Research www. AJOG.

org

OBSTETRICS
Folic acid supplementation in early second trimester
and the risk of preeclampsia
Shi Wu Wen, MB, PhD; Xi-Kuan Chen, MD, PhD; Marc Rodger, MD; Ruth Rennicks White, RN, BScN;
Qiuying Yang, MD, PhD; Graeme N. Smith, MD, PhD; Ronald J. Sigal, MD, MPH;
Sherry L. Perkins, PhD; Mark C. Walker, MD, MSc

OBJECTIVE: The objective of the study was to evaluate the association
between folic acid supplementation in early second trimester and the
risk of developing preeclampsia.
STUDY DESIGN: We carried out a prospective cohort study between
October 2002-December 2005. We recruited women who had their pre-
natal care visit (12-20 weeks’ gestation) at the Ottawa Hospital and
Kingston General Hospital. All charts for participants with a diagnosis
of preeclampsia were audited and blindly adjudicated by 4 study inves-
tigators to validate the diagnosis.
RESULTS: A total of 2951 pregnant women were included in the final analysis.
Supplementation of multivitamins containing folic acid was associated with in-
creased serum folate (on average 10.51 ␮mol/L), decreased plasma homocys-
teine (on average 0.39 ␮mol/L), and reduced risk of preeclampsia (adjusted
odds ratio, 0.37; 95% confidence interval, 0.18-0.75).
CONCLUSION: Supplementation of multivitamins containing folic acid in the
second trimester is associated with reduced risk of preeclampsia.
Key words: 5, 10-methylenetetrahydrofolate reductase, folic acid,
homocysteine, preeclampsia, supplementation

Cite this article as: Wen SW, Chen X-K, Rodger M, et al. Folic acid supplementation in early second trimester and the risk of preeclampsia. Am J Obstet
Gynecol 2008;198:45.e1-45.e7.

P reeclampsia is hypertension and

proteinuria that develop during
pregnancy, affecting at least 5% of preg-
cardiovascular disease and diabetes in
the offspring of the affected mothers
through fetal origins of adult diseases.3
duce vascular integrity, resulting in the
pathophysiologic changes of preeclamp-
sia.4 However, which factors produced
nancies worldwide.1 It is a leading cause The current hypothesis for the patho- by the poorly perfused placenta are re-
of maternal and neonatal morbidity and genesis of preeclampsia is that factors sponsible for the development of pre-
mortality.1 Women with a history of pre- produced by the poorly perfused pla- eclampsia and how they interact with
eclampsia are at increased risk of cardio- centa enter the systemic circulation and maternal predisposing factors to induce
vascular disease in later life.2 Preeclamp- alter vascular sensitivity to circulating the clinical syndrome of preeclampsia
sia may also increase the risks of pressors, activate coagulation, and re- remain elusive.4
Recent studies have found that supple-
mentation of multivitamins containing
From the OMNI Research Group, Department of Obstetrics and Gynecology (Drs Wen,
folic acid was associated with reduced
Chen, Yang, and Walker and Ms White), and the Department of Epidemiology and
Community Medicine (Dr Wen), University of Ottawa Faculty of Medicine; the Clinical risk of preeclampsia.5 Folic acid may re-
Epidemiology Program, Ottawa Health Research Institute (Drs Wen, Chen, Rodger, Yang, duce the risk of preeclampsia by improv-
Sigal, and Walker and Ms White); and the Division of Hematology, Department of ing placental and systemic endothelial
Medicine (Dr Rodger), the Department of Medicine (Dr Sigal), and the Division of functions and directly or indirectly by
Biochemistry (Dr Perkins); Queen’s Perinatal Research Unit, Kingston General Hospital, lowering blood homocysteine levels.6-10
and the Department of Obstetrics and Gynecology, Queen’s University School of Medicine, The objective of this study was to com-
Kingston, ON (Dr Smith); and the Division of Endocrinology, University of Calgary, prehensively evaluate the association be-
Calgary, AB (Dr Sigal), Canada. tween folic acid supplementation, serum
Received Nov. 20, 2006; revised April 17, 2007; accepted June 29, 2007. folate, homocysteine, and 5, 10-methyl-
Supported in part by a grant from the Canadian Institute for Health Research (Grant MOP enetetrahydrofolate reductase (MTHFR)
53188). S.W.W., G.N.S., R.J.S., and M.C.W. are recipients of a New Investigators’ Award from thermolabile variant gene with the risk of
the Canadian Institute for Health Research. M.R. is a Clinical Investigator of the Heart and Stroke
preeclampsia.
Foundation of Canada, and X.-K.C. and Q.Y. are Canadian Institute for Health
Research/Strategic Training Initiatives of Research in Reproductive Health Sciences postdoctor
fellows.
M ATERIALS AND M ETHODS
Reprints: Shi Wu Wen, MB, PhD, OMNI Research Group, Department of Obstetrics and
The Ottawa and Kingston (OaK) Birth
Gynecology, University of Ottawa, Faculty of Medicine, 501 Smyth Rd, Box 241, Ottawa,
Ontario, Canada K1H 8L6; swwen@ohri.ca Cohort recruited nontransferred, con-
0002-9378/$34.00 • © 2008 Mosby, Inc. All rights reserved. • doi: 10.1016/j.ajog.2007.06.067 senting women between 12-20 weeks’
gestation during their prenatal visit at

