﻿ Vitamins: Critical Enzyme Co-Factors

Vitamins: Water and Fat Soluble

Minerals: Critical Micronutrients
RDA Values for Vitamins and Minerals
Water Soluble Vitamins Fat Soluble Vitamins
Thiamine (B1)
Thiamine Deficiency and Disease
Riboflavin (B2)
Riboflavin Deficiency and Disease
Niacin (B3)
Niacin Deficiency and Disease
Pantothenic Acid (B5)
Pyridoxal, Pyridoxamine, Pyridoxine (B6)
B6 Deficiency and Disease
Differential Diagnosis: Microcytic Anemia
Clinical Significance of Biotin
Cobalamin (B12)
Cobalamin Deficiency and Disease
Folic Acid
Folate Deficiency and Disease
Differential Diagnosis: Folate versus B12 Deficiency
Ascorbic Acid: Vitamin C
Ascorbic Acid in Iron Homeostasis
Vitamin C Deficiency and Disease
Vitamin A
Gene Control by Vitamin A
Role of Vitamin A in Vision
Clinical Significances of Vitamin A

Vitamin D
Clinical Significances of Vitamin D

Vitamin E
Clinical Significances of Vitamin E

Vitamin K
Clinical Significance of Vitamin K

Alpha-Lipoic Acid, LA

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Introduction to Vitamins and Minerals

Vitamins are organic molecules that function in a wide variety of capacities within the body. The most prominent function of the vitamins is to serve as cofactors (co-enzymes) for enzymatic reactions. The distinguishing feature of the vitamins is that they generally cannot be synthesized by mammalian cells and, therefore, must be supplied in the diet. The vitamins are of two distinct types, water soluble and fat soluble. For more information on the food products that are good sources of the individual vitamins visit the Supplement Science page.












The minerals that are considered of dietary significance are those that are necessary to support biochemical reactions by serving both functional and structural roles as well as those serving as electrolytes. The use of the term dietary mineral is considered archaic since the intent of the term "mineral" is to describe ions not actual minerals. There are both quantity elements required by the body and trace elements. The quantity elements are sodium, magnesium, phosphorous, sulfur, chlorine, potassium and calcium. The essential trace elements are manganese, iron, cobalt, nickel, copper, zinc, selenium, molybdenum, and iodine. Additional trace elements (although not considered essential) are boron, chromium, fluoride, and silicon.

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Thiamine (Thiamin)

Structure of thiamine

Thiamine structure

Thiamine (also written thiamin) is also known as vitamin B1. Thiamine is derived from a substituted pyrimidine and a thiazole which are coupled by a methylene bridge. Thiamine is rapidly converted to its active form, thiamine pyrophosphate, TPP, by the enzyme thiamine pyrophosphokinase 1, TPK1. The TPK1 gene is located on chromosome 7q34–q35 and is composed of 27 exons that generate two alternatively spliced mRNAs encoding isoforms of 243 amino acids (isoform a) and 194 amino acids (isoform b). Expression of the TPK1 gene is highest in the liver and brain.

Structure of thiamine pyrophosphate

Thiamine pyrophosphate

TPP is necessary as a cofactor for three critical dehydrogenases. These enzymes are the pyruvate dehydrogenase complex (PDHc) and 2-oxoglutarate (α-ketoglutarate) dehydrogenase (both of which are associated with the TCA cycle), and branched-chain ketoacid dehydrogenase (BCKD) necessary for metabolism of the branched-chain amino acids, leucine, isoleucine, and valine. These three dehydrogenases also require the co-factors derived from the vitamins lipoic acid, pantothenic acid (CoA), riboflavin, and niacin. For this reason these three dehydrogenases are often referred to as the Tender (thiamine) Loving (lipoic acid) Care (CoA) For (flavin) Nancy (niacin) enzymes. In addition to these three dehydrogenases, TPP is a required co-factor for the transketolase catalyzed reactions of the pentose phosphate pathway. A deficiency in thiamine intake leads to a severely reduced capacity of cells to generate energy and to carry out reductive biosynthetic reactions as well as to synthesize nucleotides because of its role in each of these enzymes.

The dietary requirement for thiamine is proportional to the caloric intake of the diet and ranges from 1.0–1.5 mg/day for normal adults. If the carbohydrate content of the diet is excessive then an increase in  thiamine intake will be required.

The richest sources of vitamin B1 include yeasts and animal liver. Additional sources include whole-grain cereals, rye and whole-wheat flour, navy beans, kidney beans, wheat germ, as well as pork and fish.

Food source

Thiamine content (mg)

Yeast, brewer's, 2 tbls 2.3
Pork chop, lean, 3.5 oz 0.9
Ham, lean, 3.5 oz 0.7
Catfish, 3.5 oz cooked 0.4
Bagel, 2 oz enriched 0.4
Milk, soy, 1 cup 0.4
Beans, baked, 1 cup 0.34
Oatmeal, 1 cup cooked 0.26
Rice, white, cooked, 1 cup 0.26
Green peas, ½ cup cooked 0.23
Potato, one medium baked 0.22
Orange juice, 1 cup 0.20
Black beans, ½ cup cooked 0.21
Navy beans, ½ cup cooked 0.19
Soy nuts, ½ cup 0.20
Cashews, ½ cup 0.15
Peanuts, ½ cup 0.10
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Clinical Significances of Thiamine Deficiency

The earliest symptoms of thiamine deficiency include constipation, appetite suppression, and nausea. Progressive deficiency will lead to mental depression, peripheral neuropathy and fatigue. Chronic thiamine deficiency leads to more severe neurological symptoms including ataxia, mental confusion and loss of eye coordination (nystagmus). A highly diagnostic physical test of thiamine deficiency is vertical nystagmus. Vertical nystagmus is characterized by spontaneous upbeating or downbeating of the eyeball. There are numerous causes or horizontal nystagmus but vertical is only seen due to the CNS damage associated with thiamine deficiency or with phencyclidine (PCP) intoxication. Additional clinical symptoms of prolonged thiamine deficiency are related to cardiovascular and musculature defects.

Dietary thiamine deficiency is known as beri beri, is most often the result of a diet that is carbohydrate rich and thiamine deficient. An additional thiamine deficiency related syndrome is known as Wernicke syndrome which is most often associated with chronic alcohol consumption. This disease is most commonly found in chronic alcoholics due to the fact that alcohol impairs thiamine uptake from the small intestine as well as the fact that these individuals generally have poor dietetic lifestyles. Wernicke syndrome is also referred to as dry beri beri. Prolonged dietary deficiency in thiamine leads to wet beri beri. The wet form of the disease is the result the cardiac involvement in the deficiency. At this stage in the deficiency all four chambers of the heart enlarge due to loss of energy generation and fluid retention resulting in what is called dilated cardiomyopathy. The result of the enlarged chambers is that they can't fill completely resulting in systolic failure. Systole relates to the force associated with cardiac contraction expelling blood to arteries. Blood pumped from the left ventricle enters the aorta and is delivered to the body, whereas blood pumped from the right ventricle is sent to the lungs.

When thiamine deficiency manifests with CNS involvement it is called Korsakoff encephalopathy (or Korsakoff psychosis) and is also commonly referred to as Wernicke-Korsakoff syndrome (WKS). WKS is characterized by acute encephalopathy progressing to chronic impairment of short-term memory. Thiamine supplementation can reverse the symptoms of beri beri and Wernicke syndrome, however, the consequences of severe deficiency (WKS) are irreversible. The confabulation of Korsakoff psychosis is due to destruction of the mammillary bodies in the brain. The mammillary bodies are composed of two small round structures at the underside of the brain that are part of the limbic system, specifically they are part of the Papez circuit. This circuit is also called the hippocampal-mammillo-thalamo-cortical pathway. The consequence of destruction of the mammillary bodies is retrograde amnesia.

