Countercurrent approach to the enrichment of Δ9c,11t-and Δ10t,12c-18:2 isomers by urea complexation

CLA refers to a group of geometrical and positional isomers of linoleic acid. CLA has been shown to have potentially beneficial effects on cancer, atherosclerosis, and body metabolism in animals. Mixtures containing equal amounts of these isomers are commonly used in many research studies because of their greater availability and lower cost relative to pure isomers. This has hindered progress in elucidating the biological properties of specific isomers and their relevance in animal and human biology. A method was developed that offers a compromise between cost and utility to make available enriched mixtures of either the Δ9c,11t- or Δ10t,12c-18:2 isomers for use in a wide range of experimental applications. A countercurrent approach was developed to separate the Δ9c,11t- and Δ10t,12c-18:2 isomers from an equal mixture of these two isomers by urea complexation. After three successive rounds of complexation using an equal amount of CLA and urea, a fraction enriched in Δ9c,11t-18:2 containing 42.5 and 17.4% of Δ9c,11t-and Δ10t,12c-18:2, respectively, was recovered. After a single round of complexation using 2.5 g urea/g CLA, a fraction enriched in Δ10t,12c-18:2 was recovered containing 29.7 and 69.1% of Δ9c,11t- and Δ10t,12c-18:2, respectively.     

Maternal supplementation with CLA decreases milk fat in humans

CLA refers to isomers of octadecadienoic acid with conjugated double bonds. The most abundant form of CLA (rumenic acid (RA): c9,t11-18∶2) is found in milk and beef fat. Further, CLA supplements containing RA and t10,c12−18∶2 are now available. Consumption of commercially produced CLA has been shown to decrease adipose accretion in growing laboratory and production animals and cause milk fat depression in cows. We tested the hypothesis that CLA supplementation would increase milk CLA concentration and decrease milk fat content in humans. Breastfeeding women (n=9) participated in this double-blind, placebo-controlled, crossover study divided into three periods: intervention l (5 d), washout (7 d), and intervention II (5 d). Women were randomized to treatment order. During each intervention period, women consumed 1.5 g of CLA supplement or placebo (olive oil) daily; during the washout period, no supplements were consumed. Milk was collected by complete breast expression on the final day of each period; milk output was estimated by 24-h weighing on the penultimate day of each intervention period. Milk RA and t10,c12−18∶2 concentrations were greater (P<0.05) during the CLA treatment period as compared to the placebo period. Milk fat content was significantly lower during the CLA treatment, as compared to the placebo treatment (P<0.05). Data indicate no effect of treatment on milk output. Therefore, it would be prudent that lactating women not consume commercially available CLA supplements at this time. This paper was published in part in Masters, N., McGuire, M.A., and McGuire, M.K. (1999) Conjugated Linoleic Acid Supplementation and Milk Fat Content in Humans, FASEB J. 13, A697.     

Contrasting effects of t10,c12- and c9,t11-conjugated linoleic acid isomers on the fatty acid profiles of mouse liver lipids

The purpose of this study was to examine the effects of two purified isomers of CLA (c9,t11-CLA and t10,c12-CLA) on the weights and FA compositions of hepatic TG, phospholipids, cholesterol esters, and FFA. Eight-week-old female mice (n=6/group) were fed either a control diet or diets supplemented with 0.5% c9,t11-CLA or t10, c12-CLA isomers for 8 wk. Weights of liver total lipids and those of individual lipid fractions. did not differ between the control and the c9,t11-CLA groups. Livers from animals fed the t10,c12-CLA diet contained four times more lipids than those of the control group; this was mainly due to an increase in the TG fractions (fivefold), but cholesterol (threefold), cholesterol esters (threefold), and FFA (twofold) were also significantly increased. Although c9,t11-CLA did not significantly alter the weights of liver lipids when compared with the control group, its intake was associated with significant reductions in the weight percentage (wt% of total FAME) of 18∶1n−9 and 18∶1n−7 in the TG fraction and with significant increases in the weight percentage of 18∶2n−6 in the TG, cholesterol ester, and phospholipid fractions. on the other hand, t10,c12-CLA intake was linked with a significant increase in the weight percentage of 18∶1n−9 and a decrease in that of 18∶2n−6 in all lipid fractions. These changes may be the result of alterations in the activity of Δ9-desaturase (stearoyl CoA desaturase) and the enzymes involved in the metabolism of 18∶2n−6. Thus, the two isomers differed not only in their effects on the weights of total liver lipids and lipid fractions but also on the FA profile of the lipid fractions.     

