Modulation of prostaglandin H synthase activity by conjugated linoleic acid (CLA) and specific CLA isomers

Conjugated linoleic acid (CLA) has been shown to inhibit tumorigenesis in animal models and is cytostatic to numerous cell lines in vitro. However, the mechanism of action is unknown. In the current study, we determined the effects of CLA and specific isomers of CLA on the rate of oxygenation of arachidonic acid by prostaglandin H synthase (PGHS) in ram seminal vesicle microsomes. The enzyme was incubated with 0.1 to 100 μM CLA or specific isomers of CLA for 2 min prior to the addition of 44 to 176 μM arachidonate. The isomers tested were 9(E),11(E) CLA; 9(Z),11(E) CLA; 9(Z),11(Z) CLA, and 10(E),12(Z) CLA. For a positive inhibitor control, flurbiprofen was used at 0.75 to 2.50 μM. Enzyme activity was assessed by measuring the rate of oxygen consumption. Inclusion of CLA or specific isomers of CLA in the incubation mixtures inhibits PGHS. The efficacy differs for each isomer, with the 9(Z),11(E) CLA isomer being the most effective and the 9(Z),11(Z) CLA isomer being the least effective inhibitor among the four CLA isomers tested. The K _i values obtained by Dixon replots range from 18.7 μM for the most effective isomer, 9(Z),11(Z) CLA, to 105.3 μM for the least effective isomer, 9(Z),11(Z) CLA. The K _i value for flurbiprofen with ram seminal vesicle microsomes was 0.33 μM. As the concentration of arachidonate was increased, the CLA-dependent inhibition of PGHS decreased, suggesting competitive inhibition. The results of this study demonstrate the potential of CLA and specific isomers of CLA to modulate prostaglandin biosynthesis.     

Trans-geometrical and positional isomers of linoleic acid including conjugated linoleic acid (CLA) in German milk and vegetable fats

A representative number of different milk fats based on a wide range of feeding and lactation conditions as well as 123 German margarines, shortenings, cooking and dietetic fats were analyzed for a variety of trans-C18:2 isomers (exhibiting at least one trans double bond) by means of gas chromatography on a 100m Sil 88 capillary column. In milk fats contents of trans Δ9,trans Δ12,cis Δ9,trans Δ13 (+ trans Δ8,cis Δ12),trans Δ8,cis Δ13,cis Δ9,trans Δ12,trans Δ9,cis Δ12 and trans Δ11,cis Δ15 amounted to 0.09%, 0.11%, 0.11%, 0.10%, 0.07% and 0.33% on average and the content of total trans-C18:2 isomers (without cis Δ9, trans Δ11) was 0.99% in the mean. The content of conjugated linoleic acid cis Δ9,trans Δ11 amounted to 0.81% on average in 238 milk fats. In margarine among others the isomers trans Δ9,trans Δ12,cis Δ9,trans Δ13 (+ trans Δ8,cis Δ12), cis Δ9,trans Δ12 and trans Δ9,cis Δ12 were determined and quantified to 0.03%, 0.04%, 0.29% and 0.23% on average. The mean total content of trans-C18:2 isomers in margarines was 0.61%. Moreover, for all trans-C18:2 isomers the frequency distributions as well as the correlation coefficients towards the trans-C18:1 isomers trans Δ6 to trans Δ16 were derived.     

Conjugated linoleic acid (CLA) isomers: formation,analysis, amounts in foods,and dietary intake

The analysis, content, and daily intake of conjugated linoleic acid isomers (CLA) are presented in the following review. Modern analytical techniques such as capillary gas chromatography (GC), silverion high performance liquid chromatography (Ag⁺-HPLC) combined with different detection methods (flame ionisation, mass spectroscopic, ultra violet) are mandatory for the unequivocal determination of geometric and positional CLA isomers. An overview is given on the CLA contents in 139 German foods, e. g. milk/dairy products, meat/meat products, edible oils, margarines, fish, and deep fried products. The dietary intake for men and women is estimated using consumption data.     

