Lipid profile of lamb meat from different genotypes submitted to diets with different energy levels

To assess the influence of genotype and diets with different energy levels on the lipid profile of sheep meat, 54 lambs were used (18 animals from each genotype: Morada Nova, Santa Inez and 1⁄2Dorper + 1⁄2Santa Inez crossbred), distributed in a completely randomized experimental design in a 3 × 2 factorial arragement (three genotypes and two diets). The animals were submitted to two diets, 2.5 Mcal ME/kg DM and 3.0 Mcal ME/kg DM, receiving feed and water to reach the desired average live weight of 30 kg, when they were slaughtered. Total lipids, cholesterol, phospholipids and fatty acids profile of the Longissimus muscle were assessed. Diet and genotype did not influence the total lipids, cholesterol and phospholipids contents. The diet with the highest energy value presented higher values for the concentration of C12:0, C14:0, C18:0, C19:0, C22:0 acids and monounsaturated fatty acids (MUFA) in relation to that with the lowest energy value. However, for the concentrations of C15:0 acid and polyunsaturated fatty acids (PUFA), the highest values were recorded for diet with the lowest energy value. Genotype and diet influenced significantly the MUFA concentration, in which 1⁄2Dorper + 1⁄2Santa Inez crossbred genotype and diet with 2.5 Mcal ME/kg DM showed the highest values. Genotype influenced the saturated fatty acids (SFA) concentration, with 1⁄2Dorper + 1⁄2Santa Inez crossbred obtaining the lowest mean, behavior similar to that observed for PUFA concentrations. Desirable fatty acids (DFA), atherogenicity index as well as PUFA:SFA, MUFA:SFA, (C18:0 + C18:1):C16:0 ratios were influenced by diet and genotype. 1⁄2Dorper + 1⁄2Santa Inez crossbred and diet with 2.5 Mcal ME/kg DM obtained the best results.


Introduction
Sheep meat is a source of protein with high biological value, deserving special attention by the different segments of the production chain, considering the increase in the consumers' demand level and the expansion of markets.
Morada Nova and Santa Inez native breeds have great importance for the Brazilian Northeastern economy and meat fitness with rusticity characteristics, being well adapted to the inhospitable conditions of semi-arid regions.The genetic group Dorper, from South Africa, has stood out due to its importance for the crossing with native breeds, such as Santa Inez, resulting in the production of lambs with rapid growth and high-quality carcass (Garcia et al., 2000).
Additionally, studies have been directed to assess the influence of diet on the chemical quality of sheep meat.The results obtained in relation to the effect of diet on the fat content of sheep meat have been contradictory, an example is the research of Madruga et al. (2005), who reported the effect of the use of diets with different compositions on the fat content (2.74 and 8.38 g/100g) of sheep meat; however, Bonagúrio et al. (2004) verified opposite effect working with Texel lambs fed with several diets (2.4 and 4.5 g/100g).Wood et al. (2008) on his extensive review on fat and meat quality recommended to consumers a reduction in the fat ingestion, especially those rich in cholesterol and saturated fatty acids (SFA), and an increasing intake of monounsaturated (MUFA) and polyunsaturated fatty acids (PUFA), with the aim of reducing the risk of obesity, cancer and cardiovascular diseases.
The composition of fatty acid (FA) present in muscle and fat tissue reflects the breeding conditions of animals and have a remarkable effect on the meat quality, being worth stressing that lambs raised on pasture produce meat with quality more favorable to the consumers' health than those raised in stalls, because their FA profile show higher concentrations of conjugated linoleic acid (C18:2 -CLA), C18:3 n-3 (linolenic acid), long chain PUFA n-3 and a higher C18:2 n-6:C18:3 n-3 ratio (Aurousseau et al., 2007).
Considering the importance of native breeds and the difficult conditions that the sheep breeding activity finds to develop in semi-arid regions, the present research was conducted with the objective of evaluating the effect of diets with different energy concentrations used in the termination of native and crossbred sheep on the lipid profile of the meat from these animals.

