Fatty Acid Composition of Lamb Liver, Muscle, And Adipose Tissues in

Article pubs.acs.org/JAFC

Fatty Acid Composition of Lamb Liver, Muscle, And Adipose Tissues in Response to Rumen-Protected Conjugated Linoleic Acid (CLA) Supplementation Is Tissue Dependent Stefano Schiavon,† Matteo Bergamaschi,*,† Erika Pellattiero,‡ Alberto Simonetto,† and Franco Tagliapietra† †

Department of Agronomy, Food, Natural Resources, Animals and Environment, University of Padua, Viale Università 16, 35020 Legnaro, PD, Italy ‡ Department of Animal Medicine, Production and Health, University of Padua, Viale Università 16, 35020 Legnaro, PD, Italy ABSTRACT: The tissue-specific response to rumen-protected conjugated linoleic acid supply (rpCLA) of liver, two muscles, and three adipose tissues of heavy lambs was studied. Twenty-four lambs, 8 months old, divided into 4 groups of 6, were fed at libitum on a ration supplemented without or with a mixture of rpCLA. Silica and hydrogenated soybean oil was the rpCLA coating matrix. The lambs were slaughtered at 11 months of age. Tissues were collected and analyzed for their FA profiles. The dietary rpCLA supplement had no influence on carcass fatness nor on the fat content of the liver and tissues and had little influence on the FA profiles of these tissues. In the adipose tissues, rpCLA increased the proportions of saturated FAs, 18:0 and 18:2t10c12, and decreased the proportions of monounsaturated FAs in the adipose tissues. In muscles, the effects were the opposite. The results suggest that Δ9 desaturase activity is inhibited by the rpCLA mixture in adipose tissues to a greater extent than in the other tissues. KEYWORDS: adipose tissues, conjugated linoleic acid, lamb, liver, muscles

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INTRODUCTION The quality of meat is largely influenced by its fat content and fatty acid (FA) profile.1 Meat fat content and fatty acid profile are also of importance for consumer health.2−4 Ruminant products, rich in saturated FAs, are not entirely acceptable to consumers.1 Consequently, strategies to manipulate the fat content and the FA composition of ruminant meat and milk have been studied.5,6 Ruminant products are rich in conjugated linoleic acids (CLAs), which have been found to be involved in many biological activities. For example, they have anticarcinogenic, antiobesity, antioxidant, and anti-inflammatory properties.7 Lipid, energy, and nitrogen metabolism in monogastric and ruminants were found to be influenced by small amounts of CLA.7,8 In cattle, to prevent rumen hydrogenation, CLA is supplied in rumen-protected forms. Various are the forms of protection proposed,9 but few comparisons were completed.10,11 Administration of a few grams per day of rumen-protected CLA isomers, in particular 18:2c9,t11 and 18:2t10,c12, to lactating cows and sheep has been found to notably reduce the milk fat content, with the short-chain saturated FA content more affected than the longer chain polyunsaturated FAs.12−14 CLA mixtures have also been found to reduce body fat in growing animals of many species, resulting from a reduction in body fat deposition rather than mobilization of body fat already deposited.15 However, the results are controversial. Schiavon et al.16,17 found that long-term rumen-protected CLA supply notably increased the CLA contents in the meat and fat tissues of growing double-muscled Piemontese young bulls. However, this treatment had small or negligible effects on growth © 2017 American Chemical Society

performance, carcass fatness, and the lipid content of these tissues. Lambs are an interesting model for studying lipid metabolism in growing ruminants. The lamb lipid depots and their FA profiles can be altered within a short span of time after weaning. Thus, the confounding effects arising from previous lipid depots can be avoided.18 In fact, milk and solid feeds differ greatly in the amount and composition of the lipid compounds contained in them, and this leads to progressive differentiation of the body fat content and composition during the early stages of growth.19 Feeding systems based on grass or, as in the case of indoor diets, on concentrates have been found to be responsible for large differences in the FA contents of lamb tissues, particularly n-3 and n-6 polyunsaturated FAs.20 Tissues and organs are known to have varying FA compositions,21 reflecting their differing physiological roles and metabolisms,22 and they should, therefore, respond differently to rumenprotected CLA supply. A tissue-dependent response to rumenprotected CLA would be evidenced by different changes in the FA profiles of the different tissues. In the present experiment, the tissue-specific response to rumen-protected CLA supplementation was studied by analyzing the FA profiles of the liver, various muscles, and the subcutaneous and internal fat of lambs fed on indoor diets. Received: Revised: Accepted: Published: 10604

October 3, 2017 November 6, 2017 November 15, 2017 November 15, 2017 DOI: 10.1021/acs.jafc.7b04597 J. Agric. Food Chem. 2017, 65, 10604−10614

Article

Journal of Agricultural and Food Chemistry Table 1. Groups of Fatty Acids and FA Indices group of FA/index SFA MUFA PUFA FA with FA with FA with FA with FA with

14 15 16 17 18

carbons carbons carbons carbons carbons

FA with 19 carbons FA with 20 carbons FA with 22 carbons ∑15:1others ∑16:1others ∑17:1others ∑18:1others ∑19:1others ∑16:2others ∑18:2others ∑20:2others ∑20:3others even chain SFA odd chain SFA branched chain SFA CLA ∑CLAothers n-6 n-3 n-6/n-3 Desaturasesa: C14 C16 C18 CLAc9t11 a

