Part two

1. TUMS Vitamins and Coenzymes Part two Dr. Azin Nowrouzi, PhD Tehran University of Medical Sciences

2. 9. FOLIC ACID (folacin) • Folacin includes several derivatives of folic acid (monopteroylglutamic acid). • Folic acid is obtained primarily from yeasts and leafy vegetables as well as animal liver. Animals cannot synthesize PABA nor attach glutamate residues to pteroic acid, thus, requiring folate intake in the diet. “Microorganisms Only can synthesize Folacin” Sulphonamides and antibiotics block PABA incorporation by competitive inhibition

3. Structure Folic acid exists in a polyglutamate form. Intestinal mucosal cells remove some of the glutamate residues through the action of the lysosomal enzyme, conjugase.

4. Active functional form is: Tetrahydrafolic acid (THF). Humans and microorganisms: Folic acid is reduced within cells (principally in the liver where it is stored) to tetrahydrofolate (THF or H4folate) through the action of folate reductase [or dihydrofolate reductase (DHFR) ] which is an NADPH-requiring enzyme.

5. Active center (N5 and N10)

6. Active center of tetrahydrofolate (THF). The N5 position is the site of attachment of methyl  and  formimino groups, the N10 the site for attachment of formyl  group and that both N5 and N10 bridge the methylene and methenyl groups.

7. Folate conversions • Carrier of one-carbon (e.g. methyl) groups • The one-carbon units are added to, or removed from, metabolites such as: • histidine • serine • methionine • purines.

8. Functions • Role of N5,N10-methylene-THF in dTMP synthesis is the most metabolically significant function for this vitamin. • Vitamin B12 and N5-methyl-THF in the conversion of homocysteine to methionine is important in helping cells to regenerate needed THF.

9. Participation of H4folate in dTMP synthesis ______Deoxyuridine______________________Deoxythymidine____Monophosphate (dUMP)_______________Monophosphate (dTMP)_______

10. Deficiency symptoms • Deficiency is rare due to the adequate presence of folate in food. • Deficiency can occur when there is: • Poor dietary habits as those of chronic alcoholics. • Impaired absorption or metabolism • Certain drugs such as anticonvulsants and oral contraceptives and methotrxate can impair the absorption of folate. • An increased demand for the vitamin. • Pregnancy • Folate will nearly double by the third trimester of pregnancy. • Identical to those for vitamin B12 deficiency: • Effect of folate deficiency on cellular processes is upon DNA synthesis. • Impairment in dTMP synthesis and purine synthesis • Cell cycle arrest in S-phase of rapidly proliferating cells, especially hematopoietic cells. • The result is megaloblastic leukemia as for vitamin B12 deficiency. • The inability to synthesize DNA during erythrocyte maturation leads to abnormally large erythrocytes termed macrocytic anemia. • Deficiency during pregnancy can cause neural tube defects such as spina bifidia.

11. VITAMIN B12 (cobalamin) • Vitamin B12, is also called cobalamin, cyanocobalamin and hydroxycobalamin. • It is built from : • A nucleotide and • A complex tetrapyrrol ring structure (corrin ring) • A cobalt ion in the center. • A R- group • When R is cyanide (CN), vitamin B12 takes the form of cyanocobalamin. • In hydroxycobalamin, R equals the hydroxyl group (-OH). • In the coenzyme forms of vitamin B12, • R equals an adenosyl group in adenosylcobalamin. • R equals a methyl (-CH3) group in methylcobalamin. • Vitamin B12 is synthesized exclusively by microorganisms (bacteria, fungi and algae) and not by animals and is found in the liver of animals bound to protein as methycobalamin or 5'-deoxyadenosylcobalamin.

