2.1. Tworzenie krwinek (komórek krwi) (ID 79)

The claimed effect is “blood formation”. The Panel assumes that the target population is the general population.
The Panel considers that normal blood formation is beneficial to human health.

2.2. Metabolizm homocysteiny (ID 80)

The claimed effect is “homocysteine metabolism”. The Panel assumes that the target population is the general population.
The Panel considers that normal homocysteine metabolism is beneficial to human health.

2.3. Metabolizm energetyczny (ID 90)

The claimed effect is “the role of water-soluble vitamins in energy metabolism / transformation of food into physiological energy”. The Panel assumes that the target population is the general population.
The Panel considers that normal energy-yielding metabolism is beneficial to human health.

2.4. Funkcjonowanie układu odpornościowego (ID 91)

The claimed effect is “the role of vitamins and minerals in immunity”. The Panel assumes that the target population is the general population.
The Panel considers that a normal function of the immune system is beneficial to human health.

2.5. Funkcjonowanie naczyń krwionośnych (ID 94, 175, 192)

The claimed effects are “vascular function/cardiovascular health”, “cardiovascular health” and “folate helps keep arteries/ blood vessels healthy”. The Panel assumes that the target population is the general population.
The claimed effect “cardiovascular health” is not sufficiently defined in the list. From the proposed wordings the Panel assumes the claimed effect relates to normal function of the blood vessels.
The Panel considers that normal function of blood vessels is beneficial to human health.

2.6. Podziały komórek (ID 193)

The claimed effect is “cell division”. The Panel assumes that the target population is the general population.
The Panel notes that cell division is a crucial process for tissue growth and development and for tissue maintenance through cell turnover.
The Panel considers that normal cell division is beneficial to human health.

2.7. Prawidłowy wzrost matczynych tkanek podczas ciąży (ID 2882)

The claimed effect is “effect on a normal pregnancy”. The target population is women planning to become pregnant and pregnant women.
The Panel interprets the claimed effect “effect on a normal pregnancy” as normal maternal tissue growth during pregnancy.
The Panel considers that normal maternal tissue growth during pregnancy is beneficial to human health.

3.1. Tworzenie krwinek (komórek krwi) (ID 79)

In folate deficiency, a net decrease in 5,10-methlyene-tetrahydrofolate interrupts the reaction mediated by thymidylate synthase, which converts deoxyuridine monophosphate to deoxythymidine monophosphate. The excess deoxyuridine monophosphate is then phosphorylated to deoxyuridine triphosphate and is incorporated into DNA. This faulty incorporation is immediately recognised by an editorial enzyme and is excised. When this process is repeated several times, it leads to DNA fragmentation and to perturbation of the cell cycle giving rise to megaloblastosis. All proliferating cells exhibit megaloblastosis; however the changes are most striking in the blood and bone marrow (Chitambar and Anthony, 2006). The main clinical expression of folate deficiency is megaloblastic anemia characterised by large, abnormally nucleated erythrocytes that accumulate in the bone marrow. There are also decreased numbers of white cells and platelets as a result of the general impairment of cell division (Bailey and Gregory, 2006).
The Panel concludes that a cause and effect relationship has been established between the dietary intake of folate and normal blood formation.

3.2. Metabolizm homocysteiny (ID 80)

5-methyl-tetrahydrofolate is an important functional and regulatory component of the folate- dependent pathway for the production of methionine from homocysteine which is catalysed by methionine synthase. In this reaction, a methyl group is sequentially transferred from 5-methyl- tetrahydrofolate to the cobalamin coenzyme of the methionine synthase, and then to homocysteine to form methionine (Bailey and Gregory, 2006).
Under conditions of maximal metabolic efficiency, plasma concentrations of homocysteine range from 4 to 10 µmol/L. Metabolic blocks in homocysteine metabolism lead to accumulation of intracellular homocysteine with subsequent export into the blood. Depending on the magnitude of the metabolic impairment, plasma homocysteine can rise to varying degrees. Hyperhomocysteinemia is also caused by B vitamin deficiencies. Deficiencies of folate, vitamin B6 and vitamin B12 lead to impaired remethylation of homocysteine causing mild, moderate, or severe elevations in plasma homocysteine, depending on the severity of the deficiency, as well as coexistence of genetic or other factors that interfere with homocysteine metabolism (Miller, 2005).
The Panel concludes that a cause and effect relationship has been established between the dietary intake of folate and normal homocysteine metabolism.

3.3. Metabolizm energetyczny (ID 90)

Folate coenzymes are involved in the interconversion of serine to glycine and in the catabolism of histidine to glutamic acid (McPartlin, 2005).
The Panel notes that these reactions do not lead to intermediates that can be considered as significant source of energy for the body.
The Panel concludes that a cause and effect relationship has not been established between the dietary intake of folate and normal energy-yielding metabolism.

3.4. Funkcjonowanie układu odpornościowego (ID 91)

Folate plays a crucial role in nucleotide synthesis, and thus may affect immune cell proliferation and responsiveness. Folate deficiency has been shown to reduce proliferation of various cell types. Cells lacking folate accumulate in the S-phase owing to nucleotide imbalance and slow DNA synthesis; such cells also have increased uracil misincorporation and DNA damage. When folate is added back
to folate-deficient cells, there is a reversal of the S-phase accumulation, and proliferation is restored. Folate deficiency has been shown to reduce the proportion of circulating T lymphocytes and their proliferation in response to mitogen activation. In addition, folate deficiency induced in PHA- activated human T lymphocytes induced apoptosis and increased the ratio of CD4+ to CD8+ T cells because of a marked reduction in CD8+ cell proliferation. All these effects were reversible in vitro by either folate addition or nucleotide repletion, and suggest that folate status may affect the immune system by inhibiting the capacity of CD8+ T lymphocyte cells to proliferate in response to mitogen activation (Courtemanche et al., 2004).
The Panel concludes that a cause and effect relationship has been established between the dietary intake of folate and a normal function of the immune system.

