Ewa Ostrowska

Conjugated linoleic acid decreases fat accretion in pigs: evaluation by dual-energy X-ray absorptiometry

Thirty female Large White x Landrace pigs (average weight 57.2 (sd 1.9) kg) were allocated to one of six dietary treatments containing 0, 1.25, 2.5, 5.0, 7.5 or 10.0 g 55 % conjugated linoleic acids (CLA) isomers (CLA-55)/kg diet and fed for 8 weeks. Each pig was scanned at 0, 28 and 56 d and again at post slaughter using dual-energy X-ray absorptiometry (DXA) to determine the temporal pattern of body composition responses. Values determined by DXA were adjusted using regression equations generated from validation experiments between chemically and DXA-predicted values. Overall, there was a significant linear reduction in fat content with the increasing levels of CLA in the diet (P=0.007, P=0.011, P=0.008 at week 4, week 8 and for the carcass, respectively). The greatest improvement was recorded at the early stages of CLA supplementation and for the highest dose of CLA (week 4, -19.2 % compared with week 8, -13.7 %). In the first 4 weeks of feeding CLA, pigs receiving 10 g CLA-55/kg diet deposited 93 g less fat/d than pigs fed basal diets (P=0.002) compared with only 6 g less fat than control animals in the final 4 weeks. Lean content and lean deposition rate were maximised at 5 and 2.5 g CLA-55/kg diet for the first 4 weeks (P=0.016) and the final 4 weeks of treatment respectively. DXA estimates of bone mineral content and bone mineral density were not affected by CLA supplementation throughout the experiment. These data demonstrate that dietary CLA decreases body fat in a dose-dependent manner and that the response is greatest over the initial 4 weeks of treatment.

Accuracy of dual energy X-ray absorptiometry (DXA), weight and P2 back fat to predict whole body and carcass composition in pigs within and across experiments

Abstract An Hologic QDR4500 dual energy X-ray absorptiometer (DXA) was used to measure body composition in 151 pigs ranging from 10 to 120 kg live weight. Large White×Landrace pigs of mixed sexes were selected from five different experiments to evaluate DXA accuracy within and across experiments. Values predicted by DXA including total tissue mass, fat tissue mass, lean tissue mass and bone mineral content, for the live animal, carcass and half carcass were evaluated by comparison with chemically-determined values. Relationships between chemically-determined values and measurements of weight and backfat at the P2 site were also evaluated. Chemically-determined values were strongly related with DXA-derived values, more so than with weight and P2 or a combination of both, particularly in the measurement of fat composition. In contrast to estimates derived from weight and P2, DXA-derived estimates remained accurate when between experiment variation was included. Incorporation of subregions into a whole body software analysis influenced DXA’s ability to predict fat tissue mass with the most accurate measurements achieved by placing the entire scan image in the left arm region of the regional analysis grid. These results demonstrate the efficacy of DXA as a nondestructive method for determining body composition in the live animal and carcass, and its greater accuracy than current routinely used methods.

Comparison of silver-ion high-performance liquid chromatographic quantification of free and methylated conjugated linoleic acids.

Silver-ion high-performance liquid chromatography was used to fractionate a mixture of conjugated linoleic acid (CLA) isomers (as the free fatty acids, CLAFFA) in commercial CLA mixtures and biological samples. Due to the unchanged retention mechanism, it was assumed that the elution order of the isomers remained the same as that of methyl esters separated on the same column. The most abundant isomers, cis/trans 10,12-18:2 and cis/trans 9,11-18:2, were separated better as free acids on a single column than in the methyl ester form. Quantification of the CLA standard was used as the reference profile to evaluate different methylation methods commonly used to prepare CLA methyl esters for quantitation. Acid- and vuigi base-catalyzed derivatization methods resulted in CLA intraisomerization and losses in total conjugated dienes content. Acid (HCl and BF3) methylations significantly elevated the level of trans,trans isomers and significantly reduced the cis/trans isomers. Base methylation, tetramethylguanidine/methanol, resulted in loss of trans,trans isomers, and a substantial loss of total underivatized conjugated dienes. Other catalysts such as the trimethylsilyldiazomethane produced additional peaks of unidentified artifacts. The analysis of CLAFFA appears to provide more accurate quantification of CLA isomers in commercial and biological samples.

