![[Feedback]](/icons/banner/feedbackACT.gif){kind=icon}
![[For Subscribers]](/icons/banner/subscriberACT.gif){kind=icon}
![[Archive]](/icons/banner/archiveACT.gif){kind=icon}
![[Search]](/icons/banner/searchACT.gif){kind=icon}
![[Contents]](/icons/banner/tocACT.gif){kind=icon}
|
|
|
|
|
||
3 Nutrition and Health Sciences Program, and 4 Hubert Department of Global Health, Rollins School of Public Health, Emory University, Atlanta, GA 30322; 5 Pan American Health Organization, Washington, DC 20037; and 6 Institute of Nutrition of Central America and Panama, Calzada Roosevelt, Guatemala City, Guatemala
* To whom correspondence should be addressed. E-mail: rmart77{at}sph.emory.edu.
 |
ABSTRACT |
|---|
 TOP ABSTRACT IntroductionMaterials and MethodsResultsDiscussionLITERATURE CITED |
|---|
|
Introduction |
|---|
|
TOPABSTRACTIntroductionMaterials and MethodsResultsDiscussionLITERATURE CITED |
|---|
0.1% of births or 300,000 infants are affected globally each year (1). NTD contribute considerably to the global burden of fetal and infant mortality, morbidity, and disability.
Although there are no national data about folic acid (FA) deficiency in Guatemala, different studies have reported a high incidence of NTD (2,3), suggesting that FA deficiency is a significant public health concern. The incidence of NTD is particularly high in the western highlands of Quetzaltenango, Guatemala, with an estimated rate of 106/10,000 live births (3). Even though this rate is already high, it is substantially underestimated because stillbirths, which are known to include many malformations, were not considered. Vitamin B-12 deficiency is also common in Guatemala. A study of school children in peri-urban areas of Guatemala reported that 11% had low plasma vitamin B-12 and that an additional 22% had marginal vitamin B-12 concentrations (4). Findings from a study in lactating women also showed that plasma vitamin B-12 was deficient or low in 46.7% of the mothers and was associated with depletion of the vitamin in their infants (5). Additionally, 60% of childbearing age women in peri-urban areas of Guatemala had low vitamin B-12 concentrations (6).
FA and vitamin B-12 play a critical role in the methylation pathways that convert homocysteine (Hcy) to S-adenosylmethionine, a compound that has been shown to be involved in several transmethylation reactions, including DNA methylation. Deficiency of FA can inhibit DNA biosynthesis or methylation reactions, which in turn prevent the proper closure of the neural tube during fetal development (7). In addition, deficiencies of FA and/or vitamin B-12 can lead to high Hcy levels. Elevated Hcy is a risk factor for various chronic diseases such as cardiovascular and cognitive diseases (8,9). Furthermore, some studies report associations between elevated Hcy and increased rates of eclampsia and other poor pregnancy outcomes (10,11). Finally, maternal Hcy concentrations may also play a role in the etiology of fetal malformations such as NTD (12,13).
Several intervention studies have confirmed the efficacy of daily supplementation of FA in improving folate status, lowering Hcy concentrations, and reducing the risk of having a child with NTD. A 4000-µg daily FA supplement has been shown to reduce the risk of recurrence of NTD by 72% in a multicenter study (14) and 400 µg FA taken daily has been proven to reduce the risk of occurrence of NTD by 79% in the northern provinces and by 41% in the southern provinces of China (15). However, daily supplementation programs are likely to have low effectiveness because of more complicated logistical factors and poorer adherence. Little is known about whether weekly FA supplementation has the same impact on folate status as daily supplementation. One study of New Zealand women compared the effect of weekly vs. daily FA supplementation (16,17) and reported that a weekly FA supplement was as effective as a daily supplement in lowering Hcy concentrations (16) but that weekly was less effective than daily in improving folate status (17).
The objective of our study was to compare the efficacy of different regimens of FA supplementation, namely, very high-dose weekly FA (5000 µg), high-dose weekly FA (2800 µg), high-dose daily FA (400 µg), and low-dose daily FA (200 µg), in improving folate and vitamin B-12 status and lowering Hcy concentrations in healthy women of reproductive age in Guatemala.
|
Materials and Methods |
|---|
|
TOPABSTRACTIntroductionMaterials and MethodsResultsDiscussionLITERATURE CITED |
|---|
Study site. The study was conducted in the village of Concepción, Chiquirichapa, in the department of Quetzaltenango, Guatemala. This village is located 2600 m above sea level and lies 40 km to the west of Guatemala's second largest city, Quetzaltenango. Village residents are primarily of indigenous Mayan descent, particularly of the Mam culture, and of low socioeconomic status (SES). The community is agrocentric and key crops grown include potatoes and broccoli.
