Meat Science 89 (2011) 469–477

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The effects of feeding flaxseed to beef cows given forage based diets on fatty acids oflongissimus thoracis muscle and backfat

R.T. Nassu a,b, M.E.R. Dugan a,⁎, M.L. He c,d, T.A. McAllister c, J.L. Aalhus a, N. Aldai a,e, J.K.G. Kramer f

a Lacombe Research Centre, Agriculture and Agri-Food Canada, Lacombe, AB, Canadab Embrapa Pecuaria Sudeste, Sao Carlos, SP, Brazilc Lethbridge Research Centre, Agriculture and Agri-Food Canada, Lethbridge, AB, Canadad University of Saskatchewan, Saskatoon, SK, Canadae Instituto de Ganadería de Montaña, CSIC-ULE, Finca Marzanas, 24346, Grulleros, Leon, Spainf Guelph Food Research Centre, Agriculture & Agri-Food Canada, Guelph, ON, Canada

⁎ Corresponding author at: Lacombe Research CentCanada, 6000 C & E Trail, T4L1W1, Lacombe, AB, Canada. T403 782 6120.

E-mail address: duganm@agr.gc.ca (M.E.R. Dugan).

0309-1740/$ – see front matter. Crown Copyright © 20doi:10.1016/j.meatsci.2011.05.016

a b s t r a c t

a r t i c l e i n f o

Article history:Received 8 January 2011Received in revised form 11 May 2011Accepted 16 May 2011

Keywords:Cull cowFlaxseedOmega-3 fatty acidsConjugated linoleic acids

This study was conducted to investigate changes in fatty acid profiles of beef cows fed grass hay or barleysilage based diets, with or without flaxseed supplementation. Both flaxseed and hay feeding increased levelsof α-linolenic acid (LNA; 18:3n-3) in longissimus thoracis and backfat (Pb0.001). A forage type by flaxseedlevel interaction was observed for most LNA biohydrogenation intermediates (Pb0.05) that indicated feedinghay combined with flaxseed led to the greatest levels of total conjugated linolenic acid, total conjugatedlinoleic acid, total non-conjugated dienes and total trans-18:1. Predominant biohydrogenation intermediatesincluded t11,c15-18:2, rumenic acid (c9,t11-18:2) and vaccenic acid (t11-18:1).

Crown Copyright © 2011 Published by Elsevier Ltd. All rights reserved.

1. Introduction

Over the past 100 years then-6/n-3 fatty acid ratio in the humandietin North America has increased from1–2 to 1 up to 20–30 to 1, a changewhich has coincidedwith an increase in the incidence of related chronicdiseases including cardiovascular disease (Simopoulos, 1999). As aresult of their linkage to human health, regulatory authorities haverecently approved food labeling claims for total n-3 fatty acids, forexample in Canada this level is ≥300 mg per serving (CFIA, 2003).Although this claim applies to all n-3 fatty acids, these compounds differin their biological activity with long-chain (LC; N18 carbon) n-3polyunsaturated fatty acids (PUFA) typically having greater potency(Bailey, 2009; Lunn & Theobald, 2006). The LC n-3 PUFA are mostcommonly associated with marine sources, but due to their limitedintakes in several countries, alternative sources contribute substantiallyto their intake. For example, in Australia, beef accounts for 28% of LC n-3PUFA intake (Howe, Meyer, Record, & Baghurst, 2006). Development ofbeef with enhanced levels of total n-3 fatty acids could, therefore, resultin substantial increases in LC n-3 PUFA intake for humans, and providean opportunity to add value to beef.

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Flaxseed contains~40%oil, andof this 50–60% is linolenic acid (LNA),making it one of the richest plant sources of n-3 fatty acids. Feedingflaxseed is known to increase levels of n-3 fatty acids in pork, poultryand dairy products, and consumption of these products has beendemonstrated to help maintain red blood cell n-3 fatty acid levels inhumans (Legrand et al., 2010). Feeding cattle flaxseed or flaxseedproducts also increases n-3 fatty acids in beef (Kronberg, Barcelo-Coblijn, Shin, Lee, &Murphy, 2006; Scollan et al., 2001), but enrichmentin adipose tissue and meat is limited by bacterial biohydrogenation inthe rumen (Raes,DeSmet,&Demeyer, 2004). Biohydrogenation leads toextensive loss of unsaturated fatty acids and the accumulation of partialhydrogenation products such as vaccenic acid (VA, trans (t)11-18:1)and rumenic acid (RA, cis (c)9,t11-18:2) which have many purportedhealth benefits (Field, Blewett, Proctor, & Vine, 2009; Park, 2009). Thusfeedingflaxseedmay also present opportunities for producingbeefwithenhanced levels of partial biohydrogenation intermediates of LNA.

In Canada, finishing youthful cattle (i.e. under 30 months of age)on forage is not common, but mature breeding animals are typicallyfed greater levels of forage. Among Canadian beef grades, cull cowshave been shown to have higher levels of VA, RA and n-3 fatty acidscompared to beef finished on high concentrate diets, which maypresent an economically viable opportunity to produce beef withenhanced levels of beneficial fatty acids (Dugan, Rolland, Aalhus,Aldai, & Kramer, 2008). The objective of the present experiment wasto feed flaxseed to cull cows in a relatively high forage diet (i.e. 50:50forage: concentrate, DM basis) and measure the accumulation of n-3

hts reserved.

470 R.T. Nassu et al. / Meat Science 89 (2011) 469–477

fatty acids and partial biohydrogenation products of PUFA in beef.Moreover, we wished to establish whether feeding barley silage orgrass haywould influence beef fatty acid composition, as the influenceof forage type on the fatty acid composition of ruminant products hasreceived little attention (Chilliard, Ferlay, & Doreau, 2001), althoughthere is some indication that forage type can interact with flaxseed tomodulate milk fatty acid composition in dairy goats and cows(Chilliard & Ferlay, 2004; Shingfield et al., 2005) as well as affectbeef fatty acid composition (Aharoni, Orlov, & Brosh, 2004; Mir et al.,2003).

2. Materials and methods

2.1. Animals and diets

Sixty-four British by continental crossbred (N30 months of age)non-lactating, non-pregnant beef cows with body weight (BW)averaging 620±62 kg were used, and the feeding trial was conductedat the Lethbridge Research Centre. Animals were cared for accordingto the Canadian Council on Animal Care guidelines (CCAC, 1993).Cows were randomly assigned to four diets, with four pens of fourcows per diet. Cows had ad libitum access to feed and water.