12. Known as the "red" vitamin because it exists as a dark red crystalline compound, Vitamin B12 is unique in that it is the only vitamin to contain cobalt (Co3+) metal ion, which, gives it the red color. • The vitamin must be hydrolyzed from protein in order to be active. • Intrinsic factor, a protein secreted by parietal cells of the stomach, carries it to the ileum where it is absorbed. • It is transported to the liver and other tissues in the blood bound to transcobalamin II. • It is stored in the liver attached to transcobalamin I. • It is released into the cell as Hydroxocobalamin (see the next slide) • In the cytosol it is converted to methylcobalamin. • Or it can enter mitochondria and be converted to 5’-deoxyadenosyl cobalamin. Dorothy Crowfoot Hodgkin (1910-1994) Dr. Stadtman in her lab

13. In the cytosol In mitochondria

14. Functions • Only two reactions in the body require vitamin B12 as a cofactor: • During the catabolism of fatty acids with an odd number of carbon atoms and the amino acids valine, isoleucine and threonine the resultantpropionyl-CoA is converted to succinyl-CoA for oxidation in the TCA cycle. • methylmalonyl-CoA mutase, requires vitamin B12 as a cofactor in the conversion of methylmalonyl-CoA to succinyl-CoA. • 5'-deoxyadenosine derivative of cobalamin is required for this reaction • The second reaction catalyzed by methionine synthase converts homocysteine to methionine • This reaction results in the transfer of the methyl group from N5-methyltetrahydrofolate to hydroxycobalamin generating tetrahydrofolate and methylcobalamin during the process of the conversion.

15. Methionine and Folate cycles are interrelated Methyl acceptor Methyl acceptor Methionine cycle Folate cycle Methionine SAM THF MS methyl transferases CH2-THF B12 CH3-THF CH3- Homocysteine SAH CBS B6 Transulfuration pathway cystathionine B6 cysteine

16. Deficiency symptoms • Pernicious anemia in humans (inability to absorb B12 because of lack of gastric intrinsic factor). • Neurological disorders due to progressive demyelination of nerve cells. • This results from increase in methylmalonyl-CoA. • Methylmalonyl-CoA is a competitive inhibitor of malonyl-CoA in fatty acid biosynthesis. • Can substitute malonyl-CoA in any fatty acid biosynthesis and create branched-chain fatty acid altering the architecture of normal membrane structure of nerve cells. • Sources • Synthesized only by microorganisms, so traces only are present in plants; liver is a rich source. • B12 is found in organ and muscle meats, fish, shellfish, dairy products, eggs and in fortified foods like breakfast cereals.

17. Vitamin C (Chemical nature) • It is derived from glucose via uronic acid pathway. Enzyme • L-gluconolactone oxidase is responsible for conversion of gluconolactone to ascorbic acid. • This enzyme is absent in primates, including humans, some bats…. • The active form is ascorbic acid itself.

18. VITAMINC • Vitamin C is L-ascorbic acid, which is a colorless, crystalline acid with strong reducing properties. • Functions • Vitamin C has antioxidant properties similar to those of vitamin E, • Protects cells from free radicals. • Protects iron from oxidative damage, thus enhancing iron (Fe2+) absorption in the gut. • The main function is as a reducing agent. • It has the potential to reduce cytochrome a and c of the respiratory chain and molecular oxygen and nitrates. • It is required for various hydroxylation reactions e.g. proline to hydroxypoline for collagen synthesis (see next slide).

19. Hydroxylation of proline and lysine residues in collagen • Vitamin C is required for the maintenance of normal connective tissue as well as for wound healing because synthesis of connective tissue is the first event in wound tissue remodeling.

20. Other activities • Several other metabolic reactions require vitamin C as a cofactor: • The catabolism of tyrosine and the synthesis of epinephrine from tyrosine and the synthesis of the bile acids. • It is also believed that vitamin C is involved in the process of steroidogenesis. • The adrenal cortex contains high levels of vitamin C which are depleted upon adrenocorticotropic hormone (ACTH) stimulation of the gland.

21. Roles in the body Sources • Citrus fruits and green leafy vegetables • Vitamin C is readily absorbed and so the primary cause of vitamin C deficiency is poor diet and/or an increased requirement. Deficiency symptoms 1. Scurvy • Bleeding gums • Small red spots on skin • Rough skin • Wounds fail to heal • Weak bones and teeth • Anemia and infections 2. Stress (e.g., infections, smoking) • Mechanism unknown, but vitamin C requirements increase during stress 3. Common cold? 4. Disease prevention? • Cancer, heart disease