Material and Methods
This experiment was conducted at the Pendência Experimental Station, which belongs to the State of Paraiba Agriculture and Cattle Raising Research Company (EMEPA), located at the municipality of Soledade -Paraiba, Brazil.
Fifty-four whole male animals were used, 18 from each genotype: Morada Nova, Santa Inez and ½Dorper + ½Santa Inez crossbred, in a feedlot system.Morada Nova animals had approximately 150 days of age and an average weight of 14.98 kg; while Santa Inez and ½Dorper + ½Santa Inez genotypes were 100 days olds and presented an average weight of 17.63 and 17.80 kg, respectively.At the beginning of the experiment, all 54 lambs were divided into two groups, each group containing nine animals from each genotype, and submitted to two diets containing different energy levels: 2.5 Mcal ME/kg DM and 3.0 Mcal ME/kg DM, until they reached the average live weight of 30 kg.
The period of adaptation to stalls and diets was 14 days, during which the animals were vaccinated against clostridiosis and received oral moxidectin-based parasiticide at 1%.For the eimeriosis control, sulfaquinoxalin sodium 25g was used during four days.
Ingredients used in diets were: Tifton hay (Cynodum sp), soybean meal, corn, urea, mineral salt and enriched palm meal, this latter obtained through fermentation process in the presence of mineral mix (Table 1).The diets in the form of full mixture were made for a daily gain of 250 g/day (NRC, 1985).The feed and water supply were ad libitum.Diet samples were collected for chemical analysis regarding dry matter (DM), crude protein (CP) and ether extract (EE) as methodologies described by AOAC (2000), and neutral detergent fiber (NDF) and acid detergent fiber (ADF) as described by Goering & Van Soest (1970).
The animals were slaughtered when the desired final weight was reached, being submitted to a previous period of solid fasting and hydric diet for a period of 16 hours.The slaughter method used was stunning with brain concussion, followed by bleeding, skinning and gutting.The carcasses were washed, weighed and chilled in a cold room at 4°C for 24 hours.After that, samples from the Longissimus muscle (LM) were desiccated, vacuum packed, frozen and stored at -20°C for later laboratory tests for a period no longer than 90 days until the beginning of the analyses.
For the total lipids, cholesterol, phospholipids and FA profile analyses, the subcutaneous and epimisium fat tissue was removed from LM samples.The samples were thawed overnight and completely minced and homogenized before starting the tests, and three replicated per sample were used (Madruga et al., 2001).
The total lipids were extracted in chloroform: methanol solution (2:1 vol/vol), according to procedures of method proposed by Folch et al. (1957) and extracts used in the fatty acids, cholesterol and phospholipids analyses.The phospholipids content was determined in accordance with methodology described by Pikul et al. (1985) and adapted by Madruga et al. (2001), with colorimetric reading in spectrometer (METERTEC,Brazil), at wavelength of 660 nm of absorbance (Rangana, 1991).The cholesterol dosage was carried out in accordance with methodology described by Bragagnolo & Rodriguez-Amaya (1992), through spectrometer (MICRONAL, model B395, Sao Paulo, Brazil), in absorbance with wavelength of 490 nm.The analytical curve was performed with standard cholesterol P.A. (VETEC, Rio de Janeiro, Brazil), based on a standard solution prepared with 10 mg of cholesterol in 100 mL of hexane, with concentrations ranging from 0.1 to 0.6 mg/mL.
In determining the FA profile of sheep meat, the lipids, after extraction, were esterified and methylated according to method described by Hartmam & Lago (1973).The methyl esters of fatty acids were separated and quantified in a gas chromatograph (GC-QP5050A, Shimadzu, Brazil) equipped with polar-type cast-silicon capillary column Carbowax 20M (SUPELCO, USA) wrapped in polyethylene, with the following dimensions: 60 m x 0,53 mm i.d.x 1 μm of film thickness, using H 2 as carrier gas at a flow rate of 40 cm/sec.Samples of methyl esters (1.0 μ L) were introduced in a split/ splitless type injector at a temperature of 230°C.The initial and final temperatures were, respectively, 120 and 210°C, with intermediate ramp of 8°C/min, and the detector temperature was maintained at 250°C.In order to be quantified, the areas under the peaks were recorded in Peaksimple-type software (ARI Instruments, USA).FA were identified by comparing the retention times of methyl esters from samples with authentic patterns of esters from fatty acids (Merck, USA).
The animals were distributed in a completely randomized experimental design, in a 3 × 2 factorial scheme, with nine replicates per treatment.Three genotypes (Morada Nova, Santa Inez and ½Dorper + ½Santa Inez) and two diets (2.5 Mcal ME/kg DM and 3.0 Mcal ME/kg DM) were tested.The mathematical model included genotype, diet, and the genotype × diet interaction as sources of variation.Data were submitted to analysis of variance and, whenever necessary, the means were compared through the Tukey test at 5% probability.