Persons afflicted with an inherited form of Wernicke-Korsakoff syndrome appear to have an inborn error of metabolism that is clinically important only when the diet is inadequate in thiamine. These individuals are thought to harbor an abnormality in the enzyme, transketolase. Although a variant transketolase enzyme has been proposed to be associated with Wernicke-Korsakoff syndrome, no mutations have been found in the gene (symbol: TKT) encoding this enzyme when cloned from patients exhibiting the syndrome. It has been speculated that the protein encoded by a transketolase-related gene (transketolase-like 1: TKTL1) may be involved in the inherited propensity for the development of WKS. However, the TKTL1 encoded protein lacks 38 amino acids, compared to the TKT protein, in the TPP-binding region. All TPP-dependent enzymes contain a highly similar TPP-binding domain and its lack in the TKTL1 protein strongly suggests that it is unlikely that TKTL1 is a TPP-dependent protein capable of catalyzing the transketolase reaction. Indeed, recent evidence has confirmed that the TKTL1 protein does not catalyze the transketolase reaction of the PPP. Intense interest in the TKTL1 gene, and its encoded protein, was stimulated because it was shown that the level of TKTL1 expression correlated with poor patient outcomes and metastasis in many solid tumours. In addition, specific inhibition of TKTL1 mRNA has been shown to inhibit cancer cell proliferation in functional studies.

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Structure of riboflavin

Riboflavin structure

Riboflavin is also known as vitamin B2. Riboflavin is the precursor for the coenzymes, flavin mononucleotide (FMN) and flavin adenine dinucleotide (FAD). Synthesis of these two cofactors occurs in a two step process. FMN is synthesized from riboflavin via the ATP-dependent enzyme riboflavin kinase (RFK). RFK introduces a phosphate group onto the terminal hydroxyl of riboflavin. FMN is then converted to FAD via the attachment of AMP (derived from ATP) though the action of FAD pyrophosphorylase which is also known as FMN adenylyltransferase (FMNAT).

The enzymes that require FMN or FAD as cofactors are termed flavoproteins. Several flavoproteins also contain metal ions and are termed metalloflavoproteins. Both classes of enzyme are involved in a wide range of red-ox reactions and includes the same critical thiamine-dependent enzymes described above, the pyruvate dehydrogenase complex (PDHc), 2-oxoglutarate (α-ketoglutarate) dehydrogenase, and branched-chain α-ketoacid dehydrogenase (BCKD). Additional important metabolic regulatory enzymes that require flavin as a co-factor include, succinate dehydrogenase (TCA cycle and complex II of oxidative phosphorylation), glycerol-3-phosphate dehydrogenase (involved in the glycerol phosphate shuttle and triglyceride synthesis), and xanthine oxidase involved in purine nucleotide catabolism. During the course of the enzymatic reactions involving the flavoproteins the reduced forms of FMN and FAD are formed, FMNH2 and FADH2, respectively. The hydrogens of FADH2 are on nitrogens 1 and 5 as indicated in the Figure. The normal daily requirement for riboflavin is 1.2–1.7 mg/day for normal adults.

Structure of FAD

Structure of FAD

Riboflavin is found in dairy products, lean meats, poultry, fish, grains, broccoli, turnip greens, asparagus, spinach, and enriched food products.

Food source

Riboflavin content (mg)

Beef liver, 3.5oz cooked 4.14
Mackerel, 3.5 oz canned 0.54
Pork, loin, 3 oz cooked 0.24
Hamburger, lean, 3.5 oz 0.21
Chicken, dark, 3 oz cooked 0.19
Steamed clams, 3.5 oz 0.43
Yogurt, low-fat, 1 cup 0.37
Egg, cooked 0.25
Cheese, cottage, ½ cup 0.21
Milk, nonfat, 1 cup 0.34
Pasta, 1 cup cooked 0.23
Bagel, plain 0.22
Spinach, ½ cup cooked 0.16
Wheat germ, raw, 2 tbls 0.12
Soy nuts, ½ cup 0.65
Almonds, ½ cup 0.78
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Clinical Significances of Flavin Deficiency

Riboflavin deficiencies are rare in the United States due to the presence of adequate amounts of the vitamin in eggs, milk, meat and cereals. Riboflavin deficiency is often seen in chronic alcoholics due to their poor dietetic habits.

Symptoms associated with riboflavin deficiency include itching and burning eyes, angular stomatitis and cheilosis (cracks and sores in the mouth and lips), bloodshot eyes, glossitis (inflammation of the tongue leading to purplish discoloration), seborrhea (dandruff, flaking skin on scalp and face), trembling, sluggishness, and photophobia (excessive light sensitivity). Riboflavin decomposes when exposed to visible light. This characteristic can lead to riboflavin deficiencies in newborns treated for hyperbilirubinemia by phototherapy requiring dietary supplementation in these infants.

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Structures of nicotinamide and nicotinic acid

Niacin (nicotinic acid and nicotinamide) is also known as vitamin B3. Both nicotinic acid and nicotinamide can serve as the dietary source of vitamin B3. Niacin is required for the synthesis of the active forms of vitamin B3, nicotinamide adenine dinucleotide (NAD+) and nicotinamide adenine dinucleotide phosphate (NADP+). Both NAD+ and NADP+ function as cofactors for numerous dehydrogenases including the same critical thiamine-dependent enzymes described above [the pyruvate dehydrogenase complex (PDHc), 2-oxoglutarate (α-ketoglutarate) dehydrogenase, and branched-chain α-ketoacid dehydrogenase (BCKD)] as well as lactate dehydrogenase and malate dehydrogenase, for example.

Structure of NAD

Structure of NAD+. The –OH phosphorylated in NADP+ is indicated by the red arrow. NADH is shown in the box insert.

Niacin is not a true vitamin in the strictest definition since it can be derived from the amino acid tryptophan. However, the ability to utilize tryptophan for niacin synthesis is inefficient (60 mg of tryptophan are required to synthesize 1 mg of niacin). Also, synthesis of niacin from tryptophan requires vitamins B1, B2 and B6 which would be limiting in themselves on a marginal diet. The recommended daily requirement for niacin is 13–19 niacin equivalents (NE) per day for a normal adult. One NE is equivalent to 1 mg of free niacin).

Niacin is found in liver, meat, peanuts and other nuts, and whole grains. In addition, foods that are rich in protein, with exception of tryptophan-poor grains, can satisfy some of the demand for niacin.

Food source

Niacin content (mg)

Beef liver, 3.5oz cooked 14.4
Chicken, white meat, cooked 13.4
Tuna, canned in water, 3 oz 11.8
Salmon, 3.5 oz cooked 8.0
Ground beef, 3.5 oz cooked 5.3
Peanuts, ½ cup 10.5
Almonds, ½ cup 1.4
Potato, baked with skin 3.3
Mushrooms, raw, ½ cup 1.7
Barley, ½ cup cooked 1.6
Corn, yellow, ½ cup 1.3
Lentils, ½ cup cooked 1.4
Sweet potatoes, ½ cooked 1.2
Carrot, raw, medium 0.7
Peach, raw, medium 0.9
Mango, 1 medium 1.5
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Clinical Significances of Niacin and Nicotinic Acid

A diet deficient in niacin (as well as tryptophan) leads to glossitis of the tongue (inflammation of the tongue leading to purplish discoloration), dermatitis, weight loss, diarrhea, depression and dementia. The severe symptoms, depression, dermatitis and diarrhea (referred to as the "3-D's"), are associated with the condition known as pellagra. Several physiological conditions (e.g. Hartnup disorder and malignant carcinoid syndrome) as well as certain drug therapies (e.g. isoniazid) can lead to niacin deficiency. In Hartnup disorder, tryptophan absorption is impaired and in malignant carcinoid syndrome tryptophan metabolism is altered resulting in excess serotonin synthesis. Isoniazid (the hydrazide derivative of isonicotinic acid) was, at one time, a primary drug for chemotherapeutic treatment of tuberculosis.

Nicotinic acid (but not nicotinamide) when administered in pharmacological doses of 2–4 g/day lowers plasma cholesterol levels and has been shown to be a useful therapeutic for hypercholesterolemia. The major action of nicotinic acid in this capacity is a reduction in fatty acid mobilization from adipose tissue. Although nicotinic acid therapy lowers blood cholesterol it also causes a depletion of glycogen stores and fat reserves in skeletal and cardiac muscle. Additionally, there is an elevation in blood glucose and uric acid production. For these reasons nicotinic acid therapy is not recommended for diabetics or persons who suffer from gout.

