CLINICAL TRIALS

Dietary Docosahexaenoic Acid Supplementation in

Children With Autism

Robert G. Voigt, yMichael W. Mellon, zSlavica K. Katusic, Amy L. Weaver, Dietrich Matern,


Bryan Mellon, jjCraig L. Jensen, and William J. Barbaresi

ABSTRACT
Objective: The aim of the study was to determine whether docosahexaenoic acid (DHA) supplementation improves the behavior of children with
autism.
Methods: A group of 3- to 10-year-old children with autism were
randomized in a double-blind fashion to receive a supplement
containing 200 mg of DHA or a placebo for 6 months. The parents
and the investigator completed the Clinical Global ImpressionsImprovement scale to rate changes in core symptoms of autism after 3
and 6 months. The parents completed the Child Development Inventory
and the Aberrant Behavior Checklist, and both parents and teachers
completed the Behavior Assessment Scale for Children (BASC) at
enrollment and after 6 months.
Results: A total of 48 children (40 [83%] boys, mean age [standard
deviation] 6.1 [2.0] years) were enrolled; 24 received DHA and 24
placebo. Despite a median 431% increase in total plasma DHA levels
after 6 months, the DHA group was not rated as improved in core
symptoms of autism compared to the placebo group on the CGI-I.
Based on the analysis of covariance models adjusted for the baseline
rating scores, parents (but not teachers) provided a higher average rating
of social skills on the BASC for the children in the placebo group
compared to the DHA group (P 0.04), and teachers (but not parents)
provided a higher average rating of functional communication on the
BASC for the children in the DHA group compared to the placebo group
(P 0.02).
Conclusions: Dietary DHA supplementation of 200 mg/day for 6 months
does not improve the core symptoms of autism. Our results may have been
limited by inadequate sample size.

Received February 26, 2013; accepted November 19, 2013.

From the
Department of Pediatric and Adolescent Medicine, the
yDepartment of Psychology and Psychiatry, the zDepartment of Health
Sciences Research, the Department of Laboratory Medicine and Pathology, Mayo Clinic, Rochester, MN, the jjDepartment of Pediatrics,
Baylor College of Medicine, Houston, TX, and the Department of
Medicine, Childrens Hospital, Boston, MA.
Address correspondence and reprint requests to Robert G. Voigt, MD,
Department of Pediatrics, Baylor College of Medicine, Meyer Center
for Developmental Pediatrics, Texas Childrens Hospital, 8080 North
Stadium Drive, Suite 180, Houston, TX 77054 (e-mail: rgvoigt@bcm.
edu).
http://clinicaltrials.gov registration no.: NCT00577447.
The study was supported by funding from the Mayo Foundation. The DHA
supplements and placebos used in this study were provided by Martek
Biosciences Corporation (Columbia, MD).
C.L.J. has been a consultant and has received payment for lectures and for
development of educational presentations from Mead Johnson Nutritional. The other authors report no conflicts of interest.
Copyright # 2014 by European Society for Pediatric Gastroenterology,
Hepatology, and Nutrition and North American Society for Pediatric
Gastroenterology, Hepatology, and Nutrition
DOI: 10.1097/MPG.0000000000000260

JPGN

Volume 58, Number 6, June 2014

Key Words: autism, complementary and alternative therapy, dietary

supplements, docosahexaenoic acid, fish oil, v-3 fatty acids, pervasive
developmental disorders

(JPGN 2014;58: 715722)

imilar to other chronic neurodevelopmental disorders,

traditional medicine does not offer any cure for autism, and
the best evidence-based treatments for this condition involve intensive early behavioral and special educational strategies (1,2).
Although psychopharmacological therapies for autism have been
studied for >50 years, there are presently no US Food and Drug
Administration-approved indications for the treatment of the core
symptoms of autism for any agent (3). As a result, complementary
and alternative treatments are widely provided to children with
autism by parents desperate for effective biomedical interventions
for their children (4). Perhaps acting on suspicion or distrust of
standard medical practices, a desire not to have their children
drugged or the desire to seek curative treatment because of
the frustration with deficiencies in traditional medical interventions,
therapies based on dietary interventions appeal to parents of children with autism as more safe, natural, and holistic approaches to
treating their children (5).
At present, there is considerable interest in the possibility that
dietary supplementation with the long chain, n-3 polyunsaturated
fatty acid, docosahexaenoic acid (DHA, 22:6n-3), will decrease the
symptoms of autism. DHA is an important structural lipid of brain
cell membranes, and it is available in the diet through consumption
of fish. It has been estimated that approximately 55% of children
with autism take psychotropic medications or dietary supplements
(6,7) and that 23% of children with autism are specifically using n-3
or fish oil supplements (8), despite the lack of any evidence to
support the safety and/or efficacy of this treatment. Thus, the
primary objective of this study was to test the hypothesis that
dietary DHA supplementation is a safe and effective treatment for
reducing symptoms of autism using a randomized, double-blind,
placebo-controlled design.