individual FA 8:0; 10:0; 11:0; 12:0; 13:0; 14:0; 15:0; 16:0; 17:0; 18:0; 19:0; 20:0; 22:0; 23:0 14:1c9; ∑15:1others; 16:1c7; 16:1t7; 16:1c9; ∑16:1others; ∑17:1others; 18:1c7; 18:1c9; 18:1t11; ∑18:1others; 19:1; 19:1others; 20:1c9; 20:1c13 ∑16:2others; ∑18:2others, 18:5c3,c6,c9,c12,c15; ∑20:2others, ∑20:3others, ∑CLA, ∑Ω 3, ∑Ω 6 14:0; 14:0iso; 14:1c9 15:0; 15:0anteiso; 15:0iso; 15:1; 15:1t; 15:1t5; three 15:1 unknown isomer 16:0; 16:0iso; 16:1; 16:1c7; 16:1t7; 16:1c9; 16:1c11; three 16:1 cis unknown isomer; three 16:1 trans unknown isomer; 16:3c7,c10,c13 17:0; 17:0anteiso; 17:0iso; 17:1; 17:1t7; 17:1c11; four 17:1 unknown isomer 18:0; 18:1c7; 18:1c9; 18:1t11; 18:1c3; 18:1c12; 18:1t13; 18:1t14; three 18:1 unknown isomers; 18:2c9,t11; 18:2t10,c12; 18:2c9,c12; 18:3c6,c9,c12; 18:3c9,c12,c15; 18:4c6,c9,c12,c15 19:0; 19:0anteiso; 19:0iso; 19:1; 19:1t12; two 19:1 unknown isomer 20:0; 20:2c11,c14; 20:3c8,c11,c14; 20:4c5,c8,c11,c14; six 20:2 unknown isomer; two 20:3 unknown isomer 22:0; 22:4c7,c10,c13,c16; 22:5c4,c7,c10,c13,c16 15:1; 15:1t; 15:1t5; three 15:1 unknown isomer 16:1; 16:1c11; three 16:1 cis unknown isomer; three 16:1 trans unknown isomer 17:1; 17:1t7; 17:1c11; four 17:1 unknown isomer 18:1c3; 18:1c12; 18:1t13; 18:1t14; three 18:1unknown isomer 19:1t12; two 19:1 unknown isomer four 16:2 unknown isomer 18:2t11,c15; 18:2c11,c14; five 18:2 unknown isomer six 20:2 unknown isomer two 20:3 unknown isomer 8:0; 10:0; 12:0; 14:0; 16:0; 18:0; 20:0; 22:0 11:0; 13:0; 15:0; 17:0; 19:0; 23:0 14:0iso; 15:0anteiso; 15:0iso; 16:0iso; 17:0anteiso; 17:0iso; 18:0iso; 19:0anteiso; 19:0iso 18:2c9,t11; 18:2t10,c12; ∑ other cis form, ∑ other trans form ∑ other cis form, ∑ other trans form 18:2c9,c12; 18:3c6,c9,c12; 20:2c11,c14; 20:3c8,c11,c14; 20:4c5,c8,c11,c14; 22:4c7,c10,c13,c16; 22:5c4,c7,c10,c13,c16 16:3c7,c10,c13; 18:3c9,c12,c15; 18:4c6,c9,c12,c15; 20:2c14,c17; 20:5c5,c8,c11,c14,c17; 22:5c7,c10,c13,c16,c19; 22:6c4,c7,c10,c13,c16,c19 n-6 FA/n-3 FA 14:1c9/(14:1c9 + 14:0) 16:1c9/(16:1c9 + 16:0) 18:1c9/(18:1c9 + 18:0) 18:2c9,t11/(18:2c9,t11 + 18:1t11)

Δ9-desaturase indices: calculated according to the method of ref 37.

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were aged about 11 months and had slaughter weights of 61.1 ± 8.6 kg (ram lambs) and 57.4 ± 8.6 kg (ewe lambs). Tissue Sampling. The liver and the perivisceral and perinephric fats were taken from all the lambs immediately after slaughter. Each carcass was divided into two sides and cold stored at 4 °C. Twentyfour hours after slaughter, the right half of each carcass was divided into five cuts (hind leg, foreleg and shoulder, ribs-loin, withers, brisket) and each cut was weighed. The ribs-loins were vacuum-packed and transported to the laboratory where they were stored at 4 °C in a chilling room for 6 days, after which they were divided into ribs and loin. The longissimus thoracis muscle, other muscles, and the subcutaneous fat of the rib cut were separated from the other components and weighed as described by Schiavon et al.26 Sample Preparation. The samples were prepared as reported in detail by Schiavon et al.27 Briefly, all the tissue samples were ground, mixed, and homogenized for 10 s at 4500g (Grindomix GM200, Retsch, Haan, Düsseldorf, Germany) then stored at −20 °C. The samples were then freeze-dried using a CoolSafe 80−90 freeze-dryer (Scanvac, Stockholm, Sweden). Each freeze-dried sample (liver, 2.21 ± 0.17 g; longissimus thoracis, 2.17 ± 0.10 g; other muscles, 2.20 ± 0.14 g; subcutaneous fat, 0.085 ± 0.014 g; perivisceral fat, 0.086 ± 0.021 g; perinephric fat, 0.088 ± 0.013 g) was subjected to a mild acid−base transesterification/methylation process as described by Jenkins.28 Two mL of sodium methoxide (0.5 M in methanol) and 2 mL of toluene containing 2 mg/mL of methyl 12-tridecenoate as internal standard (# U-35 M, Nu-Chek Prep, Inc., Elysian, MN, USA) were added to the dried tissue samples in a culture tube. The samples were incubated in a

MATERIAL AND METHODS

Animals were treated in accordance with the Guidelines for the Care and Use of Agricultural Animals in Agricultural Research and Teaching.23 This trial was part of a larger project aimed at studying the effects of CLA supplementation on the growth performance and carcass quality traits of sheep in the Veneto region of northeastern Italy24,25 and the FA profiles of lean and fat tissues. Experimental Design. Twenty-four heavy lambs (12 males and 12 females) of local breeds were assigned to 4 groups of 6 and reared on the University of Padua’s Lucio Toniolo experimental farm (Legnaro, Padua, Italy). Two groups (one of males, the other of females) were fed at libitum an identical total mixed ration (TMR) supplemented with 20 g/d of a commercial rpCLA product (SILA, Noale, Venice, Italy). The rpCLA coating material was based on a silica and hydrogenated soybean oil matrix.17 The other two groups (one of males, one of females) were fed the same diet but without the rpCLA supplement. The CLA dose provided 1.58, 1.54, 7.76, 1.60, and 1.60 g/d of 18:2c9,t11, 18:2t10,c12, 18:0, 16:0 and 18:0cis, respectively.17 The TMR contained corn grain (400 g/kg DM), corn silage (256 g/ kg DM), soybean meal (33 g/kg DM), dried sugar beet pulp (113 g/ kg DM), wheat bran (70 g/kg DM), wheat straw (66 g/kg DM), grape seed meal (20 g/kg DM), and a vitamin and mineral mix (35 g/kg DM). At the beginning of the experiment, the animals were aged 8 months, the ram lambs had a body weight of 37.4 ± 6.3 kg, the ewe lambs 37.8 ± 6.7 kg. At the end of the fattening period, the animals 10605

