Folic acid or folate (synonyms: vitamin B9, vitamin B11, vitamin M) is the generic term for a hydrophilic (water-soluble) vitamin. Scientific interest in this vitamin began in 1930, when a factor was discovered by Lucy Wills in liver, yeast, and spinach that has growth-promoting and antianemic (prevents anemia) effects. In 1938, Day demonstrated in experiments on monkeys that an appropriate deficient diet triggers the symptoms of anemia (anemia) and that these can be eliminated by administering yeast and liver preparations. This healing factor contained in yeast and liver was initially called vitamin M (monkey). The isolation of this factor from spinach leaves was achieved in 1941 by Snell et al. Derived from the Latin term folium (= leaf), this substance was given the name “folic acid“. However, in modern times it is known that the growth-stimulating and antianemic (preventing anemia) factor originally called folic acid does not occur in nature in the form it does and that its isolation was an artificial product. Folic acid has a heterocyclic structure consisting of a nitrogen-containing pteridine ring linked to the amino group of the para-aminobenzoic acid ring through its methyl group on the C6 atom – pteroic acid. A glutamic acid molecule is attached to the carboxyl end of p-aminobenzoic acid via a peptide bond (bond between a carboxyl and amino group). The chemical name of folic acid is therefore pteroylmonoglutamic acid or pteroylmonoglutamate (PteGlu). Folic acid, which does not occur in nature, can be clearly distinguished from folates [5-8, 11, 17]. Folates are part of biological systems and thus occur naturally in foods. Compared to folic acid, folates also consist of a pteridine and p-aminobenzoate molecule – pteroic acid – and a glutamate residue. However, the latter may be conjugated at its gamma-carboxyl group with further glutamate molecules, resulting in pteroylmonoglutamate (PteGlu) or pteroylpolyglutamate (PteGlu2-7), depending on the number of glutamyl residues. The pteridine ring is present in oxidized, dihydrogenated (addition of 2 hydrogen atoms) or tetrahydrogenated (addition of 4 hydrogen atoms) form, respectively. Finally, folates differ among themselves by the length of the glutamyl chain, the degree of hydrogenation (number of hydrogen atoms) of the pteridine molecule, and the substitution (exchange) of various C1 units (1-carbon units), such as methyl, formaldehyde, and formate residues, at the N5 and N10 atoms [1-3, 9, 10, 15, 18, 21]. The biologically active form of vitamin B9 is 5,6,7,8-tetrahydrofolate (THF) and its derivatives (derivatives). THF is the key coenzyme form and functions as an acceptor (receiver) and transmitter of C1 moieties, such as methyl groups, hydroxymethyl groups (activated formaldehyde), and formyl groups (activated formic acid), especially in protein and nucleic acid metabolism [1-3, 9, 15, 18]. The C1 residues originating from various metabolic reactions are bound to THF – THF-C1 compound – and with its help transferred to suitable acceptors (receivers). The various THF-C1 compounds, which differ in their oxidation state, are convertible into each other. The following THF-C1 compounds occur in the human organism.
- THF with the C1 residue formate (formic acid).
- 10-formyl THF
- 5-formyl-THF
- 5,10-Methenyl-THF
- 5-formimino-THF
- THF with the C1 residue formaldehyde (methanal).
- 5,10-methylene THF
- THF with the C1 residue methanol
- 5-methyl THF
Folic acid has the highest stability and oxidation state compared to natural folate compounds and is almost quantitatively (completely) absorbed as a pure substance. For this reason, after synthetic production, it is used in vitamin preparations, medicines and food fortification.Meanwhile, it is also possible to produce natural folates synthetically, such as the monoglutamate 5-methyltetrahydrofolate (5-MTHF, calcium L-methylfolate).According to the results of studies on bioavailability and lowering of homocysteine levels (naturally occurring amino acid, which in increased concentration can damage blood vessels), the biological active form 5-MTHF is equivalent to folic acid – 1 µg 5-MTHF is equivalent (equivalent) to 1 µg synthetic folic acid. Long-term studies investigating the influence of the administration of folic acid or 5-MTHF on the folate concentration in erythrocytes (red blood cells) even showed a significant superiority of natural 5-MTHF.Since according to the scientific panel of the European Food Safety Authority (engl. : European Food Safety Authority, EFSA 2004), there are no safety concerns against the use of 5-MTHF as a source of folate in foods, and the synthesizable natural form has been approved for use in dietary foods and supplements since February 2006, 5-MTHF can be used instead of folic acid.
