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Folic acid, vitamin B12, and DNA methylation: An update

  • All India Institute of Medical Sciences Bibinagar
  • RD Gardi medical college
  • Sri Madhusudan Sai Institute of Medical Sciences and Research


Epigenetics is one of the exciting and fastest expanding fields of biology; this is above genetics. Methylation is the process involved in the transfer of methyl group to amino acids, proteins, enzymes and DNA of all the cells, and tissues of the body. During cell-division low folate availability may result in decreased production of thymidine wherein uracil may be substituted in the place of thymidine in the DNA sequence. It was reported that folate and Vitamin B12 restricted diet resulted in aberrant methylation patterns. The current review was undertaken to explore the role of folic acid and Vitamin B12 in DNA methylation.
Vol 11, Issue 1, 2018
Online - 2455-3891
Print - 0974-2441
1Department of Biochemistry, Mediciti Institute of Medical Sciences, Medchal, Telangana, India. 2Department of Biochemistry, Apollo
Institute of Medical sciences & Research, Hyderabad, Telangana, India. 3Department of Research, Saveetha University, Chennai, Tamil
Nadu, India. 4Department of Physiology, DM Wayanad Institute of Medical Sciences, Wayanad, Kerala, India.
Received: 09 August 2017, Revised and Accepted: 07 October 2017
Epigenetics is one of the exciting and fastest expanding fields of biology; this is above genetics. Methylation is the process involved in the transfer of
methyl group to amino acids, proteins, enzymes and DNA of all the cells, and tissues of the body. During cell-division low folate availability may result
in decreased production of thymidine wherein uracil may be substituted in the place of thymidine in the DNA sequence. It was reported that folate
and Vitamin B12 restricted diet resulted in aberrant methylation patterns. The current review was undertaken to explore the role of folic acid and
Vitamin B12 in DNA methylation.
Keywords: DNA methylation, Folic acid, Vitamin B12.
Epigenetics is one of the exciting and fastest expanding fields of
biology; this is above genetics. The modifications of epigenetics play
a very important role in the regulation of many cellular processes
including DNA replication, gene expression, and recombination. This
is achieved through regulatory mechanisms such as DNA methylation,
hydroxymethylation, histone modifications, chromatin remodeling,
and RNA modifications like methylation. Misregulation of epigenetic
mechanism may have adverse effects on health and may lead to
neurological disorders, developmental abnormalities and also cancer.
Therefore, epigenetic modifications are evolving as very potent
diagnostic and prognostic biomarkers in the world of medicine.
Methylation is the process involved in the transfer of methyl group to
amino acids, proteins, enzymes and DNA of all the cells, and tissues of the
body. Donation of the methyl group is very important in the regulation
of cell energy, gene expression, neurological function, detoxification (in
liver), immunity, etc. Methylation is one of the important biochemical
processes that occur in the body and is catalyzed by different enzymes.
This is a process influenced by environmental conditions, decreases
with age. Methylation depicts the quality of life in terms of diseased and
health conditions.
The methylation processes require two cycles - Cycle A:
S- adenosylmethionine (SAM) and Cycle B- folate cycle. The most
stable synthetic form of Vitamin B9 is folic acid (FA), and the natural
form is termed as folate. The natural dietary folates are polyglutamated
consisting of six glutamate molecules linked together by peptide
bonds. Mainly there are two dietary folates, 5-methyltetrahydrofolate
(5-MeTHF) and 10- formyltetrahydrofolate (10-formyl THF). In
the gut, before to the absorption polyglutamates are hydrolyzed
to monoglutamates by the enzymes γ-glutamyl hydrolases and is
absorbed [1,2]. FA with monoglutamyl residues gets converted to
the biologically active form known as THF by the reducing reaction
(first it is converted to dihydrofolate (DHF) and then to THF). These
reduction reactions are catalyzed by the single NADPH-dependent
enzyme DHF reductase (DHFR). THF receives the one-carbon (C1)
units from various donors such as serine, glycine, and histidine during
catabolic reactions and can transfer them to specific acceptors for the
synthesis of various compounds such as purines, methionine, choline,
formyl-methylated tRNA, thymidylate, and serine. The addition of one
carbon unit with a simultaneous reduction will produce 5-MeTHF, the
main circulating form in the blood. Protein carriers and the tissue-
specific folate receptors carry 5-MeTHF into the cells, where they get
accumulated and are transformed to polyglutamates. Polyglutamates
cannot traverse biological membranes by passive diffusion. Thus,
polyglutamylation serves to sequester folate in the cells in which it is
required. The enzymes involved in folate metabolism have a higher
affinity for polyglutamates than to monoglutamates. FA is involved in
remethylation process wherein, 5-MeTHF donates a methyl group for
homocysteine in the presence of the enzyme methionine synthase,
converting homocysteine to methionine liberating THF. This THF
directly gets converted into 5,10-methylene THF by the action of the
enzyme serine hydroxymethyltransferase which is present both in
mitochondria and cytosol.
5-MeTHF is used in remethylation of homocysteine to methionine in
the presence of methionine synthase in all the tissues except red blood
cells. This process requires a cofactor the Vitamin B12 (cobalamin).
