1. Objectives

To assess the effects of vitamin C supplementation, alone or in combination with other supplements, on pregnancy outcomes, adverse events, side effects and the use of health resources

2. How studies were identified

The following databases were searched in March 2015:

• Cochrane Pregnancy and Childbirth Group’s Trials Register
• CENTRAL (The Cochrane Library 2015)
• MEDLINE
• Embase
• CINAHL

Relevant journals, conference proceedings and reference lists were also searched

3. Criteria for including studies in the review

3.1 Study type

Randomized controlled trials, including quasi-randomized trials

3.2 Study participants

Pregnant women living in areas where dietary vitamin C intakes were considered either adequate or inadequate

3.3 Interventions

Vitamin C supplementation, alone or in combination with other supplements, compared with placebo or no supplementation

(Studies using multivitamin supplements containing more than two micronutrients in one tablet were excluded)

3.4 Primary outcomes

Maternal

• Preterm birth (<37 weeks’ gestation)
• Prelabour rupture of fetal membranes (PROM, preterm or term)
• Clinical preeclampsia

Neonatal

• Stillbirth
• Neonatal death, perinatal death
• Intrauterine growth restriction (birth weight <3^rd centile or the most extreme centile reported)

Secondary outcomes for the mother included death up to six weeks postpartum, bleeding episodes (placental abruption, antepartum haemorrhage, postpartum haemorrhage, complications of epidural anaesthesia, need for transfusion), measures of serious maternal morbidity (eclampsia, liver failure, renal failure, disseminated intravascular coagulation, pulmonary oedema, peripheral neuropathy), elective birth (induction of labour or elective caesarean section), caesarean section (emergency plus elective), measures of iron and folate status (such as iron-deficiency anaemia, megaloblastic anaemia, serum iron, serum ferritin, haemoglobin concentrations, cord serum ferritin, cord serum haemoglobin), placental weight, measures of wound healing, maternal satisfaction with care, antenatal hospital admission, visits to day care units, use of intensive care, ventilation, and dialysis. Secondary outcomes for the infant included infant death, birth weight, gestational age at birth, congenital malformations, Apgar score less than seven at five minutes, jaundice requiring phototherapy, respiratory distress syndrome, chronic lung disease, periventricular haemorrhage, bacterial sepsis, necrotising enterocolitis, retinopathy of prematurity, bronchopulmonary dysplasia, haemolytic anaemia, infantile scurvy, disability at childhood follow-up (cerebral palsy, intellectual disability, hearing or visual impairment), poor childhood growth, admission to neonatal intensive care unit, duration of mechanical ventilation, length of stay in hospital, development, and special needs after discharge. Maternal and infant side effect outcomes included adverse events related to vitamin C sufficient to cease supplementation, diarrhoea, kidney stone formation, excessive iron overload, gastrointestinal disturbances, and vitamin B12 deficiency

4. Main results

4.1 Included studies

Twenty-nine randomized controlled trials, enrolling 24,300 women, were included in this review

• Nine trials recruited women at high or increased risk of preeclampsia and two trials included women with established preeclampsia; one trial included women with a history of preterm birth, and four trials involved women with established preterm PROM or a history of preterm PROM; three trials enrolled women with no acute or chronic diseases; one study enrolled women who smoked during pregnancy; one trial enrolled pregnant and non-pregnant women with abnormal vaginal flora; one trial enrolled pregnant women with leg cramps; and one trial enrolled women with planned caesarean section after 35 weeks’ gestation
• Supplementation was initiated in the second trimester in most trials; however, the timing of initiation ranged from four weeks’ gestation to 35 weeks’ or more
• In 12 trials vitamin C was provided alone; in 17 trials vitamin C was given in combination with vitamin E, and in two of these trials another intervention was included (aspirin and fish oil or allopurinol); in six trials other additional supplements were given to both vitamin C and control groups, including iron, folic acid, vitamin B and/or calcium or a “standard prenatal vitamin”
• More than half of all trials (15/29 trials) provided 1000 mg of vitamin C per day; six trials used 500 mg/day; four trials provided 100 mg/day; two trials used 2000 mg/day; one trial provided 400 mg/day; one trial provided 250 g per day for six days followed by 250 g per week; and in one trial vitamin C was administered vaginally