Dietary conjugated linoleic acid differentially alters fatty acid composition and increases conjugated linoleic acid content in porcine adipose tissue

Conjugated linoleic acids (CLA) have been shown to decrease body fat content in pigs. It is possible that feeding pigs diets rich in CLA may increase carcass lipid CLA to levels that could provide health benefits when included as a part of a healthy diet. Therefore, the aim of the present study was to determine whether dietary CLA supplementation has any effect on the fatty acid composition of subcutaneous and intramuscular adipose tissue in pigs. Thirty-five female cross bred (Large White x Landrace) pigs (initial weight 57.2 kg and initial P2 back fat 11.5 mm) were used in the present study. Pigs were housed individually and randomly allocated to one of six dietary treatments (0.00, 1.25, 2.50, 5.00, 7.50 and 10.00 g CLA55 (55 g CLA isomers/100 g total fatty acids; Natural Lipids Ltd, Hovdebygda, Norway)/kg) and fed their respective diets for 8 weeks. Twelve CLA isomers in the diet and in pig tissue lipids were separated by Ag+-HPLC. CLA was incorporated at fivefold higher levels in subcutaneous fat as compared with intramuscular fat and in a dose-dependant manner. Overall, the transfer efficiency of CLA was maximized at 5.00 g CLA55/kg. However, there was clear selectivity in the uptake or incorporation of cis,trans-9,11 isomer over the trans,cis-10,12 isomer. In general, CLA supplementation produced significant changes in skeletal muscle and adipose tissue fatty acid composition, indicating that dietary CLA had a potent affect on lipid transport and metabolism in vivo. Significant increases in myristic, palmitic and palmitoleic acids and a reduction in arachidonic acid were observed, suggesting an alteration in activity of delta5-, delta6- and delta9-desaturases in pig adipose tissue. In conclusion, feeding pigs diets supplemented with CLA increases carcass lipid CLA, but also results in changes in the fatty acid profile in pig fat that could potentially outweigh the benefits of CLA.

Mixed mode retention and the use of competing acid for the Ag+-HPLC analysis of underivatized conjugated linoleic acids.

Summary Fatty acids (FAs) and fatty acid residues are generally methylated and analyzed by GC. The reasons for this are partly historic and partly because of the sensitivity advantage of flame ionization detection over UV absorption by the carboxylic acid functionality in saturated FAs. However, for strongly absorbing unsaturated acids such as the conjugated linoleic acids (CLAs), the sensitivity advantage is greatly reduced. Hence there seems little reason to waste time and introduce errors associated with methylation. Remarkably, this appears not to have been recognized. In this paper we describe our method development for the analysis of underivatized CLAs by silver ion HPLC separation on the ChromSpher Lipids column. Using mobile phases previously optimized for the analysis of the methylated CLAs, retention is excessive and a competing acid is required. Various combinations of small concentrations of acetic acid (3.0 ‐ 2.5%) with acetonitrile (0.0 ‐ 0.025%), respectively, yield similar resolution and run times. As well as its role as a competing acid, acetic acid acts as a general strong solvent and thus can be used alone as a modifier (without acetonitrile). However, for slightly shorter run times a mobile phase of 2.5% acetic acid and 0.025% acetonitrile was chosen as the optimum mobile phase for analysis. The separation of the free CLAs is clearly superior to those previously published and obtained in this study for the methylated CLAs. The additional specific strong interactions of the underivatized CLAs seem certain to be due to hydrogen bonding between the CLA carboxylic acid functionality and the large number of residual silanols on the surface of the silica support of the stationary phase.

Consumption of brown onions ( Allium cepa var. cavalier and var. destiny ) moderately modulates blood lipids, haematological and haemostatic variables in healthy pigs

Although garlic and onions have long been associated with putative cardiovascular health benefits, the effects of different commercially available onions and level of intake have not been studied. Therefore, the aim of the present study was to evaluate the potential health benefits of raw onions using the pig as a biomedical model. Twenty-five female (Large White x Landrace) pigs were used in a (2 x 2)+1 factorial experiment. Pigs were fed a standard grower diet supplemented with 100 g tallow/kg with the addition of Allium cepa var. cavalier or var. destiny at 0, 10 or 25 g/MJ digestible energy for 6 weeks. Overall, the consumption of onions resulted in significant reductions in plasma triacylglycerol; however, the reductions were most pronounced in pigs fed destiny onions (-26 %, P=0.042). Total plasma cholesterol and LDL:HDL ratios were not significantly different. Onion supplementation, regardless of the variety, resulted in dose-dependent reductions in erythrocyte counts and Hb levels, while the white blood cell concentrations, particularly lymphocytes, were increased in pigs that consumed onions. Furthermore, indices of blood clotting were largely unaffected by onion consumption. In conclusion, dietary supplementation with raw brown onions has moderate lipid-modulating and immunostimulatory properties. However, daily onion intake >25 g/MJ digestible energy could be detrimental to erythrocyte numbers.

Effects of dietary fat and conjugated linoleic acid on plasma metabolite concentrations and metabolic responses to homeostatic signals in pigs.