Analytical techniques for conjugated linoleic acid (CLA) analysis

Interest in conjugated linoleic acid (CLA) research has increased significantly in recent years because various CLA isomers have been reported to exhibit different beneficial physiological effects in laboratory animals. These findings are detailed in the first book on this subject entitled Advances in CLA Research, Volume 1, edited by M. P. Yurawecz, M. M. Mossoba, J. K. G. Kramer, M. W. Pariza, and G. J. Nelson, and published by the American Oil Chemists Society, Champaign, Illinois, USA, in 1999. Various mixtures of CLA isomers are sold as dietary supplements today, but the biological activities observed in animal models have yet to be demonstrated in humans. Hurdles to achieving this goal include the commercial unavailability of many individual CLA isomers, and the challenge of isolating, separating and confirming the identity of low levels of CLA isomers and their metabolites in complex chemical and/or biological matrices. Therefore, there has been a critical need for analytical methodologies that can meet these difficult objectives. This dossier is about a collection of useful analytical techniques and procedures some of which are well known, while others have only recently been developed and successfully applied to CLA research. These include GC, Ag⁺‐HPLC, RP‐HPLC, GC‐MS, GC‐IR, and NMR.     

Effect of dietary docosahexaenoic acid on desaturation and uptake in vivo of isotope-labeled oleic, linoleic, and linolenic acids by male subjects

The effect of dietary docosahexaenoic acid (22∶6n−3, DHA) on the metabolism of oleic, linoleic, and linolenic acids was investigated in male subjects (n=6) confined to a metabolic unit and fed diets containing 6.5 or <0.1 g/d of DHA for 90 d. At the end of the diet period, the subjects were fed a mixture of deuterated triglycerides containing 18∶1n−9[d6], 18∶2n−6[d2], and 18∶3n−3[d4]. Blood samples were drawn at 0, 2, 4, 6, 8, 12, 24, 48, and 72 h. Methyl esters of plasma total lipids, triglycerides, phospholipids, and cholesterol esters were analyzed by gas chromatography-mass spectrometry. Chylomicron triglyceride results show that the deuterated fatty acids were equally well absorbed and diet did not influence absorption. Compared to the low-DHA diet (LO-DHA), clearance of the labeled fatty acids from chylomicron triglycerides was modestly higher for subjects fed the high DHA diet (HI-DHA). DHA supplementation significantly reduced the concentrations of most n-6[d2] and n-3[d4] long-chain fatty acid (LCFA) metabolites in plasma lipids. Accumulation of 20∶5n−3[d4] and 22∶6n−3[d4] was depressed by 76 and 88%, respectively. Accumulations of 20∶3n−6[d2] and 20∶4n−6[d2] were both decreased by 72%. No effect of diet was observed on acyltransferase selectivity or on uptake and clearance of 18∶1n−9[d6], 18∶2n−6[d2], and 18∶3n−3[d4]. The results indicate that accumulation of n−3 LCFA metabolites synthesized from 18∶3n−3 in typical U.S. diets would be reduced from about 120 to 30 mg/d by supplementation with 6.5 g/d of DHA. Accumulation of n−6 LCFA metabolites synthesized from 18∶2n−6 in U.S. diets is estimated to be reduced from about 800 to 180 mg/d. This decrease is two to three times the amount of n−6 LCFA in a typical U.S. diet. These results support the hypothesis that health benefits associated with DHA supplementation are the combined result of reduced accretion of n−6 LCFA metabolites and an increase in n−3 LCFA levels in tissue lipids.     

Safety of conjugated linoleic acid (CLA) in overweight or obese human volunteers

The main objective of the study was to investigate the safety of conjugated linoleic acid (CLA) in healthy volunteers. The effect of CLA on body composition was also investigated. The trial design was a randomized, double‐blind placebo controlled study including 60 overweight or obese volunteers (body mass index (BMI) 27.5—39.0 kg/m²). The subjects were divided into two groups receiving 3.4 g CLA or placebo (4.5 g olive oil) daily for 12 weeks. The safety was evaluated by analysis of blood parameters and by clinical examinations at baseline and week 12. Vital signs and adverse events were registered at baseline, week 6, and week 12. Bio Impedance Assessment was applied for body composition measurements. 55 subjects completed the study. Adverse events occurred in 10% of the subjects. No difference in adverse events or other safety parameters was found between the treatment groups. Small changes in the laboratory safety data were not regarded as clinically significant. Moreover, no clinically significant changes in vital signs were observed in any of the groups. In the CLA group, mean weight was reduced by 1.1 kg (paired t‐test p = 0.005), while mean BMI was reduced by 0.4 kg/m²(p = 0.007). However, the overall treatment effect of CLA on body weight and BMI was not significant. There were no differences found between the groups with regard to efficacy parameters. The results indicate that CLA in the given dose is a safe substance in healthy populations with regard to the safety parameters investigated.     

