The term conjugated linoleic acid (CLA) refers to several positional and geometric conjugated dienoic isomers of linoleic acid (LA). LA, or cis,cis-9, 12-octadecadienoic acid, is an 18-carbon omega-6 fatty acid (18:2, 9c–12c) abundant in seed oils, such as sunflower oil. LA is converted to CLA when a chemical or microbial reaction shifts the double bonds to form alternating double and single bonds, hence the term conjugated. These double bonds can be located in different positions along the 18-carbon chain (7,9; 8,10; 9,11; 10,12; 11,13). This conjugation of the double bonds can alter the geometrical isomery of the acid, so that one or both double bonds can adopt trans isomery. The two isomers with known biological activity are cis-9,trans-11 and trans-10,cis-12 CLA.
CLA potential effects on health include anticarcinogenic, antiatherogenic, antidiabetogenic and immune modulating propertiesReference Belury1, Reference McLeod, LeBlanc, Langille, Mitchell and Currie2. CLA has also attracted interest in the scientific community because of its potential effects on body composition, reducing body fat mass and increasing the lean mass. Because of the increasing prevalence and incidence of obesity, a significant proportion of consumers are interested in CLA supplements with purported body fat-lowering effects, perceived as a ‘natural’ compound devoid of harmful effects.
Sources of conjugated linoleic acid
Rumiant animals are able to synthesise CLA through the microbial isomerisation (Butyrinvibrio fibrisolvens and other anaerobic bacteria) of dietary LA in their gastrointestinal tractsReference Kepler, Hirons, McNeill and Tove3. Rumiant meat (beef, veal, lamb) and dairy products are rich in cis-9,trans-11 CLA, also known as rumenic acid. Rumenic acid is the first intermediate step in the biohydrogenation of LA to stearic acid (C18:0).
CLA isomers can also be industrially produced by heating LA in the presence of alkali or by partial hydrogenation of LA. CLA that is typically produced for experimental purposes (MI-CLA) consists of the cis-9,trans-11 (40.8–41.1%), trans-10,cis-12 (43.5–44.9%) and trans-9,trans-11/trans-10,trans-12 (4.6–10%) isomersReference Pariza, Park and Cook4. It should be noted that some commercial CLA preparations contain additional isomers with conjugated double bonds at the 8,10 or 11,13 positionsReference Christie, Dobson and Gunstone5.
The major dietary sources of CLA for humans are beef and dairy products. Cis-9,trans-11 CLA is the principal CLA isomer in milkfatReference Pariza, Park and Cook4. Several factors influence CLA content of food products, such as animal’s diet, age, breed or seasonal factors, reaching the highest levels in cow’s milk during the spring and summer, as cows are allowed to graze in pasturesReference Rainer and Heiss6. CLA in milk or meat is a stable compound under normal cooking and storage conditionsReference Dhiman, Nam and Ure7.
Total CLA content in milk or dairy products ranges from 0.34% to 1.07% of total fatReference Dhiman, Nam and Ure7, whereas in raw or processed beef it ranges from 0.12% to 0.68% of total fatReference Dhiman, Nam and Ure7. CLA daily intakes are 212 mg for men and 151 mg for women not consuming CLA commercial preparations. Most dietary CLA is obtained from dairy products, accounting for the cis-9,trans-11 CLA isomer for more than 90% of the total CLA intakeReference Ritzenthaler, McGuire, Falen, Schultz, Dasgupta and McGuire8.
Evidence of conjugated linoleic acid effects on body weight in animals
Interest in CLA started in 1987, when Ha et al. Reference Ha, Grimm and Pariza9 reported that CLA present in fried ground beef, also synthetically produced by base-catalysed isomerisation of LA, was an effective inhibitor of benzopyrene-initiated mouse epidermal neoplasia.
Regarding obesity and body composition, several studies in animals have shown the ability of CLA to reduce adiposity and increase lean mass. Park et al.Reference Park, Albright, Liu, Storkson, Cook and Pariza10 were the first to report that dietary CLA could alter body composition. A diet containing 0.5% MI-CLA administered to the ICR (Institute for Cancer Research) line of mice caused a 60% decrease in body fat, coupled with enhanced lean body mass, after about 4–5 weeks of feeding.
