A Chiral-LC-MS Method for the Simultaneous Quantification of Short-Chain Fatty Acids and D/L-Lactate in the Ruminal Fluid of Dairy Cows.

Short-chain fatty acids (SCFA) and lactate in ruminal fluid are products resulting from the microbial fermentation of substrates and can be used to reflect the composition and activity of the ruminal microbiome. Determination of SCFA and D-/L-lactate in ruminal fluid currently requires two separate protocols, which is time-consuming and costly. In this study, we have optimised and validated a simple and unified 3-nitrophenylhydrazine (3-NPH) derivatisation protocol and a 20 min chiral-LC-MS method for the simultaneous quantification of all SCFA and D- and L-lactate in ruminal fluid. This method, which requires no sample pretreatment or purification shows adequate sensitivity (limit of detection (LOD): 0.01 µg/mL), satisfactory accuracy (recovery: 88-103%), and excellent reproducibility (relative standard deviation (RSD) for repeated analyses < 3% for most analytes). The application of this method to a cohort of 24 animals allowed us to reveal a large inter-cow variation in ruminal SCFA and lactate level, the concentration range for each species, the widespread correlation between different SCFA, and the strong correlation between D- and L-lactate.

The Effect of Direct-Fed Lactobacillus Species on Milk Production and Methane Emissions of Dairy Cows.

Using direct-fed microbials to mitigate enteric methane emissions could be sustainable and acceptable to both consumers and producers. Forty lactating, multiparous, Holstein-Friesian cows were randomly allocated one of two treatments: (1) a base of ad libitum vetch (Vicia sativa) hay and 7.0 kg DM/d of a grain mix, or (2) the basal diet plus 10 mL of MYLO® (Terragen Biotech Pty Ltd., Coolum Beach, Queensland, Australia) delivering 4.17 × 108 cfu of Lactobacillus per mL. Neither feed intake (25.4 kg/d vs. 24.8 kg/d) nor milk yield (29.9 vs. 30.3 kg/d) were affected by treatment. Feed conversion efficiency was not affected by treatment when expressed on an energy-corrected milk basis (1.15 vs. 1.18 kg/kg DMI). Neither methane yield (31.6 vs. 31.1 g/kg DMI) nor methane intensity (27.1 vs. 25.2 g/kg energy corrected milk) were affected by treatments. While these results are contrary to our expectations and not significant, all were numerically in a favorable direction. Given there are reports that diet and dose rate may impact the size of any effect, we recommend a dose-response study be undertaken using a basal diet that is commonly used in pasture-based dairy systems.

Effects of Heat Stress in Dairy Cows Offered Diets Containing Either Wheat or Corn Grain during Late Lactation.

Cereal grains that differ in the rate and extent of ruminal fermentation differ in heat increment and may be used to improve thermoregulation during heat stress. This experiment investigated the responses of dairy cows in late lactation to a heat challenge when offered wheat-grain or corn-grain. Eighteen lactating cows, 220 ± 94 (mean ± standard deviation) days in milk, 3.7 ± 0.17 years of age and 558 ± 37 kg bodyweight, were allocated treatments containing 6 kg dry matter (DM)/day of wheat grain or 6 kg DM/day corn grain (9 per treatment) plus 14 kg DM/day of alfalfa hay. Measurements were made during a 7-day pre-challenge period at ambient conditions in individual stalls, during a 4-day heat challenge (temperature humidity index of 74 to 84) in individual controlled-climate chambers, then during a 7-day recovery period at ambient conditions in individual stalls. During the heat challenge, cows offered corn had lower respiration rates (p = 0.017) and greater feed intake (p = 0.021) but energy-corrected milk (p = 0.097) was not different to that of cows offered wheat. Feeding corn grain to dairy cows during a heat challenge reduced some of the negative impacts of heat stress, enabling the cows to consume more forage compared with supplementing with wheat grain.

The Impact of Plant Phytochemicals on the Gut Microbiota of Humans for a Balanced Life.

