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ANIMAL NUTRITION |
Department of Animal Nutrition, Estación Experimental del Zaidín (CSIC), Camino del Jueves s/n, 18100 Armilla, Granada, Spain
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Abstract |
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 Top Abstract INTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
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Key Words: betaine • carcass deposition • conjugated linoleic acid • growth • Iberian pig
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INTRODUCTION |
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TopAbstractINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
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). More recent studies in finishing pigs have also demonstrated decreases in some indicators of body fat (Lawrence et al., 2002), with a potential reduction in feed intake (Matthews et al., 2001a) under some conditions. In contrast, other studies in finishing pigs have reported either minimal effects or no effect of betaine on growth, feed intake, or body fat (Matthews et al., 1998; Overland et al., 1999). Conjugated linoleic acid is a collective name that refers to a mixture of positional and geometric conjugated isomers of linoleic acid. Interest in feeding CLA to pigs has increased in the last decade as a result of its potential to increase rate of gain (Thiel-Cooper et al., 2001), to improve feed efficiency and protein accretion rate (Ostrowska et al., 1999), and to reduce body fat (Dugan et al., 1997; Thiel-Cooper et al., 2001). The results have not been consistent. Most of the studies investigating the effects of betaine or CLA were made with finishing lean pigs. With this background, we hypothesized that purebred Iberian pigs, an obese porcine genotype, would be an adequate model to analyze the effect of these growth promoters, and that a synergy of betaine and CLA, incorporated as a dietary mixture, might occur. The Iberian pig (Sus mediterraneus) is a slow-growing obese porcine breed with a limited capacity for protein deposition compared with conventional or high-performing pigs (Nieto et al., 2002). The objective of the current study was to evaluate the effect of adding betaine, CLA, or both to the feed of growing Iberian pigs with regard to growth performance and carcass composition. |
MATERIALS AND METHODS |
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TopAbstractINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
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The experimental protocol was reviewed and approved by the Bioethical Committee of the Spanish National Research Council (CSIC), Spain. The animals used in this experiment were cared for in accordance with European Union guidelines (no. 86/609/EEC).
A total of 25 purebred Iberian (Silvela strain) gilts from the same farrowing group were used in the study. Before the beginning of the growth trial, all pigs were group-housed and given ad libitum access to the control diet between 16 and 20 kg of BW. To obtain the initial body composition by chemical analysis, 5 pigs were killed at the beginning of the experiment. The remaining 20 animals were housed in individual 2 m2 pens located in a controlled environment room (20 ± 1.5°C) and randomly assigned to 1 of 4 experimental treatments. Treatments consisted of the incorporation of 0.5% betaine (Betafin S1, crystalline, 96% purity, Danisco, Copenhagen, Denmark), 1% CLA (60% CLA isomers, half cis-9, trans-11 and half trans-10, cis-12 in FFA form, BASF, Ludwigshafen, Germany), or 0.5% betaine + 1% CLA, to a basal diet, at the expense of sunflower oil (for CLA) and cornstarch (for betaine). The basal diet was used as a control. Diets were barley and soybean meal based and formulated to contain 12.0% CP, 0.81% Lys, and 14.8 MJ of ME/kg of DM (Table 1). Crude protein concentration was fixed to match the low potential for protein deposition of the Iberian breed, as determined previously by our group (Nieto et al., 2002), and the remaining nutrients met or exceeded NRC (1998) requirements. Lysine, Thr, and His were included in crystalline form. Pigs were fed at 95% of ad libitum intake, calculated as a function of BW following the linear regression model described by Nieto et al. (2001) for Iberian pigs. Animals were fed once daily (0900), and the daily feed allocation was adjusted weekly to accommodate the increasing requirement according to their individual BW. Water was provided for ad libitum consumption. The comparative slaughter technique was used, and a classical digestibility trial was carried out at approximately 30 kg of BW.