4.2 Study settings

• Australia (2 trials), Brazil, India (2 trials), Iran (4 trials), Latvia, Mexico (2 trials), South Africa (2 trials), Sweden, Turkey, Uganda (2 trials), the United Kingdom of Great Britain and Northern Ireland (2 trials), the United States of America (4 trials), Venezuela, and multi-site (3 trials: India, Peru, South Africa, and Viet Nam; Canada and Mexico; and the United Kingdom of Great Britain and Northern Ireland and the Netherlands)
• In seven trials assessing vitamin C intakes or status at baseline, overall intakes were regarded as adequate. Two trials were conducted in populations regarded to have low nutritional status, including low vitamin C intakes; in the remaining trials, vitamin C intakes at baseline were not assessed or reported

4.3 Study settings

How the data were analysed
One comparison was made: vitamin C in combination with other supplements versus placebo or no supplementation. Dichotomous data were summarized using risk ratios (RR) and 95% confidence intervals (CI), and continuous data were summarized using mean differences (MD) and 95% CI. When 10 or more trials were included in a meta-analysis, funnel plot asymmetry was assessed for publication bias. Sensitivity analyses were conducted by excluding studies with overall high or unclear risk of bias. Where substantial clinical or statistical heterogeneity (I²>30%) was present, random effects models were used. To explore potential sources of heterogeneity, the following subgroup analyses were planned for primary outcomes:

• Dosage of vitamin C: ≥60 mg vitamin C (the recommended daily intake) versus <60 mg vitamin C
• Gestational age at trial entry: <20 weeks versus ≥20 weeks
• Dietary adequacy of vitamin C intakes: low intake (less than the recommended daily intake in that setting as measure by dietary questionnaire or plasma vitamin C <11 μmol/L at trial entry) versus adequate intake
• Vitamin C alone versus in combination with other micronutrients
• Risk of adverse pregnancy outcomes at study entry (as defined by trialists)

Results
Vitamin C alone or in combination with other supplements
Primary outcomes
No evidence of a difference between vitamin C and control groups was found for the risk of preterm PROM (RR 0.98, 95% CI [0.70 to 1.36], 10 trials/16,825 women), term PROM (RR 1.23, 95% CI [0.61 to 2.47], 3 trials/3230 women), preterm birth (RR 0.99, 95% CI [0.90 to 1.10], 16 trials/20,250 women) or clinical preeclampsia (RR 0.92, 95% CI [0.80 to 1.05], 16 trials/21,956 participants). No statistically significant difference between groups was found for the infant primary outcomes stillbirth (RR 1.15, 95% CI [0.89 to 1.49], 11 trials/20,038 participants), neonatal death (RR 0.79, 95% CI [0.58 to 1.08], 11 trials/19,575 participants), perinatal death (RR 1.07, 95% CI [0.77 to 1.49], 7 trials/17,271 participants), or intrauterine growth restriction (RR 0.98, 95% CI [0.91 to 1.06], 12 trials/20,361 participants).

Assessment of reporting bias, sensitivity analyses and subgroup analyses for primary outcomes
Funnel plots for preterm birth, stillbirth, neonatal death, preterm PROM and intrauterine growth restriction did not show any evidence of publication bias, while the funnel plot for clinical preeclampsia was asymmetric. In sensitivity analysis of trials at low risk of bias, a statistically significant 73% increase in the risk of term PROM was found with vitamin C supplementation (RR 1.73, 95% CI [1.34 to 2.23], p=0.00003; 2 trials/3060 participants). Results did not alter meaningfully in any other sensitivity analyses excluding trials at risk of bias, although heterogeneity was reduced for the outcomes preterm birth, term PROM, and clinical preeclampsia. The risk of preterm PROM was reduced in subgroup analysis of one trial in which women were recruited exclusively after 20 weeks’ gestation (RR 0.31, 95% CI [0.11 to 0.89], p=0.03; 109 participants; p=0.09 for subgroup differences). In further subgroup analysis by gestation at trial entry, a reduced risk of clinical preeclampsia was found in one trial in which gestational age at enrolment was not specified (RR 0.28, 95% CI [0.10 to 0.74], p=0.01; 580 women; p=0.02 for subgroup differences). The risk of clinical preeclampsia was reduced in subgroup analysis of women with low baseline serum vitamin C concentrations, but this finding did not reach statistical significance (RR 0.14, 95% CI [0.02 to 1.05], p=0.05; 1 trial/28 women), and no effect on preeclampsia was found in subgroup analysis of trials conducted in populations with low nutritional status (RR 1.01, 95% CI [0.84 to 1.22], 2 trials/2188 participants). In subgroup analysis of trials in which vitamin C was given alone, a 34% reduction in the risk of preterm PROM (RR 0.66, 95% CI [0.48 to 0.91], p=0.0098; 5 trials/1282 women; p=0.01 for subgroup differences), and a 45% reduction in the risk of term PROM was found (RR 0.55, 95% CI [0.32 to 0.94], p=0.028; 1 trial/170 women; p=0.0002 for subgroup differences). For trials in which vitamin C was given with other supplements, no clear effect was found on preterm PROM (RR 1.28, 95% CI [0.86 to 1.92], 5 studies/15,543 women), but for term PROM, a 73% increase in risk was found (RR 1.73, 95% CI [1.34 to 2.23], p=0.000031; 2 trials/3060 participants). Subgroup analyses by baseline risk of adverse pregnancy outcome did not reveal any substantial differences in results. Subgroup analyses by dosage of vitamin C were not conducted as all trials provided more than 60 mg of vitamin C per day.