Absorption
Folates are found in both animal and plant foods, where they are present as pteroylmonoglutamates, but mainly as pteroylpolyglutamates (60-80%). These must be enzymatically cleaved in the duodenum and proximal jejunum before absorption. Hydrolysis (cleavage by reaction with water) occurs by a gamma-glutamyl carboxypeptidase (conjugase) at the brush border membrane of enterocytes (cells of the intestinal epithelium), which converts polyglutamylfolate to monoglutamylfolate. The latter is taken up into intestinal mucosa cells (mucosal cells of the intestine) by an active glucose– and sodium-dependent carrier mechanism following saturation kinetics. 20-30% of monoglutamylfolates are absorbed (taken up) via a passive transport mechanism independent of folate dose [1-3, 10, 18, 20, 21]. While pteroylmonoglutamates, such as synthetic folic acid, are almost completely absorbed (>90%), polyglutamate compounds have an absorption rate of only about 20% due to incomplete enzymatic cleavage resulting from limited conjugase activity [2, 5-8, 10-12, 16, 18]. Since the folate content and the ratio of mono- to polyglutamates in individual foods vary greatly and vitamin losses during food preparation are difficult to calculate, it is not possible to give precise information on actual folate absorption. According to current reference values, a bioavailability of about 50% can be assumed for the folate compounds contained in food. The different absorption rate of mono- and polyglutamic acid compounds gives rise to the term folate equivalent (FE). The equivalent term is defined as follows.
- 1 µg FÄ = 1 µg dietary folate.
- 1 µg dietary folate = 0.5 µg synthetic folic acid
- 1 µg synthetic folic acid = 2 µg dietary folate (or 2 µg FÄ).
The absorption of vitamin B9 is a pH-dependent process with a maximum absorption at pH 6.0. In addition to pH, the release of folates from the cell structure, the type of food matrix (food texture), and the presence of other dietary ingredients, such as organic acids, folate-binding proteins, reducing substances, and conjugase-inhibiting factors, also influence the bioavailability of vitamin B9. Thus, folates from animal foods are better absorbed than from foods of plant origin because of their binding to proteins. Absorbed monoglutamylfolate is converted in enterocytes (cells of the intestinal epithelium) by two reduction steps via 7,8-dihydrofolate (DHF) to the metabolically active 5,6,7,8-THF, which reaches the liver via the portal vein partly in methylated (5-MTHF) and formylated (10-formyl-THF) forms, but mainly without C1 substituent as free THF.
Transport and distribution in the body
In the liver, methylation of tetrahydrofolate occurs. Minor formylation reactions also occur, so that vitamin B9 circulates in the blood predominantly as 5-MTHF (>80%) and to a lesser extent as 10-formyl-THF and free THF. While the 10-formyl-THF concentration in serum is constant in healthy adults, it is elevated in rapidly growing tissues. In blood serum, 50-60 % of folate compounds with low affinity (binding strength) are non-specifically bound to albumin, alpha-macroglobulin and transferrin.In addition, a specific folate binding protein exists that binds serum folates with high affinity but only in very small amounts (picogram (pg) range). The main function of this binding protein is to transport oxidized folates to the liver, where reduction to the biologically active THF occurs. The observation that women taking oral contraceptives (birth control pills) and during pregnancy have higher levels of folate binding proteins than men and children suggests a hormonal influence.Serum folate levels range from 7-17 ng/ml under basal conditions and are determined by the time of the last food intake (duration of food abstinence), level of folate intake, and individual folate supply. Circulating monoglutamyl folates in the blood, primarily 5-MTHF, are taken up into erythrocytes (red blood cells) and peripheral cells according to the laws of saturation kinetics, with a special carrier protein localized in the cell membrane mediating transport. Reduced folates have a significantly higher affinity for this transmembrane transport protein than oxidized folates. The passage of monoglutamate compounds of vitamin B9 through the blood-brain barrier (physiological barrier present in the brain between the blood circulation and central nervous