Methylcobalamin is the donor of a methyl group to homocysteine and
the transfer mediated through THF getting converted to 5-MeTHF.
After donating the methyl group, methylcobalamin is converted back
to cobalamin. Methylcobalamin is reproduced from cobalamin by
receiving the methyl group from 5-MeTHF. Methionine is an essential
amino acid converted to active methionine, i.e., SAM also known
as the universal donor. SAM is the active donor of a methyl group
for various methylation reactions that occur in the body such as
methylation of nucleic acids, proteins, lipids, neurotransmitters, and
creatine synthesis [3]. SAM gets converted to S-adenosyl homocysteine
(SAH) after donating the methyl group to the acceptors further, gets
hydrolyzed to adenosine and homocysteine by the action of the enzyme
SAH hydrolase (SAHH). The decrease in 5-MeTHF or cobalamin made
lead to the accumulation of homocysteine the potent inhibitor of various
methyltransferases. Thus, cobalamin and folate are together involved in
methylation process. An absence of cobalamin leads to the cessation of
the reaction and build-up of methyltetrahydrofolate (MeTHF) known
as “folate trap.” Therefore, MeTHF tends to accumulate in Vitamin B12
(cobalamin) deficiency, leads to depletion of other coenzyme forms that
are needed for nucleotide synthesis. This is how folate trap hypothesis
© 2018 The Authors. Published by Innovare Academic Sciences Pvt Ltd. This is an open access article under the CC BY license (http://creativecommons.
org/licenses/by/4. 0/) DOI:
Review Article
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Varma et al.
explains the anemia of cobalamin deficiency, but it cannot account for
the neurological manifestations of pernicious anemia.
A detailed review of published literature from Google, PubMed, and
MEDLINE was performed and analyzed.
Folate DNA methylation
FA, when consumed in fortified foods or supplements, is primarily
metabolized to 5-methyl THF that behaves similar to natural dietary
folate. Initially, FA is reduced to DHF in the presence of the enzyme
DHF reductase, further converted to THF and enters the folate pool. In
some cases, the oxidized form of FA may appear in circulation with an
increase in DHF reductase enzyme [4]. The coenzyme THF is converted
to 5,10-methyleneTHF by the enzyme serine hydroxymethyltransferase
requires Vitamin B6. Further MTHFR irreversibly reduces it to 5-methyl
THF. This is a key reaction for the maintenance of the methyl flux
essential for the remethylation of homocysteine to methionine in the
presence of Vitamin B12-dependent methionine synthase. Methionine
is converted to SAM, an active methyl donor wherein numerous
SAM-dependent reactions play regulatory roles by affecting gene
transcription, genome stability [5] and localization of protein [6], etc.
Along with folate, many other dietary nutrients such as Vitamin B6,
Vitamin B12, riboflavin (Vitamin B2), and choline are required for
the maintenance of one carbon flux and normal formation of SAM,
homocysteine remethylation, and DNA methylation. DNA methylation
and one carbon metabolism work under tight regulatory control.
Homocysteine remethylation is folate-dependent and requires
SAM as an important regulator for this process. Increase in SAM
inhibits MTHFR this reduces 5-methylTHF synthesis further hinders
homocysteine remethylation. In contrast, remethylation is favored with
low concentrations of SAM and SAM-dependent methyltransferase is
inhibited by SAH [7,43,44]. Therefore, for the maintenance of normal
DNA methylation, there should be a continual conversion of SAH to
homocysteine [8] and increased plasma concentration of homocysteine
is associated with increased concentration of SAH which in-turn
associated with hypomethylation of global DNA [9]. The common
genetic variant 677C-T modifies the activity of MTHFR and reduces the
formation of 5-methylTHF [10].
DNA methylation and low folate status
The studies on the association of low folate with increased risk of
NTDs, cardiovascular disease and multiple cancers are well established
but, the mechanism leading to these disorders is yet unclear [11-13].
During cell-division low folate availability may result in decreased
production of thymidine wherein uracil may be substituted in the place
of thymidine in the DNA sequence. This may increase the frequency of
chromosomal breaks to repair the defect made by the mutagenic event.
This was studied by a tissue culture where the MTHFR-TT genotype
shows the formation of increased micronuclei as a result of multiple
chromosomal breakages occurred under low folate conditions [14].
The effects of supplementation of FA may promote or prevent cancers,
stated in many studies. In human cancers, the DNA methylation is
dysregulated. It shows either hypermethylation or hypomethylation
stating that the association of DNA methylation with tumor is cell or
tissue or organ-specific. A study showed genome-wide hypomethylation
but found 5% hypermethylated patterns defining the characteristics of
the specific type of human tumor. These altered DNA methylation cause
chromosomal instability and silencing of tumor suppressor genes [16].
However, it is very important to note that decrease in folate status may
result either in hyper or hypomethylation leading to the misregulation
in the complex system. (Shelnutt et al., 2004) On controlled feeding
studies observed the association of low folate concentration with
reduced DNA methylation in older women but not in younger female
adults (Rampersaud et al., 2000; Jacob et al., 1998). Jacob and Shelnutt
both had observed that the folate intake during repletion resulted in
increased DNA methylation and the increase was limited to MTHFR
TT genotype. Friso et al. (2002) and Pufulete et al. (2005) also found
the same trend of lowered DNA methylation with low serum folate
concentration. These studies taken together suggest that the genotype,
age, duration, and magnitude of exposure should be considered as the
response of global DNA methylation to the folate status is different at
different conditions. In the above studies, variations in DNA methylation
were found between the sexes and ages (El-Maarri et al., 2007); (Fraga
et al., 2005) justifying the observations of different methylation
patterns with low and high folate status.