Design. The study design was approved by the Ethical Committee of Hospital Roosevelt, Guatemala City, Guatemala and Emory University's Institutional Review Board, Atlanta, Georgia. There were 4 arms to the double-blind, randomized trial (Table 1). The doses of FA were 400 and 200 µg for daily and 5000 and 2800 µg for weekly supplementation. The 400 µg daily was chosen because it is the current recommended dietary allowance (RDA) for women of reproductive age for NTD risk reduction (18) and the 200 µg daily was chosen because evidence from previous studies shows improvement with this dose in folate status (19,20) and Hcy levels (21,22). The 2800 µg weekly was a simple calculation of 7 times the 400-µg daily RDA, whereas the 5000 µg weekly was selected based on the recommendation for women of reproductive age by the Guatemalan Ministry of Public Health (23). Additionally, there was 1 daily and 1 weekly dose of iron sulfate, zinc sulfate, and vitamin B-12 (Table 1). Supplementation lasted 12 wk, between March and June 2006.
|
Subject recruitment, randomization to treatment, and compliance
A census of the community was conducted by trained personnel to identify women between the ages of 15 and 49. Women were not admitted into the study if any of the following conditions were present: 1) pregnancy; 2) lactation (having had a child within the last 3 mo); 3) consumption of FA supplements; 4) chronic diseases that interfere with FA metabolism; and 5) severe anemia (Hb < 70 g/L). Women with severe anemia were provided 90 tablets containing 120 mg of iron for daily use and referred to the local health clinic.
A total of 459 women were enrolled in the study. Computer-generated random numbers were used to assign women to 1 of 4 supplemental groups. Supplementation was double-blinded. All supplements were identical in appearance and taste and were coded with lot numbers at the factory (Industrias Bioquímicas), corresponding to 1 of 4 treatment arms. The code allocation was kept secure at Emory University and was only revealed after completion of the trial. Trained field workers from the community visited each woman 7 d/wk to deliver and observe the ingestion of the supplements for the entire 12-wk duration of supplementation. All women received 7 pills per week. The weekly dose groups received 6 placebos and 1 active pill on the 3rd day of the week. Daily records were kept to track the participants' health. The trial was registered in the U.S. NIH Clinical Trials registry (identification no. NCT00394862).
Data collection
Blood collection. Venous blood samples were collected after overnight fasting at baseline and at the end of 12 wk of supplementation. Venous blood was centrifuged at 1500 x g; 10 min at 4°C. The serum was separated and stored at –70°C at the Institute of Nutrition of Central America and Panama (INCAP) in Guatemala City until analysis at the National Institute of Public Health, Cuernavaca, Mexico.
Dietary intake information. We collected dietary intake information at baseline using a semiquantitative FFQ developed by INCAP. This FFQ was shown to be valid for use in Guatemalan adults (24), and was adapted for use in the Mam indigenous region.
Other data collection. Participants were interviewed at baseline for information on their demographic background, level of education, current marital status, reproductive history, and health status. SES was assessed at baseline using a structured questionnaire that included questions related to household size, occupation, and educational level of the participants, water and sanitation, and household possessions. All interviewers were bilingual in Mam and Spanish and interviews were conducted in Mam or Spanish as required.
Laboratory analysis
Serum and RBC folate were determined by microbiological assays (25,26). The CV for this assay was 
16%. Unfortunately, the samples intended for analyses of endpoint RBC folate concentrations were improperly prepared such that reliable measures could not be obtained. We measured serum vitamin B-12 using the nefelometric immunoassay reactive kit (27). The CV for this method was between 5 and 10%. Serum Hcy was measured by HPLC and flourometric detection (27,28). The intra-assay CV of this assay was <6%.
