Genome-wide scans identify biological and metabolic pathways regulating carcass and meat quality traits in beef cattle.

Genome association studies (GWAS) provides knowledge about the genetic architecture of beef-related traits that allow linking the target phenotype to genomic information aiding breeding decision. Thus, the present study aims to uncover the genetic mechanism involved in carcass (REA: rib eye area, BF: backfat thickness, and HCW: hot carcass weight) and meat quality traits (SF: shear-force, MARB: marbling score, and IMF: intramuscular fat content) in Nellore cattle. For this, 6910 young bulls with phenotypic information and 23,859 animals genotyped with 435 k markers were used to perform the weighted single-step GBLUP (WssGBLUP) approach, considering two iterations. The top 10 genomic regions explained 8.13, 11.81, and 9.58% of the additive genetic variance, harboring a total of 119, 143, and 95 positional candidate genes for REA, BF, and HCW, respectively. For meat quality traits, the top 10 windows explained a large proportion of the total genetic variance for SF (14.95%), MARB (17.56%), and IMF (21.41%) surrounding 92, 155, and 111 candidate genes, respectively. Relevant candidate genes (CAST, PLAG1, XKR4, PLAGL2, AQP3/AQP7, MYLK2, WWOX, CARTPT, and PLA2G16) are related to physiological aspects affecting growth, carcass, meat quality, feed intake, and reproductive traits by signaling pathways controlling muscle control, key signal metabolic molecules INS / IGF-1 pathway, lipid metabolism, and adipose tissue development. The GWAS results provided insights into the genetic control of the traits studied and the genes found are potential candidates to be used in the improvement of carcass and meat quality traits.

Environmental Impacts of High-Quality Brazilian Beef Production: A Comparative Life Cycle Assessment of Premium and Super-Premium Beef.

When individual purchasing power increases in society, there is a trend toward a quantitative and qualitative increase in the consumption of products. Considering the magnitude of beef production in Brazil, environmental impacts are important factors for the domestic and international markets. This study assessed a Brazilian feedlot system characterized by high animal welfare standards that produces high-quality beef that is more marbled than that produced in grass-fed systems. We assessed the environmental impacts and compared premium and super-premium beef produced in a feedlot system using a partial Life Cycle Assessment (LCA). Data were collected directly on the property analyzed, considering environmental inputs and outputs and economic factors associated with the production stages of each line (premium versus super-premium). The results show that high-quality beef has, beyond a greater financial cost, an environmental cost, with the super-premium line producing a 286% higher carbon footprint, 297% more eutrophication, and three times higher acidification potential and land use than the premium line. The results of the environmental impacts agree with the results of production costs, reflecting a 282.82% higher production cost in super-premium than in premium animals. Footprints of 5.0323 kg, 4.7746 kg, and 8.8858 kg CO2 eq./kg live weight gain at the feedlot were found in the three lines.

Changes in the Lipid Metabolism of the Longissimus thoracis Muscle in Bulls When Using Different Feeding Strategies during the Growing and Finishing Phases.

The objective was to evaluate the supplementation strategy's effect on beef cattle during the growing phase and two systems during the finishing phase. One hundred and twenty young bulls were randomly divided in a 2 × 2 factorial design to receive either mineral (ad libitum) or protein + energy (3 g/kg body weight (BW)/day) during the growing phase and pasture plus concentrate supplementation (20 g/kg BW/day) or feedlot (25:75% corn silage:concentrate) during the finishing phase. Feedlot-fed bulls had meat (Longissimus thoracis-LT) with a higher content of lipids and saturated and monounsaturated fatty acids and a greater upregulation of stearoyl-CoA desaturase and sterol regulatory element-binding protein-1c than animals that fed on pasture (p < 0.05). On the other hand, pasture-fed bulls had meat with a higher content of α-linoleic acid, linolenic acid, and n6 and a greater n6:n3 ratio compared to the feedlot-fed group (p < 0.05). In addition, meat from pasture-fed bulls during the finishing phase had 17.6% more isocitrate dehydrogenase enzyme concentration than the feedlot group (p = 0.02). Mineral-fed and pasture-finished bulls showed down-regulation of peroxisome proliferator-activated receptor gamma (p < 0.05), while the bulls fed protein + energy and finished in the feedlot had higher carnitine palmitoyltransferase 2 expression (p ≤ 0.013). In conclusion, mineral or protein + energy supplementation in the growing does not affect the fatty acid composition of intramuscular fat of LT muscle. In the finishing phase, feeding bulls in the feedlot upregulates the lipogenic genes and consequently improves the intramuscular fat content in the meat.