The gastrointestinal tract of humans is a complex microbial ecosystem known as gut microbiota. The microbiota is involved in several critical physiological processes such as digestion, absorption, and related physiological functions and plays a crucial role in determining the host's health. The habitual consumption of specific dietary components can impact beyond their nutritional benefits, altering gut microbiota diversity and function and could manipulate health. Phytochemicals are non-nutrient biologically active plant components that can modify the composition of gut microflora through selective stimulation of proliferation or inhibition of certain microbial communities in the intestine. Plants secrete these components, and they accumulate in the cell wall and cell sap compartments (body) for their development and survival. These compounds have low bioavailability and long time-retention in the intestine due to their poor absorption, resulting in beneficial impacts on gut microbiota population. Feeding diets containing phytochemicals to humans and animals may offer a path to improve the gut microbiome resulting in improved performance and/or health and wellbeing. This review discusses the effects of phytochemicals on the modulation of the gut microbiota environment and the resultant benefits to humans; however, the effect of phytochemicals on the gut microbiota of animals is also covered, in brief.

Dietary Fat and Betaine Supplements Offered to Lactating Cows Affect Dry Matter Intake, Milk Production and Body Temperature Responses to an Acute Heat Challenge.

Supplementing the diet of lactating cows with ingredients that increase energy density, or reduce internal heat production, may reduce some of the negative impacts of hot weather on milk yield. Thirty-two dairy cows were assigned either: (1) basal diet only, (2) basal diet plus canola oil, (3) basal diet plus betaine, or (4) basal diet plus canola oil and betaine. The basal diet was lucerne hay, pasture silage, and grain. Cows were exposed to a four-day heat challenge (temperature-humidity index 74 to 84) in controlled-environment chambers. Canola oil supplementation increased milk production (22.0 vs. 18.7 kg/d) across all periods of our experiment and increased body temperature (39.6 vs. 39.0 °C) during the heat challenge. Betaine supplementation reduced maximum body temperature during the pre-challenge period (39.2 vs. 39.6 °C) but not during the heat challenge (40.3 °C). Cows fed canola oil had greater declines in dry matter intake (5.4 vs 2.7 kg DM) and energy corrected milk (1.3 vs. 1.0 kg) from the pre-challenge to the heat challenge than other cows. Contrary to our expectations, the combination of fat and betaine supplements did not result in a clear benefit in terms of milk production or body temperature. Further work is warranted to understand the interactions between diet and hot weather.

Lipid Oxidation and Colour Stability of Lamb and Yearling Meat (Muscle Longissimus Lumborum) from SheepSupplemented with Camelina-Based Diets after Short-,Medium-, and Long-Term Storage.

This study investigated the impact of feeding pelleted diets containing camelina (Camelina sativa L. Crantz) hay (CAHP) or camelina meal (CAMP) as a supplement compared with a control pellet (CONP) diet, without vitamin E fortification. The fatty acid profile, retail colour, and lipid oxidative stability of lamb and yearling meat (m. longissimus lumborum) stored for short-, medium-, or long-periods (2 days (fresh), 45 days and 90 days) under chilled to semi-frozen conditions were determined. The CAMP diet altered key fatty acids (p < 0.05) in a nutritionally beneficial manner for human health compared to the other diets, with increased total omega-3, decreased omega-6 fatty acids and decreased omega-6/omega-3 ratio of muscle. Muscle vitamin E concentration was lower (p < 0.05) for both camelina diets (CAMP and CAHP) when compared with the CONP diet, with the average concentrations less than 1 mg/kg muscle for all three treatments. Animal type and storage length were factors that all affected (p < 0.05) colour and lipid oxidative stability of meat. These results emphasise the importance of vitamin E concentration in meat stored for extended periods under semi-frozen conditions to maintain desirable meat colour during retail display, and to avoid off-flavour development of the cooked meat.

An alternative approach for sustainable sheep meat production: implications for food security.