22. Periodontal disease

23. Vitamin B-complex

24. 8. Inositol (Vitamin Bh) • Inositol is part of the vitamin B-complex. • Since it is not essential in the human diet, it cannot be considered a vitamin. • Inositol is naturally present in foods high in fiber (wheat, legumes, bran, etc). • It is required for: • For proper formation of cell membranes. • For formation of lecithin • It is not a vitamin per se, but it works synergistically with many vitamins and nutrients. • functions closely with a B complex vitamins, Choline, folacin, Vitamins B-6 and B-12. • Inositol is necessary for proper function of nerves, brain, and muscles in the body. • This nutrient is also often used to counter depression; indeed, serotonin needs inositol for proper functioning. • Exists in all human cells, where it plays an important role in cell proliferation and differentiation. • Low levels of this nutrient may result in depression and some research has shown that increased levels of Inositol appear to be a promising treatment for depression.

25. 10. P-amino benzoic acid (Bx) • A substance required for the synthesis of folic acid by many organisms. • PABA is an essential nutrient for some bacteria and is sometimes called Vitamin BX. • Para-aminobenzoicacid is included in the B-vitamin complex, although it is not an essential nutrient for humans and it varies in its activity from other B vitamins. • Although humans lack the ability to synthesize folate from PABA, it is sometimes marketed as an essential nutrient under the premise that it can stimulate intestinal bacteria. • It also absorbs ultraviolet light and is used as aminobenzoic acid, as a topical sunscreen. Abbreviated PAB or PABA. • The potassium salt is used as drug against fibrotic skin disorders.

26. Structure Folic acid PABA (vitamin Bx)

27. 11. L-CARNITINE (B11) • L-carnitine • Is made in the body from the amino acids lysine and methionine, • Is needed to release energy from fat. • It transports fatty acids into mitochondria, the powerhouses of cells. • In infancy, and in situations of high energy needs, such as pregnancy and breast-feeding, the need for L-carnitine can exceed production by the body. • Therefore, L-carnitine is considered a "conditionally essential" nutrient.

28. L-carnitine L-carnitine’s actions appear to be particularly important in the heart. As an example, patients with diabetes and high blood pressure were given 4 grams of L-carnitine per day in a preliminary study. After 45 weeks, irregular heartbeat and abnormal heart functioning decreased significantly compared with nonsupplemented patients.

29. Sources of L-carnitine • Dairy and red meat contain the greatest amounts of carnitine. Therefore, people who have a limited intake of meat and dairy products tend to have lower L-carnitine intakes. • Carnitine deficiencies are rare, even in strict vegetarians, because the body produces carnitine relatively easily. • Rare genetic diseases can cause a carnitine deficiency. • Also, deficiencies are occasionally associated with other diseases, such as diabetes and cirrhosis. • Most people do not need carnitine supplements. For therapeutic use, typical amounts are 1–3 grams per day.

30. Fat Soluble Vitamins

31. Vitamin classification

32. Fat-soluble Vitamins • Absorbed with dietary fat in small intestine • 40-90% absorption efficiency • Absorption typically regulated by need • need absorption • Transported away from small intestine in chylomicra via blood and lymph (depending on size) • They are not easily excreted through urine. • Substantial amounts are stored in liver and fat tissue. • Therefore, toxicity is possible.

33. Vitamin A Compounds with 20-carbon structure. Contain a methyl substituted cyclohexenyl ring (-ionone ring), and an isoprenoid side chain with either a hydroxyl group, and aldehyde group, a carboxylic acid group, or an ester group (retinyl ester) at the terminal C15. All-trans-retinal 11-cis-retinal Retinol Retinoic Acid Can’t be reduced to retinol or retinal in the body.

34. Vitamin A • Active vitamin A- Preformed vitamin A can be obtained either directly from foods that are substantial in vitamin A (beef liver, fish liver oils, egg yolks and butter) • The active form of vitamin is retinol, an alcohol which can be converted to other forms (e.g. vitamin A esters) for storage in liver and tissues. • much the body's vitamin A is stored in the liver as retinyl palmitate • Provitamin A-provitamins, substances that are transformed into vitamins in the body • Beta-carotene is the most abundant and widespread provitamin A. • Beta-carotene comes from a group of compounds called the "carotenoids (C40 polyisoprenoids). • One need to eat approximately six times as much beta-carotene to get the same amount of vitamin A as in retinol. • Dark-green leafy vegetables (spinach) and yellow-orange fruits (apricots and mango) and vegetables (carrots, yellow squash and sweet potatoes) are high in beta-carotene and other carotenoids (-carotene, -cryptoxathin, etc.) • Carotenoids are not toxic even at high doses for long times.

