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This study examined the relationship between collagen content as determined by hydroxyproline assay with other measures of connective tissue, shear force and tenderness in lamb muscle. Samples were taken from both m. longissimus lumborum (LL, loin) and the m. semimembranosus (SM, topside) of 99 lambs. Sensory tenderness and compression of the LL were not correlated to any measure of collagen or connective tissue, with one exception where compression was correlated (r. =. -. 0.28; P

This study determined the extent to which bovine longissimus lumborum muscle (LLM) myofibers are influenced by nutrition for 120 d from weaning and the time-course of recovery after severe postweaning nutritional restriction. After weaning, 3 groups of Belmont Red cattle, a tropically adapted breed, were fed to achieve rapid growth (RG,or =0.6 kg of BW gain/d; n = 16), slow growth (SG, 0.2 kg of BW gain/d; n = 17), or BW loss (WL, 10% loss of weaning weight; n = 17) over 120 d. They were then grazed as 1 group at pasture with forage supplementation for 600 d until slaughter at approximately 500 kg of BW. Samples of LLM were taken from 8 to 12 animals per treatment 6 d before (baseline) and 115, 204, 324, and 476 d after commencement of the study and from all cattle at slaughter (d 721). Myofiber characteristics were determined by immunocytochemical staining of myosin heavy chains. Cross-sectional areas (CSA) of the major myofiber types 1, 2A, and 2X in WL were reduced at d 115 compared with baseline and with the growth groups (all P0.001); however, there was little difference in the percentage of the different myofiber types (all P0.10). Differences in CSA of the major myofiber types between WL and the growth groups at 115 d were smallest for type 1 (slow oxidative) and greatest for type 2X (fast glycolytic). Consequently, the relative area (percentage of total myofiber area) of type 1 myofibers in WL was significantly greater at 115 d than in the growth groups (P0.001). During recovery from postweaning nutritional restriction, significant differences in major myofiber type percentages were not evident (all P0.10), and by 721 d CSA of myofiber types differed little between the treatment groups, although SG had greater CSA of type 1 (P0.05) and type 2A (P0.01) myofibers than WL and RG. At 721 d, the relative area of type 2A myofibers was less in WL compared with SG (P0.01) and RG (P0.05) and of type 2X myofibers greater (P0.05) in WL compared with SG. It is concluded that in the LLM of cattle undergoing severe nutritional restriction immediately postweaning, the size of the more glycolytic fiber types is more adversely affected than the more oxidative types, resulting in an increased relative area of type 1, slow oxidative myofibers. However, given adequate time and nutriment at pasture, LLM myofiber characteristics of cattle recovered to near normal after severe, chronic nutritional restriction immediately postweaning, consistent with earlier findings for beef quality.

Collagens are highly abundant mammalian proteins that contain a high content of hydroxylated amino acids such as hydroxyproline. We have exploited the high hydroxyproline content of collagen and developed a method for hydroxyproline quantification as a measure of collagen content in muscle samples. The novel method utilizes a highly selective and sensitive method of multiple reaction monitoring (MRM) by mass spectrometry. The analytical method is simple, rapid (5min), convenient (no derivatization), precise (17% RSD), accurate (90-108%), sensitive (4.88nmol/L) and linear (R(2)0.999) over three orders of magnitude (5-5000nmol/L).

The high molecular weight glycosaminoglycan hyaluronan (HA) is an essential component of the extracellular matrix (ECM), however, the link between HA regulation and development of the adipocyte ECM, which is essential for differentiation, remains undefined. Hyaluronan synthase gene expression, HA synthetic rate and molecular weight during differentiation of 3T3-L1 pre-adipocytes were compared to undifferentiated 3T3-L1 pre-adipocytes and non-adipogenic NIH/3T3 fibroblasts. In the 3T3-L1 pre-adipocytes, the predominant genes associated with HA metabolism were found to be HA synthase-2 (Has-2) and hyaluronidase-2 (Hyal-2) demonstrating a co-regulation of expression which was stimulated by adipogenic induction consequently resulting in increased synthesis of high molecular weight HA (>10 MDa) and its simultaneous degradation. Accumulation of HA correlated positively with cell number, although synthetic rate was inversely related suggesting a regulatory feedback mechanism. Within 24 h post-induction, pre-adipocytes responded with a higher HA synthetic rate and later, accumulated cytoplasmic lipid. In contrast, undifferentiated pre-adipocytes had a reduced HA synthetic rate during clonal expansion and did not accumulate lipid. HA was continuously and rapidly metabolised throughout 3T3-L1 adipogenesis, where terminal differentiation coincided with the increased generation of low molecular weight, angiogenic HA fragments, a likely prerequisite for concurrent neovascularisation of adipose tissue. This study has highlighted a relationship between HA metabolism and adipocyte differentiation, suggesting that the balance between the formation and regulation of the adipocyte extracellular matrix is finely coordinated in a growth phase-specific dependent manner.