system) probably also occurs according to saturation kinetics. The cerebrospinal fluid (CSF, cerebrospinal fluid) has a folate level two to three times higher than the blood serum. Intracellularly, the pteroylmonoglutamates are converted into the polyglutamate form (PteGlu2-7), mainly into penta- or hexaglutamates, since they can only be retained or stored in this form. For this purpose, 5-MTHF must first be demethylated (enzymatic cleavage of the methyl group) – a process that is vitamin B12-dependent – so that it can then be converted by polyglutamate synthetase (enzyme that transfers glutamate groups). In erythrocytes (red blood cells), polyglutamyl-THF, which has a high affinity for deoxyhemoglobin (oxygen-deficient form of hemoglobin), consists mostly of 4-7 glutamic acid molecules. The folate concentration of erythrocytes exceeds the folate content in serum about 40-fold (200-500 ng/ml). In mature erythrocytes, vitamin B9 has no metabolic functions, but only storage functions. Unlike reticulocytes (“juvenile” erythrocytes), which incorporate (absorb) substantial amounts of folate, mature erythrocytes (red blood cells) are largely impermeable (impermeable) to folate. For this reason, the erythrocyte folate level reflects vitamin B9 status more reliably than the highly fluctuating (fluctuating) serum folate level. Vitamin B9 is found in all tissues, and the distribution pattern shows a dependence on the mitotic rate (cell division rate) of the tissues – cell systems with high division rates, such as hematopoietic and epithelial cells, have high folate concentrations. The total body content of folate in humans is 5-10 mg, half of which is localized in the liver, primarily in the form of 5-MTHF and slightly as 10-formyl-THF. The liver is the main storage organ and regulates the supply to other organs. The biological half-life (time during which the concentration of a substance has decreased by half due to biological processes) of vitamin B9 is about 100 days.Due to low body reserves, serum vitamin B9 levels can be maintained for only 3-4 weeks on a folate-free diet. If deprivation of dietary folate continues, after a drop in serum folate concentration, oversegmentation (“right shift”) of neutrophil granulocytes (white blood cells that are part of the innate immune defense) occurs within 10-12 weeks, after 18 weeks, a decrease in the erythrocyte folate level, and after 4-5 months, the manifestation of megaloblastic anemia (anemia with larger-than-average erythrocyte precursor cells containing nuclei and hemoglobin in the bone marrow), which shows up in the blood count as hyperchromic, macrocytic anemia (synonym: megaloblastic anemia; anemia (anemia) due to vitamin B12, thiamine, or folic acid deficiency, resulting in impaired erythropoiesis (red blood cell production)).
Excretion
The amount of 10-90 µg monoglutamylfolate/day excreted in bile is subject to enterohepatic circulation (liver-gut circulation) and is almost quantitatively reabsorbed.Diseases of the small intestine or resection (surgical removal) of certain intestinal segments impair enteral reabsorption. The rapidly available, comparatively large biliary (affecting the bile) folate monoglutamate pool – folate concentration in the bile exceeds that in the blood plasma by a factor of 10 – together with the small intracellular folate pool (storage in liver and extrahepatic tissues) regulates short-term fluctuations in the alimentary vitamin B9 supply – folate homeostasis (maintenance of a constant folate serum level). With physiological (normal for metabolism) folate intake, only 1-12 µg (about 10-20% of the absorbed amount of folate monoglutamate) is eliminated daily by the kidney in the form of folic acid, 5-MTHF, 10-formyl-THF, and inactive degradation products, such as pteridine and acetamide benzoylglutamate derivative; most of the vitamin is tubularly reabsorbed (reabsorption by the renal tubules). An undersupply of vitamin B9 causes renal (affecting the kidney) excretion to decrease by stimulating tubular reabsorption. The amount of folate compounds excreted in feces (stool) is difficult to assess because microbially synthesized folates (vitamin B9 formed by bacteria in distal (lower) portions of the intestine) are always fecally excreted in addition to unabsorbed vitamin B9. Feces are thought to contain 5- to 10-fold higher amounts of folate than in the ingested diet.