Lipase-catalyzed fractionation of conjugated linoleic acid isomers

The abilities of lipases produced by the fungus Geotrichum candidum to selectively fractionate mixtures of conjugated linoleic acid (CLA) isomers during esterification of mixed CLA free fatty acids and during hydrolysis of mixed CLA methyl esters were examined. The enzymes were highly selective for cis-9,trans-11–18∶2. A commercial CLA methyl ester preparation, containing at least 12 species representing four positional CLA isomers, was incubated in aqueous solution with either a commercial G. candidum lipase preparation (Amano GC-4) or lipase produced from a cloned high-selectivity G. candidum lipase B gene. In both instances selective hydrolysis of the cis-9,trans-11–18∶2 methyl ester occurred, with negligible hydrolysis of other CLA isomers. The content of cis-9,trans-11–18∶2 in the resulting free fatty acid fraction was between 94 (lipase B reaction) and 77% (GC-4 reaction). The commercial CLA mixture contained only trace amounts of trans-9,cis-11–18∶2, and there was no evidence that this isomer was hydrolyzed by the enzyme. Analogous results were obtained with these enzymes in the esterification in organic solvent of a commercial preparation of CLA free fatty acids containing at least 12 CLA isomers. In this case, G. candidum lipase B generated a methyl ester fraction that contained >98% cis-9,trans-11–18∶2. Geotrichum candidum lipases B and GC-4 also demonstrated high selectivity in the esterification of CLA with ethanol, generating ethyl ester fractions containing 96 and 80%, respectively, of the cis-9,trans-11 isomer. In a second set of experiments, CLA synthesized from pure linoleic acid, composed essentially of two isomers, cis-9,trans-11 and trans-10,cis-12, was utilized. This was subjected to esterification with octanol in an aqueous reaction system using Amano GC-4 lipase as catalyst. The resulting ester fraction contained up to 97% of the cis-9,trans-11 isomer. After adjustment of the reaction conditions, a concentration of 85% trans-10,cis-12–18∶2 could be obtained in the unreacted free fatty acid fraction. These lipase-catalyzed reactions provide a means for the preparative-scale production of high-purity cis-9,trans-11–18∶2, and a corresponding CLA fraction depleted of this isomer.     

Safety of dietary conjugated linoleic acid (CLA) in a 12-weeks trial in healthy overweight Japanese male volunteers

A study was conducted to investigate the short-term safety of dietary conjugated linoleic acid (CLA) in overweight Japanese male volunteers. The study design was a randomized, double-blind placebo-controlled trial including 60 healthy overweight volunteers (body mass index (BMI), 25 approximately 35 kg/m(2)). The subjects were randomly assigned to three groups: 5.4 g CLA-triacylglycerol (3.4 g as CLA), 10.8 g CLA-triacylglycerol (6.8 g as CLA) and placebo (10.8 g safflower oil) daily for 12 weeks. The CLA-triaclyglycerol contained 9c,11t- and 10t,12c-isomers at an equal proportion. The safety was evaluated by analyses of blood parameters and by clinical examinations at the baseline, and at 6 and 12 weeks, including vital signs and adverse effects. All subjects completed the study. The occurrence of adverse events tended to be higher in the CLA groups than in the placebo group, but all of the adverse events were mild to moderate, within normal ranges, and temporary. Serum aspartate aminotransferase (AST) activity did not differ significantly between the groups at 12 weeks, but in the high CLA group it was slightly increased from the baseline. Serum alanine aminotransferase (ALT) activity was higher in the high CLA group than in the placebo group after 12 weeks and was higher than at the baseline in both CLA groups. However, statistical analysis of the population of apparently healthy volunteers who had normal blood parameters at the baseline revealed that AST and ALT levels did not differ significantly among the 3 groups at 12 weeks. Moreover, no clinically significant changes in vital signs were observed in any of the groups. These results indicate that CLA at a dose of 3.4 g/day is a safe dietary level in healthy Japanese populations in terms of the parameters examined.     