MTHFR is an essential enzyme involved in the irreversible conversion
of 5,10 MethyleneTHF to 5-methylTHF thus, playing important role
in DNA synthesis, DNA methylation and maintenance of balanced
nucleotide pool (Friso et al., 2002; Kim et al., 1999). Analyses of MTHFR
polymorphisms are included to investigate the folate status and DNA
methylation in humans. As many observational studies had established
the association of low folate with hypomethylated DNA in subjects with
homozygous MTHFR C677TT genotype. The biochemical mechanism
behind this is the MTHFR C677TT polymorphism causes thermolability
thereby reducing the MTHFR activity by lowering 5-methylTHF levels
and leading to the accumulation of 5,10methyleneTHF, increase
in plasma homocysteine levels and finally changing the cellular
composition of one-carbon folate derivatives stating that there is a
greater risk of global DNA hypomethylation in the women carrying
MTHFR T allele as it is involved in the impairment of enzyme activity
modulating both gene and genome-specific DNA methylation
(La Merrill et al. 2012).
High folate and DNA methylation
Many studies on folate insufficiency and the effect of low folate in
humans are well established. Low folate has detrimental effect on
the embryo and increases the risk of NTD’s, increases the possibility
of long-term risk of diabetes and also may lead to many other health
outcomes [26-29]. High folate concentrations and increased folate
supplementation had shown contradicting results in different in
controlled feeding trial studies. Certain studies have shown the
association of high folate with high global DNA methylation and reduced
risk of cancer [30,31]. A recent study had shown reversal effect that
is, the increased folate supplementation resulted in stimulation and
progression of existing tumors and altered normal DNA methylation
patterns [32-34]. As DNA methylation is a regulatory process depends
on tissues, sequence of DNA, genome region, stage of transformation,
degree, duration and exposure of folate intervention, timing, and other
regulatory proteins and enzymes involved in the process. Hence, we
should include all these factors to study the effect of high folate, whether
high folate leads to increased risk or benefit.
Folate, Vitamin B12, and placental DNA methylation
Placenta is an important organ. Proper development and function of
placenta are crucial for the growth, health, and survival of developing
fetus. Many studies had established links between epigenetic changes
in the placenta and the risk of disease in gestation and early life (Kim
et al., 2009). On examination of the epigenetic changes occurred in the
placenta had evolved interest in the research of biomarkers of exposure,
pathogenesis of the disease and the biology of the development of the
disease [36]. Presently, many studies on nutrition during pregnancy
and placental outcomes are taken up to understand the basis of disease
seen in early or later in life. Nutrition and epigenetic changes are the
emerging topic of interest in the present scenario to understand the
effects of increased supplementation of micronutrients like FA and
learning the importance of balancing the different micronutrients in
the diet to avoid unbalanced nutritional disorders and other health
complications later in life. One such study is carried on rats to know the
effect of FA supplementation in utero on the epigenetic changes in the
offspring. It was observed that maternal dietary folate during pregnancy
led to placental DNA hypomethylation and showed that there is a
significant correlation between folate levels of placenta and placenta
genomic DNA methylation. On the other hand, the study also stated
the importance of maintaining the ratio of folate and Vitamin B12. As
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Varma et al.
it has a significant role in determining genomic methylation patterns.
Wherein, high folate in the absence of Vitamin B12 resulted in placental
DNA hypomethylation [37]. It has also shown the association of
maternal folate deficiency with DNA hypomethylation [38]. Yet, another
animal study in sheep with maternal folate and Vitamin B12 restricted
diet resulted in aberrant methylation patterns, i.e., only 4% of cytosine-
guanine dinucleotide islands (CpG islands) out of 1400 CpG islands
were methylated. Along with hypomethylation the adult male offspring
also showed increased adiposity, altered immune function, high blood
pressure, and insulin resistance [39].
A study conducted by Kulkarni et al. [37] on the adverse effects
caused by excess FA supplementation in the presence or absence of
Vitamin B12 deficiency and correlated it with global DNA methylation
patterns. The team observed a reduction in the levels of global DNA
methylation on excess maternal FA supplementation with low plasma
Vitamin B12 concentration. They also observed the effect of Vitamin
B12 deficiency with excess or normal FA levels on docosahexaenoic
acid (DHA) levels. In this study, the team for the first time had identified
the DHA plays an important role in one-carbon metabolism, influencing
the placental global DNA methylation. Further learned that on the
supplementation of omega three fatty acids, the DHA levels in placenta
got increased and the DNA methylation levels were reverted back to
that of the control group. This study suggested that the altered ration of
FA and Vitamin B12 during pregnancy have effect on DNA methylation
thereby influence imprinting in the embryo and could be associated
with adverse pregnancy outcomes. These epigenetic changes caused,
may alter the gene expression and could be carried throughout the
lifespan of an individual [40].
