DFA: Conjugated Linoleic Acid (CLA) Supplementation and Intermediate Health Outcomes (2011)

Study Design:
- Click here for explanation of classification scheme.
Quality Rating:
Research Purpose:

Comparison of the effects on plasma lipoproteins and body composition of the consumption of a modified butter naturally enriched with CLA (CLA-B: 4.22g CLA per 100g butter fat) by the addition of sunflower oil to the diet of dairy cows with the consumption of a control butter (CON-B) that was low in CLA (0.38g CLA per 100g butter fat).

Inclusion Criteria:
  • Men
  • BMI>26 and waist circumference>90cm
  • Good health
  • Non-smokers
  • Between 18 and 55 years of age.
Exclusion Criteria:
  • Presence of a monogenic dyslipoproteinemia
  • Use of medication known to affect lipid metabolism
  • Chronic, metabolic or acute disease
  • Significant weight change in the six months before the experiment
  • Regular alcohol intake (more than one drink per day or more than seven drinks per week)
  • Unusual dietary habits such as vegetarianism, food allergies or a dislike of foods included in the experimental diets.
Description of Study Protocol:


Recruited from the Quebec City area 


  • Randomized cross-over trial
  • Eight subjects were randomly assigned to the CLA-B diet for the first four-week period and to the CON-B diet for the second four-week period. The other eight subjects were assigned to the nutritional treatment sequence in reverse.

Blinding used



  • Both butters were manufactured from milk produced by dairy cows from a Laval University dairy herd. Cows were initially fed the same diet, then after three weeks, milk samples were obtained and the CLA content measured.
  • Cows with the lowest concentration of CLA were identified, their milk was collected to make the control
  • Once the production of CON-B was completed, the same cows were fed a similar diet to which 5% sunflower oil was added. Milk samples were taken after three weeks, and cows with the greatest concentration of CLA continued consuming the diet for the purpose of milk collection and the manufacture of the CLA-B.
  • The butters were incorporated into two experimental diets
  • Figure 1 shows relative isomeric distribution of CLA isomers in the butters; the percentages below were estimated by the reviewer from the figure
  • Control butter (CON-B): 0.38g CLA per 100g fatty acids: About 52% cis-9, trans-11; cis-10, trans-11 undetectable; about 15% trans-10, cis-12+21:0
  • CLA-B: 4.22g CLA per 100g fatty acids; About 80% cis-9, trans-11; less than 5% cis-10, trans-11; less than 5% trans-10, cis-12+21:0
  • Each intervention period for was four-weeks. Experimental periods were separate by an eight-week washout period, during which the subjects resumed their usual diets.

 Statistical Analysis

  • Repeated-measures analysis of variance, adjusted for cross-over designs, using the general linear model was performed to identify differences between experimental treatments 
  • In the analysis of variance model, the carryover effects for each of the variables studied were tested by introducing a term referring to the sequence in which the dietary treatments were given. Although no significant sequence effect was found, a significant interaction was observed between treatment and sequence for VLDL-triacylglycerol and LDL apo B, which suggested a different response to the two diets across time. Therefore, only the results of the first period were analyzed for those two variables.
  • Paired T tests were used to identify differences within the experimental diets only when significant differences were found between dietary treatments
  • Group averages are reported as means ± SDs
  • CRP values were logarithmically transformed before statistical analysis to achieve normal distribution. Two subjects who were found, during the course of the study, to have CRP concentrations >10mg/L, which suggested the presence of bacterial infection or inflammation, were excluded from statistical analysis for that variable.
  • Differences were considered significant at P<0.05.
Data Collection Summary:

Timing of Measurements

  • Food intake over three days was measured prior to the start of the study to estimate subject's usual energy intakes and thereby set proper energy levels for the experimental period
  • Body weight taken every weekday before lunch
  • On weekdays, subjects came to the metabolic kitchen daily to consumer their lunch meal, and they were then given their next dinner and breakfast meals in a package to take home
  • At the beginning (day one) and at the end (day 28) of each experimental period, waist circumference was measured
  • In the week after each experimental period (day 30 or 31), visceral adipose tissue accumulation was measured
  • Fasting blood samples were collected on days one and 28 of each experimental period  
  • Experimental periods were separated by an eight-week washout period, during which the subjects resumed their usual diets to remove the residual effects of the preceding experimental diet on the tested variables.

Dependent Variables

  • Waist circumference, height and weight
  • Visceral adipose tissue accumulation was measured by computed tomography, which was performed on a Siemens Somatom DRH Scanner
  • Plasma lipoproteins: Plasma and lipoprotein cholesterol, triacylglycerols
  • LDL apolipoprotein (apo) B and HDL apo A-I were measured by rocket immunoelectrophoresis
  • LDL particle size was quantified by densitometric scanning of Sudan Black-stained gels
  • Plasma leptin concentrations were determined with the use of a highly sensitive, commercial, double-antibody radioimmunoassay.
  • Plasma CRP concentrations were measured by using a commercially available, highly sensitive immunoassay.