JANUARY 2008 American Journal of Obstetrics & Gynecology 45.e1

Research Obstetrics
the Ottawa Hospital and Kingston Gen-
eral Hospital. The current analysis in-
cluded subjects from phase I of the OaK
Birth Cohort, which started in October
2002 and ended in December 2005. The
research nurses explained to pregnant
women the purpose of the study, what
would be expected from them, and what
they could expect from the study. For
participants who gave signed informed
consent, blood was drawn for genetic
and biochemical analyses. Twins or
higher order of multiples or subjects
with missing information on gestational
age or birth weight were excluded.
Demographic and clinical data were
collected by structured interview and
chart review. Additional chart review or
participant contact was performed if am-
biguities or missing data were encoun-
tered. Information on supplementation
of folic acid and other vitamins, includ-
ing brand name, date of initiation, and
date of discontinuation, was collected
both at recruitment and at delivery. Par-
ticipants were told that this study was
observation only and that during the
study we did not want them to change
anything regarding their daily life or
health care. Only women with regular
(daily) supplementation were counted.
These same questions were asked again
at the time of delivery to determine
whether there was any change in
supplementation.
Laboratory testing was performed to
determine serum folate levels and
plasma homocysteine levels and for the
presence of the MTHFR thermolabile
variant gene. Blood for MTHFR geno-
typing and homocysteine measurement
was collected in K2EDTA Vacutainer
tubes (Becton Dickinson, Lincoln Park,
NJ). Homocysteine specimens were put
on ice immediately after collection,
transported to the laboratory within 30
minutes, and centrifuged at 4°C for 10
minutes at 3000 g. Plasma was removed
and stored at -20°C until analysis. Sam-
ples were assayed in batches. Blood for
serum folate was collected in serum sep-
arator tubes (SST; Becton Dickinson).
The specimens were allowed to clot and
then be centrifuged for 10 minutes at
3000 g to separate serum, which was
stored at -20°C until analysis. Serum fo-
45.e2 American Journal of Obstetrics & Gynecology JANUARY 2008
late was measured on the Beckman
Coulter Access II using manufacturer’s
reagents (Beckman Coulter Inc, Fuller-
ton, CA). Homocysteine was measured
on the Abbott Ax Sym II (Abbott Labo-
ratories, Abbott Park, IL) using fluores-
cence polarization immunoassay tech-
nology. MTHFR genotyping was
conducted using the method of Don-
nelly and Rock.11
Preeclampsia was defined as having a
blood pressure of 140/90 mm Hg or
30/15 mm Hg above baseline with pro-
teinuria of 2⫹ on dipstick or 300 mg in
24-hour urine collection in women
greater than 20 weeks’ gestation. All
charts for participants with a diagnosis of
preeclampsia were audited and blindly
adjudicated by 4 study investigators
(S.W.W., M.R., R.W., and M.W.) to val-
idate the diagnosis.
The patterns of supplementation of
folic acid and other vitamins in preg-
nancy and the distribution of maternal
demographic and clinical characteristics
of the study participants were ascer-
tained. The effects of folic acid supple-
mentation on serum folate and plasma
homocysteine levels was then deter-
mined overall and stratified by gesta-
tional age at recruitment and by MTHFR
genotype. Finally, the effect of folic acid
supplementation on preeclampsia was
determined. Adjusted odds ratios
(aORs) and 95% confidence intervals
(CIs) for folic acid supplementation
were estimated by multiple logistic re-
gression analysis, with no supplementa-
tion as the reference.
Potential confounding variables in-
cluded in the regression models were
maternal age, ethnic background, educa-
tional level, parity, previous preeclamp-
sia, chronic hypertension, diabetes,
prepregnancy body mass index, house-
hold income, gestational age at recruit-
ment, and cigarette smoking during
pregnancy. The effects of low serum fo-
late, hyperhomocystinemia, and the
presence of the MTHFR thermolabile
variant gene on preeclampsia were also
examined. Low serum folate was defined
as folate concentration below the 10th
percentile, and hyperhomocystinemia
was defined as concentration higher than
the 90th percentile of the study popula-
www.AJOG.org
tion, after stratification by gestational
week at which the blood sample was
taken.
Additional analyses assessing the effect
of supplementation initiation time (be-
fore vs after conception) and discontin-
uation of supplementation in the third
trimester (yes vs no) were conducted. All
analyses were performed using SAS (ver-
sion 9.1; SAS Institute Inc, Cary, NC).
R ESULTS
A total of 4024 women were approached
to participate in the study; 3134 (78%)
agreed and were recruited. Among them,
70 women were excluded because of twin
births and 113 women were excluded be-
cause of missing information such as
gestational age at delivery, birthweight,
or sex (59) and lost to follow-up because
of the participant’s relocation outside
the study center (54), leaving 2951 sub-
jects for final analysis.
Ninety-two percent of the study sub-
jects were taking folic acid supplementa-
tion in the early second trimester, most
by taking multivitamins containing folic
acid at a dose of 1.0 mg or higher (Table
1). More than half of the women initi-
ated supplementation before conception
and approximately 20% discontinued
supplementation in the third trimester
partly or completely (Table 1).
The majority of the study participants
were white with high socioeconomic sta-
tus. Women with no supplementation
were more likely to be younger, multip-
arous, non-whites; to have lower educa-
tion level and household income; and to
smoke cigarettes during pregnancy than
women with supplementation (Table 2).
Folic acid supplementation was asso-
ciated with increased serum folate and
decreased plasma homocysteine (Table
3). The association between supplemen-
tation and serum folate and plasma ho-
mocysteine was stronger in blood sam-
ples taken at later gestation and
(especially) in participants with MTHFR
thermolabile variant genes (Table 3).
The rate of preeclampsia was lower in
the supplementation group than in the
no supplementation group, with an aOR
of 0.37 and 95% CIs 0.18-0.75 (Table 4).
Women with supplementation of folic
www.AJOG.org Obstetrics Research