Till date, there are only a few studies performed to learn the association
of FA and Vitamin B12 and together their effect on thyroid hormones.
The well- established knowledge regarding the two micronutrients
(folate and Vitamin B12) is (i) deficiency of Vitamin B12 is very common
in hypothyroidism (ii) both the micronutrients are very important
for fetal development and the imbalance ratio of Vitamin B12 and FA
(high folate with low Vitamin B12) may lead to small for gestational
age infants [42-44] and other detrimental effects on the growing fetus,
and (iii) both folate and Vitamin B12 are important along with other
micronutrients such as Vitamin B6, choline to prevent NTD’s, and
proper methylation process to occur.
Nutrition and epigenetic changes are the emerging topic of interest in the
present scenario to understand the effects of increased supplementation
of micronutrients like FA. The available information is that the increased
folate supplementation had a suppressive effect on thyroid hormones
(T3 and T4) and may possibly lead to motivational deficits and memory
impairments in adolescent rats and also observed that the animals
were functionally hypothyroid during the administration of folate. The
increased levels of folate supplementation alone may be harmful not
only during fetal development and on growing infant but also in the
adolescent period. If the same effect implies in humans with increased
folate supplementation during pregnancy, i.e., high folate intake may
lead to suppression of maternal plasma thyroid hormonal levels, would
have alarming implication on the health of the fetus.
1. Stover PJ. Physiology of folate and vitamin B12 in health and disease.
Nutr Rev 2004;62:S3-12.
2. Beaudin AE, Stover PJ. Insights into the metabolic mechanisms
underlying folate-responsive neural tube defects: A mini review. Birth
Defects Res 2009;85:274-84.
3. Finkelstein JD. Metabolic regulatory properties of S-adenosylmethionine
and S-adenosylhomocysteine. Clin Chem Lab Med 2007;45:1694-9.
4. Bailey RL, Mills JL, Yetley EA, Gahche JJ, Pfeiffer CM, Dwyer JT, et al.
Unmetabolized serum folic acid and its relation to folic acid intake from
diet and supplements in a nationally representative sample of adults
aged > or =60 y in the United States. Am J Clin Nutr 2010;92:383-9.
5. Miranda TB, Jones PA. DNA methylation: The nuts and bolts of
repression. J Cell Physiol 2007;213:384-90.
6. Winter-Vann AM, Kamen BA, Bergo MO, Young SG, Melnyk S,
James SJ, et al. Targetting ras signaling through inhibition of carboxyl
methylation: An unexpected property of methotrexate. Proc Natl Acad
Sci U S A 2003;100:6529-34.
7. Hoffman DR, Marion DW, Cornatzer WE, Duerre JA. S-adenosylmethionine
and S-adenosylhomocystein metabolism in isolated rat liver. Effects
of L-methionine, L-homocystein, and adenosine. J Biol Chem
8. James SJ, Melnyk S, Pogribna M, Pogribny IP, Caudill MA. Elevation
in S-adenosylhomocysteine and DNA hypomethylation: Potential
epigenetic mechanism for homocysteine-related pathology. J Nutr
9. Yi P, Melnyk S, Pogribna M, Pogribny IP, Hine RJ, James SJ. Increase
in plasma homocysteine associated with parallel increase in plasma
S-adenosylhomocyateine and kymphocyte DNA hypomethylation.
J Biol Chem 2000;102:29318-23.
10. Bailey LB, editor. Folate in Health and Disease. Boca Raton, FL: Taylor
and Francis; 2009.
11. Erickson JD. Folic acid and prevention of spina bifida and anencephaly.
10 years after the U.S. public health service recommendation. MMWR
Recomm Rep 2002;51:1-3.
12. Lamprecht SA, Lipkin M. Chemoprevention of colon cancer by
calcium, vitamin D and folate: Molecular mechanism. Nat Rev Cancer
13. Holmquist C, Larsson S, Wolk A, Faire U. Multivitamin supplements
are inversely associated with risk of myocardial infarction in men
and women-stockholm heart epidemiology program (SHEEP). J Nutr
14. Kimura M, Umegaki K, Higuchi M, Thomas P, Fenech M. Methylene-
tetrahydrofolate reductase C677T polymorphism, folic acid and
riboflavin are important determinants of genome stability in cultured
human lymphocytes. J Nutr 2004;134:48-56.
15. Herman JG, Baylin SB. Gene silencing in cancer in association with
promoter hypermethylation. N Engl J Med 2003;349:2042-54.
16. Rooij SR, Painter RC, Phillips DI, Osmond C, Michels RP, Godsland IF,
et al. Impaired insulin secretion after prenatal exposure to the dutch
famine. Diabetes Care 2006;29:1897-901.
17. Roseboom T, Rooij S, Painter R. The dutch famine and its long-term
consequences for adult health. Early Hum Dev 2006;82:485-91.
26. Berry RJ, Li Z, Erickson JD, Li S, Moore CA, Wang H, 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
27. Painter RC, Rooij SR, Bossuyt PM, Simmers TA, Osmand C,
Barker DJ, et al. Early onset of coronary artery disease after prenatal
exposure to the dutch famine. Am J Clin Nutr 2006;84:322-7.
