1. Absorption, transport and metabolism of folate
Domina Petric, MD

2. Absorption I.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

3. Hydrolysis of folyl polyglutamates
Under fasting conditions, folic acid, 5-methyl-FH4 and 5-formyl-FH4 are virtually completely absorbed.
Most polyglutamyl folates are absorbed at efficiencies in the general range of 60-80%.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

4. Hydrolysis of folyl polyglutamates
The majority of food folates occur as reduced polyglutamates.
They must be cleaved to the mono- or diglutamate forms for absorption.
This is accomplished by the action of an exocarboxypeptidase folyl γ-glutamyl carboxypeptidase, more commonly called folyl conjugase.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

5. Folyl conjugase
Conjugase activity is widely distributed in the mucosa of the proximal small intestine, both intracellularly and in association with the brush border.
These appear to be different enzymes:
75 kDa intracellular enzyme is localized in the lysosomes and has an optimum of pH 4,5-5,0.
700 kDa brush border enzyme has an optimum of pH 6,5-7,0.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

6. Folyl conjugase
Folyl conjugase activities have also been found in bile, pancreatic juice, kidney, liver, placenta, bone marrow, leukocytes and plasma.
In the uterus, conjugase activity is induced by estrogen.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

7. Loss of conjugase activity results in impaired folate absorption.
Folyl conjugase
Loss of conjugase activity results in impaired folate absorption.
Conjugase activity is reduced by nutritional zinc deficiency or by exposure to naturally occurring inhibitors in foods.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

8. Folyl conjugase
Natural conjugase inhibitors are contained in certain foods: cabbage, oranges, yeast, beans (red kidney, pinto, lima, navy, soy), lentils and black-eyed peas.
The presence of conjugase inhibitors reduces folate bioavailability.
This effect appears to be the basis for the low availability of the vitamin in orange juice.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

9. Folyl conjugase cholestyramine (which binds folates)
Folate absorption can also be reduced by certain drugs including:
cholestyramine (which binds folates)
ethanol (which inhibits deconjugation)
salicylazosulfapyridine
diphenylhydantoin
aspirin and other salicylates
several nonsteroidal anti-inflammatory drugs
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

10. Uptake
Dietary folates are absorbed as folic acid, 5-methyl-FH4 and 5-formyl-FH4.
The overall efficiency of folate absorption appears to be ∼50% (10-90%).
Malabsorption of the vitamin occurs in diseases affecting the intestinal mucosa.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

11. Microclimate hypothesis
Folate absorption is dependent on the pH of the proximal jejunum, with an optimum at pH 6,0-6,3.
Elevated absorption of folate in individuals with pancreatic exocrine insufficiency may be due to the low pancreatic excretion of bicarbonate in that condition.
Loss of buffering capacity renders the intestinal lumenal milieu slightly more acidic, minimizing the charge on the folate molecule, thus facilitating its diffusion across the brush border membrane.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

12. Uptake
Under more basic conditions (pH>6.0), folate absorption falls off rapidly.
Two jejunal brush border folate-binding proteins (FBPs) have been isolated.
Three mechanisms are involved in folate absorption.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

13. Folate transporter
Folic acid is actively transported across the jejunum, and perhaps the duodenum, by an Na+-coupled, carrier-mediated process that is stimulated by glucose and shows a pH maximum at about pH 6.
The transporter is a transmembrane protein with much greater affinities for folic acid than for reduced folates.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

14. Its expression is suppressed by exposure to alcohol.
Folate transporter
The active transport process is maximal at lumenal folate concentrations of µM.
Its expression is suppressed by exposure to alcohol.
Hereditary folate malabsorption, apparently involving failure of transporter expression, has been reported.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

15. Folate receptor
A high-affinity, folate-binding protein, also called the folate receptor (FR), has been identified in the intestine and other tissues.
The intestine contains three isoforms, each of which is associated with the brush border.
Two isoforms (α- and ß-) are glycosylphosphatidylinositol (GPI)-anchored to the membrane.
β-type is a soluble protein.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

16. Folate receptor
The GPI-anchored receptor FRα has been shown to cycle between intracellular and extracellular compartments while remaining bound to the membrane, thus effecting the high-affinity uptake of folate and nonmolar concentrations.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

17. Diffusion
Folic acid can also be absorbed passively, apparently by diffusion.
This nonsaturable process is linearly related to lumenal folate concentration and can account for 20-30% of folate absorption at high folate intakes.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

18. Methylation
Folic acid taken up by the intestinal mucosal cell is reduced to FH4, which can either be transferred without further metabolism to the portal circulation or alkylated (by methylation to 5-methyl-FH4) before being transferred.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

19. Transport
II.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

20. Free in plasma
Folate is transported to the tissues mostly as monoglutamate derivatives in free solution in the plasma.
Plasma folate concentrations in humans are typically in the range of nM.
Most circulates in free solution, but some is bound to low-affinity protein binders such as albumin, and others are bound to a soluble form of the high-affinity FBP.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

21. Free in plasma
Erythrocytes contain greater concentrations of folate: nM.
These stores are accumulated during erythropoiesis.
The mature erythrocyte does not take up folate.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

22. Free in plasma
The folate levels of both plasma and erythrocytes are reduced by cigarette smoking.
Habitual smokers show plasma folate levels that are more than 40% less than those of nonsmokers.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

