DFA: EPA/DHA and Cognitive Health (2011)

Study Design:
- Click here for explanation of classification scheme.
Quality Rating:
Research Purpose:
  • To assess the association of plasma cholesteryl ester and phospholipid concentrations of n-3 fatty acids with decline in three areas of cognition
  • A sub-group analysis was conducted for possible interaction between n-3 fatty acids and levels of oxidative stress and the risk of neuro-degenerative disease
  • It was hypothesized that those sub-groups with elevated physiologic oxidative stress would benefit most from higher intakes of dietary n-3 fatty acids as measured by plasma concentrations in cholesteryl esters and phospholipids.
Inclusion Criteria:
  • Participant in the Atherosclerosis Risk in Communities (ARIC) study
  • Complete plasma fatty acid data.
Exclusion Criteria:
  • Less than 50 years
  • Without cognitive testing at visits two and four.
Description of Study Protocol:


  •  In the original study, adults aged 45 to 64 years were recruited in each of four US communities (Forsyth County, NC; Jackson, MS; suburbs of Minneapolis, MN; and Washington County, MD)
  • Three of the four cohorts represented the ethnic mix of their communities, whereas in Jackson, MS, only African American residents were recruited
  • For the present study, only participants from the suburbs of Minneapolis were used.


Prospective cohort.


  • No intervention
  • Comparison groups were based on global cognitive decline (no decline or decline and on sub-group analysis of various cognitive tests). 

Statistical Analysis

  • Univariate analyses of predictor and outcome variables was performed as well as covariates
  • For bivariate analyses of exposure and outcome, we computed means of predictor variables across outcome groups (zero, no decline; one, declined) and assessed statistical significance of differences by using an independent-sample T-test for continuous predictors and the chi-square test for categorical predictors at alpha of 0.05
  • The ORs of decline were computed by each one-SD change in exposure by conducting multivariate logistic regression analysis
  • Control for confounding was done using backward elimination, retaining in the model those variables that changed the estimated effect (odds ratio) of the exposure by more than 5%. This level is more suitable than 10% because the sensitivity of odds ratios to confounding effects tends to increase with increase in sample size.
  • After obtaining a parsimonious model, the authors first presented odds ratios for global cognitive decline and plasma concentrations of various groups of fatty acids
  • Subsequently, and focusing on n-3 HUFAs, stratum-specific odds ratios were presented for each sub-group of interest (e.g., hypertensives vs. normotensives)
  • Multivariate logistic regression models were also performed adding the interaction term (n-3 HUFA x binary sub-group variable) into a model with the main effects included as well (i.e., n-3 HUFA and binary sub-group variable)
  • This model was compared with the one without the interaction term and used likelihood ratio tests to assess statistical significance of interaction between exposure and potential effect modifiers in determining the outcome
  • For this purpose, a type I error level of 0.10 was considered valid
  • Statistical analyses were conducted with STATA software (version 8.2). 
Data Collection Summary:

Timing of Measurements

1987 to 1989; plasma fatty acids and neuro-psychological tests.

Dependent Variables

Cognitive assessment:

  • Delayed Word Recall Test: Assesses verbal learning and recent memory
  • Digit Symbol Substitution Test of the Wechsler Adult Intelligence Scale-Revised: Measures psychomotor performance and, for most adults, is relatively unaffected by intellectual ability, memory or learning. It appears to be a sensitive and reliable marker of brain damage.
  • Word Fluency Test: Sensitive to linguistic impairment and early mental decline in older persons. It is also a sensitive marker of damage in the left lateral frontal lobe.
  • Cutoffs were determined for decline in each domain of cognition by using the Reliable Change Index (RCI) method to correct for measurement error and practice effects. Scoring below an RCI of -1.645 was regarded as showing a statistically reliable deterioration in test scored. A composite measure of the three RCIs (obtained from DWRT, DSST and WFT change scores) to assess global cognitive decline (GCD) was created using principal-components analysis (PCA).

Independent Variables

  • Plasma fatty acid exposures: 12-hour fasting blood and plasma fatty acids analyzed by gas chromatography
  • Test-retest reliability coefficients (subjects sampled three times at two-week intervals) for various plasma fatty acids ranged from 0.50 to 0.93 for cholesteryl esters and from 0.50 to 0.89 for phospholipids. 

Control Variables

  • Most covariates considered were measured at visits one or two, although some were defined according to measurements that spanned all four visits
  • Age, sex and education were all self-reported
  • Behavioral factors measured at visit one included smoking, alcohol and caffeine consumption and physical activity
  • A validated index of physical activity, derived at visit one, of summed sports, work and leisure indexes, which ranged from a score of one (low) to a score of five (high)
  • Body mass index was computed at visit one
  • Baseline dietary intakes of antioxidants and other micronutrients (mainly vitamins B6 and B12 and folate) were also considered. The usual dietary intake of these nutrients was estimated from an interviewer-administered semi-quantitative food frequency questionnaire modified from Willett. Daily intake of nutrients was calculated by multiplying the nutrient content of each food in the portion specified by the frequency of daily consumption and summing the results.
  • All of the covariates were considered as potential confounders in the multivariate analysis
  • Potential effect modifiers were grouped as genetic and comorbid conditions.  
Description of Actual Data Sample:

Initial N


Attrition (Final N)

  • After exclusion for less than 50 years and without cognitive testing at visits two and four, N=2,251 (approximately 50% women)
  • No decline: N=2,111
  • Decline: N=140.