Early Weaning Possibly Increases the Activity of Lipogenic and Adipogenic Pathways in Intramuscular Adipose Tissue of Nellore Calves.

This study aimed to evaluate by wide-expression profile analysis how early weaning at 120 days can alter the skeletal muscle metabolism of calves supplemented with a concentrated diet until the growth phase. Longissimus thoracis muscle samples were obtained by biopsy from two groups of calves, early weaned (EW; n = 8) and conventionally weaned (CW; n = 8) at two different times (120 days of age-T1 [EW] and 205 days of age-T2 [CW]). Next, differential gene expression analysis and functional enrichment of metabolic pathways and biological processes were performed. The results showed respectively 658 and 165 differentially expressed genes when T1 and T2 were contrasted in the early weaning group and when early and conventionally weaned groups were compared at T2. The FABP4, SCD1, FASN, LDLR, ADIPOQ, ACACA, PPARD, and ACOX3 genes were prospected in both comparisons described above. Given the key role of these differentially expressed genes in lipid and fatty acid metabolism, the results demonstrate the effect of diet on the modulation of energy metabolism, particularly favoring postnatal adipogenesis and lipogenesis, as well as a consequent trend in obtaining better quality cuts, as long as an environment for the maintenance of these alterations until adulthood is provided.

Effects of molasses supplementation on animal performance and carcass parameters of beef cattle: a meta-analysis.

Molasses must be one of the most used foods in the diet of ruminant animals; however, there is no consensus on the effect of including molasses on carcass parameters. In this context, the objective was to evaluate the effect of including molasses in the diet of feedlot cattle on performance and carcass parameters. Thirteen peer-reviewed publications with 45 treatment means were included in the dataset. The effect of molasses in beef cattle diets was evaluated by examining the weighted mean differences (WMD) between molasses treatment (diet with molasses) and control diet (diet without molasses). Heterogeneity was explored by meta-regression and subgroup analysis using genetic type and experimental period, molasses in diet (g/kg dry matter (DM)), molasses type, concentrate in diet (g/kg DM), and forage type. The inclusion of molasses in the diet increased dry matter digestibility, but reduced NDF digestibility, carcass weight, subcutaneous, and visceral fat. The main sources of variation for the responses with molasses inclusion on intake, digestibility, performance, and carcass parameters were the level of molasses inclusion and the experimental period. In general context, the inclusion of molasses in the diet between 100 to 150 g/kg of DM did not affect performance and carcass parameters. However, the inclusion of molasses above 200 g/kg reduces the average daily gain and carcass weight.

Effect of Cow-Calf Supplementation on Gene Expression, Processes, and Pathways Related to Adipogenesis and Lipogenesis in Longissimus thoracis Muscle of F1 Angus × Nellore Cattle at Weaning.

The aim of this study was to identify differentially expressed genes, biological processes, and metabolic pathways related to adipogenesis and lipogenesis in calves receiving different diets during the cow-calf phase. Forty-eight uncastrated F1 Angus × Nellore males were randomly assigned to two treatments from thirty days of age to weaning: no creep feeding (G1) or creep feeding (G2). The creep feed offered contained ground corn (44.8%), soybean meal (40.4%), and mineral core (14.8%), with 22% crude protein and 65% total digestible nutrients in dry matter. After weaning, the animals were feedlot finished for 180 days and fed a single diet containing 12.6% forage and 87.4% corn-based concentrate. Longissimus thoracis muscle samples were collected by biopsy at weaning for transcriptome analysis and at slaughter for the measurement of intramuscular fat content (IMF) and marbling score (MS). Animals of G2 had 17.2% and 14.0% higher IMF and MS, respectively (p < 0.05). We identified 947 differentially expressed genes (log2 fold change 0.5, FDR 5%); of these, 504 were upregulated and 443 were downregulated in G2. Part of the genes upregulated in G2 were related to PPAR signaling (PPARA, SLC27A1, FABP3, and DBI), unsaturated fatty acid synthesis (FADS1, FADS2, SCD, and SCD5), and fatty acid metabolism (FASN, FADS1, FADS2, SCD, and SCD5). Regarding biological processes, the genes upregulated in G2 were related to cholesterol biosynthesis (EBP, CYP51A1, DHCR24, and LSS), unsaturated fatty acid biosynthesis (FADS2, SCD, SCD5, and FADS1), and insulin sensitivity (INSIG1 and LPIN2). Cow-calf supplementation G2 positively affected energy metabolism and lipid biosynthesis, and thus favored the deposition of marbling fat during the postweaning period, which was shown here in an unprecedented way, by analyzing the transcriptome, genes, pathways, and enriched processes due to the use of creep feeding.