Dietary CLA Combined with Palm Oil or Ovine Fat Differentially Influences Fatty Acid Deposition in Tissues of Obese Zucker Rats

The effect of dietary conjugated linoleic acid (CLA) supplementation in combination with fat from vegetable versus animal origin on the fatty acid deposition, including that of individual 18:1 and 18:2 (conjugated and non-conjugated) isomers, in the liver and muscle of obese rats was investigated. For this purpose, 32 male Zucker rats were randomly assigned to one of four diets containing palm oil or ovine fat, supplemented or not with 1% of 1:1 cis(c)9,trans(t)11 and t10,c12 CLA isomers mixture. Total fatty acid content decreased in the liver and muscle of CLA-fed rats. In the liver, CLA increased saturated fatty acids (SFA) in 11.9% and decreased monounsaturated fatty acids (MUFA) in 6.5%. n-3 Polyunsaturated fatty acids (PUFA) relative proportions were increased in 30.6% by CLA when supplemented to the ovine fat diet. In the muscle, CLA did not affect SFA but decreased MUFA and PUFA percentages. The estimation of Δ9-indices 16 and 18 suggested that CLA inhibited the stearoyl-CoA desaturase activity in the liver (a decrease of 13–38%), in particular when supplemented to the ovine fat diet. Concerning CLA supplementation, the t10,c12 isomer percentage was 60–80% higher in the muscle than in the liver. It is of relevance that rats fed ovine fat, containing bio-formed CLA, had more c9,t11 CLA isomer deposited in both tissues than rats fed palm oil plus synthetic CLA. These results highlight the importance to further clarify the biological effects of consuming foods naturally enriched in CLA, alternatively to CLA dietary supplementation.     

Maternal dietary alpine butter intake affects human milk: Fatty acids and conjugated linoleic acid isomers

Consumption of CLA by lactating women affects the composition of their milk, but the pattern of the different CLA isomers is still unknown. We determined the effects of short maternal supplementation with CLA-rich Alpine butter on the occurrence of FA and CLA isomers in human milk. In an open randomized controlled study with a two-period cross-over design, milk FA and CLA isomer concentrations were measured on postpartum days >-20 in two parallel groups of lactating women before, during, and after consumption of defined quantities of Alpine butter or margarine with comparable fat content (10 d of butter followed by 10 d of margarine for one group, and vice versa in the other). In the 16 women who completed the study (8/group), Alpine butter supplementation, increased the C₁₆ and C₁₈ FA, the sum of saturated FA, the 18∶1 trans FA, and the trans FA with CLA. The CLA isomer 18∶2 c9, t11 increased by 19.7%. Significant increases were also found for the isomers t9,t111, t7,c9,t11,c13, and t8,c10 18∶2. The remaining nine of the total 14 detectable isomers showed no changes, and concentrations were <5 mg/100g fat. A breastfeeding mother can therefore modulate the FA/CLA supply of her child by consuming Alpine butter. Further studies will show whether human milk containing this FA and CLA isomer pattern acts as a functional food for newborns.     

Effect of dietary conjugated linoleic acid (CLA) on metabolism of isotope-labeled oleic,linoleic, and CLA isomers in women