METHODS
Study Design
Children from 3 to 10 years of age with autism were recruited
and randomized in a double-blind manner to a placebo-controlled
trial of dietary DHA supplementation. The primary outcome
measure was a significant positive response on the Clinical Global
Impressions-Improvement (CGI-I) scale (9) at the completion of the
study. Secondary outcome measures were changes in behavior or
development noted on the Aberrant Behavior Checklist (ABC) (10),
the Behavior Assessment Scale for Children (BASC) (11), and the
Child Development Inventory (CDI) (12) completed at the baseline

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Voigt et al
and at the study completion. The patterns for plasma phospholipid
fatty acid were drawn on all of the subjects at study entry and
conclusion. This study was approved by the Mayo Clinic institutional review board, and written informed consent was obtained
from a parent or guardian before enrollment of any child.

Study Subjects
Subjects were recruited through the placement of recruitment
flyers across the Mayo Clinic campus at Rochester, MN. These
flyers were also sent to local and regional autism support groups to
be distributed to members and at meetings, workshops, and conferences. The parents and/or guardians of children with autism who
responded as possible study volunteers were first screened by the
study coordinator by telephone to determine whether their children
were between 3 and 10 years of age and had been diagnosed to have
autistic disorder. The subjects were excluded if they had a diagnosis
of pervasive developmental disordernot otherwise specified or
Asperger disorder. The subjects were also excluded if they had used
a dietary supplement containing DHA within 90 days of study
inclusion or had a medical history of a disorder of lipid metabolism.
Consistent with practice guidelines for the diagnosis of
autism, which report the criterion standard for an autism diagnosis to be the clinical judgment of an experienced clinician (13),
children who met these inclusion and exclusion requirements were
scheduled for a confirmatory medical diagnostic evaluation with a
single developmental-behavioral pediatrician to confirm that each
subject met the Diagnostic and Statistical Manual of Mental
Disorders, Fourth Edition (DSM-IV) criteria for autistic disorder
(14). The Childhood Autism Rating Scale (CARS) (15), a standardized quantitative measure of autism symptomatology, was also
administered as part of this confirmatory consultation, and only
subjects who scored in the autistic range on the CARS (scores  30)
were included. Thus, only those subjects who both met DSM-IV
criteria for autistic disorder on confirmatory medical diagnostic
assessment and scored in the autistic range on the CARS were
offered enrollment in the study.

Supplementation
All of the participants, families, and study personnel (including those performing baseline and outcome assessments) were
blinded to group assignment for the entire study. All of the subjects
were randomized in a double-blind fashion to receive either a
triglyceride oil capsule containing 200 mg DHA from algal oil
with high oleic acid sunflower oil to make 500 mg total oil or a
placebo capsule containing 250 mg of corn oil and 250 mg of
soybean oil, daily for 6 months. The oils were orange flavored, and
the capsules were identical in appearance. All study capsules were
provided by Martek Biosciences Corporation (Columbia, MD).
The supplements were dispensed by the Mayo Clinic Investigational Pharmacy according to a randomization scheme stratified
by sex and generated using a block approach by the study statistician. The number of capsules dispensed was always somewhat
more than needed before the next appointment, and subjects were
instructed to return unused capsules at that time. Comparison of the
number of capsules returned to the number that should have been
returned was used to monitor compliance, which was excellent.
The choice of DHA dose involved both biochemical and
practical considerations. The dose selected (200 mg/day) has been
shown to significantly increase erythrocyte and plasma phospholipid DHA levels (16). A higher dose was considered but administration of even 1 study capsule per day is often problematic in
this population.

