BF: Dietary Factors, Breast Milk and Infant Outcomes (2008)


Jensen CL, Maude M, Anderson EA, Heird WC. Effect of docosahexaenoic acid supplementation of lactating women on the fatty acid composition of breast milk lipids and maternal and infant plasma phospholipids. Am J Clin Nutr. 2000; 71 (suppl); 292S-299S.

PubMed ID: 10617985
Study Design:
Randomized Controlled Trial
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Quality Rating:
Positive POSITIVE: See Quality Criteria Checklist below.
Research Purpose:

To determine whether docosahexaenoic acid (DHA) supplementation of breast milk and infant plasma phospholipids (PPs), breastfeeding women were randomly assigned to three DHA supplementation groups (170-260mg/d) or a control group.

Inclusion Criteria:

26 pregnant women who planned to breast-feed exclusively for at least eight weeks were recruited during the last trimester of pregnancy or at the time of delivery and randomly assigned to one of four groups.

Exclusion Criteria:
  • Maternal age at time of delivery under 19 years of age or over 35 years of age
  • Maternal diabetes
  • Maternal history suggestive egg allergy
  • Infant gestational age under 37 weeks
  • Infant birth weight less than 2,500g or over 4,200g
  • Mother-infant pairs were dropped from the study if the infant’s intake of formula, other foods or both exceeded 25% of total intake any time during the six-week supplementation period.


Description of Study Protocol:
  • Recruitment: Methods were not detailed
  • Design: Randomized controlled trial
  • Intervention: The fatty acid composition of each of the supplements is shown in table one.

Table One: Mole Percentages of Selected Fatty Acids in Docosahexaenoic Acid
(DHA; 22:6n-3 supplements)


Fatty Acid
Algal DHA
High-DHA Eggs
Low-EPA Fish Oil
mol percentage
12:0 6.2 - -
14:0 17.9 - 4.5
16:0 15.3 23.2 24.9
18:0 1.0 8.2 6.7
16:1n-7 1.5 5.6 6.5
18:1n-7 - 1.7 2.5
18:1n-9 11.1 32.5 13.7
18:2n-6 0.7 21.6 2.2
20:4n-6   1.5 1.5
18:3n-3   1.2 0.7
20:5n-3   0.15 4.8
22:5n-3   0.27 1.1
22:6n-3 44.9 3.3 26.0

Statistical Analysis

  • All data were expressed as group means±SDs
  • The statistical significance of differences in the characteristics of mothers and infantGET/newinterface/key_questions.cfm/newinterface/key_questions.cfm66.249.67.19966.249.67.199urbanchina.smartlibrary.org80HTTP/1.1FC:\InetWeb\WebSiteRoot\\newinterface\key_questions.cfm!0"#/newinterface/key_questions.cfm Connection Keep-alive Accept*/* Accept-Encoding gzip,deflate Fromgooglebot(at) If-Modified-SinceMon, 29 Dec 2008 20:56:40 GMT User-AgentHMozilla/5.0 (compatible; Googlebot/2.1;


Data Collection Summary:

Timing of Measurements

  • Mothers and infants were admitted to the Metabolic Research Unit of Children's Nutrition Research Centers at two, five and eight weeks postpartum for determination of 24-hour milk intake and for collection of milk and blood samples
  • Infant weight, length and head circumference were measured at the same time periods as noted.

Dependent Variables

  • Total daily milk output: Sum of the volume collected by pump and the amount consumed by the infant as estimated from the difference between pre- and post-feeding weight
  • The total daily DHA intake of the infants: DHA concentration of the proportional aliquot and the 24-hour milk intake
  • Fatty acid patterns of maternal and infant plasma phospholipid fractions: Plasma lipids were extracted by the method of Bligh and Dyer and the phospholipid fractions of each were separated by one-dimensional thin-layer chromatography using hexane, diethyl ether and glacial acetic acid. The fatty acid patterns of the extracted total lipids of breast milk and the supplements were determined by using the same methods.

Independent Variables

Group DHA per Day Supplement
One <230mg
Algae-produced high DHA triacylglycerol (DHASCO; Martek Biosciences Corporation, Columbia, Missouri)
Two 170mg
High DHA-egg The County Hen, Hubbard, Massachusetts)
Three 260mg
High DHA, low-eicosapentaenoic acid marine oil (ROPUFA’30’ n-3 INF Oil; Hoffman-LaRoche, Parsipanny, New Jersey)
Four <35mg
No supplement (regular eggs per two days)


Description of Actual Data Sample:

Initial N


Attrition (final N)



Group Mothers (Age at Delivery) Infants (Gestational Age)
1  29±3  40±1
2  29±4  40±1
3  30±5  39±1
4  29±5  40±1


  • Group One: Six Caucasians
  • Group Two: One Asian-American, one African-American, four Caucasians
  • Group Three: One African-American, four Caucasians, and one Hispanic
  • Group Four: Five Caucasians and one Hispanic.