DOI: 10.1021/acs.jafc.7b04597 J. Agric. Food Chem. 2017, 65, 10604−10614

Article

Journal of Agricultural and Food Chemistry

Table 2. Effect of Gender of Heavy Lambs and rpCLA on Some Slaughter Traits and on the Significant Fatty Acids (FA) Expressed as g/100 g of Total FA gender (LSM)a item slaughter traits: BW, kg BCSe carcass yield, % muscles, % of carcass fat, % of carcass liver, kg kidney, kg perinephric fat, kg tissues fat content: liver, % Longissimus thoracis, % others rib eye muscles, % FA, g/100 g of total FA: ∑MUFA ∑PUFA ∑branched-chain SFA 10:0 15:0anteiso 16:0iso ∑16:2others 17:0anteiso 17:0iso 18:0 18:1t11 18:1c9 ∑18:1others 18:2c9c12 19:1others 20:2c11c14 20:3c8c11c14 22:5c7c10c13c16c19 ∑CLA 18:2c9t11 18:2t10c12 ∑CLAothers desaturase C18

diet (LSM)d b

P-value

male

female

SEM

65.5 3.75 48.1 46.5 28.1 1.54 0.11 0.77

57.0 3.81 50.2 41.7 36.1 1.50 0.10 1.30

2.531 0.076 0.464 1.916 1.488 0.054 0.001 0.070

*

3.13 4.25 5.48

5.26 5.00 7.20

0.505 0.253 0.731

* **

32.36 9.492 2.809 0.899 0.341 0.251 0.033 1.113 0.535 23.16 4.110 23.89 0.899 4.068 0.099 0.058 0.184 0.216 0.617 0.476 0.072 0.090 0.505

33.09 8.296 2.522 0.768 0.280 0.225 0.028 0.982 0.496 22.48 3.444 22.13 0.768 3.183 0.105 0.030 0.142 0.193 0.576 0.428 0.067 0.100 0.526

0.384 0.259 0.053 0.031 0.014 0.008 0.005 0.025 0.011 0.578 0.176 0.405 0.029 0.142 0.006 0.004 0.012 0.014 0.031 0.028 0.009 0.010 0.008

** ** ***

***

** ** * * ** ** * * * *** *** *** ** *

*

c

no CLA

CLA

SEMb

61.1 3.88 49.0 44.7 32.3 1.50 0.11 1.09

61.4 3.73 48.6 43.4 31.9 1.53 0.11 0.99

2.420 0.073 0.443 1.841 1.420 0.052 0.003 0.080

3.94 4.74 7.06

4.43 4.53 5.63

0.489 0.245 0.740

33.04 8.900 2.710 0.805 0.319 0.235 0.031 1.088 0.517 21.79 3.843 24.62 0.804 3.630 0.093 0.045 0.162 0.192 0.540 0.437 0.044 0.080 0.529

32.21 8.888 2.620 0.862 0.300 0.242 0.028 1.007 0.514 23.85 3.712 24.41 0.853 3.621 0.111 0.050 0.170 0.181 0.652 0.467 0.094 0.107 0.502

0.376 0.249 0.051 0.295 0.015 0.007 0.004 0.023 0.009 0.556 0.165 0.417 0.029 0.134 0.006 0.004 0.011 0.013 0.031 0.028 0.009 0.009 0.007

P-valuec

*

* **

*

** *** *** **

Data are least-squares means (LSM) for gender, male n = 12 and female n = 12. bStandard error of the mean. c***P < 0.001; **P < 0.01; *P < 0.05. d Data are least-squares means (LSM) for diets, NO CLA n = 12, CLA n = 12. eScore according to a scale from 1 (emaciated) to 5 (obese). a

water bath at 50 °C for 10 min and then removed from the bath and cooled for 5 min. After adding 3 mL of freshly prepared methanolic HCl (1.37 M), the samples were incubated again in a water bath at 80 °C for 10 min and then removed from the bath and cooled for 7 min. Next, 5 mL of K2CO3 (0.43 M) and 2 mL of toluene were added to each tube, and these were then vortexed for 30 s and centrifuged for 5 min at 400g and 4 °C. The organic phase (upper layer) of the tube was transferred to a screw-capped tube, to which was added 0.5 g of anhydrous sodium sulfate and 0.5 g of active charcoal (Sigma-Aldrich, MO). The solution was vortexed for 5 min and rested for 1 h. After centrifugation for 5 min (400g at 4 °C), the clear upper layer containing the FAME was transferred to a gas chromatography (GC) vial and stored at −20 °C until GC analysis. Fatty Acid Analysis. Detailed FA profiles were determined using a GC × GC instrument (Agilent 7890A, Agilent Technologies, Santa Clara, CA) with two columns in series and equipped with a modulator (G3486A CFT, Agilent Technologies), an automatic sampler (7693, Agilent Technologies), and a flame ionization detector connected to the chromatography data system software (Agilent Chem Station, Agilent Technologies) at DAFNAE, University of Padua (Legnaro,

Padua, Italy). The operating conditions of the GC apparatus were as follows: first column, 75 m × 180 μm (internal diameter) × 0.14 μm film thickness (Supelco, Bellefonte, PA), H2 carrier at a flow rate of 0.22 mL/min; second column, 3.5 m × 250 μm (internal diameter) × 0.14 μm film thickness (Agilent, Agilent Technologies), and H2 carrier at a flow rate of 22 mL/min. Oven temperature program: 50 °C (held for 2 min), increased to 150 °C at 50 °C/min (held for 15 min) and then increased to 240 °C at 2 °C/min (held for 20 min). Valves were set for a modulation delay of 1 min, a modulation period of 2.9 s, and a sample time of 2.77 s. Flame-ionization detector: heater, 250 °C; H2 carrier flow, 20 mL/min; air flow, 450 mL/min. The splitless inlet temperature was 270 °C, pressure 20.80 MPa, septum purge 3 mL/ min, and split flow 35.2 mL/min. The resulting two-dimensional chromatograms were analyzed with the comprehensive GC × GC software (GC Image Software, Zoex Corp., Houston, TX) in order to calculate the cone volume of each FA. Fatty Acid Identification and Quantification. Fatty acids were identified as reported in detail by Schiavon et al.27 Briefly, the cone positions in the chromatogram were compared with the cone positions of the FAs in the pure reference standards (#674 and #463; Nu-Chek 10606


Article


Table 3. Contents of Main Categories of Fatty Acids in Liver, Muscles, and Fat Tissues of Lambs Fed Indoor Diets (g/100 g of total FA) tissuea muscles (M)

a

orthogonal contrasts (P-value)c fats (F)

L

FA

liver (L)

Longissimus (LT)

fat, % SFA MUFA PUFA FA with 14 carbons FA with 15 carbons FA with 16 carbons FA with 17 carbons FA with 18 carbons FA with 19 carbons FA with 20 carbons FA with 22 carbons short chain FA (16 C) even chain SFA odd chain SFA branched chain SFA n-6 n-3 n-6/n-3 CLA others unknown Desaturases4: C14 C16 C18 CLAc9t11

4.03 48.94 25.85 23.82 1.32 1.30 16.72 4.57 56.97 0.96 11.40 4.17 3.31 16.72 78.10 42.35 3.79 2.79 17.44 3.70 4.71 0.80 1.89

4.48 48.25 38.41 10.13 2.31 0.61 23.41 2.86 61.08 0.43 2.26 0.17 5.82 23.41 66.82 42.28 4.15 1.82 6.07 1.84 3.30 0.99 1.24