Diets were formulated to meet or exceed nutrient requirementsfor mature cows (Table 1; (NRC, 2000)). The diets had a 50:50 forageto concentrate ratio (DM basis) and were fed as total mixed rations.Diets included grass hay control (HC), barley silage control (SC), grasshay plus flaxseed (HF) and barley silage plus flaxseed (SF). Flaxseedwas ground with barley in a 7:3 ratio through a 4 mm screen in ahammermill. The groundmaterial was screened, and whole flaxseeds

Table 1Experimental diets, nutrient concentration and fatty acid profiles in lipids extractedfrom experimental diets.

Hay control(HC)

Hay+flaxseed(HF)

Silage control(SC)

Silage+flaxseed(SF)

Diet ingredients (g/kg)Barley 473.7 327.4 292.8 201.7Barley silage 0.0 0.0 678.0 682.0Grass hay 479.1 483.6 0.0 0.0Flaxseed 0.0 141.3 0.0 87.1Supplementa 47.2 47.6 29.2 29.3

Diet compositionDry matter, % 87.76 88.58 54.24 54.56Crude protein,g/kg

136.5 153.9 126.4 143.7

NEm, Mcal/kg 1.69 1.73 1.77 1.81NEg, Mcal/kg 1.06 1.08 1.15 1.17NDF, g/kg 330.9 303.9 333.2 306.3ADF, g/kg 196.6 199.6 183.9 186.9Crude fat, g/kg 17.6 63.2 18.1 62.5

Fatty acid, % total FAME14:0 0.39 0.09 0.97 0.2716:0 18.83 7.71 19.72 8.66c9-16:1 0.14 0.07 0.17 0.0818:0 2.37 2.57 1.93 2.54c9-18:1 16.50 15.10 17.12 15.3218:2n-6 (LA) 40.55 20.99 43.99 23.0618:3n-3 (LNA) 19.24 52.46 14.03 48.96SFA 22.17 10.61 23.18 11.71MUFA 17.92 15.91 18.74 16.23PUFA 59.91 73.48 58.09 72.06

NDF: Neutral detergent fiber; ADF: Acid detergent fiber; LA—linoleic acid; LNA;linolenic acid; SFA: Saturated fatty acids; MUFA: Monounsaturated fatty acids; PUFA:Polyunsaturated fatty acids.

a The supplement was composed of: 56.5% barley, 10% canola meal, 2% urea, 25%limestone, 3% salt, 0.066% VIT E 500, 1% premix, 0.05% flavor and 2.5% molasses, whichprovided to diets in 5% (in DM) and supply to 1 kg diet (in DM) with additional:14.67 mg copper, 58.32 mg zinc, 26.73 mg manganese, 0.66 mg iodine, 0.23 mg cobalt,0.29 mg selenium, 4825 IU vitamin A, 478 IU vitamin D and 32 IU vitamin E.

recovered were estimated to account for approximately 2% of theflaxseed added to the diet. The flaxseed diets contained 15% flaxseedsubstituted for dry rolled barley (DM basis), resulting in a total dietarylipid content of ~6%. Diets were fed ad libitum for 20 weeks onaverage.

At the end of the feeding period, cowswere shipped the day beforeslaughter to the Lacombe Research Centre (~6 h) and held overnightwith access to water. Cows were slaughtered in groups of 16 with onepen of animals per diet per slaughter date and all animals wereslaughtered within 30 d. Carcasses were chilled overnight at 2 °C. At24 h post mortem, carcasses were knife ribbed between the 12 and13th rib, and a 2.54 cm loin steak with overlying backfat was removedfor fatty acid analyses.

2.2. Fatty acid analysis

From the loin steak collected, 5 g of subcutaneous fat was sampledand the longissimus thoracis (LT) muscle was comminuted using a RobotCoupe Blixir BX3 (Robot Coupe USA Inc., Ridgeland, MS). Subcutaneousfat and a 15–20 g subsample of LT were stored at−80 °C for subsequentfatty acid analyses. Prior to analyses, lipid extractions from LT, fatty acidmethylations from subcutaneous fat and LT, and GC and Ag+−HPLCanalyseswere conducted as describedby Juárez et al. (2011). Specifically,subcutaneous fat (50 mg)was freeze-dried anddirectlymethylatedwith0.5 N sodium methoxide (Cruz-Hernandez et al., 2004). Intramuscularlipids were extracted from the meat samples with 2:1 chloroform:methanol using a 20:1 solvent to sample ratio (Folch, Lees, & Stanley,1957). Toderivatize allmeat lipid classes, extractsweremethylatedusing5% methanolic HCl, and to correct for CLA isomerization, separatemethylations with 0.5 N sodium methoxide were conducted. Fatty acidmethyl esters (FAME) were analyzed using GC (acid and basicmethylations) according to Kramer, Hernandez, Cruz-Hernandez, Kraft,and Dugan (2008) and CLA isomer analysis by Ag+−HPLC (basicmethylation) according to Cruz-Hernandez et al. (2004). Additionalbiohydrogenation products of LNA, specifically t11,t15-18:2 and c9,t11,t15-18:3, were identified based on their published GC/MS characteriza-tion (Gómez-Cortés, Bach, Luna, Juárez, & de la Fuente, 2009) and theirrelative retention compared to known fatty acids.

2.3. Statistical analysis

Data were analyzed using the MIXED procedure of SAS version 9.2(SAS, 2009) and the model included flaxseed supplementation, foragetype and their interaction with slaughter date and pen nested withinthe interaction as random factors. Pen was used as the experimentalunit. Significances were reported at Pb0.05 and trends were reportedat Pb0.10. Fatty acids with concentrations less than 0.05% of totalFAME were not reported in tables.

3. Results and discussion

3.1. Diets

The diet compositions are summarized in Table 1. Among the dietsthe amount of net energy, neutral detergent fiber and acid detergentfiber were similar. Crude protein contents met or exceeded cowrequirements (NRC, 2000). Crude fat contents of flaxseed containingdiets were approximately 45 g/kg greater than control diets. In the HCand SC diets, linoleic acid (LA, 18:2n-6) was the most abundant fattyacid at 40.6 and 44.0% of total fatty acids, respectively, followed byLNA, 16:0 and c9-18:1. In flaxseed diets (HF and SF), LNA was themost abundant fatty acid accounting for 52.5 and 49.0% of total fattyacids, respectively. Levels of PUFA biohydrogenation intermediateswere negligible for all diets.

