impact of muscle (Longissimus lumborum [LL] vs psoas major [PM]) and pH (normal ultimate pH [Np] vs high pH dark cutting beef [Hp]) on color stability, indicated by basic color traits, metmyoglobin reducing activity (MRA) and oxygen consumption (OC), as well as the lipid oxidation, were determined over 7 days of display at 4°C. RESULTS: Hp-LL had the highest pH (6.92), followed by Hp-PM (6.01), Np-PM (5.76), and Np-LL (5.52). Hp-LL had increased (p<0.05) a*, chroma and % oxymyoglobin during display. Hp-LL also had the highest metmyoglobin (MMb) reducing activity and OC among all the samples, thus, the greatest color stability, although very dark throughout storage, with lowest values for lightness (L*) and yellowness (b*). Np-LL also exhibited relatively high color stability, as a result of its lower % MMb and OC and higher MRA than psoas muscle samples. The 0.2 unit difference of the pH between Hp and Np psoas muscle, resulted in the difference of the color intensity, not the color stability. Interestingly, high pH psoas muscle (Hp-PM) did not have better color stability than Np-PM, and in fact had lower color stability than even Np-LL. The similar level of OC and lipid oxidation cannot explain the difference in color stability between Hp-PM and Np-LL. CONCLUSION: The Hp does not always show better color stability compared with Np beef, which depends on the muscle type. The balance of MRA and OC is important to keep the color in great intensity and stability in the meantime.