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Pigs were killed as they individually reached 51.1 ± 0.85 kg of BW, on average 54 ± 3 d after the initiation of the experiment. After an overnight fast, pigs were stunned by electrical shock and exsanguinated. Immediately after slaughter, the contents of the gastrointestinal tract were removed and discarded. The empty gut and stomach, along with the remaining visceral tissues and internal organs, were weighed. The eviscerated carcass with head, feet, and tail was chilled overnight, weighed, and standard carcass measurements were recorded. The head was then removed at the occipito-atlas joint, the feet were removed by cuts at the carpus-metacarpal and tarsus-metatarsal joints, and the carcass was divided longitudinally. The right side of each carcass was maintained at –20°C in plastic bags to avoid dehydration. The frozen right half of the carcass was thawed, cut into small pieces, ground in a mincer (Talleres Cato, Sabadell, Spain), and homogenized in a cutter (Talleres Cato), and subsamples were taken for freeze-drying and subsequent analysis. Carcass fat was calculated, assuming the energy content of 23.8 and 39.8 kJ/g for protein and fat, respectively (Wenk et al., 2001). The initial mean composition was calculated from 5 pigs (20.0 ± 0.37 kg of BW) slaughtered at the beginning of the growth trial. Carcass composition at 20 kg of BW for the experimental groups was calculated based on the relationship between BW and empty BW (EBW; obtained by summation of all body components collected), the relationship between carcass weight and EBW, and carcass chemical composition of the initial slaughter group. Increases in protein (determined as total nitrogen retention), energy, fat, and minerals were calculated as the difference between the final measured composition of experimental pigs and the estimated initial composition assessed from the initial slaughter group.
Carcass Measurements
After hanging overnight at 4°C, standard carcass measurements were recorded. The area of the LM was determined gravimetrically (precision balance AB2004, Mettler Toledo, Greifensee, Switzerland) after assuming a direct linear relationship between the area and weight of the paper used. For this purpose, the muscle surface area at the 10th rib was drawn on tracing paper, cut with a scissors following the trace mark, and weighed. Tenth-rib fat thickness was determined by measuring the fat thickness perpendicular to the outer skin surface at 3 locations over the LM (P1, 40 mm; P2, 60 mm; P3, 80 mm from the midline). Midline backfat thicknesses at the first rib, last rib, and last lumbar vertebra were also determined.
Digestibility Trial
At 30 kg of BW, animals were individually housed in 1.6 x 0.7 m, metal slatted-floor, adjustable metabolic cages placed in a controlled environment room (20 ± 1.5°C) to allow for separate collection of feces and urine. Pigs were moved to the cages 3 d before starting excreta collection and fit with a self-retaining urethral catheter to allow for quantitative urine collection. Total collection of feces and urine was performed daily for 4 d. Aliquot (20% of total) samples of feces and urine representative of total daily fecal output and of total urine (collected into 200 mL of 4 M H2SO4) were stored at –20°C. After thawing overnight, these samples were mixed thoroughly to build up a single fecal or urine sample per animal. The fecal sample was then freeze-dried and finely ground for further analysis. Urine samples were used to calculate ME intake (MEI) and nitrogen retention during the nitrogen balance experimental period.
Chemical Analysis
All analyses were performed in duplicate on feeds and freeze-dried carcass samples. Dry matter contents of the feeds, feces, and carcass were determined gravimetrically after heating powdered samples to 100°C for 18 h (AOAC, 1990).
Total nitrogen in feeds and in freeze-dried feces and carcass samples was determined by the Kjeldahl procedure using mineralization (Digestor Block Selecta S-509, Barcelona, Spain), distillation units (Model B-324, Büchi Laboratoriums Technik AG, Flawil, Switzerland), and titration units (Model 702 SM Titrino, Metrohm AG, Herisau, Switzerland). Total lipids in feeds were determined by ether extraction according to standard procedures (AOAC, 1990). Total ash determinations in samples of the feeds, carcass, and feces were carried out by standard procedures (AOAC, 1990). Gross energy determinations, measured in an adiabatic bomb calorimeter (Gallenkamp Autobomb CBA 305, Gallenkamp, Loughborough, UK), were performed on feeds and freeze-dried samples of carcass, feces, and urine. The latter was freeze-dried in a polyethylene sheet of known energy value, and the GE values were obtained by the difference. When an analysis was made on freeze-dried material, a DM determination was performed on an aliquot sample by standard procedures to establish the residual water content after freeze-drying, and the corresponding analytical result was expressed on a DM basis. Lean tissue was estimated as the sum of water and protein.