Occurrence of conjugated linoleic acid isomers in beef

The amounts of Δ9,Δ11-conjugated linoleic acid (CLA) isomers were determined in loin-associated fat samples of bulls (n=6) and steers (n=7) by capillary gas chromatography of fatty acid methyl ester (FAME) derivatives. The main CLA-isomer—18:2 c9,t11—provided approximately 0.76 ± 0.15% and 0.86 ± 0.15% of total FAME in bulls and steers, respectively. No differences (P>0.05) were observed between the CLA isomer distribution of bulls (t9,c11, 0.026 ± 0.014%; c9,c11, 0.015 ± 0.008%; and t9,t11, 0.029 ± 0.003%) and steers (t9,c11, 0.027 ± 0.014%; c9,c11, 0.015 ± 0.005%; and t9,t11, 0.030 ± 0.007%).     

Analysis,occurrence, and physiological properties of trans fatty acids (TFA) with particular emphasis on conjugated linoleic acid isomers (CLA) – a review

The present review provides an outline of the current knowledge of trans fatty acids (TFA) including their structure and formation, analysis, occurrence in foods, estimation and evaluation of daily intake, contents in human adipose tissues and fluids, and physiological properties. Special emphasis is put on conjugated linoleic acids (CLA), which are related to unique beneficial physiological properties. The effects of CLA on carcinogenesis in in vivo and human cancer cell culture studies, on cancer inhibition via the immune system, and further physiological properties are briefly reported. Ways to affect the CLA contents in foods, e.g. influence of feeding regimens or processing conditions, are also discussed.     

Distributions of conjugated linoleic acid (CLA) isomers in tissue lipid classes of pigs fed a commercial CLA mixture determined by gas chromatography and silver ion-high-performance liquid chromatography

Pigs were fed a commercial conjugated linoleic acid (CLA) mixture, prepared by alkali isomerization of sunflower oil, at 2% of the basal diet, from 61.5 to 106 kg live weight, and were compared to pigs fed the same basal diet with 2% added sunflower oil. The total lipids from liver, heart, inner back fat, and omental fat of pigs fed the CLA diet were analyzed for the incorporation of CLA isomers into all the tissue lipid classes. A total of 10 lipid classes were isolated by three-directional thin-layer chromatography and analyzed by gas chromatography (GC) on long capillary columns and by silver-ion high-performance liquid chromatography (Ag⁺-HPLC); cholesterol was determined spectrophotometrically. Only trace amounts (<0.1%; by GC) of the 9,11–18∶2 cis/trans and trans, trans isomers were observed in pigs fed the control diet. Ten and twelve CLA isomers in the diet and in pig tissue lipids were sepatated by GC and Ag⁺-HPLC, respectively. The relative concentration of all the CLA isomers in the different lipid classes ranged from 1 to 6% of the total fatty acids. The four major cis/trans isomers (18.9% 11 cis, 13 trans-18∶2; 26.3% 10 trans, 12 cis-18∶2; 20.4% 9 cis, 11 trans-18∶2; and 16.1% 8 trans, 10 cis-18∶2) constituted 82% of the total CLA isomers in the dietary CLA mixture, and smaller amounts of the corresponding cis,cis (7.4%) and trans,trans (10.1%) isomers were present. The distribution of CLA isomers in inner back fat and in omental fat of the pigs was similar to that found in the diet. The liver triacylglycerols (TAG), free fatty acids (FFA), and cholesteryl esters showed a similar patterns to that found in the diet. The major liver phospholipids showed a marked increase of 9 cis,11 trans-18∶2, ranging from 36 to 54%, compared to that present in the diet. However, liver diphosphatidylglycerol (DPG) showed a high incorporation of the 11 cis,13 trans-18∶2 isomer (43%). All heart lipid classes, except TAG, showed a high content of 11 cis,13 trans-18∶2, which was in marked contrast to results in the liver. The relative proportion of 11 cis,13 trans-18∶2 ranged from 30% in the FFA to 77% in DPG. The second major isomer in all heart lipids was 9 cis,11 trans-18∶2. In both liver and heart lipids the relative proportions of both 10 trans,12 cis-18∶2 and 8 trans,10 cis-18∶2 were significantly lower compared to that found in the diet. The FFA in liver and heart showed the highest content of trans,trans isomers (31 to 36%) among all the lipid classes. The preferential accumulation of the 11 cis,13 trans-18∶2 into cardiac lipids, and in particular the major phospholipid in the inner mitochondrial membrane, DPG, in both heart and liver, appears unique and may be of concern. The levels of 11 cis,13 trans-18∶2 naturally found in foods have not been established.     