Statistical analysis
Statistical analyses were performed based on 370 subjects who provided both baseline and endpoint blood samples. Data were checked for normal distribution using the Kolmogorov-Smirnov test of normality. Log transformation was used to normalize the distribution of serum Hcy and vitamin B-12 values. Baseline characteristics across treatment groups were compared using ANOVA and the Kruskal-Wallis tests for continuous variables and chi-square analyses for categorical variables. Descriptive values were expressed as mean ± SD for continuous variables, and as median (inter-quartile range) for dietary intake. A socioeconomic index was created using principal components analysis according to the method used in Demographic Health Surveys and INCAP studies (29,30). Compliance was calculated as the number of tablets consumed divided by the total number of tablets provided. The effect of treatment on serum folate, Hcy, and vitamin B-12 was assessed using a generalized linear regression model (SAS Proc Mixed procedures) assuming unstructured correlation to account for the correlation among the repeated observations for a given subject (31), using treatment as a fixed factor and time as a covariate. The between-subjects factor was 4 treatment types and the within-subjects factor was treatment effects (from start to finish of supplementation). Between-group differences in treatment effect would be indicated by a significant interaction between treatment effect and treatment type. This is obtained by fitting the model below:
 |
Furthermore, we investigated whether the effects of supplementation varied depending on initial folate, vitamin B-12, and Hcy status. Differences between baseline and endpoint values were expressed as percentages rather than absolute values for Hcy and vitamin B-12 because of the log transformation. All statistical tests were 2-tailed and differences were considered significant at P < 0.05. We used SAS software, version 9.1, for statistical analysis (32).
|
Results |
|---|
|
TOPABSTRACTIntroductionMaterials and MethodsResultsDiscussionLITERATURE CITED |
|---|
|
The treatment groups did not differ in age, education, SES, weight, height, BMI, and compliance (Table 2). Reported median intakes of energy and nutrients (folate, vitamin B-12, vitamin B-6, iron, and zinc) were similar among treatment groups. The median intakes of these nutrients for the study population were lower than the RDA, with 58.7% not meeting the RDA for folate (400 µg/d), 63.3% not meeting the RDA for vitamin B-12 (2.4 µg/d), and 66.0% not meeting the RDA for vitamin B-6 (1.3 mg/d) (18).
|
14.4 µmol/L) and another 43 subjects (12.08%) were on the borderline of elevated Hcy concentrations (11.4–14.3 µmol/L). Baseline RBC folate and serum folate, vitamin B-12, and Hcy concentrations did not differ among the groups (Table 2). Efficacy of the trial. Serum folate and Hcy concentrations changed in all 4 groups (P < 0.0001), but the changes did not differ among the groups (P for interaction = 0.66 and 0.98, respectively) (Table 3). Overall, serum folate concentrations increased by 15.4 nmol/L (95% CI: 13.8, 16.9) and the geometric mean Hcy concentrations decreased by 9.8% (95% CI: –12.3, –7.1). The serum vitamin B-12 concentration increased significantly across all 4 treatment groups, but the increase did not differ among the groups (P = 0.49). When categorized as weekly and daily vitamin B-12 supplementation groups, the daily groups had a significant increase of 20% (95% CI: 8.0, 33.2; P < 0.01), whereas there was no change in the weekly group (P = 0.48) (8.1%; 95% CI: –2.7, 20.1). However, the changes in the daily and weekly groups did not differ (P = 0.17). Overall, the geometric mean serum vitamin B-12 concentrations increased by 14.1% (95% CI: 5.9, 22.9).
|
|
|
Discussion |
|---|
|
TOPABSTRACTIntroductionMaterials and MethodsResultsDiscussionLITERATURE CITED |
|---|
10% vs. 16 to 18.5%). However, the supplements in our study differed in composition from those of the New Zealand study in that ours also contained vitamin B-12. Therefore, the decline in Hcy in our study may not be due to FA alone but due in part to vitamin B-12. In terms of effects on folate status, results from the 2 studies differ; we found no difference in efficacy (serum folate) but the New Zealand study found greater efficacy (serum folate and RBC folate) for daily supplementation. We have no explanation for the differences in efficacy between daily and weekly supplementation across the studies. Compared to the New Zealand sample, women in our study had higher baseline serum folate and this may explain the lower response to supplementation in our study. Serum folate increased 47–50% in our study, whereas it increased 65–150% in the New Zealand study. Serum folate values after supplementation in our study were also similar to serum folate in the US after fortification (34) after correcting for differences in methods (35). Specifically, the Quantaphase II radio assay (Bio-Rad Diagnostics) method used in the U.S. study resulted in values that were 35% lower than those generated by the microbiological assay used in our study (35). The similar end-values observed in our study, the New Zealand study, and the U.S. study lead us to think that concentrations in these studies were converging toward a saturation point. However, due to higher baseline values in our study compared with the other studies, the response to supplementation was lower in our study.