471R.T. Nassu et al. / Meat Science 89 (2011) 469–477

3.2. Longissimus thoracis fatty acid composition

3.2.1. n-3 and n-6 fatty acidsRecovery of FAME from LT lipid averaged N96% for all diets and

there were no differences in total LT FAME between diets (Table 2).Including flaxseed in the diet increased the percentage of LNA in LTintramuscular fat (Pb0.001). Feeding hay versus barley silage alsoincreased the percentage of LNA (Pb0.001), but no flaxseed by foragetype interaction was observed. Feeding either flaxseed (Barton,Marounek, Kudrna, Bure, & Zahrádková, 2007; Kronberg et al., 2006;Raes et al., 2004) or substituting forage for concentrate in the diet(Aharoni et al., 2004 ) has previously been demonstrated to increaseLNA in beef. Forage type has also been demonstrated to influence thelevel of LNA in beef independent of the LNA concentration in theforage. Forage effects have been related to inhibition of rumenlipolysis (i.e. the first step leading to LNA biohydrogenation) bypolyphenol oxidase products present in red clover (Lee, Parfitt,Scollan, & Minchin, 2007) and also by tannins (Vasta et al., 2009). Inthe present experiment, however, rumen lipolysis of flaxseed oil didnot appear to be preferentially inhibited by components of either hayor barley silage. Overall, although higher levels of LNA were observedin LT when feeding flaxseed or hay, the overriding effects of ruminal

Table 2Effect of forage type and flaxseed feeding to cull cows on longissimus thoracis muscle total

Hay Silage

No flax Flax No flax

Total FAME (mg/g muscle) 56.80 58.75 67.72PUFA cis/methylene interrupted 5.35 5.58 4.16n-6 3.97 3.36 3.24

18:2n-6 2.60 2.40 2.1020:3n-6 0.33 0.22 0.2920:4n-6 0.84 0.56 0.63

n-3 1.19 1.89 0.7418:3n-3 0.51 1.22 0.3120:5n-3 0.24 0.27 0.1322:5n-3 0.44 0.40 0.30

n-6/n-3 3.32b 1.78c 4.39a

Total CLNA 0.08b 0.15a 0.07bc9,t11,t15-18:3 0.02b 0.06a 0.02bc9,t11,c15-18:3 0.06 0.09 0.05

Total non-CLA dienes 0.59c 2.34a 0.59c

Total CLA 0.50b 1.01a 0.41c

Total t,t-CLA 0.08c 0.19a 0.07c

Total c,t-CLA 0.42b 0.81a 0.33c

Total MUFA 48.78 49.18 51.04Total cis-MUFA 47.28 43.25 49.74c9-14:1 0.71 0.70 0.76c7-16:1 0.17 0.15 0.15c9-16:1 4.46 3.66 4.30c9-18:1 37.97 34.65 40.57c11-18:1 0.16 0.48 0.11c12-18:1 0.12c 0.45a 0.10c

c13-18:1 0.41 0.38 0.43c14-18:1 0.03c 0.08a 0.03c

c15-18:1 1.59 1.30 1.65Total trans 18:1 1.49c 5.93a 1.30c

Total SFA 43.13 40.39 42.5214:0 2.75 2.78 2.7515:0 0.30 0.31 0.2916:0 26.62 23.39 25.9617:0 0.73 0.67 0.7218:0 11.27 11.70 11.43

Means in the same rowwith different superscripts are significantly different (Pb0.05); SEM, standof18:3n−3,20:5n−3andC22:5n−3 ]; PUFA cis/methylene interrupted=sumof18:2n-6,18:3nconjugated linolenic acid; CLA, conjugated linoleic acid; t,t-CLA, total tran,-trans-CLA isomert12-18:2, t9,t11-18:2, t8,t10-18:2, t7,t9-18:2 and t6,t8-18:2; Total c,t-CLA: sum of t12,c14-18:2,and t9,c11-18:2; Total non-CLAdienes: sumof t11,t15-18:2, c9,t13-/t8,c12-18:2, t8,c13-18:2, c9tsum of c9-14:1, c9-15:1, c7-16:1, c9-16:1, c10-16:1, c11-16:1, c12-16:1, c13-16:1, c5-17:1, c7-1c11-20:1 and c13-22:1; Total trans 18:1: sumof t6-t8-18:1, t9-18:1, t10-18:1, t11-18:1, t12-18:121:0, 22:0 and 24:0.

biohydrogenation were apparent. Considerably greater LNA enrich-ments can be found in milk fat when infusing LNA post-rumenally(Khas-Erdene et al., 2010) or in monogastrics including pigs whencomparable levels of flaxseed are fed (Juárez et al., 2010).

Including LNA in the diet from either flaxseed or hay increased levelsof 20:5n-3 (P=0.005 and P=0.028, respectively), but again noflaxseedby forage type interaction effect was observed. The increase in 20:5n-3suggests increasing the availability of LNA results in enhanced synthesisof 20:5n-3 by chain elongation and desaturation (Dewhurst, Scollan,Lee, Ougham,&Humphreys, 2003; Scollanet al., 2001). Feedingflaxseeddid not increase 22:5n-3, but feeding hay as opposed to barley silageincreased 22:5n-3 (P=0.022). Feeding grass silage has been previouslyshown to increase 22:5n-3 (Warren et al., 2008), but in the presentstudy 22:5n-3 in cows fed grass hay was not further increased whenflaxseed was included in the diet. Similar to the present study, Scollanet al. (2001) found feedingflaxseed to beef cattle did not increase 22:5n-3 or 22:6n-3 (docosahexeonic acid; DHA), and they suggested this maybe due to inhibition of DHA synthesis or a failure to compete forincorporation into tissues. Li et al. (1999) alsodemonstrated that dietarysupplementation with LNA increases 20:5n-3 in blood lipid of humans,but further elongation and desaturation beyond this fatty acid arelimited, resulting in negligible increases of DHA. There are, however,

fatty acid methyl esters (FAME) and percentage (%) FAME in total FAME.

