Vitamins and Nutrition

Water-Soluble Vitamins

Vitamins are essential factors, not synthesized in the body, but required in the diet because they are needed mostly for the formation of coenzymes. Water-soluble vitamins will be discussed first. The major water-soluble vitamins are as follows: thiamine (vitamin B1, aneurine), riboflavin (vitamin B2), niacin (vitamin B3), pantothenic acid (vitamin B5), pyridoxine, pyridoxal and pyridoxamine (three forms of vitamin B6), biotin (vitamin H), folic acid (vitamin B9), cobalamin (vitamin B12), and ascorbic acid (vitamin C).

Riboflavin (Vitamin B2)

The coenzyme forms of riboflavin are flavin mononucleotide (FMN) and flavin adenine dinucleotide ( Fig. 20.4 ).

FMN is formed first by the action of riboflavin kinase on riboflavin. FMN is then converted to FAD by the action of FAD pyrophosphorylase with ATP. These reactions are shown in Fig. 20.5 .

Many flavoproteins contain Mg 2 ⁺ or other metals. Examples of such enzymes are succinate dehydrogenase and xanthine dehydrogenase catalyzing reactions shown in Fig. 20.6 .

Succinate dehydrogenase has four domains: (1) located in the mitochondrial matrix containing the binding site for succinate and is the location of the reduction to fumarate and also where FAD is converted to FADH2 (histidine is the covalent link between the flavin and the peptide chain); (2) also located in the mitochondrial matrix; (3) located in the inner mitochondrial membrane where heme is located and where ubiquinone is reduced; and (4) also in the inner mitochondrial membrane adjacent to C containing heme and the reduction of ubiquinone. Succinate dehydrogenase constitutes Complex II of the mitochondrial respiratory transport chain.

Riboflavin is degraded when exposed to light, generating lumichrome as the product as shown in Fig. 20.7 .

Riboflavin can become deficient in newborns treated with phototherapy for hyperbilirubinemia . In general, riboflavin deficiency causes aversion to light ( photophobia ), inflammation of the mouth, face, and tongue ( glossitis ), excessive oiliness of face and scalp ( seborrhea ), and angular stomatitis (fissures and inflammation of the lower lip). Recommended daily intakes for persons of various ages in milligram are as follows: babies (birth–6 months), 0.3 mg; infants (7–12 months), 0.4 mg; children (1–3 years), 0.5 mg; children (4–8 years), 0.6 mg; children (9–13 years), 0.9 mg; boys (14–18 years), 1.3 mg; girls (14–18 years), 1.0 mg; men (19 years plus), 1.3 mg; women (19 years plus), 1.1 mg; pregnant females, 1.4 mg; breastfeeding females, 1.6 mg (data from http://umm.edu/health/medical/altmed/supplement/vitamin-b2-riboflavin ). If riboflavin is given as a supplement, the best absorption is between meals.

Niacin (Vitamin B3)

There are two vitamin forms of niacin: nicotinic acid and nicotinamide ( Fig. 20.8 ).

Nicotinamide exists as two coenzyme forms: NAD ⁺ , NADH and NADP ⁺ , NADPH , structures of which are shown in Fig. 20.9 .

The formation of the coenzyme forms of nicotinic acid and nicotinamide is shown in Fig. 20.10 .

There are many enzymes that have NAD ⁺ or NADP ⁺ as their coenzymes, such as lactate dehydrogenase (LDH) and malate dehydrogenase. LDH is a good example. It catalyzes the freely reversible reaction:

as shown in Fig. 20.11 .

In Fig. 20.12 the catalytic center of LDH is pictured. In the enzymatic reaction the –CH ₃ group of pyruvate is replaced by –NH ₂ to form an oxamate (in the reaction proceeding from the left to the right in Fig. 20.11 ). The hydride transfer from NADH takes place on the C ₂ of pyruvate, and a hydrogen transfer occurs from His 195 of the enzyme protein to the pyruvate C ₂ oxygen to generate lactate and NAD ⁺ .