Results and Discussion
The lipid values obtained were quite homogeneous, ranging from 2.15 to 2.36%, with no variation (P>0.05) between genotypes and diets (Table 2).
The effect of diet on the lipids profile in sheep meat was observed by Madruga et al. (2005), who reported this interference, while Zapata et al. (2001) andAlmeida Jr. et al. (2004) found no effect of diet on the lipids profile of sheep meat.Despite this controversy, the authors found values close to those observed in this study.
A study conducted by Zapata et al. (2003) showed that the fat deposition in the carcass of sheep occurs mainly in the subcutaneous layer, and the proportion of fat is smaller in entire males, as the animals used in this study, than in castrated males or females.
According to Garcia et al. (2000), there are evidences that the Santa Inez breed, when terminated under feedlot presents potential for the production of meat with lower amounts of fat, which may explain the concentrations of total lipids found in this experiment.
Factors genotype and diet did not interfere in the cholesterol levels of the sheep meat, showing variation of the averages obtained from 65.58 to 67.88 mg/100g (Table 2).Perez et al. (2002) assessed the meat from Santa Inez and Bergamácia lambs slaughtered at different weights (15, 25, 35 and 45 kg) and observed that the cholesterol level decreased with the increased weight at slaughter, and reported value of 67.57mg/100g for animals slaughtered at 35 kg, which is in agreement with values obtained in this study, in which animals were slaughtered only when reaching 30 kg.However, there is a disagreement regarding the results presented in literature in relation to the effect of genotype and diet on the cholesterol concentrations of sheep meat.Zapata et al. (2001) observed no effect of genotype and diet on the cholesterol levels in the meat from the crossing between Brazilian breeds Somali, Criollo and Santa Inez fed with bermuda grass, leucaena hay and one group receiving, besides these, a concentrate with 20% of crude protein.On the other hand, Madruga et al. (2005) analyzed the quality of meat from Santa Inez lambs terminated with different types of feed (grass, pineapple stubbles, forage palm and corn silage) and concluded that the diet influenced this characteristic, indicating that the diet offered to the animal can improve the quality of its meat.This divergence between data is possibly due to the type of diet used.
Observing values found for phospholipids (Table 2), uniformity between genotypes and diets is verified (P>0.05) with averages ranging from 6.80 to 9.12 mg/100g of meat.Madruga et al. (2005) found phospholipids values ranging from 15.88 to 21.67 mg/100g, higher than those found in this research, the difference between values found is possibly due to diet employed.Phospholipids are structural organic compounds that have high PUFA concentrations in their composition, particularly those with three or more double bonds, such as linolenic acid (C18:3) (Fisher et al., 2000).
The FA profile in meat from ruminants was characterized by the presence of short-chain FA and small amounts of PUFA (Table 3).
The FA values observed in this evaluation show the influence (P<0.05) of genotypes and diets studied.Thirteen FA were identified, which represent the sum of fatty acids present in phospholipids and in the lipid fraction, composed of triglycerides and small quantities of free fatty acids.
Morada Nova lambs achieved the highest average for C14:1, C18:3 and SFA, and this latter value was similar to that observed for Santa Inez lambs.Still in relation to Santa Inez native breed, this genotype showed higher values for C10:0, C14:0, C18:0, C16:1 and PUFA.The meat from ½ Dorper + ½ Santa Inez crossbred obtained the highest average for MUFA and C18:1.
According to Rhee et al. (2000), Perez et al. (2002) andKesava Rao et al. (2003), the plasma cholesterol concentration is influenced by the fatty acids composition in diet, in which C16:0 increases, C18:1 decreases, and C18:0 does not exercise any influence on the blood cholesterol levels.It was observed that factor diet influenced the C18:1 concentration.This relationship may explain the homogeneity of the cholesterol dosage found in the meat from the different genotypes evaluated (Table 2).