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Volume 58, Number 6, June 2014

Our placebo capsules were designed to be identical in

appearance to the study capsules. They needed to contain oil,
similar to the DHA-containing capsules, but to serve as an appropriate control they could not contain longer-chain v-3 fatty acids.
Corn and soybean oil were used in the placebo capsules because
they have a similar texture and are composed of fatty acids, but not
longer-chain v-3 fatty acids. This allowed us to determine the
specific effects of the long-chain v-3 fatty acid, DHA, compared to
nonv-3 fatty acids. Soybean oil contains some a-linolenic acid
(5%14% of total fatty acids), which is an 18-carbon v-3 fatty
acid that is a precursor of DHA; however, the contribution of this in
terms of affecting DHA status would be negligible. Corn oil
contains virtually no a-linolenic acid (1% of total fatty acids).
Longer-chain v-3 fatty acids have been generally accepted as
essential for neurological membrane functioning. The corn and
soybean oil in the placebo capsules did not contain any of these
neurologically critical longer-chain v-3 fatty acids, and there have
never been any reports of corn or soybean oils improving symptoms
of children with autism.

Participation
The study consisted of a baseline confirmatory diagnostic
medical evaluation, along with follow-up medical evaluations after
3 and 6 months of supplementation with a single developmentalbehavioral pediatrician. Blood samples were obtained via venipuncture from the children at the baseline and after 6 months. The
parent or guardian completed a demographic questionnaire at the
baseline visit. Behavioral, developmental, and adverse effect questionnaires were completed by the parent/guardian and/or investigator at each visit as outlined below.

Plasma Phospholipid Fatty Acid Patterns

Blood samples (5 mL) were obtained at the baseline and after
6 months of supplementation to determine plasma phospholipid
fatty acid patterns. All analyses were performed by the Mayo Clinic
Biochemical Genetics Laboratory using an established assay (17).
In brief, a 2-step, acid-base hydrolysis was followed by hexane
extraction and derivatization with pentafluorobenzyl bromide.
Separation and detection were accomplished by capillary gas
chromatography electron-capture negative ion-mass spectrometry.
Quantitation was based on analysis in the selected ion-monitoring
mode by using 13 stable isotope-labeled internal standards. In this
study, the baseline plasma phospholipid levels in children with
autistic disorder were compared to pediatric reference ranges
established at the Mayo Clinic to detect nutritional deficiencies
and metabolic disorders involving C8-C26 fatty acids (17).

Behavioral and Developmental Outcome

Measures
The parents and/or guardians of all of the children enrolled in
this study completed specific questionnaires at the baseline and at 3
and 6 months after supplementation. The behavioral and developmental outcome measures selected for this study were based on
consensus recommendations in the medical literature (3), and prior
published studies investigating effects of alternative medicine (18)
and psychopharmacological (19) interventions in children with
autism. The primary aim in selecting these measures was to find
measures that have been shown to be most sensitive to behavioral
changes as a result of short-term biomedical interventions. All of the
outcome measures selected for this study are valid and reliable
measures of the childhood behavior and development. Prior studies
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JPGN

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have indicated that global measures of intelligence, adaptive behavior, language, and autistic symptomatology, which have been
designed for diagnostic purposes, are not sensitive to assessing
effects of biomedical therapeutic interventions (3). Regardless of
the objective of the study involving children with autism, the CGI-I
scale is considered to be the single measure that should be used
universally in all of the clinical trials of children with autism (3).
Thus, changes in the core symptoms of autism (social interaction,
communication, repetitive/stereotyped behavior) were assessed by
CGI-I scores derived from both parents and investigator ratings after
3 and 6 months of supplementation. The CGI-I score scale consists
of a 7-point Likert scoring system with scores ranging from 1 (very
much improved) to 7 (very much worse). Significant positive
responses were defined as either a 1 (very much improved) or a
2 (much improved).
Several secondary outcome measures of the behavior and
development were included in this study to provide data on a wide
range of developmental and behavioral outcome incident to DHA
supplementation. The ABC (10) was completed by parents at the
baseline and after 6 months of supplementation. The ABC is a
58-item parent-completed rating scale that was developed to
measure the effects of pharmacological interventions in children
with developmental disabilities, and it is a recommended outcome
measure in clinical trials of children with autism (3,18). The ABC
consists of 5 subscales: irritability, lethargy, stereotypy, hyperactivity, and inappropriate speech. The CDI (12) was completed
by parents and the BASC (11) was completed by both parents and
teachers at the baseline and after 6 months of supplementation. The
CDI consists of 9 subscales and is a 300-item yes-no questionnaire that offers a valid, reliable, and easily administered assessment
of a childs overall general development, and their development in
social, self-help, fine motor, gross motor, and expressive and
receptive language development. The BASC includes both parentand teacher-rating scales (18 and 20 subscales, respectively) that
provide reliable and valid measures of a broad sample of behaviors,
including attention span, hyperactivity, social skills, adaptability,
functional communication, and atypical behaviors (11).