A small amount of niacin can be synthesized from tryptophan , but it requires about 60 mg of tryptophan to generate 1 mg of niacin (particularly in the form of NAD) through this highly inefficient process, and the niacin generated through this mechanism ( Fig. 20.13 ) cannot satisfy the body’s requirements (about 15 mg/day), therefore, sufficient niacin must be obtained in the diet.

Deficiency of niacin in the human ( pellagra ) results in the inflammation of the tongue (glossitis), dermatitis, and diarrhea. Niacin deficiency can be the result of the malabsorption of tryptophan in the intestine and kidneys ( Hartnup disease ). Hartnup disease is an autosomal recessive disorder deriving from mutations in the gene for the Na ⁺ -dependent, Cl ⁻ -independent neutral amino acid transporter principally in the apical brush border membrane of the small intestine and the proximal tubule of the kidney. Hartnup disease has the symptoms of niacin deficiency plus cerebellar ataxia and aminoaciduria. The incidence of this disorder in New South Wales is reported to be 1 in 33,000. In malignant carcinoid syndrome (carcinoid tumors arise from neuroendocrine cells), there is the synthesis of excess serotonin and niacin deficiency can be a factor. High incidence of carcinoid tumors occurs in the gastrointestinal tract, including the small bowel and the appendix. The symptoms include flushing of the face, severe diarrhea, and asthma attacks.

Pantothenic Acid (Vitamin B5, Pantothenate)

Pantothenic acid is synthesized in microorganisms from β-alanine and pantoic acid . Pantothenic acid is a component of CoA and also the acyl carrier protein ( ACP ) of fatty acid synthase (see Chapter 9 : Lipids, on lipids for many reactions involving CoA). A great many enzymes (at least 70) require CoA or ACP rendering pantothenate critical in the metabolism of carbohydrates, fats, and proteins and the functioning of the citric acid cycle. The synthesis of CoA starting with the vitamin, pantothenate (itself synthesized in microorganisms from β-alanine and pantoic acid), is shown in Fig. 20.14 .

Adequate intake of pantothenate is 1.7 mg/day in infants to 5 mg/day in adults. Pantothenate is easily obtained in a normal diet. Good sources are avocado, yogurt, cooked chicken, sweet potatoes, cereals, meats and legumes, and many others containing lesser amounts. Pantothenic acid deficiency in humans is rare.

Pyridoxine, Pyridoxal, and Pyridoxamine (Vitamin B6)

The three forms of the vitamin in the diet are convertible in the body to the coenzyme form, pyridoxal-5′-phosphate ( PLP ) as shown in Fig. 20.15 .

The kinase for the three vitamin forms phosphorylates them (using ATP) to their respective 5′-phosphate derivatives. Pyridoxamine phosphate oxidase converts pyridoxine-5′-phosphate to the coenzyme form, PLP. Pyridoxamine-5′-phosphate is converted by an oxidase and an aminotransferase to PLP . The transaminase mechanism in which an amino acid is bound to the enzyme and forms an aldimine ( Schiff base ) that is converted to an enzyme ketimine; releases the keto acid product as the coenzyme is converted to pyridoxamine phosphate that becomes the enzyme ketimine, then the aldimine, and finally the original form that binds a second amino acid molecule to start the reaction cycle again (see Fig. 13.6 ).

The reaction between enzyme and pyridoxal phosphate forming the Schiff base aldimine is shown in Fig. 20.16 . This figure also shows the attachment of pyridoxal to pyridoxal kinase (PDXK) through lysine and aspartate residues of the enzyme.

PLP is a coenzyme for a great many enzymes, including aminotransferases, amino acid racemases, amino acid decarboxylases, and others, including glycogen phosphorylase.

The daily requirement for vitamin B6 in adults is in the range of 1.4–2.0 mg/day. A slightly higher amount is required by pregnant or lactating women.