Although changes in vitamin B-12 did not differ between daily and weekly groups, the changes in daily groups (20%) were significant, whereas there was no change in the weekly groups (P = 0.48). This suggests greater efficacy for daily than for weekly supplementation. The change in vitamin B-12 status was most likely due to the vitamin B-12 in the supplements. Further studies comparing the effects of weekly and daily supplementation on vitamin B-12 status are needed to confirm our findings.
The effect of the supplements was more pronounced in women with lower folate concentrations at baseline. Women with folate deficiency or marginal deficiency benefited the most, with mean increases of 40 and 36 nmol/L, respectively, 3 times the increase observed in women with normal baseline folate (13 nmol/L). Women with elevated Hcy concentrations at baseline showed the largest reduction after supplementation, a mean decrease from baseline Hcy of 45%. Among women with normal Hcy concentrations, there were no changes after FA supplementation (P = 0.8). Similar patterns were seen for vitamin B-12 effects in which women starting with vitamin B-12 deficiency improved the most (mean increase of 81.5%) compared with a 38.4% increase in women with marginal vitamin B-12 deficiency and no significant changes in women with normal vitamin B-12. Clearly, effects of supplementation depended on baseline values, with greater increases in those with lower values for folate and B-12 and greater reductions in those with higher Hcy concentrations. However, it was not possible to stratify the deficiency groups by dosage type due to the limited number of participants in each category.
Our study included multiple aims. Besides examining the effect of supplementation on folate status, we were also interested in testing whether providing iron and zinc in combination is less efficacious in improving iron status than is providing iron alone (see Table 1 for design). The addition of zinc to some of the supplements raises the concern that there may be an interaction between folate and zinc. It has been proposed that FA supplements may impair zinc absorption (36,37), possibly by forming an insoluble chelate in the lumen (36) or through a mutually inhibitory effect of zinc and folate on intestinal transport mechanisms (38). However, subsequent studies have failed to prove the inhibitory effect of folate on zinc absorption (39) or on zinc status (40–42). The evidence that zinc supplements affect folate status is limited. In one study in middle-aged and older European adults, supplementation with 15 or 30 mg zinc/d for 6 mo did not affect RBC folate status (43). Thus, we do not think that our interpretation of the findings is influenced by the presence of zinc in some of the supplement formulations. Rather, the results of our study suggest that adding zinc does not affect folate status, because all 4 groups responded similarly to various specifications of FA supplementation. Similarly, we are not concerned by the inclusion of vitamin B-12. The comparisons of groups within daily and weekly arms were unbiased, because the supplements contained similar amounts of vitamin B-12 within these arms (i.e. 2.4 µg for both daily and 16.8 µg for both weekly groups). Weekly doses of vitamin B-12 were 7 times greater than those in daily groups but were identical when expressed per day. The evidence that vitamin B-12 supplementation alone influences folate concentrations is limited and contradictory (44,45). Thus, we do not expect the difference in vitamin B-12 concentrations for daily and weekly supplements to confound our findings.
This study has several strengths. The study design was robust (a randomized controlled trial) and supplementation intake was supervised daily. Implementation of the study was careful and rigorous, from the training of the interview team to direct supervision of supplementation ingestion to ensure high quality data. Several regimens of supplementation were investigated: depending on frequency of supplementation (weekly compared with daily), and dose (weekly, 4000 µg compared with 2800 µg FA; daily, 400 µg compared with 200 µg FA). The 4 experimental groups had similar time lags between the timing of the last supplement consumption and blood draw (mean of 2.5 ± 1.1 d), therefore avoiding bias when comparing the effects of supplements on biochemical indicators.
Our study has a number of limitations. Our study was powered to detect a difference of 0.4 SD units, a medium size effect. We cannot claim with certainty, therefore, that smaller differences did not exist. Further, data for the final analysis were not available for 20% of the sample (89 participants), slightly higher than the 15% anticipated. However, our final sample size still allowed us to detect a difference of 0.4 SD, the specified target, with a power of 76%, only slightly less than the standard 80%.