BACKGROUND: A pelleted diet containing camelina hay (CAMH) or camelina meal (CAMM) as a supplement along with a control pellet (CONT) diet formulated with commonly available feeds during summer was used to investigate an alternative pathway for sustainable meat production. Sustainable meat production was based on a simple estimation of income from meat produced versus feed costs if animals were fed for an extended period over summer compared to early slaughter at the beginning of summer. Eighty maternal composite wether lambs (Composite) based on Coopworth genetics and 80 pure Merino wether yearlings were divided into 10 groups within breed (n = 8) using stratified randomisation based on liveweights. Following 1 week of adaptation to experimental diets, animals were fed experimental diets for up to10 weeks. RESULTS: Animals were slaughtered after either 8, 9 or 10 weeks of full feeding when the average liveweight of diet/genetic combination reached a weight appropriate for either 'heavy lamb' or 'heavy hogget' production, which occurred between 8 and 10 weeks of full feeding. There was no diet × breed interactions except for dressing percentage (DP), where Composite lambs fed the CAMH diet had the greatest DP (48.1 ± 0.35) and the Merino yearlings fed the CAMM diet the lowest DP (45.8 ± 0.33). Composite lambs gained 17.6-20.3 kg and Merino yearlings gained 10.7-12.9 kg liveweight. Based on their DP, this resulted in the production of approximately 8.3-9.5 kg additional carcass weight in Composites and 4.9-5.7 kg in Merinos, which in turn produced greater profit per Composite lamb and a small profit per Merino yearling. CONCLUSIONS: Composite lambs fed CAMM and CAMH had 5% greater carcass weights at slaughter compared to the CONT group, but dietary treatments did not change carcass weight of Merino yearlings at slaughter. The extended feeding approach offered the producer an estimated economic gain of AUD $20.00 to $25.00 when yearly average prices were used (Method 1) and AUD $40.00 to $47.70 when pre- and post-summer average prices were used (Method 2) per Composite lambs, but extra carcass gain did not result in the same profit per Merino yearling. Among the Composites, the profit for animals fed the CAMH and CAMM were AUD $2.75 to $4.50 greater than CONT group when full year average prices were applied while AUD $3.50 to $5.50 greater than CONT group when pre- and post-summer average prices were applied. However, we acknowledge a combination approach of extended feeding for a portion of animals already on ground and selling the remaining animals pre-summer with joining of additional ewes is the most likely strategy. Considering the scenario of extended feeding for 3 weeks, based on the growth rates observed for Composite lambs, if an additional 2 kg carcass weight per animal can be gained for 50% of the 22 million lambs slaughtered in Australia (= 11 million animals), it would potentially supply an additional 22 million kg of lamb carcasses produced per annum. This is equivalent to producing an extra 1 million lamb carcasses per annum weighing 22 kg each. Feeding Composite lambs for an extended period and selling Merino yearlings pre-summer may be a good option due to faster growth rate of Composites that may help quick turn-over to market.

Effects of Feeding either Red or White Grape Marc on Milk Production and Methane Emissions from Early-Lactation Dairy Cows.

Globally, annual production of grape marc (GM), the residue of skins, seeds and stems remaining after making wine, has been estimated to be approximately nine million tonnes. No previous studies have compared effects on milk production and methane emissions when GM from either red or white grapes was fed to dairy cows. This experiment examines the effects of partial replacement of a perennial ryegrass (Lolium perenne L.) based diet with GM from either red or white grapes on yield and composition of milk and methane emissions. Thirty-two Holstein dairy cows in early lactation were offered either a control diet containing 15.0 kg dry matter (DM) of freshly harvested perennial ryegrass and 5.2 kg of a concentrate mix, or a diet similar to the control diet but with 5 kg DM of ryegrass replaced with 5 kg DM of GM from red grapes (RGM), or a diet similar to the RGM diet except the GM was from white grapes (WGM). Individual cow feed intakes, milk yields, and methane emissions were measured. Both diets containing GM decreased milk yields by approximately 10% and methane emissions by 15%. When fed to dairy cows, GM reduces methane emissions but at the cost of decreased milk production.

Understanding the action of muscle iron concentration on dark cutting: An important aspect affecting consumer confidence of purchasing meat.

We investigated the association of muscle iron concentration, in addition to ultimate pH (pHU), on dark meat formation in sheep of different breeds fed forage-based diets. At 1 h simulated display, redness of meat (a*-value) increased (P < .0001) by about 3 units as the iron concentration increased from 10 to 22 mg/kg of meat, whereas the a*-value decreased by 2 units as pHU increased from 5.5 to 6.2 in fresh meat (P < .0001). After 90 days storage the corresponding responses were about 2 units increase for iron concentration and about 1 unit decrease for pHU, respectively. The results clearly show that increased muscle iron concentration was strongly associated with reduced dark cutting in fresh and stored meat evaluated at 1 h simulated display. We conclude that it may be desirable to measure iron concentration, along with pHU, for evaluation of the potential for carcasses to produce dark cutting meat, and for the meat to turn brown during display.

Volatile Fatty Acids in Ruminal Fluid Can Be Used to Predict Methane Yield of Dairy Cows.