5.84 51.30 37.75 8.54 2.63 0.71 25.01 2.99 60.92 0.42 1.90 0.18 5.23 25.01 66.45 45.85 3.56 1.89 5.31 1.24 4.28 0.78 1.21

59.90 34.52 3.58 4.02 1.80 29.44 5.98 55.14 0.27 0.17 0.01 6.80 29.44 61.57 51.92 4.64 3.33 2.27 0.29 7.83 0.36 0.67

64.57 31.02 3.94 4.10 1.56 29.10 5.02 58.22 0.25 0.19 0.01 6.53 29.10 63.71 57.55 3.85 3.17 2.52 0.32 7.88 0.39 0.71

65.89 29.98 3.73 3.80 1.45 28.39 4.85 59.65 0.24 0.16 0.01 6.16 28.39 64.92 59.00 3.83 3.06 2.45 0.25 9.80 0.37 0.66

0.552 0.547 0.344 0.225 0.058 0.568 0.147 0.613 0.021 0.152 0.090 0.245 0.568 0.662 0.342 0.219 0.060 0.157 0.043 0.181 0.034 0.048

0.03 0.07 0.40 0.15

0.03 0.05 0.63 0.21

0.04 0.05 0.60 0.15

0.02 0.04 0.56 0.07

0.01 0.03 0.46 0.06

0.01 0.03 0.44 0.06

0.013 0.012 0.040 0.029

others (OM)

subcutaneous (SF)

perivisceral (PV)

perinephric (PN)

SEMb

vs M+F

M vs F

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LT

SF

PV

vs OM

vs PV + PN

vs PN

* ***

*** ***

*** *

*

* ***

** *** ***

** ** ** **

*

* *** *** *** ***

**

*** ** ***

* ** ***

*** ** ***

** *** ***

Data are least-squares means (LSM) for tissue, n = 24. bStandard error of the mean. c***P < 0.001; **P < 0.01; *P < 0.05. fixed effect of the ikth CLA × sex interaction; birthl is the fixed effect of the lst single or twin birth (l = 1 to 2); agem is the linear covariate of the lamb’s age; and eijklmn is the random effect of animal. The experimental unit was the animal. To evidence the differential response of the tissues to rpCLA supplementation data obtained for each individual FA and FA category, expressed as g/100 g total FAs to avoid heteroskedasticity among tissues, was performed with the PROC MIXED SAS procedure according to a model:

Prep, Inc., Elysian, MN), 47080-U Bacterial Acid Methyl Esters (BAMEs; Sigma-Aldrich, St. Louis, MO), 47085-U Ω3 Menhaden Oil (Supelco, St. Louis, MO), and 5 CLAs: 18:2c9,t11 (#UC-60M; NuChek Prep, Inc., Elysian, MN, USA), 18:2t10,c12 (#UC-61M; NuChek Prep, Inc., Elysian, MN), 18:2c9,c11 (#1256; Matreya LLC., Pleasant Gap, PA), 18:2t9,t11 (#1257; Matreya LLC, Pleasant Gap, PA), and 18:2c11,t13 (#1259; Matreya LLC, Pleasant Gap, PA). In addition, other FAs were identified by elution order and their position in the two-dimensional chromatography grid using GC Image Software (Zoex Corp.) in accordance with Vlaeminck et al.29 Some FAs aligned in specific areas of the chromatogram were classified according to their main characteristics, length of carbon chain, and degree of unsaturation. Fatty acid proportions were calculated by dividing the cone volume of each FA by the total volume of FAs and were expressed as g/100 g total FAs. In accordance with other experiments by our team,30 we assumed a limit of detection for all FAs of 0.015 mg (0.0015 g/g total FAs). All values below this limit were taken to be undetectable and considered missing for the statistical analyses. The FAs were also summed in groups according to the criteria set out in Table 1. Statistical Analysis. The statistical analyses were performed with the SAS software package (SAS Institute Inc., Cary, NC) using two different models. Meat characteristics were analyzed with PROC GLM according to the following model:

yijklmno = μ + CLA i + sex k + CLA × sex ik + birthl + age m + lamb(CLA × sex)n:ijk + tissueo + CLA × tissue io + eijklmno where yijklmno is the observed trait; μ is the overall intercept of the model; CLAi is the fixed effect of the ith dietary treatment (i = 1 to 2); sexk is the fixed effect of the kth sex (k = 1 to 2); CLA × sexik is the fixed effect of the ikth diet × sex interaction; birthl is the fixed effect of the lth birth (l = 1 to 2); agem is the linear covariate of the lamb’s age; lamb(CLA × sex)n:ijk is the random effect of the nth animal (l = 1 to 24) within diet and sex; tissueo is the fixed effect of the oth tissue (m = 1 to 6); CLA × tissueio is the fixed effect of the ioth CLA × tissue interaction; and eijklmno is the residual error. The random effect of animal was used to test CLA, sex, the CLA × sex interaction, and birth, whereas the effects of tissue and the CLA × tissue interaction were tested on the residual. Orthogonal contrasts were fitted to test liver versus other tissues, fatty tissues versus muscles, longissumus thoracis versus other muscles, subcutaneous fat versus internal (perivisceral plus perinephric) fat, and perivisceral fat versus perinephric fat.

yijklmn = μ + CLA i + sex k + CLA × sex ik + birthl + age m + eijklmn

where yijklmn is the observed quality trait; μ is the overall intercept of the model; CLAi is the fixed effect of the ith CLA treatment (i = 1 to 2); sexk is the fixed effect of the kth sex (k = 1 to 2); CLA × sexik is the 10607


Article


Table 4. Short and Medium Chain Fatty Acid Contents in Liver, Muscles, and Fat Tissues of Lambs Fed Indoor Diets (g/100g of total FA) tissuea

orthogonal contrasts (P-value)c

muscles (M)

a

FA

liver (L)

Longissimus (LT)

8:0 10:0 11:0 12:0 13:0 14:0 14:0iso 14:1c11 15:0 15:0anteiso 15:0iso ∑15:1others 16:0 16:0iso 16:1c7 16:1t7 16:1c9 ∑16:1others ∑16:2others 16:3c7c10c13 (n-3) 17:0 17:0anteiso 17:0iso ∑17:1others

0.01 0.01 n.d. 0.04 0.62 1.20 0.05 0.04 0.69 0.35 0.20 0.07 14.99 0.21 0.59 0.13 1.07 0.37 n.d. 0.02 2.17 1.01 0.48 0.92

0.07 0.20 n.d. 0.16 2.47 2.22 0.03 0.08 0.39 0.14 0.09 0.02 21.38 0.14 0.31 0.07 1.18 0.36 n.d. 0.04 1.16 0.67 0.41 0.63

fats (F)

others (OM)

subcutaneous (SF)

perivisceral (PV)

perinephric (PN)