Adverse Effect Assessment

The parents and/or guardians were instructed to contact the
study coordinator at any time throughout the study with concerns
regarding possible adverse effects. To monitor for safety of the
supplementation, the Treatment Emergent Symptoms Scale (TESS)
(20) was completed via parent interview after 3 and 6 months of
supplementation. The TESS is a standardized rating scale that
measures the occurrence or nonoccurrence of 24 potential adverse
effects of medication in 6 categories (gastrointestinal, urinary,
respiratory, skin, neurological, and psychological). Each adverse
effect is scored on a 3-point Likert scale, with 2 indicating severe, 1
mild, and 0 absent adverse effects.

Statistical Analysis
The data were summarized using standard descriptive statistics. The baseline demographic and clinical characteristics were
compared between the 2 treatment groups, using the Fisher exact
test for categorical variables, the 2-sample t test for age, and the
Wilcoxon rank sum test for parental education and family income
categories. The questionnaire responses obtained during the followup visits at 3 and 6 months were compared between the 2 treatment
groups based on the participating subjects. The proportions with a
significant positive response on each of the CGI-I scales were
compared between the 2 groups using the Fisher exact test. In
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Trial of Dietary DHA Supplementation in Children With Autism

addition, the full 7-point CGI-I ratings were compared between the
2 groups using the Wilcoxon rank sum test. The CDI, ABC, and
BASC scores at 6 months of treatment were compared between the
2 groups based on fitting separate analysis of covariance models
to adjust for the corresponding baseline scores. All calculated
P values were 2-sided and P < 0.05 were considered statistically
significant. All analyses were performed using the SAS version 9.1
software package (SAS Institute, Cary, NC).
In adherence with the intent-to-treat principle, an additional
analysis was performed for the primary outcome variable, the
overall CGI-I scale at 6 months. In this analysis, all of the subjects
were analyzed according to their randomized treatment assignment,
regardless of their level of compliance, and subjects who dropped
out before the end of the study were classified as not having a
significant positive response.
The primary outcome variable for this study was defined as a
significant positive response on the 7-point CGI-I scale. A significant positive response was defined as either a 1 (very much
improved) or 2 (much improved) on at least 1 of the subcomponents
of this scale. Using a power of 0.8, a significance level of 0.05, and a
2-sided x2 test, the study was designed to enroll 32 subjects in each
group to have sufficient statistical power to detect a 25% difference
in the proportion of subjects, with a positive CGI-I response in the
DHA-supplemented group relative to the placebo group.

RESULTS
Study Subjects
The CONSORT (21,22) flow diagram for this study is shown
in Figure 1. A total of 48 children with autism (40 [83%] male, mean
age [standard deviation] 6.1 [2.0] years) were enrolled; 24 received
DHA and 24 placebo (Fig. 2). There were no differences between
the groups in demographic factors (age, sex, race, parental education, family income) or baseline total plasma phospholipid or n-3
fatty acid levels (Table 1). A group of 34 children completed the
study (19 in DHA group, 15 in placebo group).