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Pantothenic Acid

Structure of pantothenic acid

Pantothenic Acid

Pantothenic acid is also known as vitamin B5. Pantothenic acid is formed from β-alanine and pantoic acid. In the synthesis of coenzyme A from pantothenate there are five reaction steps. Pantothenate is phosphorylated on the hydroxyl group via the action of pantothenate kinase. The reactive sulfhydryl group is added from cysteine via the action of phosphopantothenoylcysteine synthetase. Through three more reactions the molecule is decarboxylated and then the ADP from ATP is added forming the fully functional coenzyme A shown in the Figure below.

Pantothenate is required for synthesis of coenzyme A, CoA and is a component of the acyl carrier protein (ACP) domain of fatty acid synthase. Pantothenate is, therefore, required for the metabolism of carbohydrate via the TCA cycle and all fats and proteins. At least 70 enzymes have been identified as requiring CoA or ACP derivatives for their function including the same critical thiamine-dependent enzymes described above, the pyruvate dehydrogenase complex (PDHc), 2-oxoglutarate (α-ketoglutarate) dehydrogenase, and branched-chain α-ketoacid dehydrogenase (BCKD).

Food source

Vitamin B5 content (mg)

Beef liver, 3.5 oz 5.3
Poultry, dark meat, 3.5 oz 1.3
Poultry, white meat, 3.5 oz 1.0
Salmon, 3.5 oz cooked 1.4
Low fat yogurt, 1 cup 1.5
Milk, nonfat, 1 cup 0.8
Bleu cheese, 1 oz 0.49
Cottage cheese, ½ cup 0.27
Corn, cooked, ½ cup 0.72
Potato, baked, one 0.7
Sweet potato, ½ cup 0.68
Broccoli, boiled, ½ cup 0.4
Wheat germ, raw, ¼ cup 1.2
Mushrooms, cooked, ½ cup 0.84
Peanuts, ½ cup 0.9
Avocado half 1.0
Sunflower seeds, ¼ cup 2.3
Dates, 10 0.65 
Papaya, ½ cup 0.33 
Strawberries, 1 cup 0.25 
Orange juice, 1 cup 0.24 

Deficiency of pantothenic acid is extremely rare due to its widespread distribution in whole grain cereals, legumes and meat. Symptoms specific to pantothenate deficiency are difficult to assess since they are subtle and resemble those of other B vitamin deficiencies. These symptoms include painful and burning feet, skin abnormalities, retarded growth, dizzy spells, digestive disturbances, vomiting, restlessness, stomach stress, and muscle cramps.

Structure of coenzyme A (CoA)

Coenzyme A

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Vitamin B6

Structures of pyridoxine, pyridoxal, and pyridoxamine

Pyridoxal, pyridoxamine and pyridoxine are collectively known as vitamin B6. All three compounds are efficiently converted to the biologically active form of vitamin B6, pyridoxal phosphate (PLP). This conversion is catalyzed by the ATP requiring enzyme, pyridoxal kinase. Pyridoxal kinase requires zinc for full activity thus making it a metaloenzyme.

Structure of pyridoxal phosphate

Pyridoxal Phosphate

Pyridoxal phosphate (PLP) functions as a cofactor in all of the enzymes that carry out the transamination reactions required for the synthesis and catabolism of the amino acids.  PLP is a cofactor for the synthesis of several neurotransmitters including serotonin, the catecholamines dopamine, norepinephrine, and epinephrine (critical enzyme is aromatic L-amino acid decarboxylase, AADC which is encoded by the DDC gene and is more commonly referred to as DOPA decarboxylase), and γ-aminobutyric acid, GABA (critical enzyme is glutamic acid decarboxylase, GAD). PLP is required for the synthesis of heme via the enzyme catalyzing the initial and regulated step in this pathway, δ-aminolevulinic acid synthase (ALAS). PLP is a cofactor for two enzymes involved in methionine and cysteine metabolism, cystathionine β-synthase (CBS) and cystathionase (cystathionine γ-lyase). As indicated above, PLP is also required for the conversion of tryptophan to niacin. PLP is also required for glycogen homeostasis as a cofactor for glycogen phosphorylase. This latter reaction is the only PLP-dependent process that is not associated with metabolism of amino compounds (principally amino acids).

The requirement for vitamin B6 in the diet is proportional to the level of protein consumption ranging from 1.4–2.0 mg/day for a normal adult. During pregnancy and lactation the requirement for vitamin B6 increases approximately 0.6 mg/day.

Food source

Vitamin B6 content (mg)

Beef liver, 3.5 oz 1.4
Turkey, light meat, 3.5 oz 0.5
Chicken, light meat, 3.5 oz 0.63
Salmon, 3.5 oz cooked 0.65
Halibut, baked, 3.5 oz 0.4
Potatoes, 1 cup 0.48
Sweet potatoes, ½ cup 0.3
Oatmeal, 1 cup cooked 0.74
Rice, brown, cooked, 1 cup 0.28
Brussels sprouts, ½ cup 0.23
Lentils, ½ cup cooked 0.18
Carrots, ½ cup cooked 0.18
Peanuts, ½ cup 0.18
Sunflower seeds, ¼ cup 0.26
Avocado, 1 Haas 0.48
Mango 0.28
Watermelon, 1 cup 0.22
Cantelope, 1 cup 0.18
Prunes, 10 dried 0.22
Blackstrap molasses, 2 tbsp 0.29
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B6 Deficiency and Disease

Deficiencies of vitamin B6 are rare and usually are related to an overall deficiency of all the B-complex vitamins. Like the role of chronic alcohol consumption and an associated poor diet leading to thiamine deficiency, alcoholism is the leading cause of deficiency in B6. Isoniazid (see niacin deficiencies above) and penicillamine (used to treat rheumatoid arthritis and cystinurias) are two drugs that complex with pyridoxal and PLP resulting in a deficiency in this vitamin. Deficiencies in pyridoxal kinase result in reduced synthesis of PLP and are associated with seizure disorders related to a reduction in the synthesis of GABA. Due to its role in heme biosynthesis, deficiency in vitamin B6 can result in microcytic hypochromic anemias that are similar to those caused by iron deficiency or as a result of heavy metal (e.g. lead) poisoning. Two additional critical enzymes requiring PLP are cystathionine β-synthase (CBS) and cystathionine γ-lyase (cystathionase) which are involved in the metabolism of methionine to cysteine. Due to the role of B6 in this latter reaction, deficiencies in the vitamin can lead to homocysteinemia/uria due to a resultant blockade in the CBS reaction (see the Amino Acid Metabolism page for discussion of this effect). Other symptoms that may appear with deficiency in vitamin B6 include nervousness, insomnia, skin eruptions, loss of muscular control, anemia, mouth disorders, muscular weakness, dermatitis, arm and leg cramps, loss of hair, slow learning, and water retention.

Differential Diagnosis: Several Causes of Microcytic Anemia

Deficiency/Defect Characteristics
B6 deficiency PLP required for the rate-limiting enzyme in heme biosynthesis: δ-aminolevulinic acid synthase (ALAS); deficiency results in loss of protoporphyrin IX synthesis, therefore, there will be a significant reduction in measureable ALAS product (δ-aminolevulinic acid, δ-ALA) and protoporphyrin in these patients; loss of heme production leads to hypochromic microcytic anemia; lack of protoporphyrin results in iron deposits on mitochondria in bone marrow erythroblasts resulting in the formation of ringed sideroblasts; loss of iron incorporation into protoporphyrin IX leads to increased serum and intracellular iron concentration; increase in intracellular iron results in increased translation of ferritin as a means to prevent iron toxicity
Iron deficiency iron deficiency is the leading cause of microcytic anemia; loss of iron results in reduced production of heme, thus, the result is a hypochromic microcytic anemia; lack of heme production results in loss of feed-back inhibition of ALAS, therefore these patients will have an associated increase in measureable protoporphyrin; loss of iron intake means reduced iron in the serum and reduced intracellular iron, the latter resulting in reduced ferritin translation; loss of iron for incorporation into protoporphyrin IX results in spontaneous, non-enzymatic incorporation of Zn2+ forming Zn-protoporphyrin (ZPP), ZPP causes erythrocytes to fluoresce under ultraviolet illumination and is the basis of the ZPP test for iron deficiency or lead poisoning
Heavy metal poisoning heavy metals, such as lead, inhibit several enzymes of heme biosynthesis and metabolism with the most significant toxic effects resulting from inhibition of ferrochelatase, the enzyme that incorporates iron into protoporphyrin IX generating heme; similar to B6 deficiency, lead poisoning leads to increased intracellular iron in bone marrow erythroblasts causing the formation of ringed sideroblasts; because there is no heme, the ALAS reaction is not inhibited, as in the case of iron deficiency, this results in increased production of δ-ALA and protoporphyrin; lack of iron incorporation into protoporphyrin results in increased serum and intracellular iron concentrations, with the latter leading to increased ferritin synthesis as in the case of iron-deficient anemia; loss of iron for incorporation into protoporphyrin IX results in spontaneous, non-enzymatic incorporation of Zn2+ forming ZPP as in the case of iron-deficient anemia the ZPP test is diagnostic for lead poisoning