0.05 0.18 0.09 0.18 1.56 2.52 0.04 0.10 0.45 0.16 0.10 0.03 22.68 0.16 0.32 0.07 1.31 0.48 n.d. 0.02 1.27 0.70 0.42 0.62

0.02 0.21 0.09 0.18 0.48 3.83 0.07 0.09 1.10 0.39 0.24 0.07 27.29 0.33 0.37 0.05 1.19 0.22 0.03 0.01 2.88 1.39 0.60 1.11

0.02 0.21 0.02 0.16 0.45 3.93 0.08 0.06 0.82 0.41 0.26 0.08 27.18 0.31 0.42 0.06 0.80 0.33 0.04 0.01 2.45 1.28 0.60 0.69

0.02 0.19 0.02 0.14 0.54 3.65 0.07 0.05 0.74 0.40 0.25 0.07 26.56 0.30 0.39 0.05 0.88 0.21 0.04 0.01 2.42 1.24 0.58 0.62

L

M

LT

SF

PV

SEMb

vs M + F

vs F

vs OM

vs PV + PN

vs PN

0.004 0.009 0.017 0.012 0.145 0.134 0.003 0.010 0.039 0.015 0.008 0.005 0.416 0.009 0.076 0.010 0.019 0.085 0.002 0.003 0.075 0.027 0.013 0.049

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***

** *** *** ***

Data are least-squares means (LSM) for tissue, n = 24; n.d. = not detectable. Standard error of the mean. ***P < 0.001; **P < 0.01; *P < 0.05. b

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RESULTS AND DISCUSSION Effects of Gender and CLA on Carcass Fat and Tissue Fat Content. The lambs used in the present experiment were slaughtered at about 60 kg body weight (Table 2). Females differed from males in a number of traits, having greater carcass yields, lower percentages of muscle, and greater percentages of carcass fat content and perinephric fat. The fat contents of the liver and the longissimus thoracis muscle of females were greater than males. However, despite a difference of 7 percentage points between males and females in carcass fat content (P < 0.01), there were only small differences between the genders in the average across-tissue FA profiles. Nevertheless, male lambs had higher proportions of medium-chain branched FAs, 18:1t11, 18:1c9, ∑18:1others and some medium- to long-chain PUFAs (from C16 to C22) than female lambs. The small differences between males and females in FA composition, despite different carcass and tissue fat contents, was expected given that few gender differences in intramuscular FA composition, even in carcasses of widely different fat levels, have previously been found.31−33 The major differences between males and females regarded the branched FAs and some partially hydrogenated FAs, such as 18:1t11, which reflect the contribution of the rumen microbial population.34 The results of the present experiment suggest, therefore, that there were some differences between males and females in the patterns of rumen fermentation or the feed particle passage rate. The dietary CLA mixture did not affect the weight or growth of the lambs, nor body condition scores, nor the proportions of fat and lean in the carcass. The rumen-protected CLA

c

supplement did not affect the weights of the liver, kidney, and perinephric fat, nor the fat content of the liver and muscles. It had little or no effect on the large majority of individual and groups of FAs across tissues, with some exceptions, these being the greater proportions of 18:0 (P < 0.01), ∑CLA (P < 0.01), and 18:2t10,c12 (P < 0.01) compared with the control diet. Conversely, the CLA mixture reduced the mean value of the desaturase index of FAs with 18 carbons (P < 0.01). The increase in 18:0 may have been due to its inclusion in the rpCLA mixture, which provided about 7.76 g/d of this FA, together with 1.6 g/d of 16:0 and 1.6 g/d of 18:0cis.17 Conjugated linoleic acid supplementation has been found to reduce body fat or body depots in many monogastrics, rats, mice, pigs, and humans, and the effect is mainly due to the 18:2t10,c12 isomer.35 Some studies have suggested that administering CLA would promote hepatic steatosis,36 whereas others have indicated that CLA would promote FA synthesis and oxidation in the liver.37 Reports of the effects of a dietary CLA mixture on body fatness and various tissue fat contents in ruminants are inconsistent. Zhang et al.38 found that a dietary CLA supplement increased intramuscular fat deposition and decreased subcutaneous fat deposition in Yellow Breed × Simmental cattle crosses. In other cases, no effect of CLA on body fatness and tissue fat contents was found, except for an increase in tissue CLA content.39−41 In the present experiment, the CLA mixture had no influence on fat accumulation in neither the liver nor the muscle of lambs. Fatty Acid Profiles of Tissues. The GC × GC analysis revealed the presence of 87 cones in the chromatograms of the lamb tissues, corresponding to 23 SFAs, 28 MUFAs, and 36 PUFAs. Of the 87 FAs detected, 8 were in concentrations of 10608


a

10609

24.61 0.23 1.11 3.28 16.27 1.32 7.22 0.57 0.03 0.20 1.10 0.48 0.29 0.12 0.08 0.30 0.26 0.02 0.34 0.19 0.92 0.12 0.04 0.21 0.51 0.75 0.28 7.05 0.94 0.31 0.55 1.13 0.60 1.09 0.84 0.02

18:0 18:0iso 18:1t8−10 18:1t11 18:1c9 ∑18:1others 18:2c9c12 (n-6) 18:2c9t11 (CLA) 18:2t10c12 (CLA) ∑ CLAothers ∑18:2others 18:3c6c9c12 (n-6) 18:3c9c12c15 (n-3) 18:4c6c9c12c15 (n-3) 18:5c3c6c9c12c15 (n-3) 19:0 19:0anteiso 19:0iso 19:1 ∑19:1others 20:0 20:1c9 20:1c13 20:2c11c14 (n-6) ∑20:2others 20:3c8c11c14 (n-6) ∑20:3others 20:4c5c8c11c14 (n-6) 20:2c14c17 (n-3) 20:5c5c8c11c14c17 (EPA - n-3)) 22:0 22:4c7c10c13c16 (n-6) 22:5c4c7c10c13c16 (n-6) 22:5c7c10c13c16c19 (DPA - n-3) 22:6c4c7c10c13c16c19 (DHA- n-3) 23:0

17.85 0.15 0.90 2.94 30.97 0.68 4.07 0.76 0.12 0.10 1.20 0.25 0.60 0.30 0.48 0.09 0.16 0.03 0.12 0.11 0.35 n.d. 0.05 0.02 n.d. 0.12 0.04 1.48 0.22 0.08 0.04 0.11 0.02 0.12 n.d. 0.04