Sixteen female cross-bred (Large White x Landrace) pigs (initial weight 65 kg) with venous catheters were randomly allocated to four treatment groups in a factorial design. The respective factors were dietary fat (25 or 100 g/kg) and dietary conjugated linoleic acid (CLA; 0 or 10 g CLA-55/kg). Pigs were fed every 3 h (close to ad libitum digestible energy intake) for 8 d and were bled frequently. Plasma glucose and non-esterified fatty acid (NEFA) responses to insulin and adrenaline challenges were determined on day 8. Plasma concentrations of NEFA were significantly increased (10.5 and 5.4 % for low- and high-fat diets respectively, P=0.015) throughout the experiment, suggesting that there was a possible increase in fat mobilisation. The increase in lipolysis, an indicator of ss-adrenergic stimulated lipolysis, was also evident in the NEFA response to adrenaline. However, the increase in plasma triacylglycerol (11.0 and 7.1 % for low- and high-fat diets respectively, P=0.008) indicated that CLA could have reduced fat accretion via decreased adipose tissue triacylglycerol synthesis from preformed fatty acids, possibly through reduced lipoprotein lipase activity. Plasma glucose, the primary substrate for de novo lipid synthesis, and plasma insulin levels were unaffected by dietary CLA suggesting that de novo lipid synthesis was largely unaffected (P=0.24 and P=0.30 respectively). In addition, the dietary CLA had no effect upon the ability of insulin to stimulate glucose removal.

Dairy proteins and the regulation of satiety and obesity

Over the past decade there has been growing scientific evidence and public acceptance of the role that dietary protein plays in regulation of satiety, feed intake and obesity-related disorders. Dietary protein appears to suppress food intake and delay the return of hunger more than fats or carbohydrates in a manner not due to energy content alone. Also, high-protein diets support the maintenance of muscle mass when subjects reduce their energy intake, ensuring primarily adipose tissue loss. Some protein sources, particularly dairy, contain specific peptides or proteins that may elicit direct effects on satiety. The major proteins present in milk include β-lactalbumin, α-lactoglobulin, immunoglobulins, bovine serum albumin, and the various caseins. In addition, processed whey contains glycomacropeptide, which stimulates pancreatic and gastrointestinal secretion of hormones involved in satiety to a greater extent than whey alone. In the context of the literature, we show that a glycomacropeptide-rich whey protein isolate decreases feed intake and weight gain to a greater extent than a soy protein isolate in obese pigs. Also, insulin sensitivity is improved in pigs consuming high-protein diets, with these effects being independent of protein source. While, high-protein diets may decrease calcium balance and bone strength, it appears that these effects are attenuated by dairy proteins and dairy sources of calcium. These findings suggest that high-protein diets, and in particular those that contain whey proteins, may reduce hunger and food intake, thereby reducing fat deposition and improving insulin sensitivity.

Seasonal variation in the concentrations of conjugated linoleic and trans fatty acids in milk fat from commercial dairy farms is associated with pasture and grazing management and supplementary feeding practices

A study of irrigated pasture-based dairy farms that used split calving (autumn and spring) was undertaken in northern Victoria, Australia, to examine associations between nutrition, time of year and season of calving on the concentrations of isomers of trans 18 : 1 fatty acids and conjugated linoleic acids (CLA) in milk fat. Factors associated with time of year explained most of the variation, with the highest concentrations observed in spring and summer when pasture intake by herds was high. However, there was substantial variation observed between herds and time of year. The mean total CLA concentration was 9.1 mg/g milk fatty acids (range 1.1–35.4 mg/g) with the cis,trans-9,11 accounting for ~84% of the total CLA. The mean total trans 18 : 1 concentration was 60.5 mg/g milk fatty acids (range 13.6–267 mg/g) with vaccenic acid (trans-11 18 : 1) accounting for ~53% of total trans 18 : 1 fatty acids. Total CLA and vaccenic acid were highest in August–September (southern hemisphere spring) (15.1 and 76.3 mg/g milk fat) and lowest in November–March (5.6 mg/g milk fat) and May–July (9.53 mg/g milk fat), respectively. There was no association between season of calving and milk CLA or trans 18 : 1 fatty acid concentrations. Trans-10 and -11 18 : 1 fatty acids and trans/trans-CLA were negatively correlated with milk fat concentrations. Management strategies designed to increase the concentration of CLA and trans 18 : 1 fatty acids in milk fat would not need to consider the effects of season of calving or stage of lactation, but should focus on pasture availability and quality.

Optimizing conditions for DNA isolation fromPinus radiata

SummaryGenomic DNA was isolated fromin vitro Pinus radiata seedling with five DNA isolation protocols commonly used for pines. The methods described by Jobes et al. (1995) and Nelson et al. (1994) utilize sodium dodecyl sulfate, whereas those of Murray and Thompson (1980), Doyle and Doyle (1990), and Devey et al. (1996) use cetyltrimethyl ammonium bromide for cell lysis. The quality and quantity of the isolated DNA was measured and compared. Lithium chloride was found to be more effective than RNase for minimizing the amount of RNA present in the solution. Protocols described by Jobes et al. (1995) and Devey et al. (1996) yielded a large quantity of pure DNA which was suitable for restriction enzyme digestion and polymerase chain reaction amplification. With these methods, 37 to 79 μg of DNA with an A260/280 ratio between 1.7 and 1.9 was obtained from 1 g ofPinus radiata seedlings grownin vitro.