Due to technical problems in preparing the samples, we did not have postintervention RBC folate values and therefore were able to assess the effects of supplementation on only serum folate. Serum folate is not as good an indicator of folate status at the individual level as is RBC folate, because serum folate concentrations fluctuate with recent folate intake. However, we compared group means, which would average out these fluctuations across individuals, making day-to-day variability less of a concern. In addition, we measured changes in serum Hcy, which is also an indicator of folate status, because an increase in serum or plasma Hcy reflects intracellular folate deficiency (46). Thus, although we lacked measures of RBC folate, changes in serum folate and serum Hcy were consistent in indicating similar efficacy across all 4 experimental groups. Finally, another limitation is that our design did not include a placebo group; hence, we cannot exclude the possibility that community-wide dietary improvements coincided with the beginning of supplementation. This, however, is not likely to have occurred.
Guatemala has been a pioneer in the developing world regarding food fortification initiatives. Wheat flour fortification with iron and FA has been implemented since the late 1960s (47). Since November 2002, as part of a national plan for the prevention of NTD, the amount of FA added to wheat flour in Guatemala has increased from 350–450 µg FA/kg wheat flour to 1800 µg FA/kg of wheat flour (48). However, despite these efforts, Guatemalans continue to suffer from micronutrient deficiencies. The consumption of fortified wheat flour is confined largely to Guatemala City and other urban centers. Many rural and highland communities do not have access to FA-fortified food products or find them too expensive to purchase on a regular basis. Most poor rural areas, particularly indigenous highland areas, consume mostly corn and corn flour rather than wheat-based products. Findings from a study by Imhoff-Kunsch et al. (47) suggested that the poor rural and indigenous populations benefit the least from wheat flour fortification. They estimated that wheat flour fortification provided 26% of the RDA for folate for women across all households but only 4% for women in extremely poor households. Given that situation, weekly FA supplementation represents an option for reaching women of reproductive age in poor rural areas and in indigenous regions of Guatemala. In addition, weekly supplementation with iron and FA has been shown to have fewer side effects than daily supplementation and this may enhance compliance (49). From a programmatic perspective, weekly supplementation is less costly and may be easier to manage. However, field studies are needed to demonstrate the feasibility of weekly supplementation in routine, large-scale programs.
The main concern with FA supplementation is that it may mask symptoms of vitamin B-12 deficiency and that this may lead to the progression of adverse neurological symptoms. The current study provided both FA and vitamin B-12; we would hope that public health programs would also provide vitamin B-12 along with FA. Recently, concern has been expressed that FA supplementation or fortification may have a promoting effect on the progression of preexisting, undiagnosed premalignant and malignant lesions (50–52). Evidence for this concern is not conclusive and needs further investigation. This potential, unconfirmed risk needs to be balanced against the proven benefit of reducing NTD rates, which are very common in the region.
FA supplementation may reduce NTD considerably, but not completely, because other causes may also be important in Guatemala. Fumonisins may be a potential, additional risk factor for NTD (53). Fumonisins is a family of mycotoxins, a common fungal contaminant of maize. Fumonisins inhibit ceramide synthase, causing depletion of complex sphingolipids, which interfere with the function of some membrane proteins, including the folate-binding protein, thus interfering with folate transport (53). In the highland areas of Guatemala where the staple is maize, consumption of fumonisins is likely to be high (54), further exacerbating folate deficiency. Further studies are needed to explore other factors, including genetic polymorphisms, which may contribute to the high incidence of NTD in this population.
Our results suggest that weekly FA plus vitamin B-12 supplementation provided to Guatemalan women of reproductive age may be as efficacious as daily supplementation in improving serum folate and lowering Hcy concentrations. Further studies are needed to confirm the efficacy of weekly FA supplementation on folate status in women of child-bearing age in other populations.
|
ACKNOWLEDGMENTS |
|---|
|
FOOTNOTES |
|---|
2 Author disclosures: P. Nguyen, R. Grajeda, P. Melgar, J. Marcinkevage, R. Flores, and R. Martorell, no conflicts of interest.
7 Abbreviations used: FA, folic acid; Hcy, homocysteine; INCAP, Institute of Nutrition of Central America and Panama; NTD, neural tube defect; RDA, recommended dietary allowance; SES, socioeconomic status.
Manuscript received 10 December 2007. Initial review completed 4 March 2008. Revision accepted 21 May 2008.
|
LITERATURE CITED |
|---|
|
TOPABSTRACTIntroductionMaterials and MethodsResultsDiscussionLITERATURE CITED |
|---|
1. Murray CJL, Lopez AD. Global burden of disease: a comprehensive assessment of mortality and disability from diseases, injuries, and risk factors in 1990 and projected to 2020. Vol. 1. The global burden of disease and injury. Cambridge (MA): Harvard School of Public Health; 1996.