Assessment of corn wet distillers grains fed to crossbred bulls on feeding behavior, rumen morphology, liver abscesses and blood parameters.

Corn ethanol production has been growing in Brazil in the last ten years, generating by-products to feedlot diets. This study evaluates the effects of the inclusion of low-fat corn wet distillers grains (LF-WDG) on feeding behavior, ruminal health, liver abscesses and blood parameters of F1 Angus-Nellore bulls feedlot finished. Our hypothesis is that evaluation of data from feeding behavior, rumen and liver health would help to explain animal performance. In this trail, one-hundred animals were fed for 129 days with diets containing amounts of 0 (control), 15, 30 and 45% of LF-WDG replacing corn grain and soybean meal. Evaluations of fluctuation of dry matter intake (DMI) were carried out. Additionally, feeding behavior data were assessed by monitoring (24-h period) the feeding, rumination, time spent eating (TSE), and time expended on other activities (resting and number of meals per day). Blood variables such as pH, bicarbonate, total CO2 content, and base excess in extracellular fluid (Beecf) were determined. After slaughter, rumen epithelium was classified according to the incidence of lesions (rumenitis) and abnormalities (papillae clumped), and samples were collected for morphology and histology evaluations. Moreover, livers were scored for severity of abscesses as follow: as unabscessed (0), one or two small abscesses (A-), two to four small active abscesses (A) or one or more large, active abscesses (A+). The DMI (kg/day) differed (P = 0.03) among treatments and there is a tendency of 15 and 30 LF-WDG (% DM) had lower %DMI fluctuation compared to 0 or 45%. The TSE increased linearly (P < 0.01) as the amounts of inclusion of LF-WDG increased. Moreover, neutral detergent fiber (NDF) intake, NDF consumption rate and NDF rumination efficiency increased linearly (P < 0.01) in response to LF-WDG feeding. The incidence of rumenitis tended (P = 0.08) to be greater at 45% LF-WDG, while keratin thickness decreased linearly in bulls fed LF-WDG (P < 0.01). The severity of liver abscesses (score A+) increased linearly (P = 0.02). Regarding blood parameters, only Beecf decreased linearly (P < 0.01) in response to LF-WDG feeding. Therefore, the hypothesis of the current study was confirmed. We previous reported that F1 Angus-Nellore bulls fed LF-WDG show greater weight gain (1.94 ± 0.09 kg/day) and final body weight (620 ± 18.8 kg) when compare to control (1.8 ± 0.09 kg/day and 602 ± 18.8 kg, respectively). Here, we conclude that inclusion of 15 to 30% LF-WDG in feedlot diets improved feeding behavior without impairing ruminal health and blood parameters, driving performance and weigh gain of crossbred bulls. However, bulls fed 45% LF-WDG had greater severity of liver abscesses.

Does distillers' grains supplementation affect beef cattle performance, carcass characteristics, and meat quality? A meta-analysis study.

This study evaluated, through meta-analysis, the effect of distillers' grains (DG) supplementation to beef cattle on their performance, carcass parameters, and meat fatty acid profile. Eighty-one peer-reviewed publications with 439 treatments means were included in the data set. The effects of DG supplementation to beef cattle were evaluated using weighted mean differences (WMD) between the control group (diets with no DG) and DG group (diets with DG). Heterogeneity was explored by meta-regression and subgroup analysis using genetic type, treatment period, DG type, amount of DG in diet (g DG/kg DM), sulfur in diet (g S/kg DM), ether extract in diet (g EE/kg DM), feed systems (pasture or total mixed ration), and concentrate level in the diet (g/kg DM). Meat fatty acid profile was more affected when DG was fed to crossbreed animals. In Angus animals, DG inclusion to the diets had little effect on meat fatty acid profile whereas, a pronounced reduction was reported in meat omega-6/omega-3 ratio. In response to DG inclusion to the diets, an increase in diet sulfur content up to 6.0 g/kg and ether extract content up to 110 g/kg had no adverse effects on performance, carcass parameters, and meat physicochemical characteristics. Distillers' grains inclusions in amounts between 500 and 600 g DG/kg provided the greatest responses when evaluating carcass parameters and meat fatty acid profile. Increased CLA c9 t11, linolenic acid, and total PUFA and reduced myristic acid concentration in meat were reported when animals were fed diets containing DG.