The growth paths of 36, nine-month-old Brahman-cross steers were modified to determine the effect of their growth history on tenderness of the semitendinosus (ST) muscle. Steers were assigned to one of three treatment groups. One group of steers (uninterrupted group) was grazed on improved tropical pasture for 257 days and had an average weight gain of 0.6 kg day−1. The other two groups were fed a restricted diet of low-quality grass hay and lost on average ~ 13% of their initial live weight over 100 days. These groups were then regrown for 157 days on either pasture (pasture finished) with the uninterrupted group, or on a grain-based feedlot diet (grain finished). Growth rates of the previously restricted groups during the regrowth phase were indicative of compensatory growth and were significantly different (p < 0.05) at 0.76 (sem 0.03) kg day−1 and 1.22 (0.05) kg day−1, pasture and grain finished groups, respectively. Growth rates for both restricted groups were significantly different (p < 0.05) from the uninterrupted group [0.55 (0.02) kg day−1]. At slaughter, the grain finished group had heavier carcases, higher dressing percentages and more fat coverage, than either the uninterrupted or pasture finished groups, the latter being significantly lighter than the uninterrupted group. Tenderness was assessed by shear, compression (C) and adhesion (ADH) measurements. Shear peak force (PF) values of cooked ST samples did not differ significantly between groups. However, PF values of pressure-heat treated ST samples from the grain finished group were significantly less (p < 0.05) than comparable values from the uninterrupted group suggesting a reduced contribution of connective tissue to toughness. The pasture finished group mean PF value was not significantly different from either the uninterrupted group or grain finished group means. C and ADH values were significantly less (p < 0.05) in the grain finished group compared to the pasture finished groups values, again indicating a reduced connective tissue contribution to toughness. We conclude that the physical properties of the connective tissue component of the ST muscle may be altered by rapid compensatory growth after a weight loss phase and reduce the connective tissue contribution to toughness which may enhance meat tenderness.

Multiple reaction monitoring (MRM) mass spectrometry may be regarded as the gold standard methodology for quantitative mass spectrometry and has been adopted for the analysis of small molecules especially within the pharmaceutical industry. It can also be applied to the analysis of peptides and proteins and to the measurement of the basic building blocks of proteins, amino acids. Here, we describe the application of MRM mass spectrometry to the measurement of hydroxyproline after acid hydrolysis of various animal tissues. We show that the measurement of hydroxyproline provides an accurate and reliable estimate of the collagen content of such tissues and may be a useful indicator of meat tenderness.

Connective tissue has recently been found to play a role in mediating mammalian skeletal muscle atrophy. We investigated connective tissue remodelling in the skeletal muscle of a species of the Australian burrowing frog, Cyclorana alboguttata. Despite being inactive whilst aestivating, the frog shows an inhibition of muscle atrophy. Connective tissue size and distribution was measured in histological sections of the cruralis muscle of control and aestivating C. alboguttata. Using a custom written software application we could detect no significant difference in any connective tissue morphological parameter between the two treatment groups. Biochemical measurements of gelatinase activity showed 2-fold higher activity in aestivating gastrocnemius muscle than in controls (p < 0.001). We measured the messenger RNA transcript levels for C. alboguttata metalloproteinase 2 (MMP2) and tissue inhibitor of metalloproteinase 2 (TIMP2) in cruralis skeletal muscle using quantitative real-time PCR. The trend of reduced expression of the two genes in the aestivators did not meet statistical significance. This work indicates that aestivation in C. alboguttata leads to subtle and specific changes in some extracellular matrix remodelling factors. Their main impact is to maintain proportional representation of extracellular matrix components of skeletal muscle and therefore preserve the active frog phenotype.