35. Vitamin A: Absorption and transport • Micelle formation • Inside mucosal cell -carotene is cleaved in the lumen of the intestine by -carotene dioxygenase to yield retinal. • Retinal is reduced to retinol by retinal reductase, an NADPH requiring enzyme within the intestines. • Retinol is esterified to palmitic acid and delivered to the liver with chylomicrons via lymphatic system. • Carotenoids which escape intestinal conversion to retinol can be stored in adipose tissue. • Species in which this occurs include humans, horses and cattle, which have yellow-pigmented body fat when carotenoid intake is high. • Vitamin A absorption efficiency 70-90%; carotenoids 9-22%.

36. Vitamin A: Biological functions • Role in vision. Vitamin A (retinal) is an essential precursor for formation of the visual pigment, rhodopsin, in the retina of the eye. Retinal plays an important role in vision, especially night vision. Lowers cataract incidence. • Role in growth. Helps regulate cell development, cell differentiation and cell division. • Role in Bone and teeth formation. Promotes the proper growth of bones and teeth. Bone cells (osteoblasts and osteoclasts) depend on vitamin A for their normal functioning. • Is important in the formation and maintenance of healthy hair, skin and mucous membranes. • Role in reproduction. Vitamin A holds an important place in sexual reproduction. Adequate levels of vitamin A are needed for normal sperm production. The female reproductive cycle requires sufficient amounts of vitamin A. • Boosts the body's immune system helping to increase body resistance to infectious diseases.

37. Role of Vitamin A in Vision(Wald’s visual cycle) • 1. When the rhodopsin is exposed to light it is bleached releasing the 11-cis-retinal from opsin. • 2. Isomerization of the cis-isomer of retinal to all-trans-retinal, causes conformational changes in rhodopsin, hyperpolarization of the retinal rod cell, and extremely rapid transmission of electrical activity to the brain via the optic nerve • 3. Trans-retinal is isomerized to cis-retinal in the dark, which associates with opsin to regenerate rhodopsin. All trans retinol = main circulating form of Vit A Visual Pigment

38. Vitamin A: Biological functions (Cell growth and differentiation) • Regulation of gene expression by retinoic acid, the acid form of vitamin A • Retinoic acid is formed within the target cell from retinol transported in the bloodstream. • Retinoic acid, combined with protein receptor, acts as transcription regulator by binding to specific sequences of DNA in the nucleus • Affected cells include epithelium of numerous tissues, including skin and gut, and bone, ovary and testis. • Retinoic acid is an especially important regulator of embryonic cell differentiation.

39. Vitamin A: Deficiency symptoms 1. Night blindness" - lessened ability to see in dim light. 2. Increased susceptibility to infection and cancer and anemia equivalent to iron deficient. 3. Prolonged lack of vitamin A (keratinization of the cornea, a condition known as xerophthalmia). 4. Abnormal bone development in fetal and neonatal life. 5. Various congenital defects. Retinol and its precursors are used as dietary supplements to prevent the above symptoms. Retinoic acid is used in dermatology to cure acne and psoriasis: 1. Mild acne (Darier’s disease) and aging Topical application of tretinoin (all-trans retinoic acid), along with benzoyl peroxide and antibiotics. 2. severe recalcitrant cystic acne Oral consumption of isotretinoin (13-cis retinoic acid). This drug is teratogenic. Should not be prescribed during pregnancy. It may lead to hyperlipidemia and increased LDL/HDL ratio (increased chance of heart disease) in long term.

40. Vitamin A: Toxicity • Hypervitaminosis A in humans • Symptoms include dry, pruritic skin with excessive itching and increase in intracranial pressure. • is becoming an increasing problem in Western countries because of self-medication and overprescription. • has occurred through eating polar bear or seal liver by polar explorers. • Eat polar bear liver sparingly, 30 grams contain 450,000 IU of retinol! Continued ingestion causes peeling of the skin from head to foot. • It is possible that some early Arctic explorers died from eating their sled dogs in an attempt to survive.