Vitamin B6 may have therapeutic usage. Experimentally, vitamin B6 inhibits the action of the glucocorticoid receptor in vitro in cultured cells and in vivo in animals. It is also known that the glucocorticoid receptor is essential for life. Many cancer cells, as well as normal cells, require PDXK for growth. Hepatoma cells in culture were shown to be killed by pyridoxal hydrochloride . Transformed cells that are susceptible to killing by vitamin B6 contain functional PDXK. Transformed cells, like certain human mammary tumor cells, may not contain active PDXK and, therefore, are not killed by vitamin B6. It is likely that the killing of transformed cells by vitamin B6 is the result of the conversion of large amounts of the vitamin to pyridoxal phosphate, thus reducing the pool of ATP so that these cells may die because of the inhibition of the glucocorticoid receptor plus the reduction of the cellular pool of ATP. Skin tumors, such as granulomas or melanomas , are good candidates because pyridoxal HCl or pyridoxine HCl can be applied directly, even as a powder. Although normal cells contain PDXK, they are not killed by vitamin B6, perhaps because they take up smaller amounts of the vitamin. This hypothesis is presented in Fig. 20.17 .

Biotin (Vitamin H)

Biotin is a cofactor for carboxylase enzymes. There are five human enzymes for which biotin is a coenzyme: acetyl-CoA carboxylase I (soluble cytoplasm), acetyl-CoA carboxylase II (mitochondrial fatty acid oxidation), pyruvate carboxylase , methylcrotonyl-CoA carboxylase , and propionyl-CoA carboxylase . Acetyl-CoA carboxylase II catalyzes the reaction:

a controlling reaction in the synthesis of fatty acids. Acetyl-CoA carboxylase operates in the synthesis of lipids from acetate. Pyruvate carboxylase is involved in liver gluconeogenesis, particularly from amino acids. Methylcrotonyl-CoA carboxylase is involved in the catabolism of the essential amino acid, leucine. Proprionyl-CoA carboxylase plays a role in the metabolism of certain amino acids, cholesterol, and odd-chain fatty acids.

Taking pyruvate carboxylase as an example of this class of enzymes, it catalyzes the reaction:

shown in Fig. 20.18 .

The biotin coenzyme reacts with the enzyme in the first part of the reaction; then it swivels over to the second part of the reaction. Biotin is the carrier of carbon dioxide, and biotin is linked to the enzyme via an ε-amino group of lysine ( Fig. 20.19 ).

When carboxylase holoenzymes are degraded, the biotin is cleaved from the ε-amino group of the active site of the enzyme by another enzyme, biotinidase , causing the release of free biotin. The residue of biotin still bound to the enzyme ( Fig. 20.19 , middle ) is called biocytin . The active site of pyruvate carboxylase is pictured diagrammatically in Fig. 20.20 .

Biotin also can be bound to histones, and the biotinylated histones play a role in the regulation of DNA synthesis, transcription, and cell proliferation. The free biotin released by biotinidase from carboxylases or biotinylated histones is available for binding to other apocarboxylases as shown for the “ biotin bicycle ” in Fig. 20.21 .

Biotin binds to other proteins and avidin , in particular, found in egg white is a strong binder (cooking inactivates the interaction). Other biotin-binding proteins are strepavidin , homocitrate synthase , and isopropylmalate synthase . Good sources of dietary biotin are liver, soy flour, egg yolk, cereal, and yeast. Biotin deficiency is rare since intestinal bacteria synthesize biotin. In general, normal biotin intake varies from 5 μg/day in the infant to 30 μg/day in the adult.

Cobalamin (Vitamin B12)

Vitamin B 12 has a complex ring structure (tetrapyrrole) and an atom of cobalt in the center as shown in Fig. 20.22 .