CLA has also been reported to lower fat mass in other lines of miceReference Pariza, Park and Cook4, as well as in Sprague–DawleyReference Yamasaki, Ikeda, Oji, Tanaka, Hirao and Kasai11 and Zucker ratsReference Sisk, Hausman, Martin and Azain12, though not reaching such impressive results as in mice. In swine, MI-CLA decreased fat deposition and increased lean tissue tooReference Dugan, Aalhus, Schaefer and Kramer13, Reference Ostrowska, Muralitharan, Cross, Bauman and Dunshea14. Interestingly, MI-CLA-supplemented diet increased fat deposition in obese Zucker ratsReference Sisk, Hausman, Martin and Azain12, while reducing insulin levels.
CLA does not reduce food intake in rodentsReference DeLany and West15, Reference West, Blohm, Truett and DeLany16 and its effects are independent of fat content of the dietReference DeLany and West15. Conversely, dietary protein source may alter the effects of CLA on adipocytes’ fat content and adipocytokine production, as rats fed with soy diet show more pronounced fat loss and different changes in leptin and adiponectin levels compared to their casein-fed counterpartsReference Akahoshi, Koba, Ichinose, Kaneko, Shimoda and Nonaka17.
There is substantial evidence that CLA reduces body fat gain in young growing animalsReference Park, Albright, Liu, Storkson, Cook and Pariza10, Reference Ostrowska, Muralitharan, Cross, Bauman and Dunshea14, Reference Bee18–Reference Dugan, Aalhus and Kramer20 although some study results do not strongly confirm thisReference Weber, Schinckel, Houseknecht and Richert21. CLA supplementation during gestation and lactation elicits greater body weight and feed efficiency in weanling mice relative to control animalsReference Chin, Storkson, Albright, Cook and Pariza22. This effect on feed efficiency and growth seems to be exerted by the cis-9,trans-11 isomerReference Pariza, Park and Cook4.
Reported effects of CLA on body weight and composition in animals are summarised in Table 1.
Effects of CLA isomers on body weight and composition
CLA – conjugated linoleic acid.↑ – increase; ↓ – decrease; ? – possible.
Evidence of conjugated linoleic acid effects on body weight in humans
Experiments in humans have not been able to show a significant effect on body weight related to either MI-, cis-9,trans-11 or trans-10,cis-12 CLAReference House, Cassady, Eisen, McIntosh and Odle23–Reference Terpstra26. Published studies report contradicting results. Although in the majority of them there are no significant differences on body weight, body compositionReference Terpstra26 or weight regainReference Kamphuis, Lejeune, Saris and Westerterp-Plantinga27, Reference Larsen, Toubro, Gudmundsen and Astrup28 between controls and CLA-supplemented groups, some studies suggest that CLA might have a tendency to increase lean body massReference Kamphuis, Lejeune, Saris and Westerterp-Plantinga27, Reference Smedman and Vessby29, Reference Blankson, Stakkestad, Fagertun, Thom, Wadstein and Gudmundsen30. On one hand, one study carried out in overweight/obese adult volunteersReference Blankson, Stakkestad, Fagertun, Thom, Wadstein and Gudmundsen30 report a reduction in body fat mass with doses ≥3.4 g MI-CLA per day at week 12 (150–580 g more than placebo), with a significant increase in body lean mass in the group that received 6.8 g MI-CLA per day, measured by dual-energy X-ray absorptiometry; however, this group was also the only one with a significant increase in exercise level. Similar effects were detected in other human trials with MI- and trans-10,cis-12 CLAReference Riserus, Smedman, Basu and Vessby31.
On the other hand, another recent study designed to assess the effect of 1-year supplementation with 3.4 g day−1 CLA in body weight or body fat regain found no differences against placeboReference Larsen, Toubro, Gudmundsen and Astrup28, Reference Whigham, Watras and Schoeller32. In a recent meta-analysis at a dose of 3.2 g day−1, CLA was found to produce a slight reduction in fat mass peaking at 6 months, although confidence intervals were broad.
Differences in doses (ranging from 0.7 to 6.8 g day−1)Reference Blankson, Stakkestad, Fagertun, Thom, Wadstein and Gudmundsen30–Reference Mougios, Matsakas, Petridou, Ring, Sagredos and Melissopoulou33, treatment compliance, type and/or proportion of the administered isomer/s, length of the study, characteristics of the study population, energy expenditure and nutrient and energy intakes are possible explanations for the observed discrepancies in the results. Noteworthy, there is no clear dose–response related to the body fat-lowering effect of trans-10,cis-12 CLA and in most of the studies that reported a significant decrease of body fat mass or in those suggesting an increase in lean mass, the participants were also involved in a training programme or did a considerable amount of exerciseReference Blankson, Stakkestad, Fagertun, Thom, Wadstein and Gudmundsen30, Reference Mougios, Matsakas, Petridou, Ring, Sagredos and Melissopoulou33. Therefore, exercise might enhance the body fat lowering and/or lean mass increasing effect of CLA.