The purpose of this study was to investigate the effect of dietary CLA on accretion of 9c-18∶1, 9c, 12c-18∶2, 10t, 12c-18∶2, and 9c, 11t-18∶2 and conversion of these FA to their desaturated, elongated, and chain-shortened metabolites. The subjects were six healthy adult women who had consumed normal diets supplemented with 6 g/d of sunflower oil or 3.9 g/d of CLA for 63 d. A mixture of 10t, 12c-18∶2-d ₄, 9c, 11t-18∶2-d ₆, 9c-18∶1-d ₈, and 9c, 12c-18∶2-d ₂, as their ethyl esters, was fed to each subject, and nine blood samples were drawn over a 48-h period. The results show that dietary CLA supplementation had no effect on the metabolism of the deuterium-labeled FA. These metabolic results were consistent with the general lack of a CLA diet effect on a variety of physiological responses previously reported for these women. The ²H-CLA isomers were metabolically different. The relative percent differences between the accumulation of 9c, 11t-18∶2-d ₆ and 10t, 12c-18∶2-d ₄ in plasma lipid classes ranged from 9 to 73%. The largest differences were a fourfold higher incorporation of 10t, 12c-18∶2-d ₄ than 9c, 11t-18∶2-d ₆ in 1-acyl PC and a two- to threefold higher incorporation of 9c, 11t-18∶2-d ₆ than 10t, 12c-18∶2-d ₄ in cholesterol esters. Compared to 9c-18∶1-d ₈ and 9c, 12c-18∶2-d ₂, the 10t, 12c-18∶2-d ₄ and 9c, 11t-18∶2-d ₆ isomers were 20–25% less well absorbed. Relative to 9c-18∶1, incorporation of the CLA isomers into 2-acyl PC and cholesterol ester was 39–84% lower and incorporation of 10t, 12c-18∶2 was 50% higher in 1-acyl PC. This pattern of selective incorporation and discrimination is similar to the pattern generally observed for trans and cis 18∶1 positional isomers. Elongated and desaturated CLA metabolites were detected. The concentration of 6c, 10t, 12c-18∶3-d ₄ in plasma TG was equal to 6.8% of the 10t, 12c-18∶2-d ₄ present, and TG was the only lipid fraction that contained a CLA metabolite present at concentrations sufficient for reliable quantification. In conclusion, no effect of dietary CLA was observed, absorption of CLA was less than that of 9c-18∶1, CLA positional isomers were metabolically different, and conversion of CLA isomers to desaturated and elongated metabolites was low.     

Supplemental Conjugated Linoleic Acid Consumption Does Not Influence Milk Macronutrient Contents in all Healthy Lactating Women

The term “conjugated linoleic acid” (CLA) refers to a group of positional and geometric isomers that are derived from linoleic acid and are found primarily in meat and milk products from ruminant animals. Due to the array of putative benefits associated with various forms of CLA, there has been recent interest in supplementing human diets with these fatty acids especially when weight loss is desired. However, in many animal models, CLA has been shown to decrease milk fat production. There is some concern, therefore, that maternal CLA supplementation during lactation might inadvertently decrease nutrient supply to the nursing infant. However, there is only limited research on the effect of CLA consumption on milk fat content in women. Based on previously published work from our laboratory, we hypothesized that CLA supplementation would reduce the milk fat percentage in lactating women in a dose-dependent manner. Breastfeeding women (n = 12) were assigned randomly to treatments of 4 g/day safflower oil (SFO), 2 g/day CLA plus 2 g/day SFO, or 4 g/day CLA in a double blind, 3 × 3 Latin square design. Conjugated linoleic acid supplements contained approximately equal amounts of cis9,trans11–18:2 and trans10,cis12–18:2; the two most common isoforms of CLA. Milk was collected by complete breast expression on the last day (day 5) of each intervention period and analyzed for macronutrient and fatty acid composition. On day 4 of each intervention period, infant milk consumption was estimated by 24 h weighing of the infant. Washout periods were 9 days in length. We observed a dose-dependent increase in the concentrations of cis9,trans11–18:2 and trans10,cis12–18:2 in the milk fat. However, we detected neither a change in overall macronutrient composition nor infant milk consumption. These data do not support those obtained from animal models or our previous human work suggesting that consumption of CLA mixtures necessarily reduces milk fat. It is possible that either (1) the interpretation of our previously published data should be reevaluated, and/or (2) there are important intra- and inter-species differences in this regard.     