A previous study reported an associa-

TABLE 1 tion between supplementation of multi-
Folic acid supplementation in pregnant women of the vitamins containing folic acid and re-
OaK Birth Cohort Study, October 2002-December 2005a duced risk of preeclampsia.5 Bodnar et
Number Percent al5 conducted a prospective cohort study
Category of supplementation in 1835 women in Pittsburgh, PA, be-
.....................................................................................................................................................................................................................................
Multivitamins containing folic acid 2016 74.30 tween 1997-2001 and found that regular
.....................................................................................................................................................................................................................................
use of multivitamins containing folic
Folic acid alone 250 9.22
..................................................................................................................................................................................................................................... acid at less than 16 weeks’ gestation was
Two or more types of vitamins 447 16.48 associated with a 45% reduction in pre-
..............................................................................................................................................................................................................................................
Dose of folic acid (mg) eclampsia risk, compared with nonusers
.....................................................................................................................................................................................................................................
0.1–0.9 114 4.20 (OR, 0.55; 95% CI, 0.32-0.95). This find-
.....................................................................................................................................................................................................................................
ing was consistent with ours. However, a
1.0 2152 79.32
..................................................................................................................................................................................................................................... previous study on this issue has several
1.1–1.9 178 6.56 weaknesses. The Pittsburgh study did
.....................................................................................................................................................................................................................................
ⱖ2.0 269 9.92 not conduct tests for genetics or blood
..............................................................................................................................................................................................................................................
Initiated before conception 1682 62.00 folate and homocysteine levels.5 Our
..............................................................................................................................................................................................................................................
study has the largest sample size with de-
Initiated after conception 1031 38.00
.............................................................................................................................................................................................................................................. tailed information on supplementation
Discontinued in third trimester (partly or completely) 544 20.10 patterns such as type, duration, and dose
..............................................................................................................................................................................................................................................
a
Doses of folic acid were derived from vitamin brands; doses of other vitamins in multivitamins were not presented in the of supplementation. We managed to