  • No decline: 56.21±4.22
  • Decline: 57.74±4.22.


All subjects were white and from the suburbs of Minneapolis, MN.

Other Relevant Demographics

  • Education (percentage):
    • Incomplete high school: No decline, 6.78; decline, 5.00
    • High school: No decline, 36.40; decline, 32.86
    • More than high school: No decline, 56.82; decline, 62.14
  • Smoking status (percentage):
    • Never smoker: No decline, 40.48; decline, 40.71
    • Former smoker: No decline 41.94; decline, 40.0
    • Current smoker: No decline 17.68; decline, 19.28
  • Physical activity scale:
    • No decline: 7.34±1.32
    • Decline: 7.09±1.39
  • Total energy intake (kcal per day):
    • No decline: 1,583±559
    • Decline: 1,546±568
  • Stroke or TIA (percentage):
    • No decline: 9.00
    • Decline: 13.6
  • Hypertensive (percentage):
    • No decline: 48.93
    • Decline: 54.58
  • Dyslipidemia (percentage):
    • No decline: 37.38
    • Decline: 32.14
  • Type 2 diabetes mellitus (percentage):
    • No decline: 13.64
    • Decline: 17.14
  • Depression scale:
    • No decline: 8.08±7.03
    • Decline: 9.73±8.25
  • Poor pulmonary function (FEV1/FVC less than 70) (percentage):
    • No decline: 18.82
    • Decline: 20.71.


BMI (kg/m2):

  • No decline: 27.21±4.41
  • Decline: 26.60±4.33.


Minneapolis, MN.

Summary of Results:

 Key Findings

  • Those who did not decline were on average younger and more physically active, reported less depressive symptoms and had a relatively hypocoaguable profile
  • Those who declined had higher baseline cognitive score and as expected, greater decline in cognitive function between the two visits
  • Within the cholesteryl ester fraction, those who declined had higher concentrations of palmitic and arachidonic acids and lower concentrations of linoleic acid
  • After controlling for other potentially confounding factors, including other fatty acids, total PUFAs, total n-6 PUFAs and linoleic acid, were all inversely related to decline, whereas a higher concentration of arachidonic acid remained a significant risk factor for decline
  • In the plasma phospholipid fraction, the adjusted odds ratio of decline with each SD increment in these fatty acids did not differ significantly from the null value of one, with the exception of palmitic acid (an SFA), which was consistently associated with the risk of cognitive decline
  • n-3 PUFAs in general and DHA+EPA in particular had no significant effect on global cognitive decline.
  • Sub-group analysis:
    • Greater DHA+EPA was associated with less decline in verbal fluency (WFT) for both cholesteryl ester and phospholipid fractions (OR: 0.74 (0.57, 0.97) and 0.73 (0.58, 0.93), respectively)
    • When the outcome was decline in verbal fluency, DHA+EPA was mostly protective among hypertensive subjects for both the cholesteryl ester and phospholipids fraction and among dyslipidemic subjects for the phosholipids fraction only
    • A higher plasma phospholipids concentration of n-3 HUFAs was protective against decline in verbal fluency only among subjects with a lower score on depressive symptoms.

Portions of Tables One and Two

  No Decline Decline P-value

Baseline cognitive scores (visit two)








WFT 37.05±11.28 45.66±12.09 P<0.05
Cognitive decline (visit four to visit two)


-0.10±1.42 -1.94±1.62 P<0.05


-3.88±5.79 -13.1±8.06 P<0.05
WFT -1.12±7.13 -12.77±8.91 P<0.05 


  Global Cognitive Decline
  No Decline Decline OR (95% CI)

Plasma cholesteryl esters

Palmitic acid



1.28 (1.07, 1.54)

Total n-6 PUFAs 64.39±3.82 63.76±3.92 0.54 (0.36, 0.82)
Arachidonic acid 8.27±1.67 8.72±1.77 1.21 (1.00, 1.47)
Linoleic acid 54.26±4.64 53.13±4.91 0.64 (0.49, 0.83)
Total n-3 PUFAs 1.42±0.43 1.46±0.39 1.08 (0.92, 1.26)
DHA+EPA 1.01±0.40 1.04±0.36 1.09 (0.94, 1.27)
(DHA/EPA)/AA 0.12±0.05 0.12±0.04 0.96 (0.80, 1.15)


Author Conclusion:
  • Promoting higher intakes of n-3 HUFAs in the diet of hypertensive and dyslipidemic persons may have substantial benefits in reducing their risk of cognitive decline in the area of verbal fluency
  • Clinical trials are needed to confirm this finding.
Funding Source:
Government: NHLBI contracts
Reviewer Comments:
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? N/A
  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? No
  3.3. Were concurrent controls or comparisons used? (Concurrent preferred over historical control or comparison groups.) N/A
  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? Yes
  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? N/A
  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? N/A
  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? N/A
  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? ???
  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? N/A
  6.2. In observational study, were interventions, study settings, and clinicians/provider described? Yes
  6.3. Was the intensity and duration of the intervention or exposure factor sufficient to produce a meaningful effect? ???
  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)? N/A
  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