High Dose trans-10,cis-12 CLA Increases Lean Body Mass in Hamsters, but Elevates Levels of Plasma Lipids and Liver Enzyme Biomarkers

The current study examined the efficacy of graded doses of c9,t11 and t10,c12 CLA isomers on body composition, energy expenditure, hepatic and serum lipid liver biomarkers in hamsters. Animals (n = 105) were randomized to seven treatments (control, 1, 2, 3% of c9,t11; 1, 2, 3% of t10,c12) for 28 days. After 28 days treatment, 1–3% of t10,c12 lowered (p < 0.05) body fat mass compared to the control group. The 1–3% t10,c12 and 3% c9,t11 fed groups showed higher (p < 0.05) lean mass compared to other groups. We observed unfavorable changes in plasma total cholesterol and non-HDL cholesterol levels in animals fed with 3% t10,c12 CLA isomers. The 2%, 3% t10,c12 groups presented elevated (p < 0.05) ALT levels. The present data suggest that a diet enriched with more than 2% t10, c12 led to liver malfunction and poses unfavorable changes on plasma lipid profiles. The 1% t10,c12 CLA lowered (p < 0.05) body fat mass and increased (p < 0.05) lean body mass. The c9,t11 CLA has less potent actions than t10,c12 CLA. We conclude that the actions of CLA on energy and lipid metabolism are form and dose dependent in the hamster model.     

Analysis of conjugated linoleic acid isomers and content in french cheeses

Conjugated linoleic acid (CLA) occurs in food as a result of microbial enzymatic reactions, free radical-type oxidation, and heat treatment. CLA is found in animal products, such as meat and dairy products, especially in cheeses. The CLA composition of 12 different French cheeses was determined by a combination of different analytical methods: reversed-phase high-performance liquid chromatography (RP-HPLC), gas chromatography-mass spectrometry (GC-MS), GC-Fourier transform infrared (GC-FTIR), and silver nitrate thin-layer chromatography (AgNO₃-TLC). New isomers (Δ8,10- and Δ11,13-octadecadienoic acids with all possible cis and trans configurations) that co-eluted with previously identified isomers (Δ9c,11t-; Δ9t,11c-; Δ10c,12t-; Δ10t,12c-; Δ11c,13c-; Δ9c,11c-; Δ10c,12c-; Δ9t,11t-; Δ10t12t-octadecadienoic acids) were detected. Δ9c,11t-Octadecadienoic acid was the major CLA isomer in these cheeses. All isomers were present in each product, whatever the production process. However, CLA content in the cheeses varied from 5.3 to 15.80 mg/g of cheese fat, which depended primarily on the origin of the milk (season, geography) and somewhat on the production process.     

Effect of <Emphasis Type="Italic">β</Emphasis>-Cyclodextrin on <Emphasis Type="Italic">trans</Emphasis> Fats,CLA, PUFA and Phospholipids of Milk Fat

The aim of this study was to evaluate the effect of β-cyclodextrin (β-CD) on trans C18:1 fatty acid isomers, conjugated linoleic acid (CLA), polyunsaturated fatty acids (PUFA) and phospholipids in pasteurized milk. The individual trans C18:1 isomers were not significantly affected by the β-CD. trans-11 C18:1 (vaccenic acid) was found to be the major isomer (1.31 ± 0.12%) followed by trans-15 C18:1 (0.35 ± 0.06%). Individual trans linoleic acids did not show differences from the effect of β-CD, representing the high amount of the isomer trans-11 cis-15 C18:2 (0.433 ± 0.087%). The main CLA isomer cis-9 trans-11 C18:2 (rumenic acid) did not show differences between the control milk (0.672 ± 0.080%) and β-CD milk (0.663 ± 0.074%). PUFA and omega-3 and -6 fatty acids were not also significant by the effect of β-CD. Total phospholipids were not significantly affected by effect of the β-CD (0.023 ± 0.001% vs. 0.022 ± 0.001%). β-CD is a effective oligosaccharide for cholesterol removal from pasteurized milk and does not significantly affect the lipid components of the milk fat.     