Houston, Texas

Summary of Results:

The mean mole percentage of selected n-3 and n-6 polyunsaturated fatty acids in breast milk at baseline (two weeks postpartum) and after six weeks of supplementation. Note that the breast milk contents of 22:5n-6 were significantly lower in group two (high-DHA eggs) than in the other groups at the end of the study period.

Table Two: Fatty Acid Composition of Breast Milk Total Lipids1


Two Weeks Postpartum
Eight Weeks Postpartum2
mol percentage of total fatty acids
mol percentage of total fatty acids
 Group One  1.00±0.18  0.95±0.23
 Group Two  1.4±0.17  1.20±0.47
 Group Three  0.96±0.21  1.11±0.33
 Group Four  1.16±0.27  1.05±0.36
20: 5n-3    
 Group One  0.07±0.02  0.07±0.02
 Group Two  0.07±0.02  0.06±0.01
 Group Three  0.08±0.02  0.09±0.02
 Group Four  0.08±0.02  0.07±0.02
 Group One  0.23±0.04  0.44±0.17a
 Group Two  0.22±0.05  0.29±0.10a,b
 Group Three  0.27±0.11  0.39±0.07a
 Group Four  0.23±0.06  0.19±0.06b
 Group One  15.1±1.4  15.2±1.5
 Group Two  16.6±3.0  15.9±3.2
 Group Three  15.4±2.3  16.1±3.8
 Group Four  15.4±3.8  15.2±3.2
 Group One  0.58±0.12  0.53±0.08
 Group Two  0.61±0.11  0.46±0.06
 Group Three  0.60±0.06  0.55±0.04
 Group Four  0.59±0.11  0.53±0.14
 Group One  0.09±0.02  0.11±0.02
 Group Two  0.09±0.03  0.05±0.01
 Group Three  0.09±0.03  0.12±0.03
 Group Four  0.09±0.03  0.10±0.03


  1. Mean ± SD, DHA (docosahexaenoic acid. Group One (n=6) consumed algal DHA; Group Two (n=6) consumed high-DHA eggs; Group Three (n=6) consumed fish oil; Group Four (n=6; control) consumes two regular eggs
  2. Values with different superscripts letters indicate that the change from two to eight weeks postpartum was significantly different, P<0.05.

Other Findings

  • The maternal and infant plasma-phospholipid and breast milk contents of 22:5n-6 were significantly lower in Group Two (high-DHA eggs) than in the other groups at the end of the study period
  • There was a significant correlation between the contents of all long-chain (containing more than 18 carbons) n-3 and n-6 polyunsaturated fatty acids in maternal plasma phospholipids and the contents of these fatty acids in milk
  • The correlation between maternal plasma phospholipid DHA and milk DHA was particularly strong.


Author Conclusion:
  • The maternal and infant plasma-phospholipid and breast milk contents of 22:5n-6 were significantly lower in Group Two (high-DHA eggs) than in the other groups at the end of the study period
  • There was a significant correlation between the contents of all long-chain (containing more than 18 carbons) n-3 and n-6 polyunsaturated fatty acids in maternal plasma phospholipids and the contents of these fatty acids in milk
  • The correlation between maternal plasma phospholipid DHA and milk DHA was particularly strong
  • DHA supplementation increased the plasma and breast DHA concentrations of lactating women, resulting in higher phospholipid DHA concentrations in infants
  • It should be noted that a statistical significance (P<0.05) difference of only less than 1.6 SD is detectable with a sample size of only six subjects. Thus, if the small number of women in this study was typical of US women in general, it appears that any of the supplements evaluated would be acceptable as DHA supplements for lactating women.


It is unclear whether the higher plasma phospholipid DHA of the infants had functional benefits.

Funding Source:
Government: US Department of Agriculture
University/Hospital: Baylor Coleg of Medicine;University of Oklahoma
Reviewer Comments:

Randomized Clinical Trials (RCTs) are considered the gold standard of research practice and are accepted as producing the most reliable scientific evidence. The use of the RCT methodology contributes to improving the overall quality and rigor of the research. Within an RCT, participants are allocated at random to either a control group or experimental group, thereby eliminating all forms of spurious or additional causality. This ensures that both groups are statistically equivalent enabling valid conclusions to be drawn regarding the effect of a treatment or intervention on the experimental group. 

The limitations and critique of the study, as stated by the authors appear to be very appropriate.


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? Yes
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? Yes
  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? No
  5.1. In intervention study, were subjects, clinicians/practitioners, and investigators blinded to treatment group, as appropriate? No
  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.) No
  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? Yes
  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? Yes
  6.6. Were extra or unplanned treatments described? Yes
  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? Yes
  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)? Yes
  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