28. Hsiung DT, Marsit CJ, Houseman EA, Eddy K, Furniss CS,
McClean MD, et al. Global DNA methylation level in whole blood
as a biomarker in head and neck squamous cell carcinoma. Cancer
Epidemoil Biomarkers Prev 2007;16:108-14.
29. Piyathilake CJ, Macaluso M, Alvarez RD, Chen M,
Badiga S, Siddiqui NR, et al. A high degree of LINE-1 methylation
in peripheral blood mononuclear cells, a one-carbon nutrient related
epigenetic alteration, is associated with a lower risk of developing
cervical intraepithelial neoplasia. Nutrition 2011;27:513-9.
30. Smith AD, Kim YI, Refsum H. Is folic acid good for everyone? Am J
Clin Nutr 2008;87:517-33.
31. Ulrich CM, Potter JD. Folate and cancer-timing is everything. J Am
Med Assoc 2007;297:2408-9.
Kim YI. Folic acid supplementation and cancer risk: Point. Cancer
Epidemiol Biomarkers Prev 2008;17:2220-5.
36. Maccani MA, Marsit CJ. Epigenetics in the placenta. Am J Reprod
Immunol 2009;62:78-89.
37. Kulkarni A, Dangat K, Kale A, Sable P, Chavan-Gautam P, Joshi S.
effects of altered maternal folic acid, vitamin B12 and docosahexaenoic
acid on placental global DNA methylation patterns in wistar rats. PLoS
One 2011;6:e17706.
38. McKay JA, Waltham KJ, Williams EA, Mathers JC. Folate depletion
during pregnancy and lactation reduces genomic DNA methylation in
murine adult offspring. Genes Nutr 2011;6:189-96.
39. Sinclair KD, Allegrucci C, Singh R, Gardner DS, Sebastin S,
Bispham J, et al. DNA methylation, insulin resistance, and blood
pressure in offspring determined by maternal periconceptional B vitamin
and methionine status. Proc Natl Acad Sci U S A 2007;104:19351-6.
40. Watreland RA, Jirtle RL. Early nutrition, epigenetic changes at
Asian J Pharm Clin Res, Vol 11, Issue 1, 2018, 17-20
Varma et al.
transposons and imprinted genes, and enhanced susceptibility to adult
chronic diseases. Nutrition 2004;20:63-8.
42. Dwarkanath P, Barzilay JR, Thomas T, Thomas A, Bhat S, Kurpad AV.
High folate and low vitamin B-12 intakes during pregnancy are
associated with small-for-gestational age infants in South Indian
women: A prospective observational cohort study. Am J Clin Nutr
43. Sarheed O, Ramesh K, Shah F. In vitro evaluation of dissolution profile
of two commercially available folic acid preparations. Int J Pharm
Pharm Sci 2015;7:473-5.
44. Antakli S, Sarkees N, Sarraf T. Determination of water-soluble vitamins
B1, B2, B3, B6, B9, B12 and C ON C18 column with particle size
3 µm in some manufactured food products by HPLC with UV-dad/FLD
detection. Int J Pharm Pharm 20157:219-24.
... However, no ocular phenotype evolved in retina ∆Mmachc mice. This finding is surprising as MMACHC has an essential function in Cbl processing required for odd-chain fatty acid and branched-chain amino acid metabolism [55][56][57] as well as methionine and SAM synthesis [58][59][60]. Although cells that underwent recombination may still express a shorter mRNA at reduced levels, it seems unlikely that a functional protein was generated. ...
Full-text available
Combined methylmalonic aciduria with homocystinuria (cblC type) is a rare disease caused by mutations in the MMACHC gene. MMACHC encodes an enzyme crucial for intracellular vitamin B12 metabolism, leading to the accumulation of toxic metabolites e.g. methylmalonic acid (MMA) and homocysteine (Hcy), and secondary disturbances in folate and one-carbon metabolism when not fully functional. Patients with cblC deficiency often present in the neonatal or early childhood period with a severe multisystem pathology, which comprises a broad spectrum of treatment-resistant ophthalmological phenotypes, including retinal degeneration, impaired vision, and vascular changes. To examine the potential function of MMACHC in the retina and how its loss may impact disease, we performed gene expression studies in human and mouse, which showed that local expression of MMACHC in the retina and retinal pigment epithelium is relatively stable over time. To study whether functional MMACHC is required for retinal function and tissue integrity, we generated a transgenic mouse lacking Mmachc expression in cells of the peripheral retina. Characterization of this mouse revealed accumulation of cblC disease related metabolites, including MMA and the folate-dependent purine synthesis intermediates AICA-riboside and SAICA-riboside in the retina. Nevertheless, fundus appearance, morphology, vasculature, and cellular composition of the retina, as well as ocular function, remained normal in mice up to 6 or 12 months of age. Our data indicates that peripheral retinal neurons do not require intrinsic expression of Mmachc for survival and function and questions whether a local MMACHC deficiency is responsible for the retinal phenotypes in patients.