Testing different devices to assess the meat tenderness: preliminary results.

Meat tenderness is one of the principal attribute associated with consumer preferences. This study describes tenderness measurements at three final endpoint cooking temperatures (51, 61 and 71 °C) using a mechanical Warner-Bratzler (WBSF) as the standard instrument versus digital texturometer (CT3) and penetrometer (FHT) devices. Thirty-six cross-breed heifers (Bos indicus) with initial body weight 330 ± 40 kg, 20-24 months of age, were slaughtered after 100 days on feed. Subsequently, 48 h post-slaughter, Longissimus thoracis (LT) samples were collected between the 10th and 13th ribs. Six LT samples from each animal were used to evaluate tenderness and cooking losses through analysis of variance and regression analyses. No interaction between device × temperature was observed (p = 0.57). Shear force values were greater (p < 0.05) as endpoint cooking increased and the results from CT3 were close to the ones using the WBSF (R2 = 0.76; p < 0.0001). In conclusion, the digital CT3 can replace the mechanical WBSF because these devices were strongly correlated (r = 0.85; p < 0.00). However, the results from FHT were underestimated (R2 = 0.19; p < 0.006), indicating that FHT device should not be used for the evaluation of meat tenderness.

Comparison of Dental Carcass Maturity in Non-Castrated Male F1 Angus-Nellore Cattle Finished in Feedlot.

Dental classification of carcasses is used as a parameter of cattle maturity at slaughter, and it can influence carcass and meat quality traits. Brazilian beef-packing companies use the number of permanent incisor (PI) teeth as a parameter for bonus and certification of carcasses with superior quality. However, when non-castrated male such as F1 Angus-Nellore (Bos taurus×Bos indicus) are slaughtered, only animals without PI teeth are subsidized by the breed association. We evaluated these animals finished in feedlot for 180 days with zero versus two PI teeth on the carcass and meat quality traits. At the time of slaughter, 88 carcasses were selected, forming two treatments according to dental carcass maturity (0 versus 2 PI teeth; 44 animals per category). It was demonstrated that the number of PI teeth (0 versus 2 PI) did not influence (p>0.05) carcass (weights, yield, cooling loss, ribeye area and the backfat thickness) and meat quality traits (Longissimus thoracis chemical composition, color, cooking losses, shear force and pH). Thus, dental carcass maturity (zero versus two PI teeth) does not influence non-castrated male F1 Angus-Nellore finished in feedlot for 180 days. This is the first study to demonstrate that carcasses of non-castrated male F1 Angus-Nellore with two PI teeth should be subsidized in a similar way to those with zero PI teeth. Moreover, Brazilian beef-packing companies could produce heavier and leaner carcasses of acceptable quality though the use of crossbred cattle such as non-castrated F1 Angus Nellore.

Gene expression profiling and identification of hub genes in Nellore cattle with different marbling score levels.

The marbling rate evaluation is difficult and expensive, requiring slaughter of the animal or ultrasound measurement. Thus, this trait is generally not included in animal breeding programs. The use of molecular techniques to elucidate intramuscular fat deposition may help improve this trait. In this respect, transcriptome studies and differential gene expression analysis by RNA-Seq can contribute to advances in this area. The objective of this study was to use RNA-Seq to identify differentially expressed genes (DEGs) in muscle tissue (longissimus thoracis) of Nellore cattle divergently ranked on marbling, in order to increase our understanding of genes involved in the expression of this trait. The results revealed 49 DEGs and three hub genes (CISH, UFM1, TSHZ1), all of them involved in insulin and diabetes mellitus metabolism. These results indicating key genes and pathways, which may help to develop strategies designed to select animals with greater marbling.

Transcriptome profiling of muscle in Nelore cattle phenotypically divergent for the ribeye muscle area.