42. Vitamin D • Vitamin D is a family of fat-soluble sterol compounds: • Vitamin D1 (lamisterol) • Vitamin D2 (ergocalciferol) • Vitamin D3 (cholecalciferol) • Vitamin D4 (dihydroergocalciferol) • Vitamin D5 (7-dehydrositosterol)

43. Vitamin D: Types and Sources • For humans, the two most important forms of vitamin D are: • vitamin D2 • vitamin D3. • Vitamin D2 (ergocalciferol) is derived from plants and irradiated yeast and fungi. • Vitamin D3 is synthesized in the body when skin is exposed to sunlight • Cholesterol + sunshine = Vitamin D3 • “sunshine vitamin” – UV-B rays (5-10 minutes arms and legs, mid-day sun). • Vitamin D3 can be obtained from foods like milk, fortified cereals, tuna, salmon and fish oils.

44. Sunlight Vitamin D2 (Ergocalciferol) Ergosterol (in plants) Diet Sunlight Vitamin D3 Cholecalciferol) 7-Dehydrocholesterol

45. Activation of Vitamin D • Vitamin D2 and vitamin D3 are biologically inactive but can have equal biological activity: • Both can be converted first to calcifediol in the liver and then to calcitriol, also known as1,25-dihydroxycholecalciferol, in the kidneys. • Calcitriol, which is the most active form of vitamin D3, is then transported via a carrier protein to the various sites in the body where it is needed. Calcitriol is also called 1,25-dihydroxy vitamin D3, or (1,25-(OH)2D3.

46. 1,25-dihydroxyvitamin D3 Conversion of 25-(OH)D3 to its biologically active form, calcitriol, occurs through the activity of a specific D3-1-hydroxylase present in the proximal convoluted tubules of the kidneys, and in bone and placenta.Cytochrome P450, O2 and NADPH are needed. 25-hydroxyvitamin D3 In the liver cholecalciferol is hydroxylated at the 25 position by a specific D3-25-hydroxylase generating 25-hydroxy-D3 [25-(OH)D3] which is the major circulating form of vitamin D.

47. Functions of vitamin D • Vitamin D regulates the calcium and phosphorus levels in the blood by promoting their absorption from food in the intestines, and by promoting re-absorption of calcium in the kidneys. • It promotes bone formation and mineralization and is essential in the development of an intact and strong skeleton. • It inhibits parathyroid hormone secretion from the parathyroid gland. • Vitamin D affects the immune system by promoting immunosupression, phagocytosis, and anti-tumor activity.

48. Vitamin D: Hormone for Calcium and Phosphate regulation • Nerves and muscles must function properly; calcium is vital for nerve cell transmissions and muscle fiber contractions. • Calcitriol functions in concert with parathyroid hormone (PTH) and calcitonin to regulate serum calcium and phosphorous levels by: • Increasing dietary calcium absorption from the small intestine. • Decreasing the urinary calcium excretion (increasing renal reabsorption). • Stimulating resorption of calcium from bone • Instrumental in the growth, hardening and repair of bones. • Too much vitamin D, however, can increase calcium losses from bone • Vitamin D is essential for normal insulin secretion by the pancreas and therefore control of blood sugar levels.

49. When vitamin D goes abnormal • Too little (northern latitudes) leads to Vit D deficiency: • Rickets (bone deformities in children) • Osteoporosis • Osteomalacia (weak bones) • Demineralization of bones • Too much (5 times the RDA, chronically): It is most toxic of the vitamins. • Nausea, thirst, loss of appetite, stupor • Hypercalcemia: calcium gets deposited in soft tissues, arteries and kidneys.

50. Diseases caused by deficiency • Vitamin D malnutrition may also be linked to an increased susceptibility to several chronic diseases such as • High blood pressure • tuberculosis • cancer • periodontal disease • multiple sclerosis • chronic pain • depression • schizophrenia • several autoimmune diseases • The American Pediatric Associations advises vitamin D supplementation of 200 IU/day (5μg/d) from birth onwards. • Liquid "drops" of vitamin D for infants.