2. Acevedo CR, Anzueto ER, Mendez AG, Ramirez FM. Prevalencia de anomalías congénitas mayores externas, en recién nacidos atendidos en hospitales nacionales y regionales de Guatemala. (Durante el periodo comprendido del año 2001 al 2003) [undergraduate graduation thesis]. Guatemala City (Guatemala): School of Medicine, Universidad de San Carlos de Guatemala; 2004.
3. Cifuentes G. Perfil epidemiológico de las anomalías del tubo neural en Guatemala, Enero-Diciembre de 2000 [undergraduate graduation thesis]. Guatemala City (Guatemala): School of Medicine, Universidad San Carlos de Guatemala; 2002.
4. Rogers LM, Boy E, Miller JW, Green R, Sabel JC, Allen LH. High prevalence of cobalamin deficiency in Guatemalan schoolchildren: associations with low plasma holotranscobalamin II and elevated serum methylmalonic acid and plasma homocysteine concentrations. Am J Clin Nutr. 2003;77:433–40.
5. Casterline JE, Allen LH, Ruel MT. Vitamin B-12 deficiency is very prevalent in lactating Guatemalan women and their infants at three months postpartum. J Nutr. 1997;127:1966–72.
6. Malmad JL. An evaluation of the potential impact of folic acid fortification of wheat flour on folic acid consumption and blood folate concentrations in women of reproductive age from periurban Guatemala city [master's thesis]. Atlanta: Rollins School of Public Heath, Emory University; 2006.
7. Scott JM, Weir DG, Molloy A, McPartlin J, Daly L, Kirke P. Folic acid metabolism and mechanisms of neural tube defects. Ciba Found Symp. 1994;181:180–7.[Medline]
8. Clarke R, Daly L, Robinson K, Naughten E, Cahalane S, Fowler B, Graham I. Hyperhomocysteinemia: an independent risk factor for vascular disease. N Engl J Med. 1991;324:1149–55.[Abstract]
9. Clarke R, Lewington S. Homocysteine and coronary heart disease. Semin Vasc Med. 2002;2:391–9.[Medline]
10. Cotter AM, Molloy AM, Scott JM, Daly SF. Elevated plasma homocysteine in early pregnancy: a risk factor for the development of severe preeclampsia. Am J Obstet Gynecol. 2001;185:781–5.[Medline]
11. Vollset SE, Refsum H, Irgens LM, Emblem BM, Tverdal A, Gjessing HK, Monsen AL, Ueland PM. Plasma total homocysteine, pregnancy complications, and adverse pregnancy outcomes: the Hordaland Homocysteine study. Am J Clin Nutr. 2000;71:962–8.
12. Steegers-Theunissen RP, Boers GH, Trijbels FJ, Finkelstein JD, Blom HJ, Thomas CM, Borm GF, Wouters MG, Eskes TK. Maternal hyperhomocysteinemia: a risk factor for neural-tube defects? Metabolism. 1994;43:1475–80.[Medline]
13. Mills JL, McPartlin JM, Kirke PN, Lee YJ, Conley MR, Weir DG, Scott JM. Homocysteine metabolism in pregnancies complicated by neural-tube defects. Lancet. 1995;345:149–51.[Medline]
14. MRC Vitamin Study Research Group. Prevention of neural tube defects: results of the Medical Research Council Vitamin Study. Lancet. 1991;338:131–7.[Medline]
15. Berry RJ, Li Z, Erickson JD, Li S, Moore CA, Wang H, Mulinare J, Zhao P, Wong LY, et al. Prevention of neural-tube defects with folic acid in China. China-U.S. Collaborative Project for Neural Tube Defect Prevention. N Engl J Med. 1999;341:1485–90.
16. Adank C, Green TJ, Skeaff CM, Briars B. Weekly high-dose folic acid supplementation is effective in lowering serum homocysteine concentrations in women. Ann Nutr Metab. 2003;47:55–9.[Medline]
17. Norsworthy B, Skeaff CM, Adank C, Green TJ. Effects of once-a-week or daily folic acid supplementation on red blood cell folate concentrations in women. Eur J Clin Nutr. 2004;58:548–54.[Medline]
18. Institute of Medicine. Dietary reference intakes for thiamin, riboflavin, niacin, vitamin B6, folate, vitamin B12, pantothenic acid, biotin, and choline. Washington, DC: National Academy Press; 2001.