23. Free in plasma
More than 80% of impoverished chronic alcoholics show abnormally low serum levels and some 40% show low erythrocyte levels.
This corresponds to a similar incidence (34-42%) of megaloblastosis of the bone marrow in alcoholic patients.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

24. Cellular uptake
The cellular uptake of folates occurs exclusively with monoglutamate derivatives found in the plasma, as the polyglutamates can not cross biological membranes.
The cellular uptake of folate involves the three processes outlined above for enteric absorption.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

25. The folate transporter is expressed in most tissues.
It has affinities for various folates varying both between tissues and between the apical and basolateral membranes.
Its affinities for folic acid are two orders of magnitude greater than those for reduced folates.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

26. A high-affinity FR is expressed in many tissues.
Folate receptor
A high-affinity FR is expressed in many tissues.
It mediates the uptake of folate in the kidney.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

27. Cellular uptake
Within cells, FH4 is methylated to yield 5-methyl-FH4, which is bound to intracellular macromolecules.
As a result of methylation in the mammary gland, 5-methyl-FH4 comprises three-quarters of the folates in breast milk.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

28. Cellular uptake
Folate is held in cells by conversion to folyl polyglutamates.
Polyglutamation traps folates inside cells at concentrations greater (by one to two orders of magnitude) than those of extracellular fluids.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

29. Folate-Binding Proteins
Folate-binding proteins (FBPs) have been identified in plasma, milk, erythrocytes, leukocytes, intestinal mucosa, kidney, liver, placenta, choroid plexus and urine.
Each binds folates noncovalently with high affinity, such that the complex does not dissociate under physiological conditions.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

30. Tissue distribution
The total body content of folate is 5-10 mg, about half of which resides in the liver in the form of tetra-, penta-, hexa- and heptaglutamates of methyl-FH4 and 10-formyl-FH4.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

31. Tissue distribution
In tissues with rapid cell division (intestinal mucosa, regenerating liver, carcinoma), relatively low concentrations of 5-methyl-FH4 are found, usually with concomitant elevations in 10-formyl-FH4.
In contrast, in tissues with low rates of cell division (normal liver), 5-methyl-FH4 predominates.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

32. Tissue distribution
Brain folate (mostly 5-methyl-FH4) levels tend to be very low, with a subcellular distribution (penta- and hexaglutamates mostly in the cytosol and polyglutamates mostly in the mitochondria) the opposite of that found in liver.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

33. Metabolism
III.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

34. Reduction of the pteridine ring system
Reduction of the pterin ring from the two nonreduced states, folic acid and dihydrofolic acid (FH2), to the fully reduced form tetrahydrofolic acid (FH4) that is capable of accepting a single-carbon unit is accomplished by the cytosolic enzyme 7,8-dihydrofolate reductase.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

35. Reduction of the pteridine ring system
This activity is found in high amounts in liver and kidney and in rapidly dividing cells (tumor).
The reductase is inhibited by several important drugs including the cancer chemotherapeutic drug methotrexate, which appears to exert its antitumor action by inhibiting the reductase activity of tumor cells.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

36. Reactions of the polyglutamyl side chain
The folyl monoglutamates, that are taken up by cells, are trapped therein as polyglutamate derivatives that can not cross cell membranes.
Polyglutamate forms are also mobilized by side-chain hydrolysis to the monoglutamate.
These conversions are catalyzed by two enzymes: polyglutamation and cellular conjugases.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

37. Acquisition of single-carbon moieties at certain positions (N-5 or N-10) on the pterin ring
Folate is metabolically active as a variety of derivatives with single-carbon units at the oxidation levels of formate, formaldehyde or methanol substituted at the N-5 and/or N-10 positions of the pteridine ring system.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

38. Acquisition of single-carbon moieties at certain positions (N-5 or N-10) on the pterin ring
The main source of single-carbon fragments is serine hydroxymethyltransferase, which uses the dispensable amino acid serine as the single-carbon donor.
Each folyl derivative is a donor of its single-carbon unit in metabolism.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

39. Catabolism and excretion
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

40. Catabolism
Tissue folates appear to turn over by the cleavage of the polyglutamates at the C-9 and N-10 bonds to liberate the pteridine and p-aminobenzoylpolyglutamate moieties.
This cleavage probably results from chemical oxidation of the cofactor in both the intestinal lumen (dietary and enterohepatically recycled folates) and the tissues.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

41. Catabolism
Once formed, p-aminobenzoylpolyglutamate is degraded, by the action of folyl conjugase, and is acetylated to yield p-acetaminobenzoylglutamate and p-acetoaminobenzoate.
Urinary levels of p-acetoaminobenzoate are correlated with the total body folate pool.
Folate breakdown is greatest under hyperplastic conditions: rapid growth, pregnancy.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

42. Excretion
Intact folates and the water-soluble side-chain metabolites p-acetaminobenzoylglutamate and p-acetaminobenzoate are excreted in the urine and bile.
The total urinary excretion of folates and metabolites is small: ≤1% of total body stores per day.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

43. Excretion Fecal concentrations of folates are usually high.
These concentrations represent mainly folates of intestinal microfloral origin.
November 22, 2018
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc

44. Literature
Combs GF. The Vitamins. Fundamental Aspects in Nutrition and Health. Elsevier Inc
November 22, 2018