P value

Flax SEM Forage Flax Forage⁎flax

64.13 5.185 0.116 0.873 0.5894.892 0.345 0.008 0.164 0.4692.99 0.263 0.038 0.103 0.4752.12 0.157 0.015 0.564 0.4810.17 0.028 0.137 0.001 0.9120.53 0.077 0.080 0.024 0.3341.64 0.093 0.001 b0.001 0.2601.06 0.044 0.001 b0.001 0.6740.23 0.028 0.005 0.028 0.1750.36 0.04 0.022 0.804 0.2271.83c 0.138 0.002 b0.001 0.0030.08b 0.009 0.001 0.001 0.0060.02b 0.004 0.001 b0.001 b0.0010.06 0.008 0.027 0.093 0.1931.46b 0.067 b0.001 b0.001 b0.0010.55b 0.031 b0.001 b0.001 b0.0010.10b 0.007 b0.001 b0.001 b0.0010.44b 0.027 b0.001 b0.001 b0.001

51.11 0.661 b0.001 0.653 0.75547.51 0.699 b0.001 b0.001 0.1610.75 0.062 0.408 0.848 0.9710.15 0.006 0.184 0.016 0.2333.86 0.217 0.931 0.013 0.408

38.58 0.636 b0.001 0.001 0.2860.47 0.036 0.429 b0.001 0.6220.21b 0.023 b0.001 b0.001 b0.0010.39 0.025 0.590 0.169 0.9360.06b 0.004 0.039 b0.001 0.0391.54 0.065 0.041 0.009 0.1893.59b 0.198 b0.001 b0.001 b0.001

40.70 0.634 0.800 0.003 0.4362.80 0.121 0.904 0.768 0.9610.32 0.014 0.953 0.214 0.447

23.61 0.466 0.642 b0.001 0.3570.71 0.022 0.635 0.083 0.330

11.94 0.399 0.591 0.203 0.910

ard error of themean; n−6/n−3 ratio=[(sumof 18:2n−6, 20:3n−6 and20:4n−6/(sum-6,20:2n-6, 20:3n-6,20:4n-6, 22:4n-6,18:3n-3, 18:4n-3, 20:3n-3,20:5n-3and22:5n-3;CLNA,s; c,t-CLA, total cis-trans-CLA isomers. Total t,t-CLA: sum of t12,t14-18:2, t11,t13-18:2, t10,c12,t14 18:2, t11,c13-18:2, c11,t13-18:2, t10,c12-18:2, t8,c10-18:2, t7,c9-18:2, c9,t11 18:212-18:2/c16-18:1, t9c12-18:2, t11c15-18:2, c9c15-18:2, and c12c15-18:2; Total cis-MUFA:7:1, c9-17:1, c10-17:1, c9-18:1, c11-18:1, c12-18:1, c13-18:1, c14-18:1, c15-18:1, c9-20:1,, t13-t14-18:1, t15-18:1, t16-18:1; Total SFA: sumof14:0, 15:0, 16:0, 17:0, 18:0, 19:0, 20:0,

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Fig. 1. Effect of forage type and flaxseed feeding in atypical dienes of cull cowslongissimus thoracis muscle.Means within the same fatty acid with different letters aresignificantly different (p≤0.05). α:Significant forage effect (p≤0.05). β:Significantflaxseed effect (p≤0.05).

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Fig. 2. Effect of forage type and flaxseed feeding in conjugated linoleic acids (CLA) of cullcows longissimus thoracis muscle. Means within the same fatty acid with differentletters are significantly different (p≤0.05).

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differences in elongation and desaturation between species and tissuesand differences between bovine tissues are currently a matter ofinvestigation (Gruffat et al., 2011).

Feeding hay versus barley silage increased LA in LT (Pb0.015), butLA levels were not affected by feeding flaxseed (PN0.05). Feedingflaxseed, however, reduced 20:3n-6 (Pb0.001) and 20:4n-6(P=0.024), which was also found when feeding linseed to beef cattle(Scollan et al., 2001). These changes in n-6 PUFA in beef wereattributed to the competition between LA and LNA for the sameelongation and desaturation enzymes. No forage type by flaxseedlevel interactionwas found for any n-6 fatty acid. Shingfield, Reynolds,et al. (2005), Shingfield et al. (2005) also found higher levels of LA andLNA in milk from hay compared to silage fed cows, and this wasrelated to higher transfer efficiencies from the diet to the milk whenfed hay (29 and 17%) than when fed silage (15 and 3%), respectively.Lower rates of ruminal biohydrogenation have been reported whenfeeding fresh grass versus grass silage in sheep (Doreau & Poncet,2000), and as reviewed by Chilliard et al. (2001), several studies haveshown milk from cows fed hay have higher LNA levels compared tosilage fed cows.

3.2.2. Diene and triene biohydrogenation productsDietary PUFA, unless protected, are subjected to biohydrogenation

processes by rumen bacteria. The first step in PUFA biohydrogenationis isomerization to conjugated fatty acids. In the case of LA, it isisomerized to conjugated linoleic acid (CLA) whereas LNA isisomerized to conjugated linolenic acid (CLNA). Feeding flaxseed orhay increased total CLNA (Pb0.001) and a forage type by flaxseedinteraction indicated the highest level of total CLNA was obtainedwhen feeding flaxseed combined with hay (Pb0.006, Table 2). Thepresence and structure of c9,t11,c15-18:3 as an intermediate in LNAbiohydrogenation were confirmed by Destaillats, Trottier, Galvez, andAngers (2005), and more recently c9,t11,t15-18:3 has been shown tobe a second CLNA isomer formed as an intermediate during LNAbiohydrogenation (Gómez-Cortés et al., 2009). In the present study,the forage type by flaxseed interaction for total CLNA was mainly aresult of alterations in the concentration of c9,t11,t15-18:3, as nosignificant interaction was observed for c9,t11,c15-18:3. The level ofc9,t11,c15-18:3 was, however, increased by feeding hay compared tosilage (P=0.027) and a trend for an increase was seen when flaxseedwas included in the diet (P=0.093). Feeding grass silage as opposedto concentrate has been previously shown to increase c9,t11,c15-18:3in beef (Faucitano et al., 2008), but interactions between forage typeand flaxseed level on biohydrogenation of LNA have not beenextensively investigated. Raes, De Smet, Balcaen, Claeys, and Demeyer(2003) fed flaxseed with either grass silage or maize silage, and grasssilage resulted in the accumulation of higher levels of CLA in beef butno other biohydrogenation dienes were reported. In studies wheresunflower oil (a rich source of LA) was fed to try and increase CLAlevels in beef, feeding pea hay instead of barley silage supportedgreater levels of CLA deposition in beef muscle (Mir et al., 2003).Combined, these results along with ours indicate that feeding flaxseedwith hay as opposed to barley silage results in greater accumulationsof initial intermediates of LNA biohydrogenation.