<Emphasis Type="Italic">t</Emphasis>10,<Emphasis Type="Italic">c</Emphasis>12-18:2-Induced Milk Fat Depression is Less Pronounced in Cows Fed High-Concentrate Diets

In intensively reared dairy cows, milk fat secretion is reduced in response to high-concentrate diets and it is often referred to as the “milk fat depression” (MFD) syndrome. Some trans fatty acid (FA) isomers produced in the rumen of the cows, including t10,c12-18:2, are known for their inhibitory effect on mammary lipogenesis. To study whether this effect depends on the basal diet, duodenal infusions of t10,c12-18:2 were performed on cows fed four different diets (a factorial arrangement of forage:concentrate ratio and linseed oil supplementation). The overall response obtained with t10,c12-18:2 infusion was consistent with previous studies: a decrease in milk fat content and yield without significant variations in milk yield. Mean transfer efficiency of infused t10,c12-18:2 was 19.6%. However, the decrease in milk fat and FA yields (both de novo synthesis and preformed long-chain FA) was less pronounced in cows fed high-concentrate diets (−27% of the initial level), compared with cows fed low-concentrate diets (−42% of initial level). This difference was independent of dietary oil supplementation and milk FA yield before infusion. Results pertaining to effects of dietary forage:concentrate ratio were confirmed by statistical meta-analysis of data from previously published t10,c12-18:2 infusion experiments. This study shows that in cows fed MFD diets the mammary gland becomes more resistant to or experiences a lower response potential to further inhibition of lipogenesis and/or delta-9 desaturation of FA.     

Effects of margarine and butter consumption on distribution of trans-18∶1 fatty acid isomers and conjugated linoleic acid in major serum lipid classes in lactating women

Trans FA (TFA) have at least one trans double bond and comprise several isomers and types, including many of the CLA (e.g., c9, t11–18∶2 CLA). Some TFA may have adverse effects (e.g., cardiovascular disease), whereas some are though to have beneficial effects (e.g., anticarcinogenicity). The presence of TFA in human tissues and fluids is related to dietary intake, although this relationship is not completely understood—especially in regard to serum lipid fractions. This study was conducted as part of an investigation designed to test the influence of butter (B), “low TFA” margarine (LT), and regular margarine (RM) on milk fat content. Here we tested the secondary hypothesis that consumption of B, LT, and RM by lactating women would result in differential distribution of TFA and CLA in major serum lipid classes. Breastfeeding women (n=11) participated in this randomized Latinsquare study consisting of five periods: intervention I (5 d), washout I (7 d), intervention II (5 d), washout II (7 d), and intervention III (5 d). Extracted serum lipid was separated into cholesterol ester (CE), TAG, and phospholipid (PL) fractions and analyzed for total and isomeric TFA and CLA concentrations. Data indicate that TAG consistently contained the highest concentration of total t-18∶1. No interaction between treatment and fraction was found for any of the t-18∶1 isomers identified. Absolute concentration of each t-18∶1 isomer was greatest during the RM period, regardless of fraction. On a relative basis, concentrations of t10–18∶1 and t12–18∶1 were most responsive to treatment in the CE fraction. The concentration of c9, t11–18∶2 CLA was highest in the TAG fraction and lowest in the PL fraction, regardless of treatment. In summary, these results indicate (i) that there is a differential distribution of some isomeric TFA and CLA among human serum lipid fractions and (ii) that dietary TFA intake influences absolute and relative concentrations of some of the isomers in selected fractions.     

Alterations in CPT‐1 mRNA and fatty acid profile in hepatic cell lines in response to treatment with t10,c12‐ or c9,t11‐conjugated linoleic acid

Trans10,cis12‐conjugated linoleic acid (t10,c12‐CLA) increases liver weights and hepatic lipids in mice. The purpose of this study was to determine the effects of CLA isomers (t10,c12‐CLA or c9,t11‐CLA) and carnitine palmitoyl transferase‐1 (CPT‐1) inhibitors (etomoxir or hemipalmitoylcarnitinium) on CPT‐1 mRNA, fatty acid profile, and cholesterol synthesis in AML‐12 and HepG2 cells. t10,c12‐CLA was incorporated to a greater extent in both cell lines than c9,t11‐CLA. In addition, t10,c12‐CLA increased the free cholesterol content of AML‐12 and HepG2 cells four‐ and fivefold, respectively. Cells incubated with medium containing CPT‐1 inhibitors or t10,c12‐CLA had higher levels of mRNA for CPT‐1 in both cell lines, indicating an increased fatty acid oxidation in hepatic cell lines due to t10,c12‐CLA. Following treatment withdrawal, percentages of c9,t11‐CLA or t10,c12‐CLA remained elevated in cells initially treated with c9,t11‐CLA or t10,c12‐CLA, suggesting a potential for carryover effects of the CLA isomers. The results presented here demonstrate a potential role for t10,c12‐CLA in the modulation of hepatic fatty acid oxidation and cholesterol synthesis.     