... Methylation is the process of the controlled transfer of a methyl group to amino acids, proteins, deoxyribonucleotide, and other biological intermediates [1][2][3][4][5][6][7][8]. Methyl transfer reactions involving transition metal complexes, such as cobalt-dependent cobalamins (Cbls) are considered as one of the most important biologically relevant reactions. ...
Methyl transfer reactions mediated by cobalamins (Cbls) have been considered as one of the most important biologically relevant molecular transformations for many enzymatic reactions catalyzed by Cbl-dependent enzymes. The exact mechanism of methyl transfer reactions involving Cbls is still poorly understood. To investigate the mechanistic details of Cbl mediated methylations by alkyl halides, density functional theory (DFT) along with the polarizable continuum model (PCM/water) for solvation has been applied. Two different mechanisms have been examined, namely SN2 and radical-based electron transfer (ET) to elucidate the methyl transfer reaction. The calculations have suggested that the methyl transfer from methyl halides proceeds through SN2 nucleophilic displacement. However, with more bulky alkyl substrate, namely isopropyl and tert-butyl halides the reaction followed the ET-based radical pathway which is associated with an ET from diradical form of cob(I)alamin to alkyl halides. Our proposed mechanism for alkyl transfer reaction corroborates with the experimental findings, which reported a mechanistic switch from a two-electron (SN2-type) to a one-electron mechanism for sterically demanding alkyl halides. The present theoretical contribution provides more in-depth insight into the methyl transfer reaction catalyzed by corrinoid-dependent methyltransferases.
Full-text available
Obesity is a worldwide epidemic responsible for 5% of global mortality. The risks of developing other key metabolic disorders like diabetes, hypertension and cardiovascular diseases (CVDs) are increased by obesity, causing a great public health concern. A series of epidemiological studies and animal models have demonstrated a relationship between the importance of vitamin B12 (B12) and various components of metabolic syndrome. High prevalence of low B12 levels has been shown in European (27%) and South Indian (32%) patients with type 2 diabetes (T2D). A longitudinal prospective study in pregnant women has shown that low B12 status could independently predict the development of T2D five years after delivery. Likewise, children born to mothers with low B12 levels may have excess fat accumulation which in turn can result in higher insulin resistance and risk of T2D and/or CVD in adulthood. However, the independent role of B12 on lipid metabolism, a key risk factor for cardiometabolic disorders, has not been explored to a larger extent. In this review, we provide evidence from pre-clinical and clinical studies on the role of low B12 status on lipid metabolism and insights on the possible epigenetic mechanisms including DNA methylation, micro-RNA and histone modifications. Although, there are only a few association studies of B12 on epigenetic mechanisms, novel approaches to understand the functional changes caused by these epigenetic markers are warranted.
Full-text available
The developmental origins of adult health and disease (DOHaD) hypothesis that argues for a causal relationship between under-nutrition during early life and increased risk for a range of diseases in adulthood is gaining epidemiological support. One potential mechanism mediating these effects is the modulation of epigenetic markings, specifically DNA methylation. Since folate is an important methyl donor, alterations in supply of this micronutrient may influence the availability of methyl groups for DNA methylation. We hypothesised that low folate supply in utero and post-weaning would alter the DNA methylation profile of offspring. In two separate 2 × 2 factorial designed experiments, female C57Bl6/J mice were fed low- or control/high-folate diets during mating, and through pregnancy and lactation. Offspring were weaned on to either low- or control/high-folate diets, resulting in 4 treatment groups/experiment. Genomic DNA methylation was measured in the small intestine (SI) of 100-day-old offspring. In both experiments, SI genomic DNA from offspring of low-folate-fed dams was significantly hypomethylated compared with the corresponding control/high folate group (P = 0.009/P = 0.006, respectively). Post-weaning folate supply did not affect SI genomic DNA methylation significantly. These observations demonstrate that early life folate depletion affects epigenetic markings, that this effect is not modulated by post-weaning folate supply and that altered epigenetic marks persist into adulthood.
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Potential adverse effects of excess maternal folic acid supplementation on a vegetarian population deficient in vitamin B(12) are poorly understood. We have previously shown in a rat model that maternal folic acid supplementation at marginal protein levels reduces brain omega-3 fatty acid levels in the adult offspring. We have also reported that reduced docosahexaenoic acid (DHA) levels may result in diversion of methyl groups towards DNA in the one carbon metabolic pathway ultimately resulting in DNA methylation. This study was designed to examine the effect of normal and excess folic acid in the absence and presence of vitamin B(12) deficiency on global methylation patterns in the placenta. Further, the effect of maternal omega 3 fatty acid supplementation on the above vitamin B(12) deficient diets was also examined. Our results suggest maternal folic acid supplementation in the absence of vitamin B(12) lowers plasma and placental DHA levels (p<0.05) and reduces global DNA methylation levels (p<0.05). When this group was supplemented with omega 3 fatty acids there was an increase in placental DHA levels and subsequently DNA methylation levels revert back to the levels of the control group. Our results suggest for the first time that DHA plays an important role in one carbon metabolism thereby influencing global DNA methylation in the placenta.