During biohydrogenation, LNA is isomerized to CLNA, thenreduced to non-conjugated dienes or possibly CLA and subsequentlyto trans or cis MUFAs, and finally to 18:0 (Chilliard et al., 2007). Theforage type by flaxseed interaction for CLNA was also observed forbiohydrogenation dienes, with feeding flaxseed combined with hayleading to the highest levels of total non-conjugated dienes and totalCLA including both total c/t- and total t/t-CLA (Pb0.001; Table 2).

With regard to individual non-conjugated dienes, the forage type byflaxseed interactionwasmainly attributed to t11,c15-18:2 and this wasfurther supported by similar interactions for c9,t13/t8,c12-18:2, c12,c15-18:2 and t11,t15-18:2 (Fig. 1). The level of t8,c13-18:2 and c9,t12-18:2/c16:18:1 was, however, only increased by feeding flaxseed

(Pb0.001), while t9,c12-18:2 was only increased by feeding hay versussilage (P=0.007). Feeding grass silage versus concentrate waspreviously demonstrated to increase levels of t11,c15-18:2 in beef(Faucitano et al., 2008), but again interactive effects of forage type andflaxseed onbiohydrogenation of LNAhave not been investigated in beef.

Of the individual c/t-CLA isomers, RA was the major isomer and aforage type by flaxseed interaction indicated the highest levels of RAwere attainedwhen flaxseedwas fed in combinationwith hay (Fig. 2).The remaining c/t-CLA isomers were generally affected by similarinteractions. Significantly higher levels of CLA were found in milkfrom cows fed fresh grass compared to silage and in hay compared tosilage (Shingfield, Reynolds, et al., 2005; Shingfield, Salo-Väänänen,et al., 2005; Chilliard et al., 2007). Fresh grass and hay must, therefore,create a rumen environment that favors initial steps in PUFA

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2

2.5

Fig. 3. Effect of forage type and flaxseed feeding in monounsaturated fatty acids of cullcows longissimus thoracismuscle.Means within the same fatty acid with different lettersare significantly different (p≤0.05). α:Significant forage effect (p≤0.05). β:Significantflaxseed effect (p≤0.05).

473R.T. Nassu et al. / Meat Science 89 (2011) 469–477

biohydrogenation, or inhibits the reduction from trans-18:1 to 18:0.This is further supported by Boufaïed et al. (2003) who reported ratesof 18:0 formation during incubations with mixed rumen bacteria invitro that were higher for silage than for hay prepared from the samegrass. The effects of forage type on concentrations of CLA arecomplicated by the fact that forage type can influence Δ9-desaturaseactivity (Dewhurst et al., 2003) and most RA is synthesized in thetissues after absorption of VA from the rumen (Griinari et al., 2000).

Overall, the forage type by flaxseed level interaction observed forCLNA was further reflected in CLA and non-conjugated dienes. Basedon levels of intermediates present, biohydrogenation of LNA waslikely more through pathways incorporating non-conjugated dienesas opposed to CLA which is consistent with the biohydrogenation ofLNA in vitro and in dairy cattle (Jenkins, Wallace, Moate, & Mosley,2008). However, the potential health effects of many non-conjugateddienes are still not known. Consequently, if flaxseed is to be fed toruminants at elevated levels, it will be necessary to determinewhether non-conjugated dienes have any positive or negative effectson human or animal health (Chilliard et al. (2007).

3.2.3. Monounsaturated fatty acids (MUFA)In the LT, total cis-MUFA were significantly higher when feeding

silage compared to hay (Pb0.001) and lower when flaxseed was notincluded in the diet (Pb0.001; Table 2). The cis-MUFA isomer highest inconcentration across all dietswas c9-18:1 and changes in total cis-MUFAwere reflected by changes in c9-18:1 (Pb0.001). Levels of c7-16:1 andc9-16:1 were also decreased by flaxseed feeding (P=0.016 andP=0.013, respectively). Aharoni et al. (2004) and Barton et al. (2007)found no changes in muscle c9-18:1 when flaxseed was added to beefdiets, but Kronberg et al. (2006) found a reduction in c9-18:1. In thepresent study, it would appear that LNA and/or its biohydrogenationproducts may have inhibited c9-16:1 and c9-18:1 synthesis, as feedingPUFA has previously been shown to reduce Δ-9 desaturase geneexpression in beef (Waters, Kelly, O'Boyle, Moloney, & Kenny, 2009). Inthe present experiment, feeding silage resulted in slightly higher levelsof c9-18:1, possibly the result of slightly higher c9-18:1 levels in thesilage containing diets. Alternatively, if feeding silage promoted morecomplete biohydrogenation of PUFA to 18:0 in the rumen, absorbed18:0 could be a substrate for Δ-9 desaturase and c9-18:1 synthesis(Chilliard & Ferlay, 2004).

It would be expected that cis-MUFAwith double bonds past carbon9 from the carboxyl end would be produced if LNA was biohydroge-nated, since LNA has double bonds in the Δ9, 12 and 15 positions.Consistent with this, forage type by flaxseed interactions were foundfor c12-18:1 (Pb0.001) and c14-18:1 (P=0.039) with the highestlevels found when feeding hay combined with flaxseed. The level ofc11-18:1 was also higher when feeding flaxseed (Pb0.001) but noeffect of forage type was noted. Interestingly, a forage type by flaxseedinteraction was not found for c13- or c15-18:1, but instead c15-18:1mimicked changes in c9-18:1 (Pb0.05). The pathways for LNAbiohydrogenation to its many metabolites have not been established,and it is not clear how feeding flaxseed could actually lead to adecrease in muscle c15-18:1.

Total trans-MUFA in intramuscular fat was influenced by a foragetypebyflaxseed level interaction,with thehighest level foundwhenhaywas fed with flaxseed (Table 2). This interaction was observed for t6 tot9-, t-11, t-12 and t-13/14-18:1 (Fig. 3) and a similar trendwas seen fort10-18:1 (P=0.06). Chilliard and Ferlay (2004) also found a forage typeby flaxseed interaction in milk with higher levels of VA when flaxseedwas fed togetherwithalfalfa hay as compared tomaize silage. Increasingtrans-18:1 in intramuscular and subcutaneous fat with flaxseedsupplementationwas also reported to be greater when feeding flaxseedcombined with high versus low forage diets (Aharoni et al., 2004).

Feeding flaxseed increased t15- and t16-18:1 but these were notaffected by a forage type or a forage type by flaxseed interaction.Potentially, t15- and t16-18:1 could be terminal hydrogenation

products or less prone to be influenced by forage type or a foragetype by flaxseed interaction. This would be consistent with thefindings of Kemp, Lander, and Gunstone (1984) where reduced ratesof 18:1 to 18:0 biohydrogenation were found when double bondswere located closer to the methyl end of 18:1.