Longissimus (LT) 19.86 0.15 0.80 3.26 29.69 0.70 3.68 0.58 0.10 0.10 1.08 0.21 0.40 0.20 0.28 0.12 0.12 0.04 0.10 0.12 0.33 0.05 0.10 0.02 0.06 0.09 0.07 1.21 0.18 0.06 0.04 0.09 0.01 0.10 n.d. 0.06

others (OM) 20.31 0.24 0.46 4.07 26.03 0.68 2.11 0.26 0.04 0.05 0.62 0.06 0.18 0.02 0.02 0.09 0.06 0.01 0.06 0.08 0.07 0.03 n.d 0.02 0.01 0.02 0.01 0.05 0.02 0.01 0.01 0.01 n.d. 0.03 n.d. 0.01

subcutaneous (SF) 25.95 0.18 0.63 4.59 22.34 0.81 2.35 0.28 0.06 0.05 0.65 0.08 0.21 0.03 0.02 0.10 0.04 0.01 0.10 0.04 0.09 0.01 0.02 0.02 0.01 0.01 0.01 0.05 0.02 0.01 0.01 0.01 n.d. 0.02 n.d. 0.01

perivisceral (PV)

fats (F) 28.34 0.19 0.52 4.52 21.77 0.77 2.32 0.26 0.06 0.05 0.60 0.06 0.19 0.02 0.01 0.10 0.04 0.01 0.04 0.07 0.10 0.01 n.d. 0.01 0.01 0.01 0.01 0.04 0.01 n.d. 0.01 0.01 n.d. 0.01 n.d. 0.01

perinephric (PN) 0.578 0.009 0.055 0.149 0.508 0.036 0.168 0.003 0.030 0.016 0.047 0.020 0.029 0.026 0.038 0.008 0.010 0.008 0.007 0.010 0.036 0.006 0.016 0.007 0.013 0.020 0.013 0.116 0.029 0.012 0.027 0.038 0.085 0.033 0.064 0.005

SEMb

*** *** *** ***

*** *** *** *** *** *** *** *** ***

***

***

*** ***

***

***

*** *** ***

*** *** ***

***

*** *** *** ** *** ***

*** *** *** *** *** *** *** *** ** *** *** *** *** *** *** *** *** ** *** *** ***

*** *** ***

M vs F

L

**

*** ** *** ** **

***

*

**

vs OM

LT

PV

***

**

***

** ***

***

vs PN

*

*** *** ** *** *** **

vs PV + PN

SF

orthogonal contrasts (P-value)c vs M + F

Data are least-squares means (LSM) for tissue, n = 24; n.d. = not detectable. bStandard error of the mean. c***P < 0.001; **P < 0.01; *P < 0.05.

liver (L)

FA

muscles (M)

tissuea

Table 5. Long Chain Fatty Acid (FA) Contents in Liver, Muscles, and Fat Tissues of Lambs Fed Indoor Diets (g/100 g of total FA)

Journal of Agricultural and Food Chemistry Article


Article


ties.25,45 The fat content may be much higher in oxidative than in glycolytic muscles,45 and the LT muscle contains a relatively high proportion of white glycolytic fibres.31 In ruminants, n-6 and n-3 long chain PUFAs are found mainly in the tissue phospholipids and not in the triacylglycerol fraction of the muscle and adipose tissues.5 In the present experiment, long chain n-6 PUFAs (C20−C22) were present in substantially greater amounts in muscles than in adipose tissues (P < 0.01), particularly arachidonic acid (20:4c5c8c11c14), which is produced from linoleic acid for desaturase and elongase enzyme activity.1 On the other hand, the muscles had very small proportions of n-3 FA products, which included 20:5c5,c8,c11,c17 (EPA), 22:5c7,c10,c13,c16,c19 (DPA), and 22:6c4,c7,c10,c13,c16,c19 (DHA), compared with other studies on grass-fed lambs.31,46 Adipose tissues are the major sites for the de novo synthesis of FAs, with acetate being the principal substrate for lipogenesis in these tissues.45 The fatty acids in the adipose tissues of lambs consisted of large proportions of SFAs (59.9−65.9 g/100 g total FAs) and MUFAs (30−34.5 g/100 g total FA s) and small proportions of PUFAs (3.6−3.9 g/100 g total FAs). The FA compositions of the 3 adipose tissues differed significantly. The two inner tissues had more SFAs (64.6−65.9 g/100 g total FAs) and 18:0 (26.0−28.4 g/100 g total FAs) but fewer 18:1c9 (21.8−22.3 g/100 g total FA) than the peripheral subcutaneous fat tissue (59.9, 20.3, and 26.0 g/100 g total FAs, respectively), and lower C-18 Δ9 desaturase activity than the subcutaneous fat (P < 0.01 in all cases). Moibi and Christopherson47 also found that internal adipose tissues contained more 16:0 and 18:0 than the peripheral subcutaneous fat. In the present experiment, the subcutaneous fat also contained greater proportions of some minor FAs, such as 11:0, 12:0, 14:1c11,15:0, 16:0iso, 16:1c9, 17:0, 17.0anteiso, ∑17:1others, 18:0iso, and 20:1c9, and smaller proportions of other minor FAs than the other adipose tissues. Similarly, the perivisceral and perinephric fats also differed in their proportions of a few FA groups and individuals. The proportion of branched FAs in the three fat depots was just over 3 g/100 g total FAs and somewhat higher in the subcutaneous fats than in the inner fat depots (P < 0.01). Wood et al.1 found that lamb subcutaneous fat usually contains small concentrations of branched FAs, normally below 4 g/100 g of total FAs. These FAs are responsible for the soft, oily consistency of fat depots, and higher concentrations were found in lambs fed on high-concentrate diets.1 The adipose tissues contained more trans FAs (18:1t11 and 18:1t8−10) than the muscles (P < 0.01), the overall ranking being internal fat depots > subcutaneous fat > liver > muscle, in agreement with ref 48. The predominant isomer formed in the rumen is commonly 18:1t11, but lambs fed high-concentrate diets may also have high proportions of 18:1t10 (coeluted with 18:1t8−10 in the present experiment).48 Differences in the proportions of trans FAs in the various tissues would therefore suggest differences in the uptake of these FAs from the rumen, but may also reflect metabolic differences among the tissues.49 Tissue-Dependent Response to CLA Supply. The influence of a rumen-protected CLA supplement on the fat content and FA profiles of organs and tissues has not been widely studied in sheep or lambs.25 However, Terré et al.50 found that the FA profiles of the longissimus dorsi of lambs fed a rumen-protected CLA mixture exhibited small variations, except in the content of the two CLA isomers provided.