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Structure of biotin


Biotin is the cofactor required of enzymes that are involved in carboxylation, decarboxylation, or transcarboxylation reactions in prokaryotes and eukaryotes. Biotin is sometimes referred to as vitamin H. Biotin, produced by intestinal bacteria as well as that found in the diet, is bound to lysine residues in protein forming a complex that is called biocytin. Removal of biotin from biocytin and recycling of biotin from human biotin-dependent enzymes requires the activity of the enzyme biotinidase. The biotinidase gene (gene symbol: BTD) is located on chromosome 3p25.1 and is composed of 4 exons that encode a 543 amino acid protein. The clinical significance of defects in the BTD gene are described below.

In humans, the biotin-requiring enzymes include acetyl-CoA carboxylase (ACC), pyruvate carboxylase (PC), propionyl-CoA carboxylase (PCC), and 3-methylcrotonyl-CoA carboxylase (MCC). The critically important biotin-requiring enzymes are ACC, PC, and PCC. All three of these enzymes are referred to as ABC enzymes because they require/utilize ATP, Biotin, and CO2. The enzyme ACC is a cytosolic enzyme that is the rate-limiting enzyme of fatty acid synthesis. PC is a mtiochondrial enzyme that catalyzes the critical first reaction in the pathway of gluconeogenesis. PCC is a mitochondrial enzyme that catalyzes reactions involved in the metabolism of several amino acids (valine, methionine, isoleucine, and threonine) as well as the oxidation of fatty acids with an odd number of carbon atoms. The role of PCC in the metabolism of these compounds is often referred to as the VOMIT pathway (Valine, Odd-chain fats, Methionine, Isoleucine, Threonine) for memorization purposes. MCC is a mitochondrial enzyme that catalyzes the fourth step in the catabolism of leucine.

The biotin-dependent carboxylating enzymes in mammals are multifunctional and contain three distinct enzymatic domains: the biotin carboxylase (BC) domain, the carboxyltransferase (CT) domain, and the biotin carboxyl carrier protein (BCCP) domain. The activities of biotin-requiring carboxylases were determined in bacteria where all three of these activities are encoded by separate proteins.

Biotin is found in numerous foods and also is synthesized by intestinal bacteria. Some of the richest sources of biotin are swiss chard, tomatoes, romaine lettuce, and carrots. Additional sources include onions, cabbage, cucumber, cauliflower, mushrooms, peanuts, almonds, walnuts, oat meal, bananas, raspberries, strawberries, soy, egg yolk, and cow and goat milk.

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Clinical Significance of Biotin

Given that biotin is synthesized by intestinal bacteria deficiencies of the vitamin are rare. Deficiencies are generally seen only after long antibiotic therapies which deplete the intestinal fauna or following excessive consumption of raw eggs. The latter is due to the affinity of the egg white protein, avidin, for biotin preventing intestinal absorption of the biotin. An important autosomal recessive inherited disorder that leads to biotin deficiency is biotinidase (BTD) deficiency. Profound biotinidase deficiency is characterized by mutations in the gene that result in enzyme activity that is less than 10% of the normal. Partial biotinidase deficiency is characterized by enzyme activity that is 10%–30% of normal. Profound biotinidase deficiency occurs with a frequency of 1 in 60,000 live births. Indeed, the frequency is high enough, and the resultant symptoms severe enough, that current neonatal disease testing includes analysis for defects in the activity of this enzyme. The most severe symptoms associated with biotin deficiency and profound biotinidase gene defects are the result of the accumulation of toxic metabolic intermediates. The symptoms of profound biotinidase deficiency include delayed development, seizures, hypotonia, respiratory difficulties, hearing and vision loss, ataxia, skin rashes, and alopecia. Patients with profound biotinidase deficiency are also highly susceptible the fungal infection, candidiasis. Symptoms associated with partial biotinidase deficiency can be similar to those of the profound deficiency form but often these symptoms do not appear as severe except during infections, illnesses, or stress.

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Cobalamin is more commonly known as vitamin B12. Vitamin B12 is composed of a complex tetrapyrrol ring structure (corrin ring) and a cobalt ion in the center. Vitamin B12 (as the hydroxocobalamin form) is synthesized exclusively by microorganisms and is found in the liver of animals bound to protein as methycobalamin or 5'-deoxyadenosylcobalamin. Vitamin B12 is also present in cows milk and soybeans. The vitamin must be released from these binding proteins in order to be active and this release occurs in the stomach by the gastric protease, pepsin. After being released, cobalamin is bound to an endogenous protein called haptocorrin (also known as transcobalamin I, which is encoded by the TCN1 gene). Haptocorrin is produced by oral salivary glands in response to intake of food. The binding of hatocorrin to cobalamin that is released by the action of pepsin, protects the vitamin while it transits through the stomach into the duodenum. Within the small intestine pancreatic trypsin hydrolyzes the haptocorrin-cobalamin complexes allowing binding of cobalamin to another protein called intrinsic factor, IF. Intrinsic factor is also called gastric intrinsic factor which is encoded by the GIF gene. Intrinsic factor binds cobalamin but only within the alkaline pH of the intestines. Haptocorrin and intrinsic factor are two of the three vitamin B12-binding proteins produced in humans. The third is transcobalamin II (encoded by the TCN2 gene) which is a plasma globulin that is the primary transport protein for vitamin B12 in the blood.

Intrinsic factor is a 50-kDa glycoprotein produced by the parietal cells of the stomach. The secretion of intrinsic factor is stimulated by all three of the mechanisms that lead to gastric acid secretion: histamine, acetylcholine, and gastrin. Cobalamin, bound to intrinsic factor, can then bind to a receptor complex called cubilin present in the apical membranes of enterocytes in the distal ileum. The cubilin protein is encoded by the CUBN gene. Cubilin is also found in epithelial cells and has been shown to be involved in apoA-I-mediated binding of HDL. Upon intrinsic factor-cobalamin binding to cubilin, the complex is endocytosed. The intrinsic factor in the endocytosed complexes  is degraded by lysosomal hydrolases and the cobalamin is released. Within intestinal enterocyte cobalamin is bound to transcobalamin II and transferred into the blood for transport to the tissues. The transcobalamin II-cobalamin (TCN2-cobalamin) complexes are taken up, from the blood, by all cells via a plasma membrane transcobalamin receptor (TCBLR) encoded by the CD320 gene. The TCN2-cobalamin-TCBLR complex is internalized via endocytosis. Within the lysosome the TCN2 is degraded and the cobalamin is released so that it can be used as an enzyme co-factor within the cell. In addition to being required for TCN2-cobalamin uptake by cells, the CD320 protein is involved in B-cell proliferation.

There are only two clinically significant reactions in the body that require vitamin B12 as a co-factor. During the catabolism of fatty acids with an odd number of carbon atoms and the amino acids valine, isoleucine, methionine, and threonine the resultant propionyl-CoA is converted to succinyl-CoA for oxidation in the TCA cycle. One of the enzymes in this pathway, methylmalonyl-CoA mutase, requires vitamin B12 as a cofactor in the conversion of methylmalonyl-CoA to succinyl-CoA. The 5'-deoxyadenosine derivative of cobalamin is required for this reaction. The second reaction requiring vitamin B12 catalyzes the conversion of homocysteine to methionine and is catalyzed by methionine synthase (also known as homocysteine methyltransferase). This reaction results in the transfer of the methyl group from N5-methyltetrahydrofolate to hydroxocobalamin generating tetrahydrofolate (THF) and methylcobalamin during the process of the conversion.