The effects of CLA on body weight and composition in humans are summarised in Table 1.
Effects of conjugated linoleic acid on adipocytes
Although the mechanisms by which CLA exerts its action on body weight have not been fully elucidated yet, it seems clear that trans-10,cis-12 CLA is the isomer involved in the changes of body composition.
Trans-10,cis-12 CLA and/or its metabolites is/are able to reduce lipid uptake by adipocytes due to the inhibitory effects on gene expression and enzymatic activity of stearoyl-CoA desaturase (SCD)Reference Choi, Kim, Han, Park, Pariza and Ntambi34, Reference Park, Storkson, Ntambi, Cook, Sih and Pariza35 and lipoprotein lipase (LPL)Reference Park, Albright, Liu, Storkson, Cook and Pariza10, Reference Park, Storkson, Albright, Liu and Pariza36, Reference Brown, Boysen, Jensen, Morrison, Storkson and Lea-Currie37. Trans-10,cis-12 CLA also inhibits the expression of glucose transporter-4 (GLUT-4)Reference Brown, Boysen, Jensen, Morrison, Storkson and Lea-Currie37, Reference Brown, Boysen, Chung, Fabiyi, Morrison and Mandrup38 and increases the activity of carnitine palmitoyltransferase (CPT), enhancing fatty acid oxidationReference Park, Albright, Liu, Storkson, Cook and Pariza10. Trans-10,cis-12 CLA reduces lipogenesis in humanReference Brown, Boysen, Jensen, Morrison, Storkson and Lea-Currie37 and mice adipocytes, decreasing sterol regulatory element-binding protein-1 (SREBP-1) expression in vivo Reference Tsuboyama-Kasaoka, Takahashi, Tanemura, Kim, Tange and Okuyama39. Trans-10,cis-12 CLA also induces adipocyte apoptosis in miceReference Tsuboyama-Kasaoka, Takahashi, Tanemura, Kim, Tange and Okuyama39–Reference Hargrave, Meyer, Li, Azain, Baile and Miner43, perhaps mediated by an increase in tumour necrosis factor alpha (TNFα)Reference Tsuboyama-Kasaoka, Takahashi, Tanemura, Kim, Tange and Okuyama39, but the effects of CLA on lipolysis in vivo and in vitro are conflictingReference Park, Albright, Liu, Storkson, Cook and Pariza10, Reference Brown, Boysen, Chung, Fabiyi, Morrison and Mandrup38. Data regarding its effects on preadipocyte differentiation are inconclusive, possibly depending on species, CLA isomer and experimental conditionsReference Pariza, Park and Cook4, Reference Brown, Boysen, Jensen, Morrison, Storkson and Lea-Currie37. Chronic treatment with trans-10,cis-12 CLA significantly reduces the expression of several adipocyte-specific genes, including PPARα and PPARγ target genes, partly due to an increase in nuclear factor kappa B (NFκB) activity and subsequent induction of interleukin-6Reference Chung, Brown, Provo, Hopkins and McIntosh44, therefore explaining at least part of its effects on glucose and lipid metabolism. Trans-10,cis-12 CLA inhibits insulin-stimulated glucose uptake and oxidation, reduces fatty acid uptake and alters fatty acid metabolism in differentiating human preadipocytesReference Brown, Boysen, Jensen, Morrison, Storkson and Lea-Currie37.
MI-CLA appears to increase energy expenditure in rodents by several mechanisms that are not fully understood, although an increase in uncoupling protein UCP1 and UCP3 gene expression does not seem to be involvedReference Pariza, Park and Cook4, Reference DeLany and West15, Reference West, Blohm, Truett and DeLany16, Reference West, Delany, Camet, Blohm, Truett and Scimeca45, Reference Ohnuki, Haramizu, Oki, Ishihara and Fushiki46. However, some studies show an increase in UCP2 expression (mostly with trans-10,cis-12 CLA supplementation) in brown and white adipose tissue and skeletal muscleReference Tsuboyama-Kasaoka, Takahashi, Tanemura, Kim, Tange and Okuyama39, Reference Ealey, El Sohemy and Archer47, Reference Takahashi, Kushiro, Shinohara and Ide48.