Plasma Total Fatty Acids

Table 2 shows the total plasma n-3 fatty acid levels in the 48
children with autistic disorder at the baseline. Only 3 (6%) children
were found to have low DHA levels and 1 (2%) had a low
eicosapentaenoic acid (20:5n-3) level, but none had low a-linolenic
acid (ALA; 18:3n-3), docosapentaenoic acid (DPA; 22:5n-3), or
total n-3 fatty acid levels. In addition, 10 children (21%) were found
to have high DHA levels, 11 (23%) high ALA levels, 4 (8%) high
eicosapentaenoic acid levels, and 7 (15%) high total n-3 fatty acid
levels. The mean values for each of these n-3 fatty acids were within
the normal reference ranges. Figure 2 shows the mean total plasma
n-3 fatty acid levels after 6 months of treatment. All of the children
in the DHA group evidenced a significant increase in their plasma
phospholipid DHA levels from the baseline to study conclusion,
confirming their compliance with taking the DHA supplements.
The DHA group showed a median 431% increase in total plasma
DHA levels after 6 months of supplementation.

Core Symptoms of Autism

Table 3 shows the number and percentage of children with a
significant positive response from parent and investigator CGI-I
ratings after 3 and 6 months of treatment. We found no statistically
significant differences in the percentage with a positive response
between the groups in either parental or investigator CGI-I ratings
after 3 or 6 months of treatment; however, when the ratings were

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Voigt et al

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Enrollment

Volume 58, Number 6, June 2014

Assessed for eligibility (n = 143)

Excluded (n = 95)
Not meeting inclusion criteria (n = 52)
Declined to participate (n = 43)

Randomized (n = 48)

Allocation
Allocated to placebo (n = 24)

Allocated to DHA (n = 24)

3-month follow-up
Discontinued intervention (n = 8)
- Inconvenient to participate (n = 3)
- Trouble taking supplement (n = 2)
- Parent concerned about supplement side
effects (n = 3)

Discontinued intervention (n = 2)
- Inconvenient to participate (n = 1)
- Trouble taking supplement (n = 1)

Participated in 3-month visit (n = 16)

Participated in 3-month visit (n = 22)

6-month follow-up
Discontinued intervention (n = 1)
- Trouble taking supplement (n = 1)

Discontinued intervention (n = 3)
- Inconvenient to participate (n = 3)

Participated in 6-month visit (n = 15)

Participated in 6-month visit (n = 19)

FIGURE 1. CONSORT flow diagram.

analyzed using the full 7-point scale, the DHA-supplemented group

received slightly worse investigator ratings of repetitive/stereotypic
behaviors at 6 months compared to the placebo group (P 0.02, as
per the Wilcoxon rank sum test; percentage scored as 3, 4, and 5 are

Micromole/liter

1000
800
600
400
200
0
DHA

DPA
Placebo Group

EPA

ALA

Total

DHA Group

FIGURE 2. Mean total plasma n-3 fatty acid levels after 6 months of
treatment. DHA docosahexaenoic acid.

718

0%, 83%, and 17% in the DHA group and 33%, 60%, and 7% in the
placebo group, respectively).
In addition, the overall CGI-I rating scale at 6 months was
analyzed based on considering all of the 48 randomized subjects. In
this analysis, subjects who dropped out before the end of the study
were classified as not having a significant positive response. Among
the 24 patients randomized to receive DHA, 5 (21%) subjects had a
significant positive response based on the parent ratings for the
overall CGI-I rating scale at 6 months, compared to 2 (8%) subjects
randomized to receive placebo (95% exact confidence interval for
difference in proportions 17.6 to 41.0). Based on the investigator
ratings at 6 months, none of the subjects had a significant positive
response, compared to just 1 (4%) subject randomized to receive
placebo (95% exact confidence interval for difference in proportions 33.4 to 25.6).

Developmental-Behavioral Questionnaires
The CDI, ABC, and BASC scores at 6 months of treatment
were compared between the 2 treatment groups, after accounting for
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JPGN

Volume 58, Number 6, June 2014

Trial of Dietary DHA Supplementation in Children With Autism

TABLE 1. Summary of the baseline demographic and clinical characteristics, by treatment assignment

Male, n (%)
Age at baseline, y
Mean (SD)
Range
Race, n (%)
Not recorded
White
Nonwhite
Age of parent/guardian, y
Mean (SD)
Range
Maximal parent education, n (%)
Not recorded
Diploma or GED
Tech/VoTech degree
Some college
College degree
Masters degree
Doctoral degree
Family income, n (%)
Median category
Not recorded
$1520K
$2530K
$3540K
$4075K
$75100K
$100150K
$150200K
>$200K
n-3 fatty acid, mean, mmol/L (SD)
ALA
EPA
DPA
DHA
Total n-3