Statistical Analyses
The treatment effect was assessed by ANOVA according to a completely randomized design with 4 treatments using the GLM procedure and a computer software package (Statgraphics Plus for Windows Version 2.0, Manugistics Inc., Rockville, MD). Treatment means were compared by Tukey’s procedure (Tukey, 1953), and significance was set at P < 0.05. The individual pig was the experimental unit for all traits. Least squares means and SEM are presented.
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RESULTS |
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TopAbstractINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
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There were no differences in DM, DE, or MEI among treatments (Table 2). Feeding betaine or CLA alone had no effect on the growth performance of young, growing gilts. However, gilts fed betaine + CLA had increased ADG and gain:MEI compared with control gilts (P < 0.05). Nevertheless, G:F was not affected (P > 0.10). Digestibilities of DM, OM, nitrogen, energy, and ME (0.822, 0.840, 0.770, 0.813, and 0.798, average values, respectively; data not shown) were not affected by dietary treatments (P = 0.46 to 0.75), which suggests that betaine, CLA, or both had no effect on the digestion of feed. There were no differences in nitrogen retention during the 4-d balance period among treatments (12.9 ± 1 g of nitrogen/d, P = 0.856; data not shown).
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Daily rates of carcass protein, water, and estimated lean tissue deposition increased (P < 0.01; Table 3) in gilts fed betaine + CLA compared with control gilts. In addition, protein deposition relative to MEI (P < 0.05) increased in gilts fed betaine + CLA compared with the control group. Similarly, numerically greater carcass gain was observed when pigs fed betaine + CLA were compared with control pigs (P = 0.13). No differences in fat, mineral, and energy deposition were observed among treatments (P > 0.15). No differences among treatments were obtained for protein deposition relative to protein intake (P > 0.15). Pigs fed betaine or CLA had protein deposition values intermediate between the values for control pigs and the betaine + CLA-fed pigs.
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) and carcass fat content tended to decrease (P = 0.09) for gilts fed betaine + CLA compared with control gilts (Table 4). Similarly, regarding the estimated composition of carcass gain (g/kg of carcass gain; Table 4), protein was numerically increased (10%, P = 0.12) and water and lean tissue tended to be greater in gilts fed betaine + CLA compared with control animals (P = 0.06 to 0.08), whereas a trend toward decreased fat (P = 0.08) content was observed. No difference in the mineral content of carcass or carcass gain (calculated) was observed among treatments (P > 0.10).
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) of gilts fed betaine + CLA compared with control gilts (35%; P = 0.116) was observed. There were no differences in LM area among treatments. There was a numerical decrease in backfat thickness (Table 5) for pigs fed betaine + CLA compared with control pigs (P = 0.123). Backfat values for betaine- or CLA-fed pigs were intermediate between control pigs and betaine + CLA-fed pigs.
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No differences in the weight of viscera components were observed among treatments except for an increase in liver weight relative to EBW of gilts fed betaine + CLA compared with control gilts (P = 0.019). Mean values for total viscera and digestive tract (g/kg of EBW) were 134.6 and 72.4 g/kg of EBW, respectively (Table 6).