The effect of growth path, as opposed to advancing age, on the biophysical and biochemical properties of muscle connective tissue was investigated. Nine-month old Brahman-cross steers were grown across either an uninterrupted path, or paths that incorporated weight-loss and then weight gain on two different diets: one group was realimented on pasture, whilst the other was realimented on a grain-based diet. Biophysical attributes of connective tissue toughness (Compression and Adhesion) in the semitendinosus muscle, were significantly reduced by treatment (P

A major aim of the research program known as SheepGENOMICS was to deliver DNA markers for commercial breeding programs. To that end, a resource flock was established, comprehensively phenotyped and genotyped with DNA markers. The flock of nearly 5000 sheep, born over two consecutive years, was extensively phenotyped, with more than 100 recorded observations being made on most of the animals. This generated more than 460 000 records over 17 months of gathering information on each animal. Here, we describe the experimental design and sample-collection procedures, and provide a summary of the basic measurements taken. Data from this project are being used to identify collections of genome markers for estimating genomic breeding values for new sheep industry traits.

Based on an association with extracellular matrix remodelling, mitosis, proliferation and adipogenic differentiation, the glycosaminoglycan hyaluronan (HA) was assessed as a marker for intramuscular fat (IMF) development (marbling) in bovine loin muscle (longissimus dorsi, LD). Loin samples collected from the quartering site of feedlot-finished Wagyu–Angus and Jersey–Limousin steers were assayed for percentage IMF (IMF%) and HA after assignment of AUS-MEAT marbling scores. There was a moderate phenotypic correlation (r2 = 0.69) between IMF% and marbling score but little variance was explained by HA concentration. Breed was not a significant factor in marbling score or IMF% but did influence the HA concentration of the LD, with Wagyu–Angus steers having 2-fold more HA than Jersey–Limousin steers at the same marbling score. The non-linear decline in fat-adjusted HA levels as marbling score increased suggests that HA concentration was associated with lean growth potential of the muscle rather than adipogenesis. Using a different experimental approach, differences in distribution and amount of HA could not be discerned in histological sections of LD from age-matched Wagyu–Hereford heifers allocated to a low (score 1) or medium (score 3) marbling score group. These findings were consistent with the absence of differences between the two groups for other indicators of fatness (IMF% and P8 fat depth), maturity and myofibre characteristics despite an increase in oxidative capacity of the muscle with age. The data support the conclusion that the concentration of HA in the LD alone was not predictive of development of intramuscular fat.

Eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) are the n-3 polyunsaturated fatty acids (n-3 PUFA) for which there is ample evidence of human health benefits, and these are also the fatty acids for which there are cut-off points for ‘source’ and ‘good source’ claims. Two consecutive experiments were conducted to determine the effect of finishing systems on the n-3 PUFA content of lamb meat in Western Australia. In experiment I, a 4-week feeding experiment was conducted using 48 Poll Dorset × Merino lambs. The lambs were divided into two lots of 24 (12 males and 12 females) and randomly assigned to either concentrate (C1) finishing on commercial lamb finishing pellets or pasture (P) finishing on kikuyu (Pennisetum clandestinum) pasture. In experiment II, 28 lambs were divided into two groups of 14 lambs (seven males and seven females) and assigned to finishing either on concentrate pellets (C2) or concentrate pellets plus linseed (C2-L). This second experiment was conducted indoors for 10 weeks. The initial liveweight (mean ± s.e.) of the lambs was 43 ± 0.6 and 32.5 ± 0.9 kg for experiments I and II, respectively. At the end of experiment II, three chops each were sampled from the leg, loin, forequarter and neck region of each carcass. The final liveweight (42 ± 0.8 v. 50 ± 1.2 kg), hot carcass weight (19 ± 0.5 v. 24 ± 0.7 kg) and GR depth (5.6 ± 0.6 v. 12.8 ± 0.6 mm) were lower (P < 0.05) for P than C1 lambs. In contrast, C2 and C2-L lambs had similar final liveweight (44 ± 0.7 v. 45 ± 0.9 kg), hot carcass weight (19 ± 0.3 v. 20 ± 0.5 kg) and GR depth (13 ± 1.3 v. 14 ± 1.2 mm). In experiment I, the total n-3 PUFA yields for C1 and P lambs in the M. longissimus lumborum were 67 ± 2.5 and 78 ± 3.2 mg per 100 g muscle, respectively. The EPA plus DHA yields were 17 and 21 mg per 100 g muscle, respectively. The sum of the long-chain (≥C20) n-3 PUFA EPA, docosapentaenoic acid and DHA for C1 and P lambs were 30 and 37 mg per 100 g, respectively. Sex had no effect on any of the n-3 fatty acids. In experiment II, the total n-3 PUFA yields for C2 lambs were 61, 54, 60 and 104 mg per 100 g for leg, loin, forequarter and neck chops, respectively. The respective values for C2-L lambs were 153, 138, 139 and 178 mg per 100 g muscle. The claimable EPA plus DHA yields for C2 lambs were 13, 10, 12 and 15 mg per 100 g of trimmed leg, loin, forequarter and neck chops, respectively. The respective values for C2-L lambs were around 2-fold higher at 27, 21, 25 and 23 mg per 100 g raw meat. All the samples from pasture-finished and linseed-supplemented groups met the 30 mg cut-off point for ‘source’ claim in Australia when the computation was based on 100 g cooked lamb serve (140 g raw). We conclude that pasture-finished lambs have more n-3 PUFA per serve than their counterparts finished indoors on commercial pellets. Further, supplementing indoor-finished lambs with linseed provided equivalent n-3 PUFA per serve to finishing lambs on pasture. Supplementation with an omega-3 source improved omega-3 per serve across the whole carcass irrespective of sex.

The effects of production and processing factors on tenderness, and colour of lamb meat produced from 7 locations as part of the Australian Sheep Industry CRC’s Information Nucleus flock were investigated, using data from 2052 lambs slaughtered in 2007. At 24 h post-slaughter, samples of m. longissimus lumborum (LL) and m. semimembranosus (SM) were collected for measurement of intramuscular fat (IMF), myoglobin, iron and copper and fresh meat colour (L*, a*, b*) and pH at 24 h measured on the LL. pH and temperature measurements made pre-rigor were used to calculate the pH at 18°C. Tenderness was measured by LL shear force at days 1 (SF1) and 5 (SF5) post-slaughter, the shear force difference (SF-diff) and SM compression and collagen concentration were determined. Retail colour stability was assessed using over-wrapped LL under simulated retail display for 3 days, according to the change in the oxymyoglobin/metmyoglobin ratio. All traits were affected by flock and date of slaughter (P < 0.001). After 4 days of ageing, 70–95% of the LL samples from all flocks, except for one, had acceptable tenderness for consumers based on their shear force. Low IMF, high LL pH at 18°C and high pH at 24 h increased SF1 and SF5 and also had an effect on SF-diff (P < 0.001). The retail colour of 44.8% of the samples on day 3 of retail display were lower than acceptable. Retail colour was influenced by IMF, pH18 and the concentration of iron and copper (P < 0.001). In conclusion, breeding and management practices that increase muscle IMF levels and reduce ultimate pH values and processing practices that result in moderate rates of pH fall post-slaughter, improve the tenderness of lamb. Extension of retail colour stability may be antagonistic to traits associated with tenderness and nutritional traits, particularly IMF and mineral levels.