Additional outcomes
The risk of placental abruption was reduced with vitamin C, alone or combination with other supplements, in comparison to a control or no supplementation (RR 0.64, 95% CI [0.44 to 0.92], p=0.017; 8 trials/15,755 women). No clear effect of vitamin C was found for other maternal secondary outcomes, including antepartum haemorrhage, maternal death, any caesarean section, prelabour caesarean section, induction of labour, eclampsia, renal failure or renal insufficiency, disseminated intravascular coagulation, pulmonary oedema, antenatal hospitalization or admission to adult intensive care unit. Gestational age at birth was increased with vitamin C supplementation (MD 0.31 weeks, 95% [0.01 to 0.61], p=0.044; 9 trials/14,062). No significant effect of vitamin C supplementation was found for other infant secondary outcomes, including infant death, mean birth weight, bacterial sepsis, necrotising enterocolitis, bronchopulmonary dysplasia, congenital malformations, Apgar score less than seven at five minutes, respiratory distress syndrome, jaundice requiring phototherapy, periventricular haemorrhage, periventricular leukomalacia, retinopathy of prematurity, admission to neonatal intensive care unit, or use of mechanical ventilation. No other pre-specified secondary outcomes were reported on.

Adverse events and side effects
None of the included trials reported on adverse events sufficient to cease vitamin C supplementation. In one trial involving 1877 participants, the risk of abdominal pain was increased in the treatment group (RR 1.66, 95% CI [1.16 to 2.37], p=0.0059). No evidence of a difference between treatment and control groups was found for other adverse outcomes, including elevated liver enzymes, acne, transient weakness, skin rash, heartburn and nausea, or any side effect.

5. Additional author observations*

Of the 29 included trials, 11 were judged to be at low overall risk of bias, 10 were assessed as being at unclear risk of bias, and eight were judged to be at high risk of bias. Using GRADE quality of evidence assessment criteria, the outcomes intrauterine growth restriction, preterm birth and placental abruption were rated as high quality, the outcomes stillbirth and clinical preeclampsia were rated as moderate quality, and the outcome preterm PROM was rated as low quality. While there were no clear differences in findings when subgrouped by vitamin C status or intake, few trials contributed data to these analyses.

Supplementation with vitamin C in pregnancy was associated with a reduced risk of placental abruption, but it is unclear whether this was due to vitamin C, vitamin E, or to both micronutrients. While the risk of preterm and term PROM was reduced when vitamin C was given alone, the risk of term PROM was increased when vitamin C was given together with vitamin E, suggesting that vitamin E may be implicated in this finding. Although mean gestational length at birth was increased with vitamin C supplementation, no difference between treatment and control groups was found for the risk of preterm birth. An increased risk of abdominal pain was found in one trial in which vitamin E was also provided. Overall, the findings do not support the routine use of vitamin C supplementation during pregnancy.

Further research into the mechanisms underlying the effect of vitamin C on the risks of preterm and term PROM and placental abruption is warranted. Furthermore, additional research is needed into the effects of vitamin C supplementation in women with inadequate intakes of vitamin C. Continuing follow-up of women and children in previous or ongoing trials would provide insight into the potential long-term effects of vitamin C supplementation during pregnancy.