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DISCUSSION |
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TopAbstractINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
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), comparable to that of Meishan gilts at 44 and 57 kg of BW (White et al., 1995) or Beltsville high-fat barrows at 40 kg of BW (Tess et al., 1986). In addition, backfat measurements in the current study compared well to those of 40 kg of BW, high-fat and 50 kg of BW, genetically obese gilts (Yen and Pond, 1985; White et al., 1995). The selection of Iberian pigs was based on an expected greater effect of betaine and CLA in this pig breed compared with conventional pigs because of their greater adiposity and slower growth rate (Nieto et al., 2002). Our study is the first to demonstrate an effect of supplementing betaine + CLA on the growth, carcass composition, and tissue deposition of animals. Growth Performance
Although some researchers have shown reduced feed intake in pigs fed CLA (Dugan et al., 1997; Bassaganya-Riera et al., 2001) or betaine (Matthews et al., 2001a), other researchers have shown no significant effects of these additives (Ostrowska et al., 1999; Ramsay et al., 2001; Thiel-Cooper et al., 2001). We restricted feed intake slightly to obtain homogeneous nutrient and energy intake among treatments and to avoid a possible confounding effect on growth performance and carcass characteristics.
Adding betaine or CLA to the diets did not affect ADG or feed efficiency in our study. With the exception of one study (Thiel-Cooper et al., 2001), no effect of CLA on ADG of pigs has been shown. However, small improvements in G:F have been described in some (Ostrowska et al., 1999; Thiel-Cooper et al., 2001; Wiegand et al., 2001), but not all, cases (O’Quinn et al., 2000; Tischendorf et al., 2002; Mitchell et al., 2005). With respect to betaine addition in swine diets, most studies have not demonstrated improved growth performance in pigs fed a variety of experimental diets, either restricted or providing ad libitum access (Matthews et al., 1998; Overland et al., 1999; Fernández-Fígares et al., 2002). Others (Siljander-Rasi et al., 2003) have shown a slight improvement in finishing pigs restrictively fed diets with low levels of betaine (0.1%).
In the current experiment, growing gilts fed diets supplemented with betaine + CLA had greater ADG and gain:MEI ratio than control gilts. The increased gain relative to MEI is likely due to an increase in lean tissue accretion (see subsequent discussion).
Digestibility of Nutrients
There is a paucity of information regarding the effects of betaine or CLA on the nutrient digestibility of pigs. From reports in the literature describing the effects of betaine (Overland et al., 1999; Schrama et al., 2003) or CLA (Muller et al., 1999) on growing and finishing pigs, no effects on the digestion of dietary nutrients and energy were anticipated. Our results using growing gilts confirmed a lack of a betaine effect, a CLA effect, or a synergistic effect on the energy and nutrient digestibility of pigs. The apparent digestibility coefficients observed in this experiment are comparable to values previously obtained for Iberian pigs (Nieto et al., 2002; Barea, 2005). The effects of betaine, CLA, or both appear to be postabsorptive in nature.
Carcass Traits
The chemical composition of carcass and the calculated carcass gain composition were not altered in pigs fed betaine- or CLA-supplemented diets, although numerical trends toward enhanced protein and decreased fat concentrations were observed in the current experiment. Bassaganya-Riera et al. (2001) also reported no change in empty body composition of nursery pigs raised in dirty and clean environments and fed graded levels of CLA, except for a linear trend toward decreased fat concentration. Similarly, no effect on carcass ash content (Ostrowska et al., 1999), bone mineral density (Ostrowska et al., 2003; Mitchell et al., 2005), or dissected bone weight (Dugan et al., 1997) was shown for finishing pigs fed CLA-supplemented diets. In contrast, finishing pigs fed CLA-supplemented diets had reduced carcass fat (Ostrowska et al., 1999, 2003; Mitchell et al., 2005) and increased carcass lean content (Dugan et al., 1997; Ostrowska et al., 1999). The effect of CLA on carcass fat and protein concentration appears to be dependent on pig age. Therefore, this effect is more pronounced in pigs that are in the final stages of finishing or accumulating fat at a greater rate. This is a possible explanation for the lack of response of growing Iberian pigs to CLA feeding shown in the current experiment. The use of CLA supplementation on pig diets has produced controversial results regarding fat and protein deposition. Most studies that have fed pigs CLA-supplemented diets have reported decreased carcass fat (Ostrowska et al., 1999; Thiel-Cooper et al., 2001; Tischendorf et al., 2002) and increased carcass lean (Dugan et al., 1997; Ostrowska et al., 1999), although no change (O’Quinn et al., 2000; Demaree et al., 2002) or increased (Gatlin et al., 2002; Joo et al., 2002) fat deposition have been reported as well. Some of this variation may be due to the measurements taken to approximate carcass composition. Furthermore, it has been suggested (Azain, 2003, 2004) that a threshold of 23 mm at 10th-rib backfat is necessary for CLA to have an effect on carcass fat, although this has recently been challenged by Weber et al. (2006), who showed a trend toward decreased 10th-rib backfat depth after 8 wk of feeding CLA to genetically lean gilts. Other reports have indicated that betaine has no effect on carcass fat or protein content in finishing pigs (Matthews et al., 1998; Overland et al., 1999; Siljander-Rasi et al., 2003). Nevertheless, Fernández-Fígares et al. (2002) reported a trend toward a linear decrease in carcass fat content in growing pigs fed increasing graded levels of betaine (maximum effect at 0.5% betaine). It should be noted that feed restriction was greater than in the current experiment.