The dry matter intake (DMI) of forage-fed cattle can be used to predict their methane emissions. However, many cattle are fed concentrate-rich diets that decrease their methane yield. A range of equations predicting methane yield exist, but most use information that is generally unavailable when animals are fed in groups or grazing. The aim of this research was to develop equations based on proportions of ruminal volatile-fatty-acids to predict methane yield of dairy cows fed forage-dominant as well as concentrate-rich diets. Data were collated from seven experiments with a total of 24 treatments, from 215 cows. Forage in the diets ranged from 440 to 1000 g/kg. Methane was measured either by open-circuit respiration chambers or a sulfur hexafluoride (SF6) technique. In all experiments, ruminal fluid was collected via the mouth approximately four hours after the start of feeding. Seven prediction equations were tested. Methane yield (MY) was equally best predicted by the following equations: MY = 4.08 × (acetate/propionate) + 7.05; MY = 3.28 × (acetate + butyrate)/propionate + 7.6; MY = 316/propionate + 4.4. These equations were validated against independent published data from both dairy and beef cattle consuming a wide range of diets. A concordance of 0.62 suggests these equations may be applicable for predicting methane yield from all cattle and not just dairy cows, with root mean-square error of prediction of 3.0 g CH4/kg dry matter intake.

Muscle Antioxidant Enzymes Activity and Gene Expression Are Altered by Diet-Induced Increase in Muscle Essential Fatty Acid (α-linolenic acid) Concentration in Sheep Used as a Model.

This study investigated the effect of dietary manipulations on muscle fatty acid composition, the activities and relative mRNA expressions of antioxidant enzymes and the relationship between muscle enzyme activity or mRNA expression and alpha linolenic acid (ALA) concentration in sheep. Eighty-four lambs blocked on liveweight were randomly allocated to four dietary treatments, lucerne pasture (Lucerne), annual ryegrass pasture (Ryegrass), feedlot pellets (Feedlot) or annual ryegrass plus feedlot pellets (RyeFeedlot). After six weeks of feeding, lambs were slaughtered and within 30 min post-mortem, samples collected from the longissimus lumborum (LL) muscle for RNA isolation and measurement of antioxidant enzyme activities. At 24 h post-mortem, LL samples were collected for determination of fatty acid concentrations. Feedlot treatment decreased ALA, eicosapentaenoic (EPA), docosapentaenoic (DPA) and docosahexaenoic (DHA) concentrations compared with other treatments and increased linoleic acid (LA) and arachidonic acid (AA) compared with Lucerne and Ryegrass (p < 0.001). The activity of Glutathione peroxidase (GPX1, p < 0.001) and Superoxide dismutase (SOD2, p < 0.001) enzymes in the muscle increased with Lucerne compared to other treatments. Lucerne increased muscle gpx1 mRNA expression by 1.74-fold (p = 0.01) and 1.68-fold (p = 0.05) compared with Feedlot and other diets, respectively. The GPX1 (r² = 0.319, p = 0.002) and SOD2 (r² = 0.244, p = 0.009) enzyme activities were positively related to ALA. There was a positive linear relationship between muscle gpx1 (r² = 0.102, p = 0.017) or sod2 (r² = 0.049, p = 0.09) mRNA expressions and ALA concentration. This study demonstrates that diet can affect concentrations of ALA and other fatty acids as well as change activities and gene expression of antioxidant enzymes in muscle. Increased antioxidant activity may, in turn, have beneficial effects on the performance, health and wellbeing of animals and humans.

The effect of diet of the donor cows on in vitro measurements of methane production from wheat and corn incubated in various forage-to-grain ratios.

BACKGROUND: Supplementation of ruminant diets with wheat and corn grains influences ruminal fermentation. In vitro fermentation is a methodology that can be used to screen feeds for their potential to produce enteric methane. However, there is evidence that the diet of the donor cows could impact the results of in vitro analysis. This research investigated the in vitro fermentation of wheat and corn grain when incubated in ruminal fluid from cows fed different grain types and different forage-to-grain ratios. RESULTS: The type of grain fed to the donor cows, as well as forage-to-grain ratio, affected the outcome of fermentation of wheat and corn grain. Differences in methane production (MP) between grains were only observed when incubated with ruminal fluid adapted to each specific grain type. Increasing proportions of wheat but not of corn decreased in vitro MP in a linear manner compared with MP produced from forage only. CONCLUSIONS: Wheat grain has a greater in vitro antimethanogenic effect than corn. However, to detect the different fermentations between wheat and corn, grains should be incubated in ruminal fluid from cows adapted to that specific grain type. © 2019 Society of Chemical Industry.