Overall, the feeding of flaxseed resulted in the accumulation oftrans-MUFA with double bonds from Δ11 to Δ16, but VA was still themost abundant isomer. VA is the precursor for RA that is synthesizedby Δ-9 desaturase in mammalian tissue (Griinari et al., 2000). Othertrans isomers such as t10-18:1 are associated with coronary arterydisease in humans (Hodgson, Wahlqvist, Boxall, & Balazs, 1996), butthe formation of this fatty acid is generally associated with feedinghigh-concentrate diets rich in PUFA (Aldai et al., 2010). Thebioactivities of many individual trans-MUFA and LNA biohydrogena-tion products have yet to be determined, and for this reason, ifflaxseed is to be fed to ruminants at elevated levels, it will beimportant to examine their potential bioactivity in animals andhumans.

3.2.4. Saturated fatty acids (SFA)There were no differences in total intramuscular SFAwhen feeding

hay or silage. Total SFA were, however reduced by feeding flaxseed(P=0.003) and this was mainly due to a reduction in 16:0, the mostabundant SFA (Pb0.001; Table 2). Levels of 14:0, 15:0, 17:0 and 18:0were unchanged by either forage type or flaxseed level in the diet(PN0.05). Scollan et al. (2001) also found feeding flaxseed to steersreduced 16:0 in both neutral lipids and phospholipids of muscle while18:0 was not affected. Barton et al. (2007) also reported lower 16:0and no change in 18:0 in intramuscular tissue of flaxseed-fed heifers.Adding flaxseed to a hay based diet also reduced levels of 10:0 to 16:0in goat milk (Chilliard & Ferlay, 2004), and reductions in all casesbeing attributed to the inhibitory effect of LNA and/or its biohydro-genation products on de novo fatty acid synthesis.

3.3. Backfat fatty acid composition

3.3.1. n-3 and n-6 fatty acidsThe incorporation of n-3 fatty acids into backfat was much lower

than in LT. The levels of LNAwere less than one third, while their long-

474 R.T. Nassu et al. / Meat Science 89 (2011) 469–477

chain PUFA metabolites were present only in very small amounts.However, the response to diet was similar to that seen in LT, withhigher levels found when feeding hay (P=0.004) compared to silagediets, and when flaxseed was included in diets (P=0.001), but noforage type by flaxseed interactions were found. LNA was the mostabundant n-3 fatty acid, with changes in its concentration beingreflected in total n-3 fatty acids (Pb0.005). Others have reportedsimilar increases in n-3 fatty acids in backfat of beef when flaxseedwas fed (Barton et al., 2007; Raes et al., 2004; Scollan et al., 2001), andwhen feeding forage as a source of LNA (Faucitano et al., 2008).

The effects of diet on total n-6 fatty acids were different both inamount and in response to the feeding of flaxseed betweenmuscle andbackfat. The incorporation of n-6 fatty acids into backfat was generallyone third of that found in muscle. While diet had no effect on the n-6fatty acids in muscle, a forage type by flaxseed level interaction wasfound in backfat (P=0.045; Table 3). Feeding flaxseed decreased totaln-6 fatty acids in adipose tissuewhen fed together with hay, but did notchangen-6 fatty acid levelswhen fedwith barley silage. LAwas themostabundant n-6 fatty acid, and changes in LA were reflected in changesseen for total n-6, and this is consistent with some reports (Raes et al.,2004; Scollan et al., 2001). Raes et al. (2004) attributed the difference inn-6 fatty acid levels betweenmuscle and adipose tissue to differences intheir phospholipid to neutral lipid ratio and the higher PUFA to SFA ratioin phospholipids. Conceivably n-6 fatty acids might be actively orselectively incorporated into muscle with its relatively high phospho-lipid to neutral lipid ratio,while levels of n-6 fatty acids in adipose tissuemight be more supply dependant and reflective of their plasma

Table 3Effect of forage type and flaxseed feeding to cull cows on backfat percentage (%) fatty acid

Hay Silage

No flax Flax No flax

PUFA-cis/methylene interrupted 1.79 2.40 1.48n-6 1.24a 1.12b 1.06b

18:2n-6 1.14a 1.06ab 0.98b

20:3n-6 0.07 0.04 0.06n-3 0.39 0.88 0.24

18:3n-3 0.35 0.79 0.20n-6/n-3 3.23b 1.28c 4.66a

Total CLNA 0.00c 0.17a 0.00c

c9,t11,t15-18:3 0.00c 0.10a 0.00c

c9,t11,c15-18:3 0.00c 0.07a 0.00c

Total non-CLA dienes 0.62c 3.49a 0.55c

Total CLA 0.70bc 1.73a 0.54c

Total t,t-CLA 0.04c 0.15a 0.04c

Total c,t-CLA 0.65bc 1.58a 0.50c

Total MUFA 53.78 54.01 55.36Total cis-MUFA 52.04 47.71 53.98c9-14:1 1.78 1.50 1.55c7-16:1 0.15 0.12 0.12c9-16:1 7.35 5.21 6.40c9-18:1 38.21 36.13 41.21c11-18:1 1.75 1.28 1.93c12-18:1 0.10c 0.43a 0.09c

c13-18:1 0.55 0.47 0.62c14-18:1 0.02 0.09 0.02c15-18:1 0.10 0.87 0.10Total trans-18:1 1.74c 6.30a 1.38c

Total SFA 42.96a 38.22c 42.02a

14:0 3.79 3.46 3.3415:0 0.49 0.43 0.4116:0 27.89 23.20 26.8917:0 0.73 0.63 0.7518:0 7.51 8.29 8.38

Means in the same rowwith different superscripts are significantly different (Pb0.05); SEM, sand 20:4n−6/(sum of 18:3n−3, 20:5n−3 and C22:5n−3 ]. PUFA cis/methylene interru20:3n-3, 20:5n-3 and 22:5n-3; CLNA: Conjugated linolenic acid; t-t-CLA: trans–trans conjugatt11,t15-18:2, c9,t13-/t8,c12-18:2, t8,c13-18:2, c9,t12-18:2/c16-18:1, t9,c12-18:2, t11,c15-1t10,t12-18:2, t9,t11-18:2, t8,t10-18:2, t7,t9-18:2 and t6,t8-18:2; Total c,t-CLA: sum of t12,cc9,t11 18:2 and t9,c11-18:2; Total cis-MUFA: sum of c9-14:1, c9-15:1, c7-16:1, c9-16:1, c10-1c12-18:1, c13-18:1, c14-18:1, c15-18:1, c9-20:1, c11-20:1 and c13-22:1; Total trans 18:1: sum

concentrations. Changes in adipose tissue n-6 fatty acid levels acrossall experiments have not, however, been consistent as Barton et al.(2007) reported a slight increase in LA when feeding flax seed.Influences of diet on LA concentrations in tissues, therefore, appear tobe complex, and although related to absolute levels in the diet, alsoappear to be influenced by tissue type and potentially to basal dieteffects on rumen (i.e. biohydrogenation) conditions.

