Determination by GC×GC of Fatty Acid and Conjugated Linoleic Acid (CLA) Isomer Profiles in Six Selected Tissues of Lambs Fed on Pasture or on Indoor Diets with and without Rumen-Protected CLAby Erika Pellattiero, Alessio Cecchinato, Franco Tagliapietra, Stefano Schiavon, Giovanni Bittante

J. Agric. Food Chem.


Chemistry (all) / Agricultural and Biological Sciences (all)


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Determination by GC×GC of Fatty Acid and Conjugated Linoleic Acid (CLA) Isomer Profiles in Six Selected Tissues of Lambs Fed on Pasture or on Indoor Diets with and without Rumen-Protected CLA

Erika Pellattiero, Alessio Cecchinato, Franco Tagliapietra,* Stefano Schiavon, and Giovanni Bittante

Department of Agronomy, Food, Natural Resources, Animals and Environment (DAFNAE), University of Padova,

Viale dell’Universita ̀ 16, 35020 Legnaro (PD), Italy *S Supporting Information

ABSTRACT: In this study GC×GC was used to study the effects of pasture, hay, concentrate (indoor), and indoor plus 8 g/day of a rumen-protected conjugated linoleic acid (indoor-CLA) diets on the detailed fatty acid (FA) profiles of six tissues (muscles, fatty tissues, and liver) collected from 36 lambs. This powerful technique allowed the quantification of 128 FAs, of which 21

SFAs, 16 MUFAs, 19 PUFAs were identified by reference standards. The diets had similar, but not identical, effects on FA profiles (g/100 g FA) in the various tissues, as both indoor diets reduced total PUFAs (from 8.91 ± 6.27 to 8.06 ± 5.97; p < 0.05) and n-3 PUFAs (from 2.70 ± 2.37 to 1.50 ± 1.69; p < 0.01) and increased n-6 PUFA (from 3.76 ± 2.46 to 4.58 ± 3.42; p < 0.01), branched (from 2.37 ± 2.05 to 3.23 ± 0.54; p < 0.01), odd-chain FAs (from 5.88 ± 5.33 to 7.07 ± 1.51; p < 0.01) compared to pasture. Indoor-CLA increased CLAc9,t11 (from 0.42 ± 0.13 to 0.53 ± 0.19; p < 0.01), CLAt10,c12 (from 0.07 ± 0.06 to 0.12 ± 0.22; p < 0.05), and CLAc11,t13 (from 0.02 ± 0.04 to 0.05 ± 0.04; p < 0.05) compared to indoor.

KEYWORDS: two-dimensional gas chromatography, fatty acids, rumen-protected conjugated linoleic acid, lambs, tissues ■ INTRODUCTION

The fatty acid (FA) profile of meat from ruminants is often associated with health problems for humans because of the high content of saturated FAs (SFA) as well as the high n-6/n-3

PUFA ratio and the trans FA content.1 Beneficial FAs, such as conjugated linoleic acid (CLA), may also be present;2 nevertheless, the effects of FA and micronutrient profiles on human health remains a controversial issue.3 New powerful analytical methodology and equipment, such as two-dimensional gas chromatography (GC×GC), makes obtaining detailed FA profiles feasible.4

Greater in-depth knowledge of the nutritional properties of meat can also be of benefit to local endangered breeds reared on pasture. Sheep breeding has an important role in rural, marginal, and mountain areas, where farmers have strong links to the land and to tradition and where there is significant use of native breeds. In the Veneto region (northeastern Italy) four native breeds, Foza, Brogna, Alpagota, and Lamon, all with different characteristics,5,6 are at risk of extinction.7 Lambing of these breeds is generally concentrated in late winter, and ewes and lambs are kept on pasture from spring to autumn. However, there is a tendency to distribute parturitions over other seasons as well to provide the market with lamb carcasses over more extended periods of time. This necessitates the use of an indoor feeding system based on hay and concentrate, which could alter carcass and meat characteristics, and these need to be evaluated with respect to their potential effects on human health.

According to guidelines for healthy nutrition, meat quality can be improved by increasing monounsaturated fatty acids (MUFAs) and polyunsaturated fatty acids (PUFAs), reducing the n-6/n-3 PUFA ratio and increasing the CLA content of red meat.8 Fatty acid composition can be affected by factors such as diet, species, breed, gender, age and weight, fatness, and fat deposit site. Some feeding systems can bring about an increase in the n-3 PUFA content, in particular C18:3 α-linoleic, and a reduction in the n-6/n-3 PUFA ratio.9 The CLA content can be increased through different dietary strategies; for example, pasture feeding resulted in a significant increase in CLAc9,t11 in muscle lipids of bulls compared with concentrate-fed bulls.10

However, pasture is subject to seasonal availability and has negative effects on animal growth, making it necessary to seek alternatives. Some feeds containing oil from soybean, linseed, and sunflower seeds, for example, were found to influence the n-3 PUFA level in tissues,11,12 and providing the animals with a commercial rumen-protected conjugated linoleic acid (rpCLA) was found to increase CLA content in the meat of growing cattle.13

The aim of this study was to use GC×GC to obtain very detailed FA profiles, including CLA isomers, of different tissues (liver, adipose tissues, and muscles) of lambs of local breeds either pasture-fed or on indoor diets with or without rpCLA supplement. ■ MATERIALS AND METHODS

Chemicals. Petroleum ether (32044, Sigma-Aldrich, St. Louis, MO,

USA), methanol (34860-2-5L-R, Sigma-Aldrich), and n-hexane (133242.1612, PanReac Applichem, Darmstadt, Germany) were used as solvents. Sodium methoxide (663234C, BDH Laboratory Supplies,

Poole, UK), oxalic acid (306007, Carlo Erba, Rodano, MI, Italy), and

Received: May 7, 2014

Revised: December 16, 2014

Accepted: December 19, 2014

Published: December 19, 2014

Article © 2014 American Chemical Society 963 DOI: 10.1021/jf504956x

J. Agric. Food Chem. 2015, 63, 963−974 ethyl ether (447534, Carlo Erba) were used as reagents. Mixtures of pure FA (674, Nu-Chek Prep Inc., Elysian, MN, USA), bacterial acid methyl esters (BAMEs) (Sigma-Aldrich) plus CLAc9,t11 (UC-60M,

Nu-Chek Prep Inc.), CLAt10,c12 (UC-61M, Nu-Chek Prep, Inc.),

CLAc9,c11 (1256, Matreya LLC, Pleasant Gap, PA, USA), CLAc11,t13 (1259, Matreya LLC), and CLAt9,t11 (1257, Matreya LLC) were used as reference standards, whereas methyl-12-tridecenoate (U-35 M,

Nu-Chek Prep Inc.) was used as internal standard.

Animals and Feeding Systems. The present study followed the “Guidelines for the Care and Use of Agricultural Animals in Agricultural Research and Teaching”.14 Data regarding growth performance and carcass traits of the lambs used in the current experiments are given in Pellattiero et al.15 Briefly, the lambs were selected from the two largest flocks on an in situ conservation program. Given the need for breed conservation, the number of lambs included in the experiment was kept to a minimum yet was sufficient to attain adequate statistical power. In total, 36 weaned lambs of three breeds,