The effects of sire genetics and growth path on the fascicular structure (FS) of lamb loin muscle (m. longissimus lumborum; LL) and the consequences for eating quality were studied in progeny (n = 299) of 20 Poll Dorset sires and Merino ewes. Sires were selected on the basis of their Australian sheep breeding values (ASBV) for post-weaning growth (PWWT), loin eye muscle depth (PEMD) and subcutaneous fat (PFAT). Growth of lambs was varied by time of weaning (early or late) and by feeding level within weaning group, where lambs were either fully fed throughout or were feed restricted for 55 days then re-alimented to achieve liveweight of ~45 kg at slaughter. The FS variables were determined in differentially stained histological sections of LL by using automated image segmentation and circle geometry methodology and were defined as percentage connective tissue, perimysial seam thickness and fascicular width. Sire ASBV was not a significant covariate to measures of FS, despite ASBV-related effects on growth, carcass leanness and fatness. Independent of weaning time, faster growth was induced by re-alimentation of feed-restricted lambs, resulting in LL with less connective tissue as a percentage of cross-sectional area and larger fascicles than for fully fed peers. The direction of the perimysial seam thickness response varied with treatment and time of weaning. Rapid growth of re-alimented early weaned restricted lambs resulted in thinner seams than for fully fed peers, whereas the opposite response was observed in the late-weaned lamb contrast. This suggests that the weight and age of the lamb at the time of nutritional compromise and re-alimentation are important factors in determining the thickness of perimysial connective-tissue seams. Whilst the evidence confirms differential accretion of the myofibrillar and connective tissues and despite reports of positive effects on tenderness brought about by such changes, variation in structural measures was not associated with variation in either objective measures (compression) or sensory scores for tenderness of the loin muscle. The data from the present study support the tenet that rapid growth before slaughter, combined with appropriate sire genetics, will not compromise the physical characteristics and sensory qualities of the meat.

The purpose of this study was to determine if estimated breeding value (EBV) of an animal’s sire and or the animal’s nutrition affected the structure of its M. longissimus thoracis et lumborum (LL) and, hence, the eating quality of meat derived from its carcass. Lambs were chosen based on the EBV of their sires in terms of post-weaning live weight (PWWT), post weaning fat at the C-site (PFAT), and post-weaning eye muscle depth (PEMD). Morphometric techniques were used to characterise muscle structure in terms of the distribution of intramuscular connective tissue; the variables together are called fascicular structure. Perimysial seam thickness and fascicular width were both influenced by sire estimated breeding values for PWWT, PFAT, and PEMD. Variation in fascicular structure was associated with an interaction between PEMD-EBV and PFAT-EBV of the sire. Fascicular width decreased with increased PEMD-EBV and increased with PFAT-EBV, but was not affected by PWWT-EBV. When the total seam thickness was adjusted to a common fascicular width, the lambs on a low plane of nutrition had relatively more intramuscular connective tissue than those on a high plane. The total seam thickness was negatively associated with PFAT-EBV and positively associated with PEMD-EBV. Warner Bratzler shear (WBS) peak force (PF) and initial yield were not associated with differences in sire EBV. The residual WBS shear force, peak force minus initial yield (PFIY), and compression values were negatively associated with nutrition but were positively associated with PWWT-EBV and PEMD-EBV of the sires. These latter 2 effects were moderated by nutrition. The data support the hypothesis that morphological characteristics of perimysium are genetically determined and nutritionally responsive. Variance in morphology accounted for some variance in the biophysical attributes of meat and may help explain why sheep with high muscling potential have tougher meat.

This study investigates pre-weaning growth of cattle and its effect on biochemical and histochemical markers of muscle development and subsequent biophysical attributes of eating quality. Combinations of cow (late pregnancy to mid-lactation) and pre-weaning (varying duration of access to a high-energy ration) supplementation were used to vary calf growth to weaning in 6 treatment groups. After weaning, calves were grazed together on pasture (backgrounding) and then grown rapidly on a feedlot ration (finishing) until slaughter. Biochemical and myofibre characteristics were determined in semitendinosus muscle samples collected just prior to weaning (7 months), at the end of backgrounding (13 months), and at slaughter (17 months). The concentration of sarcoplasmic protein and the activity of lactate dehydrogenase in the muscle at weaning were associated with differences in pre-weaning growth and both variables correlated positively with liveweight at weaning. Isocitrate dehydrogenase activity varied with sex, not treatment, at weaning and at the end of backgrounding. The size of myofibres at weaning related to differences in growth path and correlated positively with liveweight. Pre-weaning growth effects on these characteristics were not evident at slaughter. Biophysical properties of the meat were not affected by earlier growth path treatment, and were not correlated with biochemical characteristics or myofibre type profile. Variation in both shear peak force and adhesion was related to sex. We conclude that the effects of divergent early life growth do not persist 10 months after weaning, at least in meat quality characteristics.