Similar to the LT, levels of 20:3n-6 in backfat were lower whenfeeding flaxseed (Pb0.001) and this is likely related to the competitiveinhibition of elongation and desaturation of LA by LNA.

3.3.2. Diene and triene biohydrogenation productsSimilar to LT, levels of all diene and triene biohydrogenation

intermediateswere affectedbya forage typebyflaxseed level interaction(Table 3). Levels of total and individual CLNA, total non-conjugateddienes, total CLA, total t,t-CLA and total c,t-CLA were greatest whenfeeding flaxseed combined with hay (Pb0.001). All individual non-conjugated dienes were affected in the same manner (Fig. 4) except fort8,c13-18:2, which increased when feeding flaxseed (Pb0.05) but wasunaffected by forage type. Levels of all individual CLA isomers were alsogreatest when feeding flaxseed combined with hay (Fig. 5). RA was thepredominant CLA isomer and reached levels of up to 1.3% of total FAME.

Overall, the forage type by flaxseed interactions seen for diene andtriene biohydrogenation intermediates in LT were also evident inbackfat. Similar to the LT, the levels in backfat indicated that thebiohydrogenation of LNA was likely to occur more through pathwaysincorporating non-CLA dienes as opposed to CLA (Jenkins et al., 2008).

methyl esters (FAME) in total FAME.

P value

Flax SEM Forage Flax Forage⁎flax

2.13 0.07 b0.001 b0.001 0.7201.08b 0.037 0.003 0.151 0.0451.04b 0.036 0.007 0.727 0.0430.03 0.004 0.093 b0.001 0.9090.81 0.036 0.004 b0.001 0.2390.73 0.034 0.004 b0.001 0.1821.37c 0.187 b0.001 b0.001 0.0010.05b 0.006 b0.001 b0.001 b0.0010.02b 0.004 b0.001 b0.001 b0.0010.03b 0.003 b0.001 b0.001 b0.0011.96b 0.108 b0.001 b0.001 b0.0010.77b 0.068 b0.001 b0.001 b0.0010.08b 0.009 b0.001 b0.001 b0.0010.69b 0.066 b0.001 b0.001 b0.001

55.04 0.678 0.029 0.935 0.63951.41 0.758 b0.001 b0.001 0.1971.41 0.138 0.220 0.123 0.6010.12 0.007 0.013 0.020 0.0555.11 0.442 0.247 0.003 0.342

40.04 0.59 b0.001 0.016 0.4501.51 0.089 0.007 b0.001 0.6980.21b 0.016 b0.001 b0.001 b0.0010.50 0.034 0.117 0.003 0.5960.07 0.005 0.014 b0.001 0.0580.77 0.078 0.474 b0.001 0.5033.64b 0.248 b0.001 b0.001 b0.001

39.96b 0.757 0.509 b0.001 0.0303.43 0.146 0.122 0.431 0.1710.43 0.022 0.059 0.220 0.054

24.02 0.535 0.855 b0.001 0.0850.72 0.032 0.089 0.037 0.2999.25 0.556 0.050 0.076 0.913

tandard error of themean; n−6/n−3=n−6/n−3 ratio [(sum of 18:2n−6, 20:3n−6pted: sum of 18:2n-6, 18:3n-6, 20:2n-6, 20:3n-6, 20:4n-6, 22:4n-6, 18:3n-3, 18:4n-3,ed linoleic acid; c-t-CLA: cis–trans conjugated linoleic acid; Total non-CLA dienes: sum of8:2, c9,c15-18:2, and c12,c15-18:2; Total t,t-CLA: sum of t12,t14-18:2, t11,t13-18:2,14-18:2, c12,t14 18:2, t11,c13-18:2, c11,t13-18:2, t10,c12-18:2, t8,c10-18:2, t7,c9-18:2,6:1, c11-16:1, c12-16:1, c13-16:1, c5-17:1, c7-17:1, c9-17:1, c10-17:1, c9-18:1, c11-18:1,of t6-t8-18:1, t9-18:1, t10-18:1, t11-18:1, t12-18:1, t13-t14-18:1, t15-18:1 and t16-18:1.

b

c

cc

b

ca

a

a

a

aa

b

c

c c

b

cb

b

b

b

b

b

g10

0g-1

FA

ME

Hay

Hay-Flax

Silage

Silage-Flax

0

0.2

0.4

0.6

0.8

1

1.2

1.4

1.6

Fig. 4. Effect of forage type and flaxseed feeding in atypical dienes of cull cows backfat.Means within the same fatty acid with different letters are significantly different(p≤0.05).β:Significant flaxseed effect (p≤0.05).

475R.T. Nassu et al. / Meat Science 89 (2011) 469–477

3.3.3. Monounsaturated fatty acids (MUFA)Similar to LT, backfat total cisMUFAwere decreased by feeding either

hay (Pb0.001) or flaxseed (Pb0.001) and a forage type by flaxseed levelinteraction was not found (Table 3). The predominant cis-MUFA inbackfatwas c9-18:1 and itwasalso reducedwhen feedinghay compared

c

b

b c

a

a

a ac

b

b cb

b

b b

t7,c9-CLA c9,t11-CLA t11,c13-CLA t12,c14-CLA

g10

0g-1

FA

ME

Hay

Hay-Flax

Silage

Silage-Flax

0

0.2

0.4

0.6

0.8

1

1.2

1.4

1.6

Fig. 5. Effect of forage type and flaxseed feeding in conjugated linoleic acids (CLA) of cullcows backfat.Means within the same fatty acid with different letters are significantlydifferent (p≤0.05).