Comparison of five methods for the estimation of methane production from vented in vitro systems.

BACKGROUND: There are several methods for estimating methane production (MP) from feedstuffs in vented in vitro systems. One method (A; 'gold standard') measures methane proportions in the incubation bottle's headspace (HS) and in the vented gas collected in gas bags. Four other methods (B, C, D and E) measure methane proportion in a single gas sample from the HS. Method B assumes the same methane proportion in the vented gas as in the HS, method C assumes constant methane to carbon dioxide ratio, method D has been developed based on empirical data, and method E assumes constant individual venting volumes. This study aimed to compare the MP predictions from these methods to that of the gold standard method under different incubation scenarios, to validate these methods based on their concordance with a gold-standard method. RESULTS: Methods C, D and E had greater concordance (0.85, 0.88 and 0.81), lower root-mean-square error (RMSE; 0.80, 0.72 and 0.85) and lower mean bias (0.20, 0.35, -0.35) with the gold standard than did method B (concordance 0.67, RMSE 1.49 and mean bias 1.26). Methods D and E were simpler to perform than method C, and method D was slightly more accurate than method E. CONCLUSION: Based on precision, accuracy and simplicity of implementation, it is recommended that, when method A cannot be used, methods D and E are preferred to estimate MP from vented in vitro systems. © 2018 Society of Chemical Industry.

Interaction of diet and long ageing period on lipid oxidation and colour stability of lamb meat.

Eighty-four crossbred wether and ewe lambs were allocated to four finishing diets. The diets were: Lucerne pasture (n=24), Annual ryegrass with sub clover pasture (n=18), Standard commercial feedlot pellets (n=24) and Annual ryegrass based pasture and commercial feedlot pellets (500g/day/head) (n=18). After 8weeks of feeding the lambs were slaughtered and the m. longissimus (LL) and m. semimembranosus (SM) were vacuum packaged and held chilled for 5 (fresh) or 60 (long aged) days, after which samples of each were displayed for 4days under simulated retail conditions. Irrespective of muscle type the long aged samples exhibited a rapid reduction in redness (a*-values) and R630/580nm ratio values such that consumer acceptable thresholds for both traits were quickly exceeded providing limited shelf life. Long ageing also lead to high TBARS levels measured as MDA mg/kg muscle suggestive of a product likely to exhibit rancidity and off flavours from lipid oxidation.

Survey of the Variation in Grape Marc Condensed Tannin Composition and Concentration and Analysis of Key Compositional Factors.

Grape marc contains a number of compounds with potential antimethanogenic activity in ruminants, including condensed tannins (CTs). Using direct phloroglucinolysis, a survey of CT chemistry across 66 grape marc samples showed diversity in concentration (6.9 to 138.8 g/kg of dry matter). Concentration was found to be independent of CT composition, although all compositional variables were significantly correlated (P < 0.0001). Twenty samples diverse in CT were selected from a cluster analysis and analyzed for compounds relevant to ruminant digestion and methanogenesis, including metabolizable energy (6.6-12.0 MJ/kg DM), crude protein (3.2-14.4% DM), neutral detergent fiber (18.4-61.4% DM), and ethanol soluble carbohydrates (2.0-40.6% DM). Fatty acid concentrations varied throughout the 20 samples (5.2-184.5 g/kg DM), although fatty acid profile showed two distinct groups. Grape marc varies widely in nutritional value, and in compounds that have been linked with changes in ruminant digestion and methane emissions.

Muscle antioxidant (vitamin E) and major fatty acid groups, lipid oxidation and retail colour of meat from lambs fed a roughage based diet with flaxseed or algae.

The effect of feeding flaxseed or algae supplements to lambs on muscle antioxidant potential (vitamin E), major fatty acid groups, lipid oxidation and retail colour was investigated. Lambs (n=120) were randomly allocated to one of 4 dietary treatments according to liveweight and fed the following diets for eight weeks: Annual ryegrass hay [60%]+subterranean clover hay [40%] pellets=Basal diet; Basal diet with flaxseed (10.7%)=Flax; Basal diet with algae (1.8%)=Algae; Basal diet with flaxseed (10.7%) and algae (1.8%)=FlaxAlgae. Flaxseed or algae supplementation significantly affected major fatty acid groups in muscle. The addition of algae (average of Algae and FlaxAlgae) resulted in lower vitamin E concentration in muscle (P<0.003; 1.0 vs 1.3mg/kg of muscle) compared with lambs fed a diet without algae (average of Basal and Flax). Increasing muscle EPA+DHA by algae supplementation significantly increased lipid oxidation, but retail display colour of fresh meat was not affected.