According to Grundy (1994), C18:0 is considered hypolipidemic, since it plays a role in the cholesterol reduction, which results from the rapid conversion of C18:0 into C18:1, only requiring desaturation in the n-9 portion, potential beneficial agent in reducing the cholesterol levels.MUFA suffered influence from genotype and diet, with ½Dorper + ½Santa Inez crossbred presenting the lowest average for C14:1, which is in agreement with results found by Sañudo et al. (2000), who assessed the FA composition in the meat from British and Spanish breed lambs.
With regard to the diets used, the one with the highest energy concentration provided higher averages for most acids (P<0.05),except for C15:0, C18:1, C18:2 and C18:3.
The results presented in this work with respect to the total MUFA, SFA and PUFA percentage, differed from values reported by Rizzi et al. (2002) and Tshabalala et al. (2005), who found MUFA percentages higher than those of this evaluation and lower for other summations.
The genotype factor influenced the summation of MUFA, SFA and PUFA, while diet showed difference (P<0.05)only for MUFA and PUFA.Madruga et al. (2005) evaluated the quality of meat from Santa Inez lambs terminated with different diets and found values ranging from 50.51 to 47.18% for SFA; from 51.81 to 44.50% for MUFA and from 5.01 to 2.25% for PUFA.The values of the first summations are lower than those found in this research, while for MUFA and PUFA, the behavior was the opposite, stressing that this increase is due to the high linoleic acid percentage found in treatments, indicating an increase in the PUFA:SFA ratio, important to the consumers' health (Wood et al., 2003).
In this work, it was found that the meat from ½Dorper + ½Santa Inez crossbred lambs stands out in the nutritional aspect due to its higher C18:1 fatty acid and MUFA contents, while Santa Inez genotype showed higher SFA values, which is a less desirable nutritional condition.Diets had influenced the PUFA profile for the genotype studied, with the lowest energy concentration presenting the best averages.
Relations or proportions have been suggested as a way of assessing the risk factor of foods in relation to the increase in the blood cholesterol levels; since it is known that SFA increases the plasma cholesterol levels, while PUFA lower them.According to Banskalieva et al. (2000), it is possible to express the concentration of desirable fatty acids (DFA) through the sum of unsaturated fatty acid with stearic acid.The stearic acid (C18:0), although saturated, is neutral, and has fewer implications in the lipid profile, since it can be converted into oleic acid (C18:1) in the body; however, monounsaturated acids, oleic acid, polyunsaturated acids, linolenic acid and α-linolenic acid reduce the LDLcholesterol levels, and thus the risk of obesity, cancer and cardiovascular diseases (Perez et al., 2002).
The influence of genotype and diets was observed in all presented relations (Table 4).½Dorper + ½Santa Inez crossbred lambs obtained the desirable average for the atherogenicity index, and high MUFA:SFA and (C18:0 + C18:1):C16:0 ratios, while Santa Inez lambs obtained the highest average for the PUFA:SFA ratio.For DFA and n6:n3 ratio, these two genotypes were similar (Table 4).
The biological effect of essential FA depends on the PUFA:MUFA and PUFA:SFA ratios.The high C18:1 content and the consequent increase in the PUFA:SFA ratio are

Table 1 -
Composition of experimental diets

Table 2 -
Fat cholesterol percentage and phospholipids contents (mg/100g) of meat from Morada Nova, Santa Inez and ½ Dorper + ½ Santa Inez (½D+½SI) genotypes submitted to diets with different energy concentrations

Table 4 -
Averages of relations between saturated, monounsaturated and polyunsaturated fatty acids in the meat from Morada nova, Santa Inez and ½Dorper + ½Santa Inez (½D+½SI) genotypes submitted to diets with different energy concentrations