Effect of temperature on linolenic acid loss and 18:3 Δ9-cis, Δ12-cis, Δ15-trans formation in soybean oil

Oil was extracted from soybeans, degummed, alkalirefined and bleached. The oil was heated at 160, 180, 200, 220 and 240°C for up to 156 h. Fatty acid methyl esters were prepared by boron trifluoride-catalyzed transesterification. Gas-liquid chromatography with a cyanopropyl CPSil88 column was used to separate and quantitate fatty acid methyl esters. Fatty acids were identified by comparison of retention times with standards and were calculated as area % and mg/g oil based on 17:0 internal standard. The rates of 18:3ω3 loss and 18:3 Δ9-cis, Δ12-cis, Δ15-trans (18:3c,c,t) formation were determined, and the activation energies were calculated from Arrhenius plots. Freshly prepared soy oil had 10.1% 18:3ω3 and no detectable 18:3c,c,t. Loss of 18:3ω3 followed apparent first-order kinetics. The first-order rate constants ranged from .0018±.00014 min⁻¹ at 160°C to .083±.0033 min⁻¹ at 240°C. The formation of 18:3c,c,t did not follow simple kinetics, and initial rates were estimated. The initial rates (mg per g oil per h) of 18:3c,c,t formation ranged from 0.0031±0.0006 at 160°C to 2.4±.24 at 240°C. The Arrhenius activation energy for 18:3ω3 loss was 82.1±7.2 kJ mol⁻¹. The apparent Arrhenius activation energy for 18:3c,c,t formation was 146.0±13.0 kJ mol⁻¹. The results indicate that small differences in heating temperature can have a profound affect on 18:3c,c,t formation. Selection of appropriate deodorization conditions could limit the amount of 18:3c,c,t produced.     

Comparing Subcutaneous Adipose Tissue in Beef and Muskox with Emphasis on <Emphasis Type="Italic">trans</Emphasis> 18:1 and Conjugated Linoleic Acids

Muskox (Ovibos moschatus) are ruminant animals native to the far north and little is known about their fatty acid composition. Subcutaneous adipose tissue (backfat) from 16 wild muskox was analyzed and compared to backfat from 16 barley fed beef cattle. Muskox backfat composition differed substantially from beef and the most striking difference was a high content of 18:0 (26.8 vs. 9.77%). This was accompanied by higher levels of most other saturated fatty acids except beef had more 16:0. Muskox backfat also had a lower level of cis-18:1 and this was related to a lower expression of steroyl-CoA desaturase mRNA. Beef backfat had a higher level of total trans-18:1 (4.25 vs. 2.67%). The most prominent trans-18:1 isomers in beef backfat were 10t-18:1 (2.13%) and 11t-18:1 (0.77%) whereas the most prominent isomers in muskox backfat were 11t-18:1 (1.41%), 13t/14t- (0.27%) and 16t-18:1 (0.23%). The total conjugated linoleic acid (CLA) content was higher in beef backfat than muskox (0.67 vs. 0.50%) with 9c,11t-18:2 as the most abundant CLA isomer. The second most abundant CLA isomer in beef backfat was 7t,9c-18:2 (0.10%) whereas in muskox it was 11t13c-18:2 (0.04%). Muskox backfat had a higher content of 18:3n-3 and its elongation and desaturation products 20:5n-3, 22:5n-3 and 22:6n-3 and a lower n-6/n-3 ratio. Overall, the high forage diet of muskox seemed to produce a healthier fatty acid profile and highlighted the need to develop feeding strategies for intensively raising beef that will not negatively impacting fatty acid composition.     