table; the ranges (medians) of doses for these vitamins were: vitamin A, 1000-4000 IU (1800 IU); vitamin C, 60-150 mg
(95.83 mg); vitamin B1, 1.4 to 3.0 mg (2.23 mg); vitamin B2, 1.4 to 3.75 mg (2.78 mg); vitamin B3, 10-40 mg ( 25 mg);
measure several aspects of folate metab-
vitamin B5, 0.03 to 10.0 mg (5.02 mg); vitamin B6, 1.9 to 10 mg (4.98 mg); vitamin B12, 2.6 to 14 ␮g (9.77 ␮g); vitamin olism, including genetics, supplementa-
D, 200-400 IU (360 IU); vitamin E, 25-30 IU (25.58 IU); beta-carotene,1050-3000 IU (2270 IU); biotin, 30-45 ␮g (36 ␮g);
niacin, 15-20 mg (18.25 mg); pantothenic acid, 6-10 mg (9 mg). Minerals were: calcium carbonate, 175-450 mg (250 mg);
tion, and blood folate and homocysteine
copper, 1-2 mg (1.6 mg); chromium, 25-30 ␮g (27 ␮g); iodine, 0.15 to 0.22 mg (0.16 mg); iron, 10-30 mg (22 mg); lutein, levels simultaneously. We used a pro-
250-300 ␮g (275 ␮g); magnesium, 50-100 mg (60 mg); manganese, 2-50 mg ( 16 mg); molybdenum, 25-50 ␮g (38 ␮g);
phosphorous, 125 mg (125 mg); selenium, 25-30 ␮g (28 ␮g); zinc, 7.5-25 mg (16.88 mg).
spective cohort study design and blindly
Wen. Folic acid supplementation and the risk of preeclampsia. Am J Obstet Gynecol 2008. adjudicated the outcome, which helped
to minimize bias.
There are compelling biologic ratio-
acid alone also had a lower rate of pre- Supplementation of multivitamins nale to believe that folic acid may reduce
eclampsia than those women who had containing folic acid was associated with the risk of developing preeclampsia. Pre-
no supplementation, although the dif- increased serum folate, lowered plasma eclampsia is likely a 2-stage disorder: at
ference was not statistically significant homocysteine, and reduced risk of pre- stage I (most likely at late first trimester
(Table 4). No significant associations be- eclampsia (by 63%). In our study, or early second trimester), a decreased
tween low serum folate level, hyperho- women with no supplementation were placental perfusion, secondary to abnor-
mocystinemia, and MTHFR thermo- of lower socioeconomic status, which mal placental developments, develops;
labile variant genes with risk of may lead to increased risk of preeclamp- and at stage II (most likely at the early
preeclampsia were found (Table 4). sia.1 On the other hand, women with no third trimester), the maternal syndrome
Whether women initiated supplementa- supplementation were more likely to be of preeclampsia, secondary to systemic
tion before or after conception or younger and multiparous and to smoke endothelial dysfunction, develops.4
whether women discontinued supple- cigarettes during pregnancy, which may Supplementation of large doses of fo-
mentation in the third trimester, the rate lead to a decreased risk of preeclamp- lic acid in early gestation may work at
of preeclampsia was similarly lower than sia.1,13 Thus, the potential confounding both stages of preeclampsia develop-
those women who did not have any sup- effects of maternal age, parity, socioeco- ment. Folic acid, or folate, is one of the B
plementation (Table 4). nomic status, and cigarette smoking may vitamins. It is a coenzyme in the produc-
have canceled each other and may have tion of nucleic acids and therefore is re-
C OMMENT limited impact on the observed associa- quired by all cells for growth. The pla-
Our prospective cohort study in a cohort tion between supplementation and pre- centa develops from a single cell to a
of Canadian women found that 92% had eclampsia. Our study did not observe an complex entity with a weight of about
supplementation with folic acid or mul- association between MTHFR gene mu- 500 g during pregnancy. An adequate
tivitamins containing folic acid in the tation and preeclampsia, which is consis- cellular folate supply may play an impor-
early second trimester, and among them, tent with the study by Powers et al.14 We tant role in the implantation and devel-
most (⬎95%) had a supplementation of speculate that high-level supplementa- opment of the placenta. Folate may also
1.0 mg or higher, twice the recom- tion of folic acid in our study population reduce the risk of developing preeclamp-
mended level for the prevention of neu- may have suppressed the genetic effect sia by improving endothelial function at
ral tube defects by Health Canada.12 on preeclampsia. both placental and systemic levels, di-