Forms of n-3 (ALA, C18:3n-3 or DHA, C22:6n-3) Fatty Acids Affect Carcass Yield, Blood Lipids, Muscle n-3 Fatty Acids and Liver Gene Expression in Lambs.

The effects of supplementing diets with n-3 alpha-linolenic acid (ALA) and docosahexaenoic acid (DHA) on plasma metabolites, carcass yield, muscle n-3 fatty acids and liver messenger RNA (mRNA) in lambs were investigated. Lambs (n = 120) were stratified to 12 groups based on body weight (35 ± 3.1 kg), and within groups randomly allocated to four dietary treatments: basal diet (BAS), BAS with 10.7 % flaxseed supplement (Flax), BAS with 1.8 % algae supplement (DHA), BAS with Flax and DHA (FlaxDHA). Lambs were fed for 56 days. Blood samples were collected on day 0 and day 56, and plasma analysed for insulin and lipids. Lambs were slaughtered, and carcass traits measured. At 30 min and 24 h, liver and muscle samples, respectively, were collected for determination of mRNA (FADS1, FADS2, CPT1A, ACOX1) and fatty acid composition. Lambs fed Flax had higher plasma triacylglycerol, body weight, body fat and carcass yield compared with the BAS group (P < 0.001). DHA supplementation increased carcass yield and muscle DHA while lowering plasma insulin compared with the BAS diet (P < 0.01). Flax treatment increased (P < 0.001) muscle ALA concentration, while DHA treatment increased (P < 0.001) muscle DHA concentration. Liver mRNA FADS2 was higher and CPT1A lower in the DHA group (P < 0.05). The FlaxDHA diet had additive effects, including higher FADS1 and ACOX1 mRNA than for the Flax or DHA diet. In summary, supplementation with ALA or DHA modulated plasma metabolites, muscle DHA, body fat and liver gene expression differently.

The synergism of biochemical components controlling lipid oxidation in lamb muscle.

Lipid oxidation of M. longissimus lumborum in fresh or vacuum packaged (aged) lamb meat stored at 3 °C for 0 or 4 weeks, respectively and displayed under refrigerated conditions for a further 4 days was assessed by measuring the concentration of malondialdehyde (MDA) in meat using the thiobarbituric acid reactive substances procedure. The effects of vitamin E, heme iron and polyunsaturated fatty acids (n-6 and n-3) on lipid oxidation were examined. Results showed a strong positive relationship between heme iron, n-6 and n-3 fatty acids and lipid oxidation when vitamin E was below 2.95 mg/kg muscle. When lipid oxidation was related to vitamin E concentration and the other three variables, respectively, any increase in heme iron or n-6 or n-3 fatty acids concentration did not influence lipid oxidation. Management of diet to elevate muscle vitamin E concentration above 3.45 mg/kg muscle is beneficial to maintain the level of lipid oxidation below 2.4 mg MDA/kg muscle in meat stored for up to 4 weeks. This demonstrates that vitamin E concentration in muscle has a greater influence on controlling lipid oxidation in muscle tissues than do heme iron or polyunsaturated fatty acids.

Relationship between muscle antioxidant status, forms of iron, polyunsaturated fatty acids and functionality (retail colour) of meat in lambs.

The relationship between muscle vitamin E, forms of iron, polyunsaturated fatty acids (PUFA) and the redness of meat (retail display) at days 3 to 4 post slaughter from lambs offered 2 different diets was examined. Meat redness was positively related to vitamin E and heme iron and negatively related to total n-3, total n-6 and total PUFA content. However, after adjusting for the effects of vitamin E and heme iron content, there was no indication of any residual relationship between redness at days 3-4 of retail display and total n-3, total n-6 or total PUFA. This indicates that the relationship between PUFA and redness in meat is mediated through the effects of heme iron and vitamin E in the muscle. It appears that the level of highly oxidisable PUFAs in muscle tissues do not play a major role in maintenance of redness at days 3-4 of retail display, but the level of vitamin E and heme iron content are important.