Vitamin B12 is synthesized by microorganisms. In ingested meat the vitamin is bound to proteins and is released from the bound form by hydrolysis by stomach acid or by the action of trypsin in the intestine. A protein secreted by parietal cells of the stomach is the intrinsic factor that binds cobalamin . The cobalamin–intrinsic factor complex is absorbed in the ileum after attaching to cubilin , a receptor for the cobalamin–intrinsic factor complex. Cubilin facilitates the absorption of the complex. Mutation of the gene for cubilin results in impaired absorption of the cobalamin–intrinsic factor complex generating the rare megaloblastic anemia (formation of red blood cells requires vitamin B 12 , folic acid, and iron; a deficiency in any one of these causes anemia). Pernicious anemia , a form of megaloblastic anemia, is an autoimmune disorder in which parietal cells of the stomach are destroyed (by autoantibodies) so that there is insufficient intrinsic factor available to facilitate the absorption of vitamin B 12 . Normally, vitamin B 12 is bound to the transporting protein, transcobalamin II , in the intestinal cells, and the vitamin B 12 –transcobalamin II complex is carried in the bloodstream to tissue cells and to the liver where it is stored as a complex of vitamin B12–transcobalamin III (transcobalamin III is also present in blood). In the liver the stored form of vitamin B 12 can be released into the small intestine through the bile; then vitamin B 12 is released from the complex, becomes bound to intrinsic factor and cycles again through the intrinsic factor–vitamin B 12 –cubilin complex, and eventually reaches the bloodstream bound to the transporting protein.

The 5′-deoxyadenosine derivative ( Fig. 20.23 ) of cobalamin is the coenzyme for methylmalonyl-CoA mutase that catalyzes the conversion of methylmalonyl-CoA to succinyl-CoA in the system catabolizing fatty acids.

This form of cobalamin is involved in methionine synthase that catalyzes the conversion of homocysteine to methionine. In this reaction a methyl group is transferred from N 5 -methyltetrahydrofolate to hydroxycobalamin. When cobalamin is deficient because the intrinsic factor is absent or inactive, folic acid becomes trapped as the N 5 -methyltetrahydrofolate due to the inactivity of methionine synthase. Also, other tetrahydrofolate (THF) derivatives cannot be formed, and these are required for the synthesis of purines and the pyrimidine, thymidine . Additionally, there is an increase in methylmalonyl-CoA resulting in the inhibition of fatty acid synthesis essential for the turnover of the myelin sheath (of neurons), and this results in progressive demyelination .

Folic Acid (Vitamin B9, Pteroylglutamic Acid)

The structure of folic acid contains 6-methylpteridine linked to para-aminobenzoic acid ( PABA ) linked to a glutamate residue as shown in Fig. 20.24 . The human cannot synthesize PABA.

Folic acid in the diet (folic acid is synthesized in plants and yeasts) can occur as mixtures of polyglutamates containing as many as seven glutamic acid residues ( Fig. 20.25 ).

The polyglutamates are hydrolyzed by peptidases in the intestine to the natural form of folate with a single or a reduced number of glutamate residues, and these glutamate-degraded forms are recognized by the folic acid receptor ( RFC1 transporter ) in the intestine. The RFC1 transporter is structurally related to the thiamine transporter , since both derive from the same gene family; the gene-encoding information for the folate transporter (RFC1) is SLC19A1 , while the gene-encoding information for the thiamine transporter is SLC19A2 . Removal of conjugated glutamate residues reduces the negative charges on the folate molecule making it more easily passed through the basolateral membrane of the intestinal cell destined for the bloodstream. There are other receptors in some tissue cells, such as a proton-coupled folate transporter ( PCFT ) and folate receptor 1 ( FOLR1 ). PCFT is an intestinal PCFT that also transports heme from the gut lumen into duodenal epithelial cells. Subsequently, iron is released from heme and moved into the bloodstream. FOLR1 binds folic acid and its reduced derivatives and is the transporter for 5-methyltetrahydrofolate into cells.

In the hepatocyte, folic acid is reduced to THF by dihydrofolate reductase . During biosynthetic reactions, THF derivatives transfer one-carbon units (methyl, methylene, formyl, or formimino groups; see Fig. 20.26 ).

There are one-carbon transfer reactions in the biosyntheses of serine, methionine, glycine, choline, purine nucleotides, and deoxythymidylate monophosphate (dTMP). The role of N5,N10-methylene-THF in the regeneration of dTMP may be the most important role of folate . The formation of methionine from homocysteine catalyzed by methionine synthase also involves cobalamin with N5,N10-methylene-THF . Methionine synthase catalyzes the conversion of homocysteine to methionine where methyltetrahydrofolate is the coenzyme. Fig. 20.27 shows the active center of the enzyme and how the methyltetrahydrofolate is positioned in the Fol barrel of the enzyme structure.