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Research Obstetrics www.AJOG.org

TABLE 2
Demographic and clinical characteristic of the pregnant women who participated
in the OaK Birth Cohort Study, October 2002-December 2005a
Variables Overall No supplementation Supplementation P value
Number of subjects 2951 238 2713
................................................................................................................................................................................................................................................................................................................................................................................
Maternal age (y)
.......................................................................................................................................................................................................................................................................................................................................................................
⬍25 6.40 13.45 5.79 ⬍.0001
.......................................................................................................................................................................................................................................................................................................................................................................
25-29 21.82 23.11 21.71
.......................................................................................................................................................................................................................................................................................................................................................................
30-34 40.09 36.55 40.40
.......................................................................................................................................................................................................................................................................................................................................................................
ⱖ35 31.68 26.89 32.10
................................................................................................................................................................................................................................................................................................................................................................................
Maternal background
.......................................................................................................................................................................................................................................................................................................................................................................
Aboriginal 0.75 0.42 0.77 ⬍.0001
.......................................................................................................................................................................................................................................................................................................................................................................
White 83.46 74.38 84.26
.......................................................................................................................................................................................................................................................................................................................................................................
Middle Eastern 5.29 7.56 5.09
.......................................................................................................................................................................................................................................................................................................................................................................
African 3.42 10.08 2.84
.......................................................................................................................................................................................................................................................................................................................................................................
Asian 7.08 7.56 7.04
................................................................................................................................................................................................................................................................................................................................................................................
2
Prepregnancy body mass index (kg/m )
.......................................................................................................................................................................................................................................................................................................................................................................
⬍18.5 (underweight) 6.03 5.46 6.08 .1492
.......................................................................................................................................................................................................................................................................................................................................................................
18.5-24 (normal) 60.01 59.24 60.08
.......................................................................................................................................................................................................................................................................................................................................................................
25-29 (overweight) 20.40 17.23 20.68
.......................................................................................................................................................................................................................................................................................................................................................................
ⱖ30 (obesity) 13.55 18.07 13.16
................................................................................................................................................................................................................................................................................................................................................................................
Education level
.......................................................................................................................................................................................................................................................................................................................................................................
High school and below 13.15 27.97 11.85 ⬍.0001
.......................................................................................................................................................................................................................................................................................................................................................................
College/university not completed 9.34 10.17 9.27
.......................................................................................................................................................................................................................................................................................................................................................................
College/university completed 77.51 61.86 78.88
................................................................................................................................................................................................................................................................................................................................................................................
Nulliparous 34.46 23.53 35.42 .0002
................................................................................................................................................................................................................................................................................................................................................................................
Household income ($,CAD)
.......................................................................................................................................................................................................................................................................................................................................................................
⬍25,000 6.76 18.50 5.75 ⬍.0001
.......................................................................................................................................................................................................................................................................................................................................................................
25,000-49,999 14.07 22.47 13.35
.......................................................................................................................................................................................................................................................................................................................................................................
50,000-79,999 27.76 27.31 27.80
.......................................................................................................................................................................................................................................................................................................................................................................
ⱖ80,000 51.41 31.72 53.10
................................................................................................................................................................................................................................................................................................................................................................................
Smoking during pregnancy 7.73 13.03 7.27 .0014
................................................................................................................................................................................................................................................................................................................................................................................
Chronic hypertension 1.58 2.14 1.53 .4761
................................................................................................................................................................................................................................................................................................................................................................................
Type 1 diabetes 0.82 1.29 0.78 .4151
................................................................................................................................................................................................................................................................................................................................................................................
Type 2 diabetes 0.86 0.86 0.86 .9990
................................................................................................................................................................................................................................................................................................................................................................................
Previous preeclampsia 3.96 4.62 3.91 .5879
................................................................................................................................................................................................................................................................................................................................................................................
Gestational age at recruitment (wks)
.......................................................................................................................................................................................................................................................................................................................................................................
ⱕ12 33.82 27.31 34.39 .2079
.......................................................................................................................................................................................................................................................................................................................................................................
13-15 45.21 49.58 44.82
.......................................................................................................................................................................................................................................................................................................................................................................
16-20 20.98 23.11 20.79
................................................................................................................................................................................................................................................................................................................................................................................
a
Because of missing information in some variables: smoking (2), chronic hypertension (39), type 1 diabetes (40), type 2 diabetes (40), numbers in this table did not add up.
Wen. Folic acid supplementation and the risk of preeclampsia. Am J Obstet Gynecol 2008.