over 2 g/100 g total FAs, 10 were in concentrations of 0.5 to 2 g/100 g total FAs, and 69 were below 0.5 g/100 g total FA (data not shown). Although there were notable differences in the FA profiles of the 3 groups of tissues in terms of FA categories (Table 3) and individual FAs (Tables 4, 5), the FA composition was generally typical of lambs fed concentratebased indoor diets.42 The tissues differed in their desaturase indices, reflecting Δ9 desaturase activity.43 The highest value was found for the 18-C desaturase index, and the ranking was muscles > subcutaneous adipose tissue > internal adipose tissues, and liver. The tissue with the greatest proportion of PUFAs was the liver (23.8 g/100 g total FAs; P < 0.01), followed by muscle (9.3 g/100 g total FA) and adipose tissues (3.8 g/100 g total FAs). Liver had the lowest proportions of MUFAs (25.9 g/100 g total FA; P < 0.01), and FAs with 8 to 16 and 18 carbons (P < 0.01), and the highest proportion of very long chain FAs (>18 carbons; P < 0.01). Among the tissues, liver had the highest proportions of n-6 (17.4 g/100 g total FAs; P < 0.01) and n-3 (3.7 g/100 g total FAs; P < 0.01), but the n-6/n-3 ratio (4.71) was similar to that of the muscles (3.8) and lower than that of the adipose tissues (7.83−9.80). The liver contained more 18:0 (24.6 g/100 g total FAs), but fewer total FAs with 18 carbons (57.0 g/100 g total FAs), 16:0 (15.0 g/100 g total FAs), and 18:1c9 (16.3 g/100 g total FAs) than either the muscles (18.9, 61.0, 22.0, and 30.3 g/100 g total FAs, respectively) or the fats (20.3−28.4, 55−60, 26.6−27.3, and 21.8−26.0 g/100 g total FAs, respectively). This composition is consistent with the view that conversion of palmitate (16:0) to oleate (18:1c9) in the liver is rate-limited by Δ9 desaturase activity (stearoyl-CoA desaturase, SCD), which governs the conversion of 18:0 to 18:1c9, and not by elongation of 16:0 to 18:0.44 This does not contradict the observation that the liver discriminates against 18:0 in the absorption of longchain FAs from plasma NEFAs,22 so that as the liver get fatter, the proportion of 18:0 tends to fall while the proportions of 16:0 and 18:1c9 tend to increase.44 The liver contained notable concentrations of linoleic acid and derived n-6 FAs, of which arachidonic acid (20:4c5,c8,c11,c14) was the most abundant, and small concentrations of linolenic acid (18:3c9,c12,c15) and derived n-3 FAs, due to the diet being poor in linolenic acid, precursor of the n-3 FAs.25 The muscles had greater proportions of MUFAs (37.8−38.4 g/100 g total FAs) than the adipose tissues (P < 0.01), mainly because of the contribution of 18:1c9 (P < 0.01). The proportion of PUFAs was on the order of 9.3 g/100 g total FAs, intermediate between the liver and adipose tissues. The FA profiles of the muscles differed significantly from those of the adipose tissues for the majority of individual FAs, and there were also many significant differences between the longissimus thoracis (LT) and the other rib muscles (psoas major, psoas minor, multifidus dorsi, and longissimus costarum). The other rib muscles, which had more fat than the LT (6.34 vs 4.6 g/100 g total FAs, P < 0.01), also had more of the SFAs 14:0, 16:0, and 18:0 (P < 0.01) and fewer of the PUFAs 18:2c9,t11 and other n-3 FAs (P < 0.01). Scollan et al.5 and Wood et al.1 found that the fat content of the muscle influences the FA profile as the proportion of unsaturated polar lipids, constituents of the cell membrane phospholipids, is progressively diluted by saturated neutral lipids or triglycerides with increasing fat content. Differences among the muscles may also be related to the presence of muscle fibers with different oxidative, glycolytic, or oxido-glycolytic metabolic proper10610


Article

Journal of Agricultural and Food Chemistry Sinclair et al.35 reported that CLA supplementation did not alter the weight of the liver nor body fat deposition in lactating ewes and that it had little influence on the FA profile of the longissimus thoracis muscle, except for an increase in the two CLA isomers supplied. Wynn et al.51 reported a tissuedependent effect of rumen-protected CLA in sheep and also found that a rumen-protected CLA supplement resulted in lower concentrations of monounsaturated fats (palmitoleate and oleate) in adipose tissues and liver. These effects on the FA profiles of tissues were consistent with inhibition of the Δ9 desaturase enzyme, as reported for pigs by Cordero et al.52 In the present experiment, the rumen-protected CLA supplement had little influence on the major indices of fatness of the lamb carcass, and little or no influence on the fat content of the liver and tissues. On average, CLA supplementation increased the proportions of 18:2c9,t11 and 18:2t10,c12 in the tissues but had only a small effect on the relative contents of other FAs, although it interacted with the tissues for some groups or individual FAs. The most notable CLA × (muscle × fat depot) interactions (Figure 1) regarded the proportions of SFAs (P < 0.01) and

Figure 2. Influence of a commercial CLA mixture providing 1.58 and 1.54 g/d of 18:2c9,t11 and 18:2t10,c12, respectively, on the proportion of 18:0 and 18:1c9 in different lamb tissues [the interactions CLA × (muscles × fat depots) were significant at P < 0.01]. Each bar is the least-squares mean from 12 observations and vertical bars indicate the standard error of the means.

interesting finding given the susceptibility of the ruminant liver to hepatic steatosis, especially when fed a diet rich in polyunsaturated FAs, such as linoleic acid.45 The CLA × (muscle × fat depot) interaction was also significant for the 18:2t10,c12 isomer (P < 0.01), evidenced by the greater increase in this isomer in the adipose tissues than in muscle as a result of CLA supplementation (Figure 3). The CLA × (muscle × fat depot) interaction was also consistently significant for the C-18 index of desaturase activity, as the CLA mixture reduced the value of this index in the adipose tissues but not in the muscles or liver. The variations observed in the SFAs, MUFAs, 18:0, 18:1c9, 18:2t10,c12, and the C-18 desaturase index were generally consistent and suggest the existence of strict cause and effect relationships among these factors. Overall, the results of the present experiment reveal that the response of lambs fed on indoor diets to a rumen-protected CLA supplement seems to be tissue-dependent, with the 18:2c10,t12 and 18:2t10,c12 isomers possibly playing a key role in controlling the Δ9 desaturase activity. As these isomers were found to have distinct cellular and metabolic effects,53 studies to evaluate their influences, separately or in combination, on the metabolic response of different organs and tissues are desired. In conclusion, administering a rumen-protected CLA supplement to lambs fed on indoor diets had small effects on the fat content of the carcass, liver and tissues, and on the FA profiles of these body constituents. However, the CLA increased the proportions of SFAs (mainly 18:0, a component of the rpCLA mixture) and decreased the proportions of MUFAs (18:1c9) in the adipose tissues, while the muscles exhibited the opposite effect. These variations, consistent with those in the C-18 desaturase index and the 18:2t10,c12 content in the various tissues, suggest that 18:1t10,c12 and 18:2t10,c12 probably play

Figure 1. Influence of a commercial CLA mixture providing 1.58 and 1.54 g/d of 18:2c9,t11 and 18:2t10,c12, respectively, on the proportion of saturated (SFA) and monounsaturated (MUFA) fatty acids in different lamb tissues [the interactions CLA × (muscles × fat depots) were significant at P < 0.01]. Each bar is the least-squares mean from 12 observations and vertical bars indicate the standard error of the means.