Effects of conjugated linolenic acid and conjugated linoleic acid on lipid metabolism in mice

The effects of two isomers of conjugated linolenic acid (CLnA), α‐eleostearic acid (α‐ESA) and punicic acid (PA), on body fat and lipid metabolism were investigated, compared with a conjugated linoleic acid (CLA) mixture (primarily cis9,trans11‐ and trans10,cis12‐18:2) and α‐linolenic acid (ALA), a non‐conjugated octadecatrienoic acid, in the present study. ICR mice were fed either a control diet or one of four experimental diets supplemented with 1% α‐ESA, 1% PA, 1% CLA mixture and 1% ALA in the form of triacylglycerols (TAG) for 6 weeks. The weights of perirenal and epididymal adipose tissues were significantly decreased while the liver weight was significantly increased in mice fed CLA, compared with the control. In contrast to CLA, the tissue weights in α—ESA‐, PA‐ and ALA‐fed mice were not affected. No significant differences were observed in TAG, total cholesterol, high‐density lipoprotein and low‐density lipoprotein cholesterol levels among the five groups. The liver TAG level was significantly decreased in mice fed α‐ESA and PA while it was significantly increased in mice fed the CLA mixture. These results indicate that CLnA and CLA have differential effects on body fat mass and liver TAG levels in mice.     

Impact of novel methodologies on the analysis of conjugated linoleic acid (CLA). Implications of CLA feeding studies

Interest in conjugated linoleic acid (CLA) has increased in the past decade as a result of reports of several health benefits related to its consumption. Naturally occurring CLA isomers are found in milk, dairy, and meat products from ruminants. Detailed isomeric composition of CLA in different chemical and biological matrices had been hindered by the lack of adequate analytical techniques. New methodologies were developed and used to determine the distribution of major and minor geometric and positional CLA isomers in cheese, beef, cow milk, human adipose, and human milk. Base-catalyzed methylation was used. A novel silver ion high-performance liquid chromatographic procedure was developed, which successfully resolved up to 16 isomers. The double bond configuration and position for CLA isomers were confirmed by gas chromatography (GC)-direct deposition-Fourier transform infrared spectroscopy and GC-electron ionization mass spectrometry, respectively: The incorporation of CLA isomers in tissues of animals fed CLA diets was also determined. Currently available analytical data suggest the need to re-evaluate prior CLA studies and their nutritional and biological implications.     