19. Venn BJ, Mann JI, Williams SM, Riddell LJ, Chisholm A, Harper MJ, Aitken W, Rossaak JI. Assessment of three levels of folic acid on serum folate and plasma homocysteine: a randomised placebo-controlled double-blind dietary intervention trial. Eur J Clin Nutr. 2002;56:748–54.[Medline]
20. Daly S, Mills JL, Molloy AM, Conley M, Lee YJ, Kirke PN, Weir DG, Scott JM. Minimum effective dose of folic acid for food fortification to prevent neural-tube defects. Lancet. 1997;350:1666–9.[Medline]
21. Daly S, Mills JL, Molloy AM, Conley M, McPartlin J, Lee YJ, Young PB, Kirke PN, Weir DG, et al. Low-dose folic acid lowers plasma homocysteine levels in women of child-bearing age. QJM. 2002;95:733–40.
22. Ward M, McNulty H, McPartlin J, Strain JJ, Weir DG, Scott JM. Plasma homocysteine, a risk factor for cardiovascular disease, is lowered by physiological doses of folic acid. QJM. 1997;90:519–24.
23. Solomons NW. Guatemala introduces once-a-week supplementation. Nutriview. 2003;3:7.
24. Rodriguez MM, Mendez H, Torun B, Schroeder D, Stein AD. Validation of a semi-quantitative food-frequency questionnaire for use among adults in Guatemala. Public Health Nutr. 2002;5:691–9.[Medline]
25. Tamura T, editor. Microbiological assay of folates in folic acid metabolism in health and disease. New York: John Wiley & Sons, Inc.; 1990.
26. O'Broin JD, Temperley IJ, Scott JM. Erythrocyte, plasma, and serum folate: specimen stability before microbiological assay. Clin Chem. 1980;26:522–4.
27. CDC. Laboratory procedures used for the Third National Health and Nutrition Examination Survey (NHANES III) 1988-94. Atlanta: CDC, U.S. Department of Health and Human Service; 1998.
28. Araki A, Sako Y. Determination of free and total homocysteine in human plasma by high-performance liquid chromatography with fluorescence detection. J Chromatogr. 1987;422:43–52.[Medline]
29. Gwatkin DR, Rustein S, Johnsin K, Pande RP, Wagstaff A. Socio-economic differences in health, nutrition, and population in Bangladesh. Washington, DC: The World Bank's Health and Population Advisory Service; 2000.
30. Maluccio JA, Murphy A, Yount KM. Research note: a socioeconomic index for the INCAP Longitudinal Study 1969–77. Food Nutr Bull. 2005;26:S120–4.[Medline]
31. Kung-Yee L, Zeger SL. Longitudinal data analysis using generalized linear models. Biometrika. 1986;73:13–22.
32. SAS. SAS, the power to know. SAS Institue Inc.; 2002–2004.
33. Selhub J, Jacques PF, Bostom AG, D'Agostino RB, Wilson PW, Belanger AJ, O'Leary DH, Wolf PA, Schaefer EJ, et al. Association between plasma homocysteine concentrations and extracranial carotid-artery stenosis. N Engl J Med. 1995;332:286–91.
34. Pfeiffer CM, Caudill SP, Gunter EW, Osterloh J, Sampson EJ. Biochemical indicators of B vitamin status in the US population after folic acid fortification: results from the National Health and Nutrition Examination Survey 1999–2000. Am J Clin Nutr. 2005;82:442–50.
35. Pfeiffer CM, Johnson CL, Jain RB, Yetley EA, Picciano MF, Rader JI, Fisher KD, Mulinare J, Osterloh JD. Trends in blood folate and vitamin B-12 concentrations in the United States, 1988–2004. Am J Clin Nutr. 2007;86:718–27.
36. Milne DB, Canfield WK, Mahalko JR, Sandstead HH. Effect of oral folic acid supplements on zinc, copper, and iron absorption and excretion. Am J Clin Nutr. 1984;39:535–9.
37. Simmer K, Iles CA, James C, Thompson RP. Are iron-folate supplements harmful? Am J Clin Nutr. 1987;45:122–5.
38. Ghishan FK, Said HM, Wilson PC, Murrell JE, Greene HL. Intestinal transport of zinc and folic acid: a mutual inhibitory effect. Am J Clin Nutr. 1986;43:258–62.