Placebo group (N 24)

DHA group (N 24)

20 (83)

20 (83)

0.99
0.20

6.5 (2.2)
3.410.7

5.8 (1.8)
3.39.6

5
13 (68)
6 (32)

2
19 (86)
3 (14)

38.2 (6.5)
23.647.9

40.8 (4.8)
31.148.0

0.26

0.16

0.19
4
0
2 (10)
5 (25)
9 (45)
3 (15)
1 (5)

4
1 (5)
1 (5)
4 (20)
4 (20)
6 (30)
4 (20)

$4075K
4
0
0
2 (10)
8 (40)
4 (20)
3 (15)
1 (5)
2 (10)

$75100K
4
2 (10)
1 (5)
0
3 (15)
5 (25)
2 (10)
4 (20)
3 (15)

0.33

95.3
53.6
120.6
116.3
379.2

(54.5)
(31.0)
(50.7)
(80.7)
(176.9)

96.5
55.7
126.5
117.7
395.8

(48.6)
(36.6)
(43.5)
(82.5)
(157.4)

0.94
0.83
0.67
0.95
0.73

ALA a-linolenic acid; DHA docosahexaenoic acid; DPA docosapentaenoic acid; EPA eicosapentaenoic acid; GED General Educational
Development; SD standard deviation.


Comparisons were evaluated using the Fisher exact test for sex and race, the Wilcoxon rank sum test for maximal parent education and family income, and
the 2-sample t test for all other characteristics.

the baseline scores. Among the 52 subscales that were evaluated,

only 2 statistically significant differences were found (Table 4). On
average, parents (but not teachers) provided a more favorable
average change in their rating of social skills on the BASC for
the children in the placebo group compared to those in the DHA
group (mean change, 3.0 vs 0.2). In addition, teachers (but not

parents) provided a more favorable average change in their rating of

functional communication on the BASC for the children in the DHA
group compared to those in the placebo group (mean change, 1.5 vs
4.5); however, after adjusting for the multiple comparisons using
a Bonferroni correction, none of these comparisons reached statistical significance with a P < 0.01.

TABLE 2. Total plasma n-3 fatty acid levels measured in micromoles per liter in 48 children with autistic disorder at baseline
n-3 fatty acid

Mean (SD)

ALA
EPA
DPA
DHA
Total n-3

95.9
54.6
123.6
117.0
387.5

(51.1)
(33.6)
(46.8)
(80.7)
(166)

Median

Range

Reference range

85.0
45.5
120.5
90.0
400

29270
6195
37256
5364
100800

20120
890
30270
30160
100500

ALA a-linolenic acid; DHA docosahexaenoic acid; DPA docosapentaenoic acid; EPA eicosapentaenoic acid; SD standard deviation.

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Voigt et al

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Volume 58, Number 6, June 2014

TABLE 3. Parent and investigator CGI-I ratings after 3 and 6 months of treatment

Placebo group

No. subjects with significant positive responses (%)

Parent CGI-I ratings at 3 mo
Social interaction
Communication
Stereotyped behavior
Overall behavior
Investigator CGI-I ratings at 3 mo
Social interaction
Communication
Stereotyped behavior
Overall behavior
Parent CGI-I ratings at 6 mo
Social interaction
Communication
Stereotyped behavior
Overall behavior
Investigator CGI-I ratings at 6 mo
Social interaction
Communication
Stereotyped behavior
Overall behavior

DHA group

5/16
6/16
4/16
5/16

(31)
(38)
(25)
(31)

6/22
7/22
4/22
4/21

(27)
(32)
(18)
(19)

1.00
0.74
0.70
0.46

1/13
0/13
0/13
0/13

(8)
(0)
(0)
(0)

1/17
1/17
1/17
1/17

(6)
(6)
(6)
(6)

1.00
1.00
1.00
1.00

6/15
7/15
1/15
2/13

(40)
(47)
(7)
(15)

5/19
6/19
3/18
5/18

(26)
(32)
(17)
(28)

0.47
0.48
0.61
0.67

0/15
0/15
0/15
1/14

(0)
(0)
(0)
(7)

0/18
0/18
0/18
0/18

(0)
(0)
(0)
(0)

0.44

CGI-I Clinical Global Impressions-Improvement; DHA docosahexaenoic acid.