Similar to CLA, the effects of betaine on carcass protein, fat, and energy deposition in pigs are not conclusive. Schrama et al. (2003) showed a marginal increase in total energy retention, whereas energy retained as protein or fat remained unchanged in growing pigs restrictively fed diets supplemented with 0.129% betaine. Fernández-Fígares et al. (2002) showed a linear trend toward an increased carcass protein deposition rate without a change in the carcass fat deposition rate of growing pigs restrictively (70% ad libitum) fed betaine-supplemented diets. Other authors showed no difference in estimated lean gain (Matthews et al., 2001b; Lawrence et al., 2002) or fat gain (Lawrence et al., 2002) when pigs were fed ad libitum. It seems that the effects of betaine are more definitive under conditions of feed restriction.
It is common to estimate the carcass fat content by measuring backfat thickness and fat depth, whereas LM area and depth, carcass length, and the weight of LM, shoulder, or ham are used as an estimate of carcass leanness. No change in LM area (Matthews et al., 2001a,b; as in the current study) or increased carcass length (Matthews et al., 1998, 2001a) in pigs fed betaine-supplemented diets have been reported.
Viscera Traits
In pigs, the use of CLA did not alter liver weight (Dugan et al., 2002; Tischendorf et al., 2002) or spleen (Tischendorf et al., 2002) as in the current study. The weight of liver was also not increased in betaine-fed pigs (Overland et al., 1999; Fernández-Fígares et al., 2002), similar to our results. Interestingly, the association of betaine + CLA elicited an unexpected increase (19%) in liver weight in the present experiment. Enlargements of the liver and spleen of rodents fed CLA-supplemented diets have raised safety issues (DeLany et al., 1999; Tsuboyama-Kasaoka et al., 2000), although the examination of liver tissue did not show any severe pathological changes except increased lipid droplets. Betaine is a well-known lipotropic agent, and it has been shown to reduce hepatic liposis induced by carbon tetrachloride in rats (Junnila et al., 1998). Hence, we would expect that the lipotropic effect of betaine would compensate for the putative increase in hepatic lipid content described in CLA-supplemented mice.
Responses to dietary betaine in feed intake, growth performance, and carcass composition are often inconsistent in pigs, and interactions with dietary protein (Matthews et al., 1998; Lawrence et al., 2002), energy (Matthews et al., 1998), sex (Lawrence et al., 2002), time (Matthews et al., 2001b; Schrama et al., 2003), and dietary level of betaine (Siljander-Rasi et al., 2003) have been shown. It is possible that positive effects of betaine on growth performance and carcass characteristics are evident only under conditions of metabolic or nutritional stress, as suggested by Fernández-Fígares et al. (2002). It is not well established how betaine could promote growth and carcass composition. There are indications that betaine may play a role in lipid metabolism. Betaine lowered plasma FFA concentrations in untrained Thoroughbred horses (Warren et al., 1999), prevented alcoholic fatty liver accumulation in rats (Barak et al., 1993, 1994), and decreased backfat thickness in pigs (Cadogan et al., 1993; Virtanen and Campbell, 1994).