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Unmetabolized serum folic acid (UMFA) has been detected in adults. Previous research indicates that high folic acid intakes may be associated with risk of cancer. The objective was to examine UMFA concentrations in relation to dietary and supplemental folate and status biomarkers in the US population aged > or =60 y. Surplus sera were analyzed with the use of data from the National Health and Nutrition Examination Survey (NHANES) 2001-2002, a cross-sectional, nationally representative survey (n = 1121). UMFA was detected in 38% of the population, with a mean concentration of 4.4 +/- 0.6 nmol/L (median: 1.2+/- 0.2 nmol/L). The group with UMFA (UMFA+) had a significantly higher proportion of folic acid supplement users than did the group without UMFA (60% compared with 41%). UMFA+ men and women also had higher supplemental and total (food + supplements) folic acid intakes than did their counterparts without UMFA. Forty percent of the UMFA+ group was in the highest quartile of total folic acid intake, but total folic acid intake was only moderately related to UMFA concentrations (r(2) = 0.07). Serum folate concentrations were significantly higher in the UMFA+ group and were predictive of UMFA concentrations (r(2) = 0.15). Serum 5-methyltetrahydrofolate and vitamin B-12 concentrations were higher in the UMFA+ group, whereas there was no difference between the 2 UMFA groups in red blood cell folate, serum homocysteine, or methylmalonic acid concentrations. Approximately 40% of older adults in the United States have UMFA that persists after a fast, and the presence of UMFA is not easily explained in NHANES by folic acid intakes alone. Given the possibility that excessive folic acid exposure may relate to cancer risk, monitoring of UMFA may be warranted.
Chronic nutritional deficiencies in folate, choline, methionine, vitamin B-6 and/or vitamin B-12 can perturb the complex regulatory network that maintains normal one-carbon metabolism and homocysteine homeostasis. Genetic polymorphisms in these pathways can act synergistically with nutritional deficiencies to accelerate metabolic pathology associated with occlusive heart disease, birth defects and dementia. A major unanswered question is whether homocysteine is causally involved in disease pathogenesis or whether homocysteinemia is simply a passive and indirect indicator of a more complex mechanism. S-Adenosylmethionine and S-adenosylhomocysteine (SAH), as the substrate and product of methyltransferase reactions, are important metabolic indicators of cellular methylation status. Chronic elevation in homocysteine levels results in parallel increases in intracellular SAH and potent product inhibition of DNA methyltransferases. SAH-mediated DNA hypomethylation and associated alterations in gene expression and chromatin structure may provide new hypotheses for pathogenesis of diseases related to homocysteinemia.
Objectives: Folic acid is a water-soluble B vitamin that is naturally present in some foods such as dark green leafy vegetables and dairy products, added to others such as fortified breads, and available as a dietary supplement. In a study performed in the United States, out of nine multivitamin products, only three products met US Pharmacopeia (USP) standard for folic acid dissolution. The aim of this study is to evaluate the dissolution profile of folic acid supplements in the United Arab Emirates according to the USP standards. Methods: Two commercial brands were collected from the market that are commonly prescribed. Water and citrate buffer (pH 6.0) were used as dissolution media. Results: Both products passed the dissolution testing of releasing the required amount of drug substance (75%) within 60 minutes. Also, both products contain an over age of up to 150% of the labeled amount to ensure the availability of the claimed amount. Conclusion: The study indicates that dissolution test is well established, reproducible, reliable and valuable tool for characterizing a drug product at different stages in its lifecycle. The use of citrate buffer showed a significant change in the release of folic acid from the tablets.
Objective: Objective of the study was to develop a simple, precise and accurate RP-HPLC ion-pair method, for the determination of water-soluble vitamins (B1, B2, B3, B6, B9, B12, and C) in some manufactured food products. Methods: RP-HPLC with C18 BDS (100 x 4.6 mm; 3 μm) column were used. Mobile phase constituents were solvent (A): 5.84 mM of hexane-1-sulfonic acid sodium: acetonitrile (95:5) with 0.1% triethylamine at pH 2.5 and solvent (B): 5.84 mM of hexane-1-sulfonic acid sodium: acetonitrile (50:50) with 0.1% triethylamine at pH 2.5, flow rate 1.6 ml/min, at column temperature 40 °C and suitable detection wavelength. Results: Two different detectors were used: photo diode array detector (UV-DAD) and the fluorescence detector (FLD). Calibration graphs plotted with five concentrations of each vitamin where linear regression coefficients R2˃ 0.9972. LOQ values were 50.0, 81.0, 19.1, 19.0, 30.0, 9.7, 50.0 μg/l with DAD for vitamins B1, B2, B3, B6, B9, B12 and C respectively, and 5.7, 4.1 μg/l with FLD for vitamins B2, B6 respectively. LOD values were 16.5, 26.7, 6.3, 6.3, 9.9, 3.2, 16.5 μg/l with DAD for vitamins B1, B2, B3, B6, B9, B12 and C respectively, and 1.9, 1.3 μg/l with FLD for vitamins B2, B6 respectively. Conclusion: The proposed method was successfully applied to analysis mixture of seven water-soluble vitamins in pure form and in manufactured food products, with average recovery of 98.14% to 100.96%. © 2015, International Journal of Pharmacy and Pharmaceutical Sciences. All Rights Reserved.