The CGI-I scale consists of a 7-point scoring system with scores ranging from 1 (very much improved) to 7 (very much worse). Significant positive
responses were defined as either a 1 (very much improved) or a 2 (much improved).

Adverse Effect Assessment

The parents or the guardians of children in the placebo group
did not report any severe level of adverse effects on the TESS. One
parent of a child in the DHA group reported a severe level of
headaches and 1 a severe level of restlessness at 3 months but not at
6 months, and 1 parent of a child in the DHA group reported a severe
level of agitation at 6 months (but not at 3 months). Overall, there
was no significant difference between groups in treatment-emergent
adverse effects.

DISCUSSION
Autism is a neurodevelopmental disability characterized by
clinically significant impairments in social interaction and communication, in association with atypical repetitive, stereotypic, and
ritualistic behaviors (14). The prevalence of autism appears to be
increasing, likely in large part the result of increased awareness and
improved identification and the availability of federally mandated
early intervention services (23). In a study, we found the incidence
of autism to be 2.9/1000 in our geographic region (Olmsted County,
MN) (23). In 2012, the Autism and Developmental Disabilities
Monitoring Network of the Centers for Disease Control and Prevention reported the prevalence of autism to be 1 in every 88

children (24). The prevalence of autism is greater in the pediatric

population than that of diabetes, congenital heart disease, cystic
fibrosis, inflammatory bowel disease, and chronic renal disease
combined, making autism a major public health concern (25).
The epidemiological data suggest that the population with
lower dietary n-3 fatty acid consumption and, hence, lower plasma
and, presumably, brain contents of n-3 fatty acids, including DHA,
has higher rates of psychiatric disorders, including depression and
bipolar disorder (26,27). In addition, children with attention-deficit/
hyperactivity disorder (ADHD) (28) and adults with schizophrenia
(29) have been shown to have lower plasma phospholipid DHA
contents than controls. DHA is the predominant long-chain polyunsaturated fatty acid (LC-PUFA) present in the structural lipids of
brain cell membranes, especially those at the synaptic terminals
(3033). DHA and other long-chain polyunsaturated fatty acid may
influence synaptic function directly through effects on membrane
structure, and indirectly through the generation of eicosanoids/
docosanoids (prostaglandins, leukotrienes, thromboxanes, resolvins, neuroprotectins) or through immune system/cytokine interactions (34,35). Thus, there has been considerable interest in the
role of DHA in the etiology and treatment of a variety of psychiatric
and neurodevelopmental disorders, including depression (36),
schizophrenia (37), aggression (38), bipolar disorder (39),

TABLE 4. Significant differences between the treatment groups on developmental-behavioral rating scales
Placebo group

DHA group

BASC rating scale

No.

Baseline mean (SD)

6-mo mean (SD)

No.

Baseline mean (SD)

6-mo mean (SD)

Parentsocial skills
Teacherfunctional communication

12
8

30.3 (9.1)
38.5 (4.3)

33.3 (9.7)
34.0 (5.6)

13
11

26.5 (7.1)
32.2 (7.6)

26.3 (6.8)
33.6 (9.3)

0.04
0.02

BASC Behavior Assessment Scale for Children; DHA docosahexaenoic acid; SD standard deviation.


For each rating scale, an analysis of covariance model was fit to compare the 6-month scores between the 2 treatment groups, after adjusting for the
corresponding baseline scores.

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Volume 58, Number 6, June 2014