Independent Variables

  • CON-B
  • CLA-B.

Control Variables

Throughout the study, participants were asked to maintain usual level of physical activity, which was evaluated by a weekly questionnaire completed by the subjects.

Description of Actual Data Sample:
  • Initial N: 17 men
  • Attrition (final N):
    • 16 men completed the study
    • One subject withdrew before the second phase of the study due to a job relocation
  • Age: 36.6±12.4 (20-55)
  • Ethnicity: All were white
  • Other relevant demographics: Most were Canadian
  • Anthropometrics:
    • Weight (kg): 94.6±13.3 (71.2-119.5)
    • BMI (kg/m2): 31.2±4.4 (26.2-44.4)
    • Waist circumference (cm): 105.8±9.8 (92.5-129.5)
  • Location: Subjects were free living but came to the metabolic kitchen at Laval University for their lunch meal, to pick up their remaining food and for measurements. 
Summary of Results:

 Key Findings

  • The 16 participants were overweight or obese, but their mean blood lipid profiles were within the normal range at baseline
  • Table 1 of the article compares the subjects usual diet with the nutritional composition of the experimental diets. That some subjects lost weight despite the fact that their experimental energy intake was higher than their reported habitual energy intake suggested that, on average, subjects underestimated their energy intake. 
  • The diet-induced variations in body weight, waist circumference, and plasma leptin concentrations did not differ significantly between the two groups
  • No significant difference was found in the accumulation of visceral and subcutaneous adipose tissue between the diet groups.

Table 1. Body weight

  CON-B CLA-B P (between diets)
Body weight (kg)      
Day 1 94.4±13.4 94.3±13.3  
Day 28 93.1±13.0 92.5±12.9  
(day 28-day one)
-1.31±1.23 -1.77±1.46 0.18

Other Findings

  • When compared with the CON-B diet, the CLA-B diet resulted in significantly smaller reduction in plasma total cholesterol (-0.02 vs. -0.26mmol/L), but the changes in VLDL, LDL and HDL cholesterol and HDL subfractions did not differ significantly between the two diets. The greater reduction in total cholesterol after the CON-B diet was accompanied by a significantly greater decrease in total:HDL and LDL:HDL cholesterol after the CON-B diet than after the CLA-B diet.
  • The CON-B diet resulted in a significantly greater reduction in plasma apo B concentrations than did the CLA-B diet
  • Neither diet affected the LDL peak particle diameter or the concentration of plasma CRP.
Author Conclusion:
  • The results indicate that, in overweight or obese men with only a slight deterioration of their lipid profile, a 10-fold CLA enrichment of butter fat via the addition of sunflower oil to the feed of dairy cows does not induce significantly greater metabolic effects than are observed with the consumption of a control butter diet low in CLA.
  • Further human studies are needed to evaluate the individual effects of different CLA isomers on lipoprotein metabolism and in persons with a more detrimental lipid profile at study onset.
Funding Source:
Government: Natural Sciences and Engineering Research Council of Canada; FCAR fund; Quebec Ministry of Agriculture; Canada Research Chair in Nutrition
Novalait Inc,; Dairy Farmers of Canada
Food Company:
Commodity Group:
Reviewer Comments:

I think the author's recommendations for future studies is key to the effect of CLA. It appears most of the studies have been with relatively healthy people, maybe CLA has an effect in people who are not as healthy and showing effects of metabolic syndrome etc.