Preparation of conjugated linoleic acid from safflower oil

Synthetically prepared mixtures of conjugated linoleic acid (CLA) are widely used in animal and cell culture studies to investigate the potential effects of the Δ9c, 11t-18:2 isomer found in food products from ruminant animals. Alkali isomerization of linoleic acid is a common method used in the synthesis of a mixture of CLA isomers containing predominantly the Δ9c, 11t-18:2 and Δ10t, 12c-18:2 isomers. Some biological activity might also be mediated by the Δ10t, 12c-18:2 isomer. Currently few published methodologies exist describing procedures for the enrichment of these two isomers. A method is described herein to take advantage of an inexpensive oil, safflower oil, for use in synthesis of CLA and a procedure to enrich the Δ10t, 12c-18:2 isomer.     

Total Lipids of Sarda Sheep Meat that Include the Fatty Acid and Alkenyl Composition and the CLA and <Emphasis Type="Italic">Trans</Emphasis>-18:1 Isomers

The total lipids of the longissimus dorsi muscle were analyzed from commercial adult Sarda sheep in Sardina taken from local abattoirs, and in the subsequent year from three local farms in the Sassari region that provided some information on the amount and type of supplements fed to the pasture-fed sheep. The complete lipid analysis of sheep meat included the fatty acids from O-acyl and N-acyl lipids, including the trans- and conjugated linoleic acid (CLA) isomers and the alk-1-enyl ethers from the plasmalogenic lipids. This analysis required the use of a combination of acid- and base-catalyzed methylation procedures, the former to quantitate the O-acyl, N-acyl and alkenyl ethers, and the latter to determine the content of CLA isomers and their metabolites. A combination of gas chromatographic and silver-ion separation techniques was necessary to quantitate all of the meat lipid constituents, which included a prior separation of the trans-octadecenoic acids (18:1) and a separation of fatty acid methyl esters and the dimethylacetals (DMAs) from the acyl and alk-1-enyl ethers, respectively. The alk-1-enyl moieties of the DMAs were analyzed as their stable cyclic acetals. In general, about half of the meat lipids were triacylglycerols, even though excess fat was trimmed from the meat. The higher fat content in the meat appears to be related to the older age of these animals. The variation in the trans-18:1 and CLA isomer profiles of the Sarda sheep obtained from the abattoirs was much greater than in the profiles from the sheep from the three selected farms. Higher levels of 10t-18:1, 7t9c-18:2, 9t11c-18:2 and 10t12c-18:2 were observed in the commercial sheep meat, which reflected the poorer quality diets of these sheep compared to those from the three farms, which consistently showed higher levels of 11t-18:1, 9c11t-18:2 and 11t13c-18:2. In the second study, sheep were provided with supplements during the spring and summer grazing season, which contributed to higher levels of 11t-18:1 and 9c11t-18:2. The farm that provided a small amount of supplements during the spring had the better lipid profile at both time periods. The polyunsaturated fatty acid (PUFA) content was higher in the meat from Sarda sheep from the three farms than in the meat from those sheep obtained from commercial slaughter operations. The plasmalogenic lipid content ranged from 2 to 3% of total lipids, the alk-1-enyl ethers consisted mainly of saturated and monounsaturated moieties, and the trans-18:1 profile was similar to that of the FA. The n-6 (6–8%) and n-3 PUFA (2–3%) contents, the n-6/n-3 ratio (3:1), as well as the saturated fatty acid (SFA) content (42–45%) and the SFA to PUFA ratio (4:1 to 5:1) of the Sarda sheep from the three farms were comparable to sheep meat lipids found in similar commercial operations in Europe. Inclusion of small amounts of supplements for the grazing Sarda sheep resulted in improved quality of sheep meat lipids.     