39. Hansen M, Samman S, Madsen LT, Jensen M, Sorensen SS, Sandstrom B. Folic acid enrichment of bread does not appear to affect zinc absorption in young women. Am J Clin Nutr. 2001;74:125–9.
40. Butterworth CE Jr, Hatch K, Cole P, Sauberlich HE, Tamura T, Cornwell PE, Soong SJ. Zinc concentration in plasma and erythrocytes of subjects receiving folic acid supplementation. Am J Clin Nutr. 1988;47:484–6.
41. Green TJ, Skeaff CM, Whiting SJ, Gibson RS. Effect of folic acid supplementation on plasma zinc concentrations of young women. Nutrition. 2003;19:522–3.[Medline]
42. Kauwell GP, Bailey LB, Gregory JF III, Bowling DW, Cousins RJ. Zinc status is not adversely affected by folic acid supplementation and zinc intake does not impair folate utilization in human subjects. J Nutr. 1995;125:66–72.
43. Intorre F, Polito A, Andriollo-Sanchez M, Azzini E, Raguzzini A, Toti E, Zaccaria M, Catasta G, Meunier N, et al. Effect of zinc supplementation on vitamin status of middle-aged and older European adults: the ZENITH study. Eur J Clin Nutr. 2007; Jul 11.
44. Yajnik CS, Lubree HG, Thuse NV, Ramdas LV, Deshpande SS, Deshpande VU, Deshpande JA, Uradey BS, Ganpule AA, et al. Oral vitamin B12 supplementation reduces plasma total homocysteine concentration in women in India. Asia Pac J Clin Nutr. 2007;16:103–9.
45. Eussen SJ, de Groot LC, Joosten LW, Bloo RJ, Clarke R, Ueland PM, Schneede J, Blom HJ, Hoefnagels WH, et al. Effect of oral vitamin B-12 with or without folic acid on cognitive function in older people with mild vitamin B-12 deficiency: a randomized, placebo-controlled trial. Am J Clin Nutr. 2006;84:361–70.
46. Ueland PM, Refsum H, Stabler SP, Malinow MR, Andersson A, Allen RH. Total homocysteine in plasma or serum: methods and clinical applications. Clin Chem. 1993;39:1764–79.[Abstract]
47. Imhoff-Kunsch B, Flores R, Dary O, Martorell R. Wheat flour fortification is unlikely to benefit the neediest in Guatemala. J Nutr. 2007;137:1017–22.
48. Dary O. Food fortification successful in Central America. Nutriview. 2003;3:2–3.
49. Viteri FE, Berger J. Importance of pre-pregnancy and pregnancy iron status: can long-term weekly preventive iron and folic acid supplementation achieve desirable and safe status? Nutr Rev. 2005;63:S65–76.[Medline]
50. Kim YI. Role of folate in colon cancer development and progression. J Nutr. 2003;133:S3731–9.
51. Kim YI. Folate and colorectal cancer: an evidence-based critical review. Mol Nutr Food Res. 2007;51:267–92.[Medline]
52. Mason JB, Dickstein A, Jacques PF, Haggarty P, Selhub J, Dallal G, Rosenberg IH. A temporal association between folic acid fortification and an increase in colorectal cancer rates may be illuminating important biological principles: a hypothesis. Cancer Epidemiol Biomarkers Prev. 2007;16:1325–9.
53. Marasas WF, Riley RT, Hendricks KA, Stevens VL, Sadler TW, Gelineau-van Waes J, Missmer SA, Cabrera J, Torres O, et al. Fumonisins disrupt sphingolipid metabolism, folate transport, and neural tube development in embryo culture and in vivo: a potential risk factor for human neural tube defects among populations consuming fumonisin-contaminated maize. J Nutr. 2004;134:711–6.
54. Meredith FI, Torres OR, Saenz de Tejada S, Riley RT, Merrill AH Jr. Fumonisin B1 and hydrolyzed fumonisin B1 (AP1) in tortillas and nixtamalized corn (Zea mays L.) from two different geographic locations in Guatemala. J Food Prot. 1999;62:1218–22.[Medline]
55. Gibson RS. Assessment of folate and vitamin B12 status. In: Principles of nutritional assessment. New York: Oxford University Press; 2005.
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