Folate is a water-soluble B-vitamin and enzymatic cofactor that is necessary for the synthesis of purine and thymidine nucleotides and for the synthesis of methionine from homocysteine. Impairment of folate-mediated one-carbon metabolic pathways can result from B-vitamin deficiencies and/or single nucleotide polymorphisms, and increases risk for pathologies, including cancer and cardiovascular disease, and developmental anomalies including neural tube defects. Although several well validated metabolic and genomic biomarkers for folate deficiency exist, our understanding of the biochemical and genetic mechanisms whereby impaired folate metabolism increases risk for developmental anomalies and disease is limited, as are the mechanisms whereby elevated folate intake protects against these pathologies. Therefore, current initiatives to increase folate intakes in human populations to ameliorate developmental anomalies and prevent disease, while effective, lack predictive value with respect to unintended adverse outcomes.
Folic acid supplementation in those with a low vitamin B-12 intake or status may have adverse effects. These effects are unknown with regard to birth outcome in pregnant Indian women who are routinely supplemented with high doses of folic acid. The objective was to examine the association of unbalanced vitamin B-12 and total folate (folic acid supplement + dietary folate) intakes during pregnancy with outcomes in small-for-gestational-age (SGA) infants. This was a prospective observational cohort study of 1838 pregnant women in South India. Low intake of dietary vitamin B-12 in the presence of high total folate intake was examined as the ratio of vitamin B-12 intake to total folate intake. The inadequacy of vitamin B-12 intake (<1.2 μg/d) assessed by a food-frequency questionnaire in the first, second, and third trimesters of pregnancy was 25%, 11%, and 10%, respectively. Multivariate log binomial regression showed that low vitamin B-12 and folate intakes in the first trimester were independently associated with a higher risk of SGA. In a subgroup of women with high supplemental folic acid intakes in second trimester, those with the lowest tertile of vitamin B-12:folate ratio had a higher risk of SGA outcome with reference to the highest tertile (adjusted RR: 2.73; 95% CI: 1.17, 6.37). A similar trend was observed in the analysis of blood micronutrient status in a random subset (n = 316) of the sample. These findings suggest that, in addition to vitamin B-12 and folate deficiencies alone, there may be adverse birth outcomes associated with unbalanced vitamin B-12 and folate intakes or status during pregnancy. These findings have important implications for the antenatal B vitamin supplementation policy in India. This trial was registered at the Clinical Trial Registry of India as 2013/07/005342.
The objective of the study was to evaluate LINE-1 methylation as an intermediate biomarker for the effect of folate and vitamin B12 on the occurrence of higher grades of cervical intraepithelial neoplasia (CIN ≥ 2). This study included 376 women who tested positive for high-risk human papillomaviruses and were diagnosed with CIN ≥ 2 (cases) or CIN ≤ 1 (non-cases). CIN ≥ 2 (yes/no) was the dependent variable in logistic regression models that specified the degree of LINE-1 methylation of peripheral blood mononuclear cells (PBMCs) and of exfoliated cervical cells (CCs) as the independent predictors of primary interest. In analyses restricted to non-cases, PBMC LINE-1 methylation (≥ 70% versus <70%) and CC LINE-1 methylation (≥ 54% versus <54%) were the dependent variables in logistic regression models that specified the circulating concentrations of folate and vitamin B12 as the primary independent predictors. Women in the highest tertile of PBMC LINE-1 methylation had 56% lower odds of being diagnosed with CIN ≥ 2 (odds ratio 0.44, 95% confidence interval 0.24-0.83, P = 0.011), whereas there was no significant association between degree of CC LINE-1 methylation and CIN ≥ 2 (odds ratio 0.86, 95% confidence interval 0.51-1.46, P = 0.578). Among non-cases, women with supraphysiologic concentrations of folate (>19.8 ng/mL) and sufficient concentrations of plasma vitamin B12 (≥ 200.6 ng/mL) were significantly more likely to have highly methylated PBMCs compared with women with lower folate and lower vitamin B12 (odds ratio 3.92, 95% confidence interval 1.06-14.52, P = 0.041). None of the variables including folate and vitamin B12 were significantly associated with CC LINE-1 methylation. These results suggest that a higher degree of LINE-1 methylation in PBMCs, a one-carbon nutrient-related epigenetic alteration, is associated with a lower risk of developing CIN.
Epigenetics is focused on understanding the control of gene expression beyond what is encoded in the sequence of DNA. Central to growing interest in the field is the hope that more can be learned about the epigenetic regulatory mechanisms underlying processes of human development and disease. Researchers have begun to examine epigenetic alterations - such as changes in promoter DNA methylation, genomic imprinting, and expression of miRNA - to learn more about epigenetic regulation in the placenta, an organ whose proper development and function are crucial to the health, growth, and survival of the developing fetus. A number of studies are now making important links between alterations to appropriate epigenetic regulation in the placenta and diseases of gestation and early life. In addition, these studies are adding important insight into our understanding of trophoblast biology and differentiation as well as placental immunology. Examining epigenetic alterations in the placenta will prove especially important in the search for biomarkers of exposure, pathology, and disease risk and can provide critical insights into the biology of development and pathogenesis of disease. Thus, epigenetic alterations may aid in disease diagnosis and prognosis as well as in targeting new treatment and prevention strategies.