According to all these data, it seems that trans-10,cis-12 CLA elicits its de-lipidative activity through both metabolism and cell cycle control. More research is necessary to clarify conflicting results and differences between species.
Potential effects of CLA on adipocytes are summarised in Table 2.
Effects of trans-10,cis-12 CLA on adipocytes
CLA – conjugated linoleic acid; SCD – stearoyl-CoA desaturase; LPL – lipoprotein lipase; GLUT4 – glucose transporter-4; CPT – carnitine palmitoyltransferase; SREBP-1 – sterol regulatory element-binding protein-1; TNFα – tumour necrosis factor alpha; UCP2 – uncoupling protein 2; NFκB – nuclear factor kappa B; PPAR – peroxisome proliferator-activated receptor. ↑ – increase; ↓ – decrease; ? – possible.
Effects of conjugated linoleic acid on the liver and skeletal muscle
Several in vivo and in vitro studies in mice have reported an increase in both liver fatty acid synthesis and oxidation in response to MI-CLA supplementationReference Tsuboyama-Kasaoka, Takahashi, Tanemura, Kim, Tange and Okuyama39, Reference Clement, Poirier, Niot, Bocher, Guerre-Millo and Krief49–Reference Peters, Park, Gonzalez and Pariza51. CLA may mediate its effects through PPARα and PPARβ/δ (both trans-10,cis-12 and cis-9,trans-11 CLA isomers are their ligandsReference Moya-Camarena, Vanden Heuvel, Blanchard, Leesnitzer and Belury52), a decrease in SCD-1Reference Lee, Pariza and Ntambi53 expression and other alternative pathways, as suggested by studies on knockout miceReference Kang, Miyazaki, Ntambi and Pariza54, Reference Peters, Park, Gonzalez and Pariza51. However, there seem to be differences between species, as hamstersReference de Deckere, van Amelsvoort, McNeill and Jones55 on a CLA-supplemented diet developed liver hypertrophy but not lipid accumulation, and rats or pigs fed with CLA-enriched diets showed no changes in weight or lipid content in the liverReference Pariza, Park and Cook4.
There is evidence of the anabolic properties of CLA on lean mass in miceReference Park, Albright, Liu, Storkson, Cook and Pariza10, and probably in other speciesReference Ostrowska, Muralitharan, Cross, Bauman and Dunshea56, Reference Ostrowska, Suster, Muralitharan, Cross, Leury and Bauman57. However, the effects of CLA on the skeletal muscle are still unclear. CLA-fed mice showed an increase of CPT activity on skeletal muscle, improving β-oxidationReference Park, Albright, Liu, Storkson, Cook and Pariza10 and, as previously indicated, an enhancement in UCP2 expression is also evident in some studies.
Metabolic effects of conjugated linoleic acid
The effects on lipid metabolism of trans-10,cis-12 CLA, the active isomer involved in fat mass loss, are currently objects of concern. Administration of trans-10,cis-12 CLA to humans results in an increase of the LDL/HLDL ratioReference Tricon, Burdge, Kew, Banerjee, Russell and Jones24, some studies reflecting an increase in LDL-cholesterolReference Smedman and Vessby29, while others showing a decrease in HDL-cholesterolReference Smedman and Vessby29, Reference Blankson, Stakkestad, Fagertun, Thom, Wadstein and Gudmundsen30, Reference Mougios, Matsakas, Petridou, Ring, Sagredos and Melissopoulou33, Reference Riserus, Arner, Brismar and Vessby58; however, such effects are just marginal and not statistically significant compared to the control group in some of the studiesReference Smedman and Vessby29, Reference Mougios, Matsakas, Petridou, Ring, Sagredos and Melissopoulou33. These deleterious effects are not evident with the administration of MI-CLA or the cis-9,trans-11 isomerReference Tricon, Burdge, Kew, Banerjee, Russell and Jones24, Reference Tricon, Burdge, Jones, Russell, El-Khazen and Moretti25, Reference Riserus, Smedman, Basu and Vessby31, Reference Riserus, Arner, Brismar and Vessby58–Reference Moloney, Yeow, Mullen, Nolan and Roche60. Moreover, cis-9,trans-11 CLA actually appears to decrease LDL cholesterol in humansReference Tricon, Burdge, Kew, Banerjee, Russell and Jones24, Reference Blankson, Stakkestad, Fagertun, Thom, Wadstein and Gudmundsen30. A 6.4 g day−1 MI-CLA doses for 12 weeks produced a modest increase in lean mass (0.6 kg) but significant decrease in serum HDL-cholesterol, sodium, hemoglobin, and hematocrit, and significant increases in serum alkaline phosphatase, C-reactive protein, and K-6, and white blood cells.