It has also been suggested that the effects of betaine on fat and protein accretion may be mediated more through allocation of AA among lean growth, visceral growth, and metabolic breakdown than by lipid metabolism per se (Virtanen and Campbell, 1994). Furthermore, Wray-Cahen et al. (2004) established both in vivo and in vitro that betaine does not have an effect on long-chain fatty acid oxidation in pigs.
Improvements in body composition caused by CLA supplementation in pigs are generally supported, but evidence indicating that CLA improves growth rate or feed conversion is limited. Conjugated linoleic acid reduced carcass fat in pigs with thick subcutaneous fat (Ostrowska et al., 1999; O’Quinn et al., 2000; Thiel-Cooper et al., 2001) at 100 kg of BW and in gilts compared with barrows (Tischendorf et al., 2002). Nevertheless, the Iberian gilts used in the current study were not responsive to CLA in spite of their lipogenic profile compared with modern breeds. It should be noted that it may have been too early in their growth stage for CLA to elicit a significant effect on carcass composition. Furthermore, that there was no effect of CLA on the growth performance of weanling pigs (Weber et al., 2001) and growing pigs (Ramsay et al., 2001, and present study) may be the consequence of the more efficient growth rate of young pigs compared with finishing pigs.
The basis of the multiple effects of CLA likely involves the effects of CLA on eicosanoid metabolism, cytokine production, gene expression, or all the aforementioned factors (Azain, 2003). Mechanistically, CLA may decrease the adiposity of pigs through its ability to depress the activity of stearoyl CoA desaturase in porcine adipose tissue (Smith et al., 2002). Indeed, the increased abundance of saturated fatty acids found in the tissues of pigs fed CLA may be reflective of a decrease in desaturase activity (Bee, 2000). Another mechanism via which CLA may decrease adiposity in the pig is by decreased fat cell size (Corino et al., 2005). De novo lipogenesis was not altered by CLA in finishing pigs (Bee, 2001), but there are indications that it reduced the rate of lipogenesis from preformed fatty acids and reduced lipolysis in 65 kg of BW pigs (Ostrowska et al., 2002).
Age, sex, and genetics influence the capacity for lean accretion and the amount of fat deposition, and consequently might interfere with the mode of action of betaine or CLA. Metabolic modifiers that decrease carcass fat should be more effective in obese animals or animals otherwise having a tendency toward fat accretion (e.g., gilts have greater capacity for fat accretion than boars; Dunshea et al., 1993). However, the responses may vary with the genetic propensity for fat or lean accretion. We expected greater effects of betaine or CLA in Iberian pigs, which are characterized by a very high capacity for fat accretion, than in modern breeds showing maximum protein and low fat gain; however, betaine or CLA alone did not alter the growth performance or carcass characteristics of young Iberian gilts.
In summary, betaine or CLA alone did not elicit significant changes in the growth performance and body composition of growing Iberian gilts, but the addition of both did affect growth and body composition, indicating synergistic action. Further studies are needed to quantify the potential effects of a dietary mixture of betaine + CLA on lipid, protein, and energy metabolism.
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Footnotes |
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2 This research was supported by grant no. AGL2002-01562 from the Spanish Ministry of Science and Education. The authors thank BASF (Ludwigshafen, Germany) and Danisco (Copenhagen, Denmark) for the donation of conjugated linoleic acid and betaine, respectively; Encarnación Colmenero and Francisco Funes for skillful technical assistance; and Sánchez Romero Carvajal Jabugo S.A. (Seville, Spain) and Sucesores de Miguel Vílchez Riquelme S.A. (Granada, Spain) for their helpful collaboration.
3 Corresponding author: ignacio.fernandez-figares{at}eez.csic.es
Received for publication April 10, 2006. Accepted for publication September 25, 2007.
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LITERATURE CITED |
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TopAbstractINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
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9-desaturase index and stearoyl coenzyme A desaturase enzyme activity in porcine subcutaneous adipose tissue. J. Anim. Sci. 80:2110–2115.
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| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