Thermally induced formation of conjugated isomers of linoleic acid

Chemical pathways responsible of the conjugation of linoleic acid during heat treatments such as refining (deodorization), frying or cooking processes have been investigated. For this purpose, methyl linoleate was submitted to oxidative and non‐oxidative thermal conditions. The resulting degradation products were mainly composed of geometrical and conjugated fatty acid isomers. Oxidative conditions were obtained using tert‐butyl hydroperoxide under inert atmosphere, and air. The obtained results from both thermal oxidative conditions were compared to non‐oxidative thermal treatment. Higher levels of conjugated linoleic acid were found when linoleate was heated under oxidative conditions. Two distinct mechanisms responsible for the formation of CLA isomers are proposed and discussed. Evidence of formation of 9,11‐C18:2 and 10,12‐C18:2 acids from 9,12‐C18:2 by a free‐radical chain reaction is provided. The first step consists in the formation of a free radical by abstraction of an active bis‐allylic hydrogen. By delocalization of the initial free radical, two allylic free radicals were stabilized and converted into the corresponding CLA isomers via the abstraction of a hydrogen radical from other linoleic acid or oxygenated species. Kinetic observations confirmed the significance of the bimolecular mechanism. Moreover, the proposed mechanism is supported by several pieces of information from the literature on peroxidation of linoleic acid. Under pure thermal conditions and/or for diluted samples, a second pathway to the formation of CLA from heat‐treated linoleic acid is proposed via an intramolecular rearrangement of the pentadienyl structure. This thermal [1,3]‐sigmatropic rearrangement results in a mixture of 9,11 and 10,12 CLA isomers. The formed cis/trans CLA isomers were readily rearranged by a [1,5]‐sigmatropic shift to yield trans‐8,cis‐10 and cis‐11,trans‐13 CLA isomers, respectively.     

Quantitative determination of conjugated linoleic acid isomers in beef fat

The amounts of 14 conjugated linoleic acid (CLA) isomers (t12t14, t11t13, t10t12, t9t11, t8t10, t7t9, t6t8; 12,14 c/t, t11c13, c11t13, t10c12, 9,11 c/t, t8c10, t7c9‐18:2) in 20 beef samples were determined by triple‐column silver‐ion high‐performance liquid chromatography (Ag⁺‐HPLC). Quantitation was performed using an external CLA reference standard consisting of cis9,trans11‐18:2,trans9,trans11‐18:2 and cis9,cis11‐18: 2. Linearity was checked as being r > 0.9999 between 0.02 × 10^‐3 to 2 mg/ml. The determination limit (5‐fold signal/noise ratio) of the CLA reference was estimated to be 0.25, 0.50, 1.0 ng/injection for the cis/trans, trans,trans and cis,cis isomers, respectively. As expected, cis9,trans11‐18:2 was the predominant isomer (1.95 ± 0.54 mg/g fat) in beef, followed by trans7,cis9‐18:2 (0.19 ± 0.04 mg/g fat); cis,cis isomers were below the determination limit in most beef samples. Total CLA amounts determined by Ag⁺‐HPLC were compared to total CLAs determined by gas chromatography (GC, 100 m CPSil^TM 88 column). The amounts obtained by GC were generally higher than those determined by Ag⁺ ‐HPLC due to co‐eluting compounds.     

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.     

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.     

Effects of conjugated linoleic acid isomers on lipid-metabolizing enzymes in male rats

Male weanling Wistar rats (n=15), weighing 200–220 g, were allocated for 6 wk to diets containing 1% (by weight) of conjugated linoleic acid (CLA), either as the 9c,11t-isomer, the 10t,12c-isomer, or as a mixture containing 45% of each of these isomers. The five rats of the control group received 1% of oleic acid instead. Selected enzyme activities were determined in different tissues after cellular subfractionation. None of the CLA-diet induced a hepatic peroxisome-proliferation response, as evidenced by a lack of change in the activity of some characteristic enzymes [i.e., acyl-CoA oxidase, CYP4A1, but also carnitine palmitoyltransferase-I (CPT-I)] or enzyme affected by peroxisome-proliferators (glutathione S-transferase). In addition to the liver, the activity of the rate-limiting β-oxidation enzyme in mitochondria, CPT-I, did not change either in skeletal muscle or in heart. Conversely, its activity increased more than 30% in the control value in epididymal adipose tissue of the animals fed the CLA-diets containing the 10t,12c-isomer. Conversely, the activity of phosphatidate phosphohydrolase, a rate-limiting enzyme in glycerolipid neosynthesis, remained unchanged in adipose tissue. Kinetic studies conducted on hepatic CPT-I and peroxisomal acyl-CoA oxidase with CoA derivatives predicted a different channeling of CLA isomers through the mitochondrial or the peroxisomal oxidation pathways. In conclusion, the 10t,12c-CLA isomer seems to be more efficiently utilized by the cells than its 9c,11t homolog, though the Wistar rat species appeared to be poorly responsive to CLA diets for the effects measured.