Plasma triacylglycerides (TAG) did not significantly change or decrease compared to placebo in some studies with MI-CLA supplementationReference Mougios, Matsakas, Petridou, Ring, Sagredos and Melissopoulou33, Reference Noone, Roche, Nugent and Gibney59, Reference Petridou, Mougios and Sagredos62, Reference Benito, Nelson, Kelley, Bartolini, Schmidt and Simon63. Nevertheless, in another study mean plasma TAG concentration was higher during supplementation with a mixture 85% trans-10,cis-12 CLA than with another mixture 80% cis-9,trans-11 CLAReference Tricon, Burdge, Kew, Banerjee, Russell and Jones24.
Interestingly, there is no clear dose–response relationship in the relative hyperlipidemic properties of trans-10,cis-12 CLA and hypolipidemic properties of cis-9,trans-11 CLA in humansReference Tricon, Burdge, Kew, Banerjee, Russell and Jones24, suggesting that this influence could be exerted even at low dosesReference Tricon, Burdge, Kew, Banerjee, Russell and Jones24. Moreover, in apolipoprotein E knockout mice, supplementation with trans-10,cis-12 CLA had a profound pro-atherogenic effect, whereas cis-9,trans-11 CLA impeded the development of atherosclerosisReference Benito, Nelson, Kelley, Bartolini, Schmidt and Simon64.
In a similar way, there are conflicting results related to carbohydrate metabolism in humans. Although several studies show that neither cis-9,trans-11 CLA nor trans-10,cis-12 CLA supplementation significantly modify plasma glucose or insulin levels or alter insulin sensitivity (revised QUICKI) or insulin resistance (HOMA-IR)Reference Tricon, Burdge, Kew, Banerjee, Russell and Jones24, Reference Smedman and Vessby29, Reference Noone, Roche, Nugent and Gibney59, Riserus et al.Reference Riserus, Smedman, Basu and Vessby31, Reference Riserus, Arner, Brismar and Vessby58, Reference Arbones-Mainar, Navarro, Guzman, Arnal, Surra and Acin65 repeatedly report marginal but detrimental effects of both trans-10,cis-12 CLA (3.4 g day−1) and cis-9,trans-11 CLA (3 g day−1) on insulin sensitivity (measured as the insulin sensitivity index with the euglycemic clamp) and lipid peroxidation in obese subjects with metabolic syndrome. Another study in type 2 diabetic patients supplemented with MI-CLA (3 g day−1) also shows unfavourable effects on fasting glucose, insulin resistance (HOMA) and oral glucose insulin sensitivityReference Moloney, Yeow, Mullen, Nolan and Roche60. These results are in agreement with those of CLA-fed miceReference Tsuboyama-Kasaoka, Takahashi, Tanemura, Kim, Tange and Okuyama39, but not with the reported improvement on insulin sensitivity of some other studies in genetically obese miceReference Riserus, Vessby, Arner and Zethelius66 or Zucker rats supplemented with CLAReference Sisk, Hausman, Martin and Azain12.
Conclusions
Although there is evidence in animals of the ability of trans-10,cis-12 CLA to reduce adiposity and increase lean mass, such unequivocal data supporting the efficacy of either of the CLA isomers in humans are not available.
Moreover, contradictory results raise concern about the possibility of the deleterious effects of trans-10,cis-12 CLA on the lipid profile, glucose metabolism and insulin sensitivity. Therefore, it is advisable to consider with caution the use of CLA supplements containing high quantities of trans-10,cis-12 CLA, especially in obese patients with type 2 diabetes or metabolic syndrome, which constitutes a considerable proportion of the whole overweight/obese population.
Before CLA supplementation is recommended, further research on human effectiveness and safety of the different isomers of CLA is required.
Acknowledgements
Conflict of interest declaration: The authors had no conflicts of interest to report.
Authorship contributions: All the authors (M.B.S., R.C., S.M., J.T.) declare that they participated sufficiently in the work to take full and public responsibility for its content..