This study aimed to use RNA-Seq to identify differentially expressed genes (DEGs) in muscle of uncastrated Nelore males phenotypically divergent for ribeye muscle area (REA). A total of 80 animals were phenotyped for REA, and 15 animals each with the highest REA and the lowest REA were selected for analyses. DEGs found (N = 288) belonging to families related to muscle cell growth, development, motility and proteolysis, such as actin, myosin, collagen, integrin, solute carrier, ubiquitin and kelch-like. Functional analysis showed that many of the significantly enriched gene ontology terms were closely associated with muscle development, growth, and degradation. Through co-expression network analysis, we predicted three hub genes (PPP3R1, FAM129B and UBE2G1), these genes are involved in muscle growth, proteolysis and immune system. The genes expression levels and its biological process found this study may result in differences in muscle deposition, and therefore, Nelore animals with different REA proportions.

Prediction of hub genes associated with intramuscular fat content in Nelore cattle.

BACKGROUND: The aim of this study was to use transcriptome RNA-Seq data from longissimus thoracis muscle of uncastrated Nelore males to identify hub genes based on co-expression network obtained from differentially expressed genes (DEGs) associated with intramuscular fat content. RESULTS: A total of 30 transcriptomics datasets (RNA-Seq) obtained from longissimus thoracis muscle were selected based on the phenotypic value of divergent intramuscular fat content: 15 with the highest intramuscular fat content (HIF) and 15 with the lowest intramuscular fat content (LIF). The transcriptomics datasets were aligned with a reference genome and 65 differentially expressed genes (DEGs) were identified, including 21 upregulated and 44 downregulated genes in HIF animals. The normalized count data from DEGs was then used for co-expression network construction. From the co-expression network, four modules were identified. The topological properties of the network were analyzed; those genes engaging in the most interactions (maximal clique centrality method) with other DEGs were predicted to be hub genes (PDE4D, KLHL30 and IL1RAP), which consequently may play a role in cellular and/or systemic lipid biology in Nelore cattle. Top modules screened from the gene co-expression network were identify. The two candidate modules had clear associated biological pathways related to fat development, cell adhesion, and muscle differentiation, immune system, among others. The hub genes belonged in top modules and were downregulated in HIF animals. PDE4D and IL1RAP have known effects on lipid metabolism and the immune system through the regulation of cAMP signaling. Given that cAMP is known to play a role in lipid systems, PDE4D and IL1RAP downregulation may contribute to increased levels of intracellular cAMP and thus may have effects on IF content differences in Nelore cattle. KLHL30 may have effects on muscle metabolism. Klhl protein families play a role in protein degradation. However, the downregulation of this gene and its role in lipid metabolism has not yet been clarified. CONCLUSIONS: The results reported in this study indicate candidate genes and molecular mechanisms involved in IF content difference in Nelore cattle.

Influence of oxidative damage to proteins on meat tenderness using a proteomics approach.

The objective of this study was to evaluate the association between oxidative damage to proteins (represented by protein carbonylation) and beef tenderness. Three experimental groups were selected by shear force (SF): tender (38.2 ±â€¯2.9 N), intermediate (51.9 ±â€¯6.8 N), and tough meat (74.5 ±â€¯7.8 N). Two-dimensional electrophoresis with hydrazide fluorophore derivatization was used. The structural proteins actin (ACTA1), myosin (MYL1 and MYL3), desmin (DES) and troponin T (TNNT1 and TNNT3), antioxidant proteins (PRDX1, PRDX2 and PARK7) and heat shock proteins (HSPB1, CRYAB and HSPB6) showed an increase in the oxidative damage in tender meat when compared to the intermediate and tough meat (P < .05). Decrease in oxidative damage of the metabolic enzymes (TPI1, GAPDH and ENO3) were observed in tender meat group (P < .05). The present results suggest that oxidation act on the proteins of different metabolic pathways and consequently affect meat tenderness in Angus crossbred cattle.

Prediction of meat quality traits in Nelore cattle by near-infrared reflectance spectroscopy.