Quality Criteria Checklist: Primary Research
Relevance Questions
  1. Would implementing the studied intervention or procedure (if found successful) result in improved outcomes for the patients/clients/population group? (Not Applicable for some epidemiological studies) Yes
  2. Did the authors study an outcome (dependent variable) or topic that the patients/clients/population group would care about? Yes
  3. Is the focus of the intervention or procedure (independent variable) or topic of study a common issue of concern to dieteticspractice? Yes
  4. Is the intervention or procedure feasible? (NA for some epidemiological studies) Yes
Validity Questions
1. Was the research question clearly stated? Yes
  1.1. Was (were) the specific intervention(s) or procedure(s) [independent variable(s)] identified? Yes
  1.2. Was (were) the outcome(s) [dependent variable(s)] clearly indicated? Yes
  1.3. Were the target population and setting specified? Yes
2. Was the selection of study subjects/patients free from bias? Yes
  2.1. Were inclusion/exclusion criteria specified (e.g., risk, point in disease progression, diagnostic or prognosis criteria), and with sufficient detail and without omitting criteria critical to the study? Yes
  2.2. Were criteria applied equally to all study groups? Yes
  2.3. Were health, demographics, and other characteristics of subjects described? Yes
  2.4. Were the subjects/patients a representative sample of the relevant population? ???
3. Were study groups comparable? Yes
  3.1. Was the method of assigning subjects/patients to groups described and unbiased? (Method of randomization identified if RCT) Yes
  3.2. Were distribution of disease status, prognostic factors, and other factors (e.g., demographics) similar across study groups at baseline? Yes
  3.3. Were concurrent controls or comparisons used? (Concurrent preferred over historical control or comparison groups.) Yes
  3.4. If cohort study or cross-sectional study, were groups comparable on important confounding factors and/or were preexisting differences accounted for by using appropriate adjustments in statistical analysis? N/A
  3.5. If case control study, were potential confounding factors comparable for cases and controls? (If case series or trial with subjects serving as own control, this criterion is not applicable.) N/A
  3.6. If diagnostic test, was there an independent blind comparison with an appropriate reference standard (e.g., "gold standard")? N/A
4. Was method of handling withdrawals described? Yes
  4.1. Were follow-up methods described and the same for all groups? Yes
  4.2. Was the number, characteristics of withdrawals (i.e., dropouts, lost to follow up, attrition rate) and/or response rate (cross-sectional studies) described for each group? (Follow up goal for a strong study is 80%.) Yes
  4.3. Were all enrolled subjects/patients (in the original sample) accounted for? Yes
  4.4. Were reasons for withdrawals similar across groups? No
  4.5. If diagnostic test, was decision to perform reference test not dependent on results of test under study? N/A
5. Was blinding used to prevent introduction of bias? Yes
  5.1. In intervention study, were subjects, clinicians/practitioners, and investigators blinded to treatment group, as appropriate? Yes
  5.2. Were data collectors blinded for outcomes assessment? (If outcome is measured using an objective test, such as a lab value, this criterion is assumed to be met.) Yes
  5.3. In cohort study or cross-sectional study, were measurements of outcomes and risk factors blinded? N/A
  5.4. In case control study, was case definition explicit and case ascertainment not influenced by exposure status? N/A
  5.5. In diagnostic study, were test results blinded to patient history and other test results? N/A
6. Were intervention/therapeutic regimens/exposure factor or procedure and any comparison(s) described in detail? Were interveningfactors described? Yes
  6.1. In RCT or other intervention trial, were protocols described for all regimens studied? Yes
  6.2. In observational study, were interventions, study settings, and clinicians/provider described? N/A
  6.3. Was the intensity and duration of the intervention or exposure factor sufficient to produce a meaningful effect? Yes
  6.4. Was the amount of exposure and, if relevant, subject/patient compliance measured? Yes
  6.5. Were co-interventions (e.g., ancillary treatments, other therapies) described? N/A
  6.6. Were extra or unplanned treatments described? N/A
  6.7. Was the information for 6.4, 6.5, and 6.6 assessed the same way for all groups? Yes
  6.8. In diagnostic study, were details of test administration and replication sufficient? N/A
7. Were outcomes clearly defined and the measurements valid and reliable? Yes
  7.1. Were primary and secondary endpoints described and relevant to the question? Yes
  7.2. Were nutrition measures appropriate to question and outcomes of concern? Yes
  7.3. Was the period of follow-up long enough for important outcome(s) to occur? ???
  7.4. Were the observations and measurements based on standard, valid, and reliable data collection instruments/tests/procedures? Yes
  7.5. Was the measurement of effect at an appropriate level of precision? Yes
  7.6. Were other factors accounted for (measured) that could affect outcomes? Yes
  7.7. Were the measurements conducted consistently across groups? Yes
8. Was the statistical analysis appropriate for the study design and type of outcome indicators? Yes
  8.1. Were statistical analyses adequately described and the results reported appropriately? Yes
  8.2. Were correct statistical tests used and assumptions of test not violated? Yes
  8.3. Were statistics reported with levels of significance and/or confidence intervals? Yes
  8.4. Was "intent to treat" analysis of outcomes done (and as appropriate, was there an analysis of outcomes for those maximally exposed or a dose-response analysis)? No
  8.5. Were adequate adjustments made for effects of confounding factors that might have affected the outcomes (e.g., multivariate analyses)? Yes
  8.6. Was clinical significance as well as statistical significance reported? Yes
  8.7. If negative findings, was a power calculation reported to address type 2 error? N/A
9. Are conclusions supported by results with biases and limitations taken into consideration? Yes
  9.1. Is there a discussion of findings? Yes
  9.2. Are biases and study limitations identified and discussed? Yes
10. Is bias due to study's funding or sponsorship unlikely? Yes
  10.1. Were sources of funding and investigators' affiliations described? Yes
  10.2. Was the study free from apparent conflict of interest? Yes