45.e4 American Journal of Obstetrics & Gynecology JANUARY 2008

www.AJOG.org Obstetrics Research

TABLE 3
Serum folate and plasma homocysteine levels in subjects with folic acid supplementation vs those without,
by gestational age at recruitment and MTHFR genotype, OaK Birth Cohort Study, October 2002-December 2005
Variables No supplementation Supplementation P value
Folate level (mean ⫾ SD, ␮mol/L)a 25.79 ⫾ 8.60 36.30 ⫾ 7.99 ⬍ .0001
................................................................................................................................................................................................................................................................................................................................................................................
Folate level stratified by gestational weeks at recruitment (mean ⫾ SD, ␮mol/L)
.......................................................................................................................................................................................................................................................................................................................................................................
ⱕ12 25.93 ⫾ 8.78 37.00 ⫾ 7.71 ⬍ .0001
.......................................................................................................................................................................................................................................................................................................................................................................
13-15 26.42 ⫾ 8.69 36.63 ⫾ 7.82 ⬍ .0001
.......................................................................................................................................................................................................................................................................................................................................................................
16-20 24.27 ⫾ 8.15 34.35 ⫾ 8.52 ⬍ .0001
................................................................................................................................................................................................................................................................................................................................................................................
Folate level stratified by MTHFR genotype (mean ⫾ SD, ␮mol/L) b
.......................................................................................................................................................................................................................................................................................................................................................................
677CC 26.93 ⫾ 8.47 36.24 ⫾ 8.16 ⬍ .0001
.......................................................................................................................................................................................................................................................................................................................................................................
677CT 24.77 ⫾ 7.96 36.25 ⫾ 7.79 ⬍ .0001
.......................................................................................................................................................................................................................................................................................................................................................................
677TT 24.29 ⫾ 10.43 36.63 ⫾ 8.14 ⬍ .0001
................................................................................................................................................................................................................................................................................................................................................................................
Homocysteine level (mean ⫾ SD, ␮mol/L) c
4.69 ⫾ 1.24 4.32 ⫾ 1.01 ⬍ .0001
................................................................................................................................................................................................................................................................................................................................................................................
Homocysteine level stratified by gestational weeks at recruitment (mean ⫾ SD, ␮mol/L)
.......................................................................................................................................................................................................................................................................................................................................................................
ⱕ12 4.61 ⫾ 1.02 4.43 ⫾ 1.02 .1868
.......................................................................................................................................................................................................................................................................................................................................................................
13-15 4.85 ⫾ 1.36 4.36 ⫾ 1.02 .0002
.......................................................................................................................................................................................................................................................................................................................................................................
16-20 4.43 ⫾ 1.13 4.04 ⫾ 0.94 .0043
................................................................................................................................................................................................................................................................................................................................................................................
Homocysteine level stratified by MTHFR genotype (mean ⫾ SD, ␮mol/L) b
.......................................................................................................................................................................................................................................................................................................................................................................
677CC 4.58 ⫾ 1.07 4.28 ⫾ 0.99 .0019
.......................................................................................................................................................................................................................................................................................................................................................................
677CT 4.59 ⫾ 1.15 4.31 ⫾ 1.04 .0199
.......................................................................................................................................................................................................................................................................................................................................................................
677TT 5.43 ⫾ 1.83 4.48 ⫾ 0.96 .0112
................................................................................................................................................................................................................................................................................................................................................................................
a
Thirty-five subjects missing information on serum folate level.
b
Forty-four subjects missing information on genotype
c
36 subjects missing information on serum homocysteine
Wen. Folic acid supplementation and the risk of preeclampsia. Am J Obstet Gynecol 2008.