Structure of cobalamin, vitamin B12

Vitamin B12 (cobalamin)

The recommended daily intake (RDA) of vitamin B12 for adults is 2.4 micrograms (μg). Vitamin B12 is found primarily in animal products. Due to the lack of sufficient vitamin B12 in plant foods it is added to breakfast cereals and this serves as a good source of the vitamin for vegetarians. Two plant sources that are useful for obtaining vitamin B12 are alfalfa and comfrey (also written comfry). However, to ensure adequate intake vegans should use a vitamin B12 supplement that contains at least 5–10μg due to the low absorption rate of the vitamin in supplement form.

Food source

Vitamin B12 content (mcg: μg)

Beef liver, 3.5 oz 48
Rainbow trout, wild, 3.0 oz 5.4
Salmon, 3.0 oz 4.9
Beef, 3.0 oz 2.4
Tuna, white, 3.0 oz 1.0
Yogurt, plain, 1 cup 1.4
Fortified cereal, 100% RDA 6.0
Milk, 1 cup 0.9
Swiss cheese, 1 oz 0.9
Egg, 1 whole 0.6
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Clinical Significances of B12 Deficiency

The liver can store up to six years worth of vitamin B12, hence deficiencies in this vitamin are rare. Pernicious anemia is a megaloblastic anemia resulting from vitamin B12 deficiency that develops as a result a lack of intrinsic factor in the stomach leading to malabsorption of the vitamin. The anemia results from impaired DNA synthesis due to a block in purine and thymidine biosynthesis. The block in nucleotide biosynthesis is a consequence of the effect of vitamin B12 on folate metabolism. When vitamin B12 is deficient essentially all of the folate becomes trapped as the N5-methylTHF derivative as a result of the loss of functional methionine synthase (also called homocysteine methyltransferase). This trapping prevents the synthesis of other THF derivatives required for the purine and thymidine nucleotide biosynthesis pathways.

Neurological complications also are associated with vitamin B12 deficiency and result from a progressive demyelination of nerve cells. The demyelination is thought to result from the increase in methylmalonyl-CoA that result from vitamin B12 deficiency. Methylmalonyl-CoA is a competitive inhibitor of malonyl-CoA in fatty acid biosynthesis as well as being able to substitute for malonyl-CoA in any fatty acid biosynthesis that may occur. Since the myelin sheath is in continual flux the methylmalonyl-CoA-induced inhibition of fatty acid synthesis results in the eventual destruction of the sheath. The incorporation methylmalonyl-CoA into fatty acid biosynthesis results in branched-chain fatty acids being produced that may severely alter the architecture of the normal membrane structure of nerve cells.

Deficiencies in B12 can also lead to elevations in the level of circulating homocysteine. Elevated levels of homocysteine are known to lead to cardiovascular dysfunction. Due to its high reactivity to proteins, homocysteine is almost always bound to proteins, thus thiolating them leading to their degradation. Homocysteine also binds to albumin and hemoglobin in the blood. Some of the detrimental effects of homocysteine are due to its' binding to lysyl oxidase, an enzyme responsible for proper maturation of the extracellular matrix proteins collagen and elastin. Production of defective collagen and elastin has a negative impact on arteries, bone and skin and the effects on arteries are believed to be the underlying cause for cardiac dysfunction associated with elevated serum homocysteine. In individuals with homocysteine levels above ≈12μM there is an increased risk of thrombosis and cardiovascular disease. The increased risk for thrombotic episodes, such as deep vein thrombosis (DVT), associated with homocysteinemia is due to homocysteine serving as a contact activation nucleus for activation of the intrinsic coagulation cascade.

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Folic Acid

Structure of folic acid

Folic Acid: positions 7 and 8 carry hydrogens in dihydrofolate (DHF); positions 5–8 carry hydrogens in tetrahydrofolate (THF)

Folic acid is a conjugated molecule consisting of a pteridine ring structure linked to para-aminobenzoic acid (PABA) that forms pteroic acid. Folic acid itself is then generated through the conjugation of glutamic acid residues to pteroic acid. Folic acid is obtained primarily from yeasts and leafy vegetables as well as animal liver. Animal cannot synthesize PABA nor attach glutamate residues to pteroic acid, thus, requiring folate intake in the diet.

When stored in the liver or ingested folic acid exists in a polyglutamate form. Intestinal mucosal cells remove some of the glutamate residues through the action of the lysosomal enzyme, conjugase. The removal of glutamate residues makes folate less negatively charged (from the polyglutamic acids) and therefore more capable of passing through the basal lamina membrane of the epithelial cells of the intestine and into the bloodstream. Within cells (principally the liver where it is stored), folate is converted first to dihydrofolate (DHF) and then to tetrahydrofolate (THF also H4folate) through the action of dihydrofolate reductase (DHFR), an NADPH-requiring enzyme.

The function of THF derivatives is to carry and transfer various forms of one carbon units during biosynthetic reactions. The one carbon units are either methyl, methylene, methenyl, formyl or formimino groups.

Active center of tetrahydrofolate (THF)

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

These one carbon transfer reactions are required in the biosynthesis of serine, methionine, glycine, choline, the purine nucleotides, and dTMP. Indeed, the interconversion of serine and glycine, via the involvement of THF, represents a major pathway for the generation of N5,N10-methylene-THF which of a member of the active folate pool. The active folate pool includes formyl-, methenyl-, and methylene-THF. Methyl-THF is referred to as the reduced folate pool. All of the active folate pool molecules can be converted to methyl-THF but the reverse process cannot occur. For this reason, the inability to convert methyl-THF to THF (as occurs via the B12- and folate-dependent enzyme, methionine synthase) leads to trapping of folate in the reduced form. The ability to acquire choline and amino acids from the diet and to salvage the purine nucleotides makes the role of N5,N10-methylene-THF, in dTMP synthesis, a metabolically significant function for this vitamin. The role of vitamin B12 and N5-methyl-THF in the conversion of homocysteine to methionine also can have a significant impact on the ability of cells to regenerate needed THF and as such represents the physiologically most significant reaction involving folate. Deficiencies in B12 or defects in methionine synthase will result in methyl-THF trapping with the result being the development of megaloblastic anemia due to the block in nucleotide production.

The recommended daily allowance for folic acid is 400 micrograms (μg) for both men and women except that pregnant women should increase their intake to at least 600 μg/day. The best sources for folic acid are cereals that have been fortified with 100% of the RDA of folic acid. Beef liver (3 oz) contains 45% of the RDA of folic acid. Excellent vegetarian sources for folic acid are baked beans, raw spinach, asparagus, green peas, broccoli, lentils, turnip greens, egg noodles, avocado, peanuts, lettuce, wheat germ, tomato juice and orange juice.

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Clinical Significances of Folate Deficiency

Folate deficiency results in complications nearly identical to those described for vitamin B12 deficiency. The most pronounced effect of folate deficiency on cellular processes is upon DNA synthesis. This is due to an impairment in dTMP synthesis which leads to cell cycle arrest in S-phase of rapidly proliferating cells, in particular hematopoietic cells. The result is megaloblastic anemia as for vitamin B12 deficiency. The inability to synthesize DNA during erythrocyte maturation leads to abnormally large erythrocytes termed macrocytic (megaloblastic) anemia. Since both folate and vitamin B12 deficiencies result in megaloblastic anemias, it is necessary to be able to clinically distinguish the cause as it relates to vitamin deficiency. Since B12 is required for both the methionine synthase reaction and the methylmalonyl-CoA mutase reaction a megaloblastic anemia resulting from B12 deficiency is also associated with an accompanying methylmalonic acidemia, whereas megaloblastic anemias caused by folate deficiency are not. An additional critical diagnostic is related to the onset of symptoms. Because human cells cannot store any significant amount of folate but the liver can store 3-6 years worth of B12, folate deficient megaloblastic anemias manifest very quickly, whereas B12 deficient anemias take considerably longer to manifest.