3.5. Funkcjonowanie naczyń krwionośnych (ID 94, 175, 192)

A total of 73 references were cited to substantiate the claimed effect. Six references were textbook or opinions of scientific bodies and 47 references were not directly related to the claimed effect but to blood homocysteine concentrations, insulin sensitivity, markers of oxidative stress or to the relationship between hyperhomocysteinemia and the risk of cardiovascular disease. The Panel notes that the references cited did not provide any scientific data that could be used to substantiate the claimed effect.
One meta-analysis, three review papers and 16 human studies investigating the relationship between folic acid intake and endothelial function were cited. 15 of the human studies were conducted with high doses of folic acid (5 mg/d, 10 mg/d or 20 mg/d). The Panel notes that the doses used in these studies were 5 to 20 fold the Tolerable Upper Intake Level of folic acid for adults and 12.5 to 50 times the proposed conditions of use. The Panel considers that no scientific conclusions can be drawn from these studies for the substantiation of the claimed effect at the proposed conditions of use.
In a randomised double blind placebo controlled parallel design intervention, 56 subjects with coronary artery disease not exposed to folate fortification were randomised to receive either a low dose (400 µg/d) (n= 20) or high dose (5 mg/d) of folic acid (n=22) or placebo (n=14) for 7 weeks before coronary artery bypass grafting. Vascular function was quantified by high-resolution magnetic resonance imaging at baseline and at the end of the treatment period. Images of the aorta and carotid arteries were used to determine vascular distensibility and pulse-wave velocity as indices of vascular stiffness. FMD was used as a measure of endothelial function. Supplementation with both low and high doses of folic acid significantly improved aortic and carotid distensibility, reduced aortic pulse- wave velocity, and increased FMD compared with placebo, whereas no significant differences were observed between the folic acid groups for any outcome variable (Shirodaria et al., 2007). The Panel notes that this study was performed in patients with coronary artery disease before coronary artery bypass grafting and no evidence has been provided that the finding in this small study of cardiovascular disease patients can be extrapolated to the general population.
The meta-analysis included 14 randomised controlled trials. One trial had two intervention groups and thus 15 intervention groups were analysed. In total, this meta-analysis was based on 732 persons treated with folic acid (either alone or in combination with vitamin B6, vitamin B12, or both) or placebo for a median duration of 8 weeks and with a median study size of 34 participants. Most study participants were middle-aged males (median age 55.8 years, 86% males). The median folic acid dose was 5000 µg/d (range: 400–10,000 µg/d). The primary outcome was the net change in flow-mediated dilatation (FMD) induced by the supplementation with folic acid (either alone or in combination with vitamin B6, vitamin B12, or both). Supplementation with folic acid significantly improved FMD when all studies were considered together. Study design, mean age of the study population, study duration, or addition of vitamin B6 or vitamin B12 had no effect on the estimated change in FMD following folic acid supplementation, whereas the dose of folic acid used was an independent predictor of the outcome. Trials using lower doses of folic acid (400 - 800 µg/d, 4 intervention
groups) did not show a significant beneficial effect of folic acid intake on FMD, whereas the studies using doses ≥5000 µg/d (11 intervention groups) did (de Bree et al., 2007). The Panel notes that an effect of folic acid on endothelial function was only observed at doses that are 5 to 10 fold the Tolerable Upper Intake Level of folic acid for adults and 12.5 to 25 times the proposed conditions of use. The Panel considers that no scientific conclusions can be drawn from these studies for the substantiation the claimed effect at the proposed conditions of use.
The Panel concludes that a cause and effect relationship has not been established between the dietary intake of folate and normal function of blood vessels.

3.6. Podziały komórek (ID 193)

During the S-phase of the cell division cycle the DNA of the cell is replicated. This DNA synthesis is dependent on the presence of 5,10-methlyene-tetrahydrofolate, a co-factor of the thymidylate synthase, which converts deoxyuridine monophosphate to deoxythymidine monophosphate (Chitambar and Anthony, 2006). An additional function of folate in the nucleotide production involves the de novo synthesis of adenine and guanine (Bailey and Gregory, 2006).
The Panel concludes that a cause and effect relationship has been established between the intake of folate and normal cell division.

3.7. Prawidłowy wzrost matczynych tkanek podczas ciąży (ID 2882)

During pregnancy folate is needed for increasing the mother's red blood cell mass, for the formation of the placenta and for the growth of the foetus, the uterus, breasts and other maternal tissues (Chitambar and Anthony, 2006) owing to its role in cell division (see section 2.3.6).
The Panels concludes that a cause and effect relationship has been established between the dietary intake of folate and normal maternal tissue growth during pregnancy.

4.1. Tworzenie krwinek (komórek krwi) (ID 79)

The Panel considers that the following wording reflects the scientific evidence: “Folate contributes to normal blood formation.”

4.2. Metabolizm homocysteiny (ID 80)

The Panel considers that the following wording reflects the scientific evidence: “Folate contributes to normal homocysteine metabolism.”

4.3. Funkcjonowanie układu odpornościowego (ID 91)

The Panel considers that the following wording reflects the scientific evidence: “Folate contributes to a normal function of the immune system.”

4.4. Podziały komórek (ID 193)

The Panel considers that the following wording reflects the scientific evidence: “Folate contributes to normal cell division.”

4.5. Prawidłowy wzrost matczynych tkanek podczas ciąży (ID 2882)

The Panel considers that the following wording reflects the scientific evidence: “Folate contributes to normal maternal tissue growth during pregnancy.”