rectly or indirectly by its effect on lower- preeclampsia development, may be the between serum folate and plasma homo-
ing blood homocysteine level.7,15-17 most important. cysteine levels with preeclampsia col-
Endothelial dysfunction is demonstra- We did not observe an association be- lected blood sample in late gestation or at
ble within the myometrial arteries of tween serum folate or plasma homocys- delivery.19,20 For those studies that col-
women with preeclampsia, and the incu- teine level with preeclampsia. In large- lected blood samples in the early second
bation of healthy vessels within plasma scale epidemiologic studies, it is difficult trimester, the prediction of hyperhomo-
obtained from women with preeclamp- to tightly control for factors that may af- cystinemia on preeclampsia was
sia induces similar endothelial changes.6 fect the measured values of folate or ho- poor.19,2,22
Young women with folate deficiency or mocysteine. Variations in gestational age These findings suggest that variation
hyperhomocystinemia may be prone to and timing at which blood samples were by gestational period is a major obstacle
not only systemic endothelial dysfunc- taken, folate consumption by the grow- in the study of the cause-and-effect rela-
tion but also placental endovasculature.9 ing fetus, and other physical and patho- tionship between a particular metabolite
In our study, whether women initiated logical processes that may have an im- and a particular pregnancy outcome. In-
folic acid supplementation before or af- pact on the metabolism of folate/ creasing study samples to precisely mea-
ter conception or whether the women homocysteine during pregnancy may sure gestation period specific values in
discontinued supplementation in the introduce variability in the measure- future studies may help determine the
third trimester, the rate of preeclampsia ment of folate/homocysteine.18 association between metabolites and
was similarly lower than in those women It is rather difficult to distinguish pregnancy outcomes. Using biomarkers
who did not have supplementation, sug- cause and effect for biomarkers that are with longer half-life biomarkers (eg, red
gesting that for preeclampsia preven- the intermediate steps of a health prob- blood cell folate) may be helpful as well.
tion, folic acid supplementation in the lem occurring in a rather short period of These markers measure not only the cur-
late first trimester or early second trimes- time, such as preeclampsia. Most previ- rent status but also the previous status.
ter, the most critical time window for ous studies that observed an association Because the development of specific

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Research Obstetrics www.AJOG.org

TABLE 4
Occurrences of pregnancy complications according to folic acid supplementation status, serum folate level,
plasma homocysteine level, and MTHFR genotype, OaK Birth Cohort Study, October 2002-December 2005
Number of
Variables subjects Preeclampsia ORs (95% CIa)
Folic acid supplementation
.......................................................................................................................................................................................................................................................................................................................................................................
No 238 12 (5.04) Reference
.......................................................................................................................................................................................................................................................................................................................................................................
Yes 2713 59 (2.17) 0.37 (0.18-0.75)
................................................................................................................................................................................................................................................................................................................................................................................
Supplementation of folic acid alone
.......................................................................................................................................................................................................................................................................................................................................................................
No supplementation 238 12 (5.04) Reference
.......................................................................................................................................................................................................................................................................................................................................................................
Yes 421 12 (2.85) 0.46 (0.16-1.31)
................................................................................................................................................................................................................................................................................................................................................................................
Initiation time of supplementation
.......................................................................................................................................................................................................................................................................................................................................................................
Before conception 1031 24 (2.33) Reference
.......................................................................................................................................................................................................................................................................................................................................................................
After conception 1682 35 (2.08) 1.04 (0.59-1.82)
................................................................................................................................................................................................................................................................................................................................................................................
Discontinuation of supplementation in late gestation
.......................................................................................................................................................................................................................................................................................................................................................................
Yes 544 14 (2.57) Reference
.......................................................................................................................................................................................................................................................................................................................................................................
No 2169 45 (2.07) 0.77 (0.41-1.46)
................................................................................................................................................................................................................................................................................................................................................................................
b
Serum folate level
.......................................................................................................................................................................................................................................................................................................................................................................
Normal (ⱖ10th percentile for each gestational week) 2649 64 (2.42) Reference
.......................................................................................................................................................................................................................................................................................................................................................................
Low (⬍10th percentile for each gestational week) 302 7 (2.32) 1.22 (0.52-2.90)
................................................................................................................................................................................................................................................................................................................................................................................
Serum homocysteine level (␮mol/L) c
.......................................................................................................................................................................................................................................................................................................................................................................
Normal (⬍90th percentile for each gestational week) 2529 56 (2.21) Reference
.......................................................................................................................................................................................................................................................................................................................................................................
High (ⱖ90th percentile for each gestational week) 422 15 (3.55) 1.25 (0.66-2.36)
................................................................................................................................................................................................................................................................................................................................................................................
d
Genotype
.......................................................................................................................................................................................................................................................................................................................................................................
677CC 1282 29 (2.26) Reference
.......................................................................................................................................................................................................................................................................................................................................................................
677CT 1286 35 (2.72) 1.27 (0.75-2.15)
.......................................................................................................................................................................................................................................................................................................................................................................
677TT 339 6 (1.77) 0.70 (0.28-1.77)
................................................................................................................................................................................................................................................................................................................................................................................
OR, odds ratio.
a
Adjusted for maternal age, ethnic background, education level, parity, history of preeclampsia, chronic hypertension, diabetes, prepregnancy body mass index, household income, gestational age
at recruitment, and cigarette smoking.
b
Thirty-five subjects missing information on serum folate level.
c
Thirty-six subjects missing information on serum homocysteine.
d
Forty-four subjects missing information on genotype.
Wen. Folic acid supplementation and the risk of preeclampsia. Am J Obstet Gynecol 2008.