Folate deficiency (often occurring in alcoholics with inadequate diets) diminishes dTMP synthesis that can lead to cell cycle arrest in the S-phase of rapidly dividing hemopoietic cells. Folate deficiency also can occur when there is inadequate absorption of the vitamin. Anticoagulants and oral contraceptives interfere with folate absorption. As mentioned before, folate deficiency can lead to megaloblastic anemia , identical to that induced by the deficiency of vitamin B 12 . The recommended daily intake of folic acid in the adult is 400 μg/day. Fortified cereals, orange juice, spinach, asparagus, lentils, and garbanzo beans are good sources of folic acid.

Ascorbic Acid (Vitamin C)

Ascorbic acid is an important antioxidant that reduces other compounds [e.g., reactive oxygen species (ROS)] and is converted to its oxidized form, dehydroascorbic acid ( DHA ), in the process. Humans are unable to synthesize ascorbic acid, although other species are able to do so. The loss of the ability to synthesize ascorbic acid seems to parallel the inability of humans and certain other species to degrade uric acid (loss of the enzyme uricase through mutation) that is also a strong reducing agent. Inability to synthesize ascorbate is due to a defective pseudogene (ψGULO) that encodes information for the enzyme, l-gulonolactone oxidase , which is the final enzyme required for the synthesis of ascorbate. In the human and in other species that have lost the ability to synthesize ascorbate, there is a compensatory vitamin C recycling mechanism. In this system, after ascorbic acid is used to reduce some oxidized substance and ascorbic acid is, itself, oxidized to DHA can again be reduced back to ascorbic acid, either by an NADH system or by the action of glutathione ( GSH ) as suggested by Fig. 20.28 . This is of particular importance in the red blood cell.

Loss of l -gulonolactone oxidase and uricase through ancient mutations probably benefitted early primates by conferring a survival advantage in maintaining blood pressure when dietary and environmental changes were occurring. However, in modern society having moved to the Western diet and relative physical inactivity, these mutations increase the risk for hypertension and cardiovascular disease.

Ascorbic acid can be imported into cells through an Na ⁺ /ascorbate symporter , although it is more likely imported as the oxidized form (DHA) down its concentration gradient (DHA is more concentrated outside the cell) through facilitative glucose transporters , such as glucose transporter 1 (GLUT 1) . The blood–brain barrier is unable to pass ascorbic acid but does transport DHA by the way of GLUT 1, and the transported DHA is subsequently reduced back to ascorbic acid by GSH.

One of the main functions of ascorbic acid is in the hydroxylation of proline in collagen, where ascorbic acid is a cofactor for the hydroxylation reaction ( Fig. 20.29 ).

This hydroxylation is required for aggregation of the collagen molecule into a triple helix ( Fig. 20.30 ).

Ascorbate , then, is essential for the maintenance of connective tissue and also for wound healing where the synthesis of connective tissue is an early step of the process. Ascorbate is also necessary for bone remodeling because collagen is in the organic matrix of bone. In addition, ascorbate is involved in the synthesis of epinephrine from tyrosine ( Fig. 20.31 ).

Ascorbate is needed in severe stress when the adrenal store is rapidly depleted of ascorbate, and consequently it plays a role in the formation of cortisol. Vitamin C also is involved in signal transduction of nuclear factor kappa B (NF-κB). DHA transported into the cell through a glucose transporter inhibits IκB kinase β (IKKβ). ROS that are intermediates in the activation of NF-κB are quenched by ascorbate; ascorbate becomes oxidized to DHA that inhibits IKKβ and IκB kinase α enzymatic activities. In this way, ascorbate depresses NF-κB signaling ( Fig. 20.32 ).

Ascorbate can be degraded to DHA to diketogulonic acid and then to other acids, such as oxalic acid.

The adult male requires about 90-mg ascorbate/day, and the adult female requires about 75-mg ascorbate/day. Excellent sources of ascorbic acid are as follows: guava, red bell pepper, papaya, and concentrated orange juice.