Alzheimer disease (4042), dyslexia (43), ADHD (4446), and

developmental coordination disorder (43,47).
Children with autism tend to be extremely picky eaters as a
result of their ritualistic tendencies (48), and this may result in a
lack of appropriate nutrients, including DHA intake. Based on
neuroanatomical and functional MRI data, it appears that the
symptoms of autism may be mediated by the medial-temporal
lobe (hippocampus and amygdala) and the cerebellum (49,50).
Interestingly, animal studies have shown that the hippocampus and
cerebellum are the most vulnerable brain structures to effects of
malnutrition (51,52). Permanent alterations are often seen in these
structures, whereas other brain areas tend to recover. If this is also
true in humans, it can be hypothesized that the picky eating of
children with autism may result in a lack of appropriate nutrient
intake, including DHA. As DHA is a critical component of brain
cell membranes, a lack of dietary DHA, and hence, low levels of
DHA in the plasma and brain, would be expected to affect the
cognitive processes mediated by these most nutritionally sensitive
brain areas and result in autistic symptomatology. Two of 3 prior
studies have reported lower levels of v-3 fatty acids in children
with autism compared to controls (5355). We, however, did not
find children with autism to have consistently low plasma DHA or
other n-3 fatty acid levels, as 94% of our subjects had normal or
high DHA levels compared to the reference range determined by
our laboratory.
In addition to any long-term effects owing to altering the
structural membranes of neurons, DHA supplementation may act in a
more short-term pharmacological manner. A direct relation between
plasma phospholipid DHA content and metabolism of serotonin and
dopamine within the central nervous system has been reported
(56,57). In addition, limbic areas of the medial-temporal lobe (amygdala and hippocampus), critical for emotional expression and social
behavior, are richly innervated with serotonergic neurons, and while
serotonergic innervation of the cerebellum is less prominent, serotonin projections are also critical to cerebellar function (58); however,
although we found that dietary DHA supplementation was safe (few
treatment-emergent adverse effects reported), despite a median
increase in total plasma DHA levels of 431%, we did not find that
DHA supplementation improved the core symptoms of autism based
on either parental or investigator ratings. We also found that DHA
supplementation did not generally affect the development or behavior
of children with autism based on both parental and teacher ratings.
Teachers (but not parents) reported an improved functional communication for children in the DHA group compared to those in the
placebo group; however, after adjusting for multiple comparisons,
this improvement no longer reached statistical significance.
Our findings are potentially limited by our small sample size.
Although the study was designed to enroll 32 patients per group,
we were only able to enroll 24 subjects per group and retain 15
and 19 subjects per group at 6 months. Thus, our study had 80%
power to detect a difference between group means of 1 standard
deviation between the DHA-supplemented and placebo groups after
6 months. Therefore, our study could not determine whether DHA
supplementation may produce more subtle developmental or behavioral improvements in children with autism; however, although
this study may not detect smaller differences, improvements of a
magnitude less than that designed to be detected in this study may
not be clinically meaningful for children with autism. It is also
possible that supplementation with a higher dose of DHA for a
longer time period or that supplementation with a combination of
n-3 fatty acids may produce different results. Numerous prior
studies have, however, found DHA supplementation for as little
as 2 to 4 months to result in clinically significant improvements in
the neurobehavioral symptoms of ADHD (46,47), bipolar disorder
(38), and developmental coordination disorder (46), and thus our
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Trial of Dietary DHA Supplementation in Children With Autism

6-month study duration should have been long enough to capture
similar neurobehavioral changes in children with autism.
Another limitation of our study is that although we found a
median increase in total plasma DHA levels of 431% in our DHA
supplemented group, it is unknown whether plasma DHA levels
reflect levels in neuronal membranes in children, as results from
the required postmortem studies have not been reported. In 1 study,
the correlation coefficient between brain cortex DHA and erythrocyte DHA in infants who underwent postmortem examinations
because of sudden death was 0.33 (59). It is, however, unclear to
what degree these results would apply to our study population, and
extrapolation from results of animal studies is problematic.

CONCLUSIONS
Despite concerns about nutritional deficiency derivative of
their picky eating, we did not find children with autism to be
deficient in DHA or other n-3 fatty acids at the baseline. In addition,
although dietary DHA supplementation is a widely popular treatment for children with autism, we did not find this intervention to
improve the core symptoms of autism or a broad range of associated
developmental and behavioral difficulties in children with autism,
although children who received the supplementation experienced a
431% median increase in their plasma total fatty acid DHA levels;
however, although none of our comparisons reached statistical
significance after adjusting for multiple comparisons, our finding
of a favorable change in functional communication reported by
teachers in children with autism who received DHA supplementation may be confirmed by further investigation in a larger randomized, placebo-controlled clinical trial.
Acknowledgments: We thank our study coordinator Candice
Klein and all of the children and families who volunteered to
participate in this study. We also thank Martek Biosciences
Corporation (Columbia, MD) for providing the DHA supplements
and placebos used in this study.

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