health problem is often a process rather mins. Second, recent randomized taining folic acid (odds ratio, 0.37), other
than a point effect, biomarkers of accu- controlled trials found that supplemen- vitamins (eg, vitamin B6) may also play
mulation measures may be better mea- tation with vitamins C and E (without some role in the prevention of
sures of the cause-and-effect relation- folic acid) during pregnancy at doses preeclampsia.
ship than biomarkers of point measures. many times higher than the ones in our Our study has several weaknesses.
Most women in our study had supple- study had no protective effect on pre- The sample sizes for women with no
mentation of multivitamins containing eclampsia.23,24 Third, in the subgroup of supplementation or with supplemen-
folic acid. There are several reasons for us women who had supplementation of fo- tation at the doses of ⬍1.0 mg or ⬎1.0
to believe that folic acid may have played lic acid alone (n ⫽ 421), we did observe a mg folic acid or with folic acid alone
a more important role in preeclampsia statistically nonsignificant trend toward were small. As a result, we could not
than other vitamins. First, there is strong similar protective effect on preeclamp- find statistically significant association
biologic rationale to believe that folic sia. However, because the effect of sup- of supplementation with folic acid
acid may reduce the risk of developing plementation by folic acid alone (odds alone or perform a dose-response anal-
preeclampsia. No similar biologic mech- ratio, 0.46) was smaller than supplemen- ysis. Although we adjusted for several
anisms could be found for other vita- tation dominated by multivitamins con- obstetric, socioeconomic, and lifestyle

45.e6 American Journal of Obstetrics & Gynecology JANUARY 2008

www.AJOG.org
factors in our analysis, residual con-
founding may exist. We did not collect
data on folic acid intake from food.
Women with folic acid supplementa-
tion were of a higher socioeconomic
status and therefore may have higher
folic acid from food intake. Ray and
Mamdani25 studied hospital discharge
data in the Canadian province of On-
tario before (1990-1997) and after
(1998-2000) mandatory folic acid food
fortification and found no change in
the rate of preeclampsia, suggesting
that folic acid from food intake was too
low to make any impact on the risk of
preeclampsia.
The findings of our study and others
give hope of a new prevention strategy
for preeclampsia, which needs further
evaluation. Randomized, controlled tri-
als could provide definitive evidence re-
garding the relationship between folic
acid supplementation and the risk of
preeclampsia. Findings from observa-
tional studies such as the current one can
help the design of future randomized tri-
als (eg, when the supplementation
should be initiated and what dose should
be used, etc) and to establish the equi-
poise for future randomized trials.21 f
ACKNOWLEDGMENTS
We acknowledge the following people because
without their commitment and expertise, this
project would not have been possible: the
OMNI Research Nurse Group (Ottawa and Lucy
Chura, RN, BScN, study coordinator, Queen’s
Perinatal Research Unit research team
(Queen’s Perinatal Research Unit Kingston and
Lizy Kodiattu, MD); Dr. George Tawagi and
team (Ottawa Hospital–Civic Campus); and
Carol Ann Jodouin, Department of Laboratory
Medicine, Ottawa Hospital.
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