β-Oxidation of conjugated linoleic acid isomers and linoleic acid in rats

To assess the oxidative metabolism of conjugated linoleic acid (CLA) isomers, rats were force-fed 1.5–2.6 MBq of [1-¹⁴C]-linoleic acid (9c,12c-18∶2),-rumenic acid (9c,11t-18∶2), or-10trans, 12cis-18∶2 (10t, 12c-18∶2), and ¹⁴CO₂ production was monitored for 24 h. The animals were then necropsied and the radioactivity determined in different tissues. Both CLA isomers were oxidized significantly more than linoleic acid. Moreover, less radioactivity was recovered in most tissues after CLA intake than after linoleic acid intake. The substantial oxidation of CLA isomers must be considered when assessing the putative health benefits of CLA supplements.     

Fatty Acid Desaturation and Elongation Reactions of <Emphasis Type="Italic">Trichoderma</Emphasis> sp. 1-OH-2-3

The fatty acid desaturation and elongation reactions catalyzed by Trichoderma sp. 1-OH-2-3 were investigated. This strain converted palmitic acid (16:0) mainly to stearic acid (18:0), and further to oleic acid (c9-18:1), linoleic acid (c9,c12-18:2), and α-linolenic acid (c9,c12,c15-18:3) through elongation, and Δ9, Δ12, and Δ15 desaturation reactions, respectively. Palmitoleic acid (c9-16:1) and cis-9,cis-12-hexadecadienoic acid were also produced from 16:0 by the strain. This strain converted n-tridecanoic acid (13:0) to cis-9-heptadecenoic acid and further to cis-9,cis-12-heptadecadienoic acid through elongation, and Δ9 and Δ12 desaturation reactions, respectively. trans-Vaccenic acid (t11-18:1) and trans-12-octadecenoic acid (t12-18:1) were desaturated by the strain through Δ9 desaturation. The products derived from t11-18:1 were identified as the conjugated linoleic acids (CLAs) of cis-9,trans-11-octadecadienoic acid and trans-9,trans-11-octadecadienoic acid. The product derived from t12-18:1 was identified as cis-9,trans-12-octadecadienoic acid. cis-6,cis-9-Octadecadienoic acid was desaturated to cis-6,cis-9,cis-12-octadecatrienoic acid by this strain through Δ12 desaturation. The broad substrate specificity of the elongation, and Δ9 and Δ12 desaturation reactions of the strain is useful for fatty acid biotransformation.     

Effects of the individual isomers cis-9,trans-11 vs. trans-10,cis-12 of conjugated linoleic acid (CLA) on inflammation parameters in moderately overweight subjects with LDL-phenotype B

Immune-modulating effects of CLA have been reported in animals, but results are inconsistent. In humans, CLA has shown no effects or only minor effects on immune function. The objective of this study was to evaluate the immune-modulating effects of 3 g cis-9,trans-11 (c9,t11) vs. trans-10,cis-12 (t10,c12) CLA isomers in a population with a high risk of coronary heart disease characterized by moderate overweight (body-mass index, 25–32.5 kg/m²) in combination with LDL-phenotype B (≥35% small LDL cholesterol, density≥1.040 g/mL). After a run-in period of 1 wk, 42 men and women were randomly allocated to the c9,t11 CLA group, the t10,c12 CLA group, or the placebo group. Effects of 13 wk of consumption of 3 g of CLA isomers on cytokine production by ex vivo lipopolysaccharide (LPS)-stimulated peripheral blood mononuclear cells (PBMC) and whole blood, and on plasma C-remononuclear protein (CRP) concentrations were evaluated. To generate hypotheses for future studies, protein expression patterns of 42 cytokines, chemokines, and growth factors were evaluated with an antibody array in pooled, nonstimulated, fasting plasma samples. LPS induced interleukin (IL)-6, IL-8, and tumor necrosis factor-α production by PBMC, and whole blood as well as plasma CRP concentrations were not significantly changed by the c9,t11, and the t10,c12 CLA isomers. The cytokine expression profile in nonstimulated plasma suggested that both CLA isomers induced a specific inflammatory signature, in which the c9,t11 CLA group showed more activity in terms of numbers of proteins regulated. We conclude that daily consumption of 3 g of c9,t11 or t10,c12 CLA isomer did not affect LPS-stimulated cytokine production by PBMC or whole blood and plasma CRP levels. Inflammatory signatures in fasting, nonstimulated plasma as determined by an antibody array may indicate enhanced immune function by both CLA isomers.