The main definition for meat quality should include factors that affect consumer appreciation of the product. Physical laboratory analyses are necessary to identify factors that affect meat quality and specific equipment is used for this purpose, which is expensive and destructive, and the analyses are usually time consuming. An alternative method to performing several beef analyses is near-infrared reflectance spectroscopy (NIRS), which permits to reduce costs and to obtain faster, simpler, and nondestructive measurements. The objective of this study was to evaluate the feasibility of NIRS to predict shear force [Warner-Bratzler shear force (WBSF)], marbling, and color (*a = redness; b* = yellowness; and L* = lightness) in meat samples of uncastrated male Nelore cattle, that were approximately 2-yr-old. Samples of longissimus thoracis (n = 644) were collected and spectra were obtained prior to meat quality analysis. Multivariate calibration was performed by partial least squares regression. Several preprocessing techniques were evaluated alone and in combination: raw data, reduction of spectral range, multiplicative scatter correction, and 1st derivative. Accuracies of the calibration models were evaluated using the root mean square error of calibration (RMSEC), root mean square error of prediction (RMSEP), coefficient of determination in the calibration (R2C), and prediction (R2P) groups. Among the different preprocessing techniques, the reduction of spectral range provided the best prediction accuracy for all traits. The NIRS showed a better performance to predict WBSF (RMSEP = 1.42 kg, R2P = 0.40) and b* color (RMSEP = 1.21, R2P = 0.44), while its ability to accurately predict L* (RMSEP = 1.98, R2P = 0.16) and a* (RMSEP = 1.42, R2P = 0.17) was limited. NIRS was unsuitable to predict subjective meat quality traits such as marbling in Nelore cattle.

Association of CAST2, HSP90AA1, DNAJA1 and HSPB1 genes with meat tenderness in Nellore cattle.

The objective of this study was to evaluate the association of expression of CAPN1, CAPN2, CAST, HSP90AA1, DNAJA1 and HSPB1 genes with meat tenderness in Nellore cattle. Three experimental groups were selected by shear force (SF): moderately tender (SF=34.3±5.8N), moderately tough (SF=56.8±7.8N), and very tough meat (SF=80.4±15N). Gene expression was evaluated by real-time PCR. Expression of the CAPN1, CAPN2, CAST and CAST1 genes did not differ between groups. Expression of the CAST2 was up-regulated (P<0.05) in the moderately tough and very tough meat groups. Down-regulation of the HSP90AA1, DNAJA1 and HSPB1 genes (P<0.05) was observed in the moderately tender meat group. The present results suggest that meat tenderness in Nellore cattle does not directly depend on the expression of the CAPN1 and CAPN2 genes, but is associated with the expression of other genes such as CAST2, HSP90AA1, DNAJA1 and HSPB1.

Effect of the g.98535683A>G SNP in the CAST gene on meat traits of Nellore beef cattle (Bos indicus) and their crosses with Bos taurus.

The objective of this study was to estimate allele frequencies of the g.98535683A>G:BTAU7 SNP in the CAST gene in different genetic groups of beef cattle produced in Brazil (Nellore and their crosses with Bos taurus), and to evaluate associations between this polymorphism and meat traits. Five hundred animals from six different genetic groups were genotyped and phenotyped for shear force (SF), myofibrillar fragmentation index (MFI), rib eye area, backfat thickness, and total lipids. Alleles A and G of the SNP were detected in all genetic groups and the frequency of A was higher than G. Significant association (P<0.05) was observed between the polymorphism and meat tenderness (SF and MFI), in which genotype AA exhibited the best values. These results demonstrate for the first time the occurrence of the studied SNP in a Zebu breed and its potential application to the genetic improvement of meat tenderness in the Nellore breed (Bos indicus) and its crosses with Bos taurus.

Bioanalytical methods for the metalloproteomics study of bovine longissimus thoracis muscle tissue with different grades of meat tenderness in the Nellore breed (Bos indicus).

The work describes a metalloproteomics study of bovine muscle tissue with different grades of meat tenderness from animals of the Nellore breed (Bos indicus) based on protein separation by two-dimensional gel electrophoresis, the identification of calcium ions in protein spots by X-ray fluorescence (SR-XRF) and the characterisation of proteins by electrospray ionisation mass spectrometry. Forty (40) specimens were selected and divided into two experimental groups: animals with tough meat (TO) and animals with tender meat (TE). A third group (P) of Piedmontese breed animals (Bos taurus) was included to serve as a comparative model for the level of meat tenderness. The procedures were efficient and preserved the metal-protein structure, enabling calcium detection in protein spots by SR-XRF at a given molecular weight range of 14-97kDa. Two proteins (pyruvate kinase and albumin) were inferred to be related to the phenotypical differences in animals from the different groups.