to silage (Pb0001) or when flaxseed was included in the diets(P=0.016). As in the LT, changes in c9-18:1 may be related to (1)dietary levels of c9-18:1, (2) more complete biohydrogenation of PUFAto 18:0 and its availability for 18:1 synthesis, and (3) inhibition of Δ-9desaturase by LNA and/or its biohydrogenation products when feedingflax. In backfat, feeding flaxseed also reduced levels of c9-16:1(Pb0.003),whichwould be consistentwith LNA or its biohydrogenationproducts inhibiting Δ-9 desaturase. A lower level of c7-16:1 was alsofound when feeding flaxseed (Pb0.020), but feeding hay increased c7-16:1. The level of c7-16:1 ranged from 0.12 to 0.15 but reasons forchanges in its composition are not immediately apparent. In contrast tothe LT, the level of c11-18:1 was decreased in backfat when feedingflaxseed as well as when feeding hay compared to silage. Raes et al.(2004) did not find any effect of flaxseed on c11-18:1 levels in backfat.Feeding flaxseed in general increased levels of individual cis-MUFAisomers in backfat that have double bonds higher than Δ11-18:1. Theonly exception was c13-18:1 where levels decreased when feedingflaxseed (P=0.003) and this was consistent with changes found in LT.Also consistent with LT, a forage type by flaxseed level interaction wasfound for c12-18:1 with the highest levels foundwhen feeding flaxseedcombinedwith hay. This supports the theory that feedinghay can inhibitthe last step in biohydrogenation of LNA leading to an accumulation ofbiohydrogenation intermediates. Interestingly, the level of c15-18:1increased in backfat and decreased in LT when feeding flaxseed;however, this between tissue difference is not readily explained.

As in the LT, a forage type by flaxseed level interaction was foundfor most trans-MUFA (t6, t7, t8-, t9-, t11-, t13,t14-18:1) in backfat,with the highest levels found when feeding flaxseed combined withhay (Pb0.01; Fig. 6). The level of t10-18:1 was increased by feedinghay (P=0.018) and by feeding flaxseed (Pb0.001), but unlike the LT,a forage type by flaxseed level interaction was not found (P=0.124).Similar to the LT, feeding flaxseed increased the levels of t15- andt16-18:1 and no forage type effect was observed for these twoisomers. This would again suggest that the accumulation of t15- andt16-18:1 could be due to their slower rate of biohydrogenationrelative to the other trans-18:1 isomers, and their biohydrogenationwould be less likely affected by forage influences.

The diet effects on trans MUFA in backfat were similar to thoseobserved in LT, andbackfat fromanimals fedflaxseed combinedwith haywould be a good source of VA. The question is to what extent might thebeneficial health effects of VA be compromised or enhanced byaccompanying increased levels of other biohydrogenation intermediates.

cc

b

cca a

α,β

a

a

a

cc

c

ccb b

b

b

bβ

β

g10

0g-1

FA

ME

HayHay-FlaxSilageSilage-Flax

0

0.5

1

1.5

2

2.5

3

Fig. 6. Effect of forage type and flaxseed feeding in monounsaturated fatty acids of cullcows backfat.Means within the same fatty acid with different letters are significantlydifferent (p≤0.05). α:Significant forage effect (p≤0.05). β:Significant flaxseed effect(p≤0.05).

476 R.T. Nassu et al. / Meat Science 89 (2011) 469–477

3.3.4. Saturated fatty acids (SFA)Unlike the LT, total SFA in backfat was affected by an interaction

between forage type and flaxseed level (P=0.030; Table 3). Thelowest values of total SFA were observed when feeding hay combinedwith flaxseed and this was mainly due to a tendency for a 16:0interaction (P=0.085). This may be related to SFA dilution due togreater accumulations of LNA and LNA biohydrogenation intermedi-ates in backfat, or their effects (i.e. inhibition) on de novo fatty acidsynthesis (Chilliard & Ferlay, 2004). Backfat also had higher levels of18:0 when feeding barley silage as opposed to grass hay (P=0.050),again indicating that feeding silage might result in rumen conditionsconducive to more complete PUFA biohydrogenation.

3.4. n-3 source claims

The n-3 fatty acids were also calculated as mg per 114 g serving(i.e. 4 oz) to evaluate if the LT from the animals in the present studyachieved the level necessary to make an n-3 fatty acid sourceclaim (the required level in Canada is ≥300 mg of n-3 perserving; CFIA, 2003). Feeding flaxseed significantly increased(Pb0.001) the n-3 fatty acid content in both meat and backfat, butlean meat did not achieve the required level (only 116–120 mg n-3fatty acidsmg/serving). On the other hand, backfat reachedmore thanthe minimum level when hay (380 mg/114 g) but not silage(237 mg/114 g) was fed, and more than two times the minimumlevel when flaxseed was included in the hay and silage based diets(851 and 789 mg/114 g, respectively). When combining the meatwith 30% backfat in regular ground beef, the required level for anenrichment claim was achieved when feeding flaxseed combinedwith silage and hay (318 and 339 n-3 mg per serving respectively).The current labeling requirement for a source claim does not,however, distinguish between LNA and its LC n-3 PUFA metabolites,even though the latter have been shown to be more biologicallyactive, and relate more directly to improved health (Bailey, 2009).Therefore, the type of n-3 fatty acids in a product has nutritionalimplications if the extent of conversion of LNA to long-chain n-3 PUFAis limited, although a recent study suggests the ability to synthesizeDHA in humans may have been previously underestimated (Welch,Shakya-Shrestha, Lentjes, Wareham, & Khaw, 2010).

4. Conclusion

Feeding flaxseed resulted in an increase of n-3 fatty acids and thiswas mainly LNA in backfat and LT. Feeding flaxseed also increasedtotal LC n-3 PUFA in LT. A forage type by flaxseed level interactionindicated a preferential accumulation of LNA biohydrogenationintermediates (trans-MUFA, non-conjugated dienes, CLA and CLNA)in LT and backfat when feeding flaxseed combined with grass hay ascompared to barley silage. Omega-3 fatty acids in the LT did notachieve the level required in Canada for a source claim, but the levelwould have been reached for regular ground beef (70% lean, 30% fat)when feeding flaxseed combined with either hay or silage. Feedingflaxseed in a 50:50 forage to concentrate diet would thus beconsidered as positive due to increased levels of fatty acids beneficialto human health in beef (i.e. n-3, rumenic and vaccenic acids), butpotential effects of elevated levels of other biohydrogenation in-termediates require further investigation.

Acknowledgments

Dr. N. Aldai acknowledges the receipt of a research contract fromthe 7th European Community Program (Marie Curie InternationalOutgoing Fellowship).

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