MUFAs (P < 0.01). The CLA mixture resulted in an increase in SFAs and a decrease in MUFAs in all the 3 adipose tissues, while the opposite was observed for the muscles, which was primarily due to the changes observed in 18:0 and 18:1c9, the main MUFA constituents (Figure 2). The greater increase in 18:0, a component of the rpCLA mixture, and the decrease in 18:1c9 in the adipose tissue might suggest that Δ9 desaturase activity in these tissues was inhibited by the CLA mixture more than in other tissues. The liver was the least responsive to the CLA supplement in terms of 18:0 and 18:1c9 contents, an 10611


Article


(EC) no. 1698/2005-PSR 2007−2013, Regional Resolution no. 199/2008). Notes

The authors declare no competing financial interest.

■

ACKNOWLEDGMENTS The authors would like to thank Luca Grigoletto (DAFNAE, University of Padua, Legnaro, Italy) for technical work and the GC × GC analysis and Giovanni Bittante (DAFNAE, University of Padua, Legnaro, Italy) for scientific support. The authors are also grateful to SILA s.r.l. (Noale, VE, Italy) for providing the rumen-protected CLA.

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ABBREVIATIONS SFAs, saturated fatty acids; PUFAs, polyunsaturated fatty acids; MUFAs, monounsaturated fatty acids; FAs, fatty acids; CLAs, conjugated linoleic acids; rpCLA, rumen-protected conjugated linoleic acid; TMR, total mixed ration; DM, dry matter; GC × GC, two-dimensional gas chromatography−mass spectrometry; LOD, limit of detection; SEM, standard error of the mean

■

Figure 3. Influence of a commercial conjugated linoleic acid (CLA) mixture providing 1.58 and 1.54 g/d of 18:2c9,t11 and 18:2t10,c12, respectively, on the proportions of total CLA (∑CLA), 18:2c9,t11, 18.2t10,c12, and Δ9 desaturase index in different lamb tissues. The following interactions were significant (P < 0.01): [CLA × (liver × other tissues)] for ∑CLA, and 18:2c9,t11; [CLA × (muscle × fat depots)] for 18.2t10,c12 and Δ9 desaturase. Each bar is the leastsquares mean from 12 observations and vertical bars indicate SEM.

a role in regulating Δ9 desaturase activity and that this role is tissue-dependent.

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REFERENCES

(1) Wood, J.; Enser, M.; Fisher, A.; Nute, G.; Sheard, P.; Richardson, R.; Hughes, S.; Whittington, F. Fat deposition, fatty acid composition and meat quality: A review. Meat Sci. 2008, 78, 343−358. (2) Mozaffarian, D.; Wilson, P. W.; Kannel, W. B. Beyond established and novel risk factors: lifestyle risk factors for cardiovascular disease. Circulation 2008, 117, 3031−3038. (3) Siri-Tarino, P. W.; Chiu, S.; Bergeron, N.; Krauss, R. M. Saturated Fats Versus Polyunsaturated Fats Versus Carbohydrates for Cardiovascular Disease Prevention and Treatment. Annu. Rev. Nutr. 2015, 35, 517−543. (4) Chowdhury, R.; Warnakula, S.; Kunutsor, S.; Crowe, F.; Ward, H. A.; Johnson, L.; Franco, O. H.; Butterworth, A. S.; Forouhi, N. G.; Thompson, S. G.; et al. Association of dietary, circulating, and supplement fatty acids with coronary risk: a systematic review and meta-analysis. Ann. Intern. Med. 2014, 160, 398−406. (5) Scollan, N. D.; Choi, N.; Kurt, E.; Fisher, A. V.; Enser, M.; Wood, J. D. Manipulating the fatty acid composition of muscle and adipose tissue in beef cattle. Br. J. Nutr. 2001, 85, 115−124. (6) Demirel, G.; Wachira, A.; Sinclair, L.; Wilkinson, R.; Wood, J.; Enser, M. Effects of dietary n-3 polyunsaturated fatty acids, breed and dietary vitamin E on the fatty acids of lamb muscle, liver and adipose tissue. Br. J. Nutr. 2004, 91, 551−565. (7) Hur, S. J.; Kim, H. S.; Bahk, Y. Y.; Park, Y. Overview of conjugated linoleic acid formation and accumulation in animal products. Livestock Science 2017, 195, 105−111. (8) Schiavon, S.; Tagliapietra, F.; Dalla Montà, G.; Cecchinato, A.; Bittante, G. Low protein diets and rumen-protected conjugated linoleic acid increase nitrogen efficiency and reduce the environmental impact of double-muscled young Piemontese bulls. Anim. Feed Sci. Technol. 2012, 174, 96−107. (9) Moon, H. S.; Lee, H. G.; Chung, C. H.; Choi, Y. J.; Cho, C. S. Physico-chemical modifications of conjugated linoleic acid for ruminal protection and oxidative stability. Nutr. Metab. 2008, 5, 16. (10) Perfield, J.; Lock, A.; Pfeiffer, A.; Bauman, D. Effects of amideprotected and lipid-encapsulated conjugated linoleic acid (CLA) supplements on milk fat synthesis. J. Dairy Sci. 2004, 87, 3010−3016. (11) De Veth, M.; Gulati, S.; Luchini, N.; Bauman, D. Comparison of calcium salts and formaldehyde-protected conjugated linoleic acid in inducing milk fat depression. J. Dairy Sci. 2005, 88, 1685−1693. (12) Bauman, D. E.; Perfield, J. W., 2nd; Harvatine, K. J.; Baumgard, L. H. Regulation of fat synthesis by conjugated linoleic acid: lactation and the ruminant model. J. Nutr. 2008, 138, 403−409. (13) Pellattiero, E.; Cecchinato, A.; Tagliapietra, F.; Schiavon, S.; Bittante, G. The use of 2-dimensional gas chromatography to

AUTHOR INFORMATION

Corresponding Author

*E-mail: [email protected]. ORCID

Matteo Bergamaschi: 0000-0002-3983-5019 Funding

The authors would like to thank the Veneto Region’s “BionetRegional Biodiversity Network” project (Measure 214 H Reg. 10612


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Fatty Acid Composition of Lamb Liver, Muscle, And Adipose Tissues in

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