Folate deficiencies are rare due to the adequate presence of folate in food. Poor dietary habits as those of chronic alcoholics can lead to folate deficiency. The predominant causes of folate deficiency in non-alcoholics are impaired absorption or metabolism or an increased demand for the vitamin. The predominant condition requiring an increase in the daily intake of folate is pregnancy. This is due to an increased number of rapidly proliferating cells present in the blood. The need for folate will nearly double by the third trimester of pregnancy. Certain drugs such as anticonvulsants and oral contraceptives can impair the absorption of folate. Anticonvulsants also increase the rate of folate metabolism.

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Differential Diagnosis: Folate versus B12 Deficiencies

Deficiency/Defect Characteristics
B12 deficiency B12 is required for the methionine synthase and methylmalonyl-CoA mutase catalyzed reactions; deficiency in this vitamin is associated with both homocysteinemia and megaloblastic (macrocytic) anemia; the megaloblastic anemia associated with B12 deficiency is the result of folate (as N5-methyl-THF) trapping at the methionine synthase reaction; homocysteinemia results from the resultant inhibition of the methionine synthase reaction which leads to increased production of homocysteine which cannot be adequately accommodated in the catabolic direction at the cystathionine β-synthase (CBS) reaction; because methionine synthase is inhibited there will be a reduction in measureable methionine in the blood and urine; the resultant homocysteinemia is associated with a significant increase in the risk for cardiovascular disease such as deep vein thrombosis (DVT) and atherosclerosis; both the homocysteinemia and the megaloblastic anemia that result from B12 deficiency will be accompanied by dramatic increases in measurable methylmalonic acid in the serum which is NOT seen in folate deficiency; because the liver can store B12 for up to 6 years it takes a considerable amount of time for a dietary (or other cause) deficiency in B12 to manifest
Folate deficiency folate is also required for the methionine synthase reaction and, therefore, as for B12 deficiency, there will be both homocysteinemia and megaloblastic anemia with deficiency in this vitamin; in the case of folate deficiency, the megaloblastic anemia results from a direct loss of purine nucleotide and thymidine nucleotide synthesis; folate deficiency is associated with reduction in measureable serum and urine methionine as for B12 deficiency; the resultant homocysteinemia results in a significant increase in the risk for cardiovascular disease such as DVT and atherosclerosis as for B12 deficiency; since there is no associated inhibition of the methylmalonyl-CoA mutase reaction, there is no associated methylmalonic acidemia with the folate deficiency-associated homocysteinemia and megaloblastic anemia; because the human body cannot store folate, and the active folate pool is very small, the onset of symptoms associated with folate deficiency is very rapid

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Ascorbic Acid

Structure of ascorbic acid

Ascorbic Acid

Ascorbic acid is more commonly known as vitamin C. Ascorbic acid is derived from glucose via the uronic acid pathway, however, the enzyme L-gulono-γ-lactone oxidase responsible for the conversion of gulonolactone to ascorbic acid is absent in primates making ascorbic acid required in the diet. The gene encoding the enzyme (symbol: GULO) is present in the human genome but it lacks five of the twelve exons required to make a function protein. Therefore, the gene is considered a pseudogene and the designation for the gene in humans is GULOP.

The active form of vitamin C is ascorbic acid itself. The main function of ascorbate is as a reducing agent in a number of different reactions. Ascorbate is the cofactor for Cu+–dependent monooxygenases and Fe2+–dependent dioxygenases. Several critical enzymes that require ascorbate as a cofactor include the collagen processing enzymes, the lysyl hydroxylases and the prolyl hydroxylases as well as the catecholamine synthesis enzyme dopamine β-hydroxylase. Dietary ascorbate is also involved in non-heme iron absorption in the small intestine and in the overall regulation of iron homeostasis. In addition to its role in the regulation of the redox state of iron, ascorbate has the potential to reduce cytochromes a and c of the respiratory chain as well as molecular oxygen.

The most important reactions requiring ascorbate as a cofactor are the hydroxylations of lysine and proline residues in collagen. Vitamin C is, therefore, required for the maintenance of normal connective tissue as well as for wound healing since synthesis of connective tissue is the first event in wound tissue remodeling. Vitamin C also is necessary for bone remodeling due to the presence of collagen in the organic matrix of bones. Several other metabolic reactions require vitamin C as a cofactor. These include the catabolism of tyrosine, the synthesis of epinephrine from tyrosine (critical enzyme is dopamine β-hydroxylase), the synthesis of carnitine, peptide hormone amidation, and the synthesis of bile acids. It is also believed that vitamin C is involved in the process of steroidogenesis since the adrenal cortex contains high levels of vitamin C which are depleted upon adrenocorticotropic hormone (ACTH) stimulation of the gland.

Ascorbic acid also serves as a reducing agent and as an antioxidant. A critical anti-oxidant function of ascorbate is in the plasma membrane reduction of oxidized α-tocopherol (vitamin E). When functioning as an antioxidant, ascorbic acid itself becomes oxidized to semidehydroascorbate and then dehydroascorbate. Semidehydroascorbate is reconverted to ascorbate in the cytosol by cytochrome b5 reductase and thioredoxin reductase in reactions involving NADH and NADPH, respectively. Dehydroascorbate, the fully oxidized form of vitamin C, is reduced spontaneously by glutathione, as well as enzymatically in reactions using glutathione or NADPH.

Ascorbate is found at highest concentrations within nucleated cells with the intracellular concentration being found on the order of 30-100 times the concentration found in the plasma. The uptake on ascorbate from the lumen of the small intestine as well as its transport into cells is the function of two Na+-dependent vitamin C co-transporters identified as SVCT1 and SVCT2. Expression of SVCT1 is predominantly in intestinal epithelial cells and renal tubular cells where the transporter is found in the apical plasma membrane. Expression of SVCT2 is found in almost all nucleated cells. Expression of the SVCT2 transporter in the brain is critical for normal brain function and for ascorbate homeostasis within the brain. Both SVCT1 and SVCT2 are members of the SLC23 family of solute carriers and as such the gene encoding SVCT1 is SLC23A1 and that encoding SVCT2 is SLC23A2. Although the SVCTs are the primary transporters for ascorbate, cells are also capable of ascorbate uptake via the activity of the facilitative glucose transporters, GLUT1, GLUT3, and GLUT4. Indeed, erythrocyte accumulation of ascorbate occurs primarily through the action of GLUT1.

The amount of vitamin C that is recommended to consume each day (the RDA) depends upon the age and sex of the individual. Infants less than 1 year old should get 50 milligrams (mg) per day. children 1–3 years old need 15mg, 4–8 years old need 25mg, and 9–13 years old need 45mg. Adolescent girls should get 65mg per day and adolescent boys should get 75mg per day. Adult males need 90mg per day and adult women should get 75mg per day. Women who are breastfeeding should increase their intake to at least 120mg per day. Individuals who smoke should increase their daily intake by at least 35mg since smoking depletes vitamin C levels. The recommended daily intake of vitamin C to prevent conditions such as the cardiovascular disorders indicated above is reported to be between 500mg and 1000mg.

Excellent sources of vitamin C are fruits and vegetables such as oranges, watermelon, papaya, grapefruit, cantaloupe, strawberries, raspberries, blueberries, cranberries, pineapple, kiwi, mango, green peppers, broccoli, turnip greens, spinach, red and green peppers, canned and fresh tomatoes, potatoes, Brussels sprouts, cauliflower, and cabbage. Citrus juices or juices fortified with vitamin C are also excellent sources of the vitamin.

Vitamin C is sensitive to light, air, and heat, so the most vitamin C is available in fruits and vegetables that are eaten raw or lightly cooked. Natural or synthetic vitamin C can be found in a variety of forms. Tablets, capsules, and chewables are probably the most popular forms, but vitamin C also comes in powdered crystalline, effervescent, and liquid forms. An esterified form of vitamin C is also available, which may be easier on the stomach for those who are prone to heartburn. The best way to take vitamin C supplements is 2–3 times per day, with meals, depending on the dosage.

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Ascorbic Acid and Iron Homeostasis

Iron homeostasis is critical for cell survival. Indeed, iron-depletion rapidly results in cellular death. The details of dietary uptake and cellular storage and utilization of iron are presented in the Iron and Copper Homeostasis page, thus, this discussion will be limited to the role of ascorbate in the overall processes. As a brief overview, the adult human body contains 3–5 grams of iron. Of this, greater than 80% is utilized by red blood cells and is found within the hemoglobin in these cells. The vast majority of the remainder, approximately 20% of the total, is transiently stored within the liver and macrophages, with the liver being the major storage location. When inside cells, the bulk of iron is stored bound within complexes of the protein ferritin. The remainder of the iron is in the heme of cytochromes, iron-sulfur center (ISC)-containing proteins, and in non-heme and non-ISC iron-containing proteins.

When contained in proteins, iron is involved in redox reactions that are one-electron transfers between its ferric (Fe3+) and ferrous (Fe2+) forms. These electron transfers involving iron are vital for cell survival, but must be limited to the context of reactions directed by specific enzymatic reactions. Iron that is not bound in a redox-inert form is capable of generating highly deleterious reactive oxygen species (ROS) via a reaction known as the Fenton reaction. The product of this process is primarily the hydroxy free radical, the most poisonous of all ROS. Thus, as might be expected, iron homeostasis is tightly controlled through processes that regulate its import, storage and cellular efflux. Ascorbate is an essential co-factor in the overall process of iron homeostasis. Ascorbate stimulates dietary iron absorption, contributes to plasma transferrin-bound iron uptake following binding of the transferrin-iron complexes to the plasma membrane transferrin receptor, stimulates the synthesis of the iron storage protein ferritin, inhibits lysosomal ferritin degradation, and inhibits cellular iron efflux.

The absorption of non-heme iron, from the lumen of the small intestine, requires that the dietary iron, which is predominantly in the oxidized state (Fe3+), be reduced to the ferrous (Fe2+) state. Ferrous iron is then taken up into intestinal enterocytes via the action of the divalent metal transporter 1 (DMT1) protein. There are at least three mechanisms known to exist that contribute to the process of dietary iron reduction, all of which involve ascorbic acid. The major mechanism for intestinal iron reduction involves a ferrireductase that is a member of the cytochrome b561 class of transmembrane redox enzymes. This intestinal ferrireductase is known as duodenal cytochrome b561 (DCYTB) which is encoded by the CYBRD1 (cytochrome b reductase 1) gene. During the process of DCYTB-mediated reduction of lumenal ferric iron, intracellular ascorbate is oxidized to an ascorbyl radical and then to dehydroascorbate. Ascorbic acid, as an ascorbyl radical within the lumen of the gut is also capable of reducing ferric iron to ferrous iron which can then be absorbed via DMT1. A third mechanism for intestinal ferric iron reduction involves ascorbic acid being oxidized to dehydroascorbate in the lumen of the intestine concomitant with reduction of ferric to ferrous iron. The resulting ferrous iron is absorbed by the enterocyte via DMT1 while the dehydroascorbate is transported into the enterocyte most likely via the action of GLUT1.

When stored within cells, iron is complexed with a heteromeric protein called ferritin. Ferritin, that stores iron is composed of 24 subunits, that include heavy ferritin (H-ferritin) and light ferritin (L-ferritin). When stored inside ferritin, the ferrous iron that was transported into the cell is oxidized back to the ferric state. The ferroxidase activity of ferritin is associated with H-ferritin. When ferritin stored iron is released it is reduced back to the ferrous state. This intracellular reduction process involves ascorbic acid. Ferrous iron can be transported into the circulation from intestina enterocytes and from hepatocytes which represent the major iron storage cells in the body. This transport involves the transmembrane transporter identified as ferroportin. Associated with ferroportin in the membrane of intestinal enterocytes is the ferroxidase called hepheastin which oxidizes the ferrous iron to the ferric state. When iron is transported from heptocytes to the blood via ferroportin action, the ferrous iron is oxidized by the major plasma copper-dependent ferroxidase, ceruloplasmin. The oxidation of ferrous iron to ferric is required for plasma transport as the major plasma iron transport protein, transferrin, only binds the ferric state iron. The iron bound to trasnferrin is taken up by most cells after binding of the transferrin-iron complexes to the plasma membrane transferrin receptor followed by endocytosis of the ligand-receptor complex. The internalization and release of iron and its reduction to the ferrous state within the cell involves intracellular ascorbic acid.

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Clinical Significances of Ascorbate Deficiency

Deficiency in vitamin C leads to the disease scurvy due to the role of the vitamin in the post-translational modification of collagens. Scurvy is characterized by easily bruised skin, muscle fatigue, soft swollen gums, decreased wound healing and hemorrhaging, osteoporosis, and anemia. Due to its role in collagen processing, the bleeding dysfunction associated with vitamin C deficiency is characterized by the lack of effect on prothrombin time (PT) but with a prolonged bleeding time. The latter effect is the result of the reduced ability for platelets to adhere to exposed sub-endothelial extracellular matrix collagen which is required for their activation.

Vitamin C is readily absorbed and so the primary cause of vitamin C deficiency is poor diet and/or an increased requirement. The primary physiological state leading to an increased requirement for vitamin C is severe stress (or trauma). This is due to a rapid depletion in the adrenal stores of the vitamin. The reason for the decrease in adrenal vitamin C levels is unclear but may be due either to redistribution of the vitamin to areas that need it or an overall increased utilization.

Inefficient intake of vitamin C has also been associated with a number of conditions, such as high blood pressure, gallbladder disease, stroke, some cancers, and atherosclerosis (plaque in blood vessels that can lead to heart attack and stroke). Sufficient vitamin C in the diet may help reduce the risk of developing some of these conditions, however, the evidence that taking vitamin C supplements will help or prevent any of these conditions is still lacking.

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Vitamin A

Vitamin A consists of three biologically active molecules, retinol, retinal (retinaldehyde) and retinoic acid.

Structures of major forms of vitamin A

Structures of the major vitamin A compounds. The retinaldehyde forms of vitamin A are also commonly referred to as retinals. Both all-trans-retinaldehyde (all-trans-retinal) and 11-cis-retinaldehyde (11-cis-retinal) function in the process of vision. Retinoic acid is a major developmental regulating growth factor.

Each of these vitamin A compounds is derived from the plant precursor molecule, β-carotene (a member of a family of molecules known as carotenoids). Beta-carotene, which consists of two molecules of retinal linked at their aldehyde ends, is also referred to as the provitamin form of vitamin A.

Ingested β-carotene is cleaved in the lumen of the intestine by either of two β-carotene oxygenases (BCO1 and BCO2) to yield retinaldehyde (retinal). Retinal is reduced to retinol by retinaldehyde reductase, an NADPH requiring enzyme within the intestines. Retinol is esterified to palmitic acid and delivered to the blood via chylomicrons. The uptake of chylomicron remnants by the liver results in delivery of retinol to this organ for storage as a lipid ester within lipocytes. Transport of retinol from the liver to extrahepatic tissues occurs by binding of hydrolyzed retinol to retinol binding proteins, RBPs. The predominant plasma carrier of hepatic retinol is retinol binding protein 4 (RBP4). The retinol-RBP4 complex is then transported to the cell surface within the Golgi and secreted. While in the vasculature, the RBP4-retinol complexes interact with the protein transthyretin which prevents loss of retinol via renal glomerular filtration. The uptake of retinol, from the blood, occurs in response to the binding of RBP4-retinol complexes to the plasma membrane receptor identified as STRA6 (stimulated by retinoic acid 6). When bound to STRA6 the retinol is removed from RBP4 and transported across the membrane into the cell. Within extrahepatic tissues retinol is bound, primarily, to one of two RBPs, identified as RBP1 and RBP2. Within the eye, the major retinol binding protein is RBP3.

Vitamin A is found in dark green and yellow vegetables and yellow fruits, such as broccoli, spinach, turnip greens, carrots, squash, sweet potatoes, pumpkin, cantaloupe, and apricots, and in animal sources such as liver, milk, butter, cheese, and whole eggs. When determining the amount of vitamin A to consumw each day the RDA is expressed as retinol activity equivalents (RAE).

Food source

Vitamin A content (mcg: μg RAE)

Organ meats, cooked, 3 oz 1490–9126
Sweet potato with peel (1 medium), baked 1096
Carrots, cooked from fresh, ½ cup 671
Spinach, cooked from frozen, ½ cup 573
Kale, cooked from frozen, ½ cup 478
Beet greens, cooked, ½ cup 276
Dandelion greens, cooked, ½ cup 260
Cantaloupe, raw, ¼ medium melon 233
Mustard greens, cooked, ½ cup 221
Red sweet pepper, cooked, ½ cup 186
Chinese cabbage, cooked, ½ cup 180
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Gene Control Exerted by Retinoic Acid

Retinol is converted to retinaldehyde via the action of two classes of oxidizing enzymes, cytoplasmic and microsomal. The cytoplasmic enzymes are alcohol dehydrogenases (ADHs) of the medium-chain dehydrogenase/reductase family. The microsomal enzymes are retinol dehydrogenases (RDHs) of the short-chain dehydrogenase/reductase family. Conversion of retinaldehyde to retinoic acid is catalyzed by a family of retinaldehyde dehydrogenases (RALDHs) that are members of the aldehyde dehydrogenase 1 family of enzymes. The retinoic acid producing enzymes are RALDH1 (gene symbol: ALDH1A1), RALDH2 (gene symbol: ALDH1A2), and RALDH3 (gene symbol: ALDH1A3). All three RALDHs are expressed in tissue specific and developmental stage specific patterns. The degradation of retinoic acid is catalyzed by enzymes of the cytochrome P450 (CYP) family, specifically CYP26A1, CYP26B1, and CYP26C1.

Within cells retinoic acid (as well as the related compound 9-cis-retinoic acid) binds to specific receptor proteins that are members of the nuclear receptor family of lipid hormone receptors. Prior to interaction with retinoic acid, the receptor protein is bound to retinoic acid response elements (RARE) in retinoic acid responsive genes. Following binding, the receptor-retinoic acid complex alters the transcriptional activity of the associated genes. Retinoic acid responsive genes are involved in growth and differentiation pathways during embryogenesis that control organogenesis, cranio-facial development, central nervous system patterning and axial patterning. In this capacity retinoic acid is considered a hormone of the steroid/thyroid hormone superfamily. The retinoic acid receptors are abbreviated RAR and the related retinoid X receptors (RXRs) bind 9-cis-retinoic acid. Several genes, whose patterns of expression are altered by retinoic acid, are involved in the earliest processes of embryogenesis including the differentiation of the three germ layers, organogenesis and limb development. Within the adult retinoic acid is involved in processes of reproduction, immunity, memory, and learning. To date, there are more than 530 genes that have been shown to be directly regulated (either positive or negative) via interaction with retinoic acid bound RAR or RXR. The canonical retinoic acid response element (RARE) in retinoic acid target genes is the direct repeat 5'–PuG(G/T)(T/A)CA–3' separated by a spacer of five nucleotides. This response element is referred to as the DR5 element.

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Vision and the Role of Vitamin A

Photoreception in the eye is the function of two specialized neuron types located in the retina; the rod and cone cells. The rod cells are utilized only for seeing at night During the day rod cells continuously release the excitatory neurotransmitter glutamate. The released glutamate binds to the synaptic membranes of specialized cells called bipolar cells. The glutamate receptor on bipolar cells is the mGluR6 metabotropic receptor. For more information on the glutamate receptors go to the Biochemistry of Nerve Transmission page. Activation of bipolar cells induces them to release the inhibitory neurotransmitter, GABA. Release of GABA, thus leads to inhibition of the optic nerve.

Both rod and cone cells contain a photoreceptor pigment in their membranes. The photosensitive compound of most mammalian eyes is a protein called opsin to which is covalently coupled an aldehyde of vitamin A. The opsin of rod cells is called scotopsin. The photoreceptor of rod cells is specifically called rhodopsin or visual purple. This compound is a complex between scotopsin and the 11-cis-retinal (also called 11-cis-retinene) form of vitamin A. Rhodopsin is a G-protein coupled receptor (GPCR) embedded in the membrane of the discs inside the outer segment of rod cells. Coupling of 11-cis-retinal occurs at three of the transmembrane domains of rhodopsin. Intracellularly, rhodopsin is coupled to a specific G-protein called transducin. The specific Gα-subunit of transduction is identified as Gαt.

role of vitamin A in the function of visual signaling via rod cells

Vitamin A and visual signaling in rod cells: When the rhodopsin is exposed to light, the associated 11-cis-retinal is converted to all-trans-retinal, a process referred to as photobleaching. The conformational change in retinal converts rhodopsin to metarhodopsin II. The change to metarhodopsin II in turn activates the associated G-protein, tranducin. Activated transducin exhibits an increased GTP-binding by the α-subunit. Binding of GTP releases the α-subunit from the inhibitory βγ subunits. The GTP-activated α-subunit in turn activates an associated cGMP phosphodiesterase (PDE); an enzyme that hydrolyzes cyclic-GMP (cGMP) to GMP. This PDE is specifically identified as PDE6B. Cyclic GMP is required to maintain a cyclic nucleotide gated (CNG) ion channel in the open conformation. The rod cGMP-gated ion channel is a heterotetramer consisting of three α subunits and one β subunit (α3β). These subunits are encoded by the CNGA1 and CNGB1 genes, respectively. The CNG ion channel, when opened via cGMP interaction, transports both Na+ and Ca2+ ions into the rod cell maintaining the cell in a state of depolarization. The CNG ion channel is tightly associated with a member of the Na+/Ca2+–K+ exchanger (NCKX) family of transporters encoded by the SLC24 family of genes. The rod cell NCKX transporter (NCKX1) is encoded by the SCL24A1 gene and is associated with the CNGA1/CNGA2 heterotetramer as a homodimer. The primary function of the NCKX1 transporter is to ensure ionic balance in the rod cell in the dark state by transporting one Ca2+ ion and one K+ ion out of the rod cell in exchange for four Na+ ions transported into the cell. The reciprocal Ca2+ and Na+ transport via NCKX1 is referred to as the dark current and resets the rod cell following photon activation. In the dark, the continuous influx of Ca2+ through the CNG ion channel allows the rod cell to release glutamate which then binds to its receptor (mGluR6) on the bipolar cell activating the bipolar cell to release GABA which in turn inhibits the activity of the optic nerve. The PDE6B-mediated drop in cGMP concentration results in closure of the CNGA1/CNGA2 channel resulting in reduced uptake of Ca2+ ion. Since Ca2+ ions are required for the release of glutamate from rod cell pre-synaptic vesicles, the release of glutamate is inhibited. The loss of glutamate release results in less activation of the mGluR6 receptor on bipolar cells with the consequences that these cells are no longer depolarized by glutamate binding. The release of the inhibitory neurotransmitter, GABA, by the bipolar cells requires the glutamate-mediated depolarization. Therefore, in the absence of glutamate stimulation the bipolar cell no longer releases GABA. The net result is that the optic nerve become disinhibited enabling visual signaling under low light conditions.

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Clinical Significances of Vitamin A Deficiency

Vitamin A is stored in the liver and deficiency of the vitamin occurs only after prolonged lack of dietary intake. As a fat soluble vitamin any lipid absorption disorder can be associated with deficiency in the vitamin. Patients with cystic fibrosis are particularly prone to deficiencies in the fat soluble vitamins due to defective pancreatic enzyme secretion and function. The earliest symptoms of vitamin A deficiency are night blindness. Additional early symptoms include follicular hyperkeratinosis, increased susceptibility to infection and cancer and anemia equivalent to iron deficient anemia. Prolonged lack of vitamin A leads to deterioration of the eye tissue through progressive keratinization of the cornea, a condition known as xerophthalmia.

The increased risk of cancer in vitamin deficiency is thought to be the result of a depletion in β-carotene. Beta-carotene is a very effective antioxidant and is suspected to reduce the risk of cancers known to be initiated by the production of free radicals. Of particular interest is the potential benefit of increased β-carotene intake to reduce the risk of lung cancer in smokers. However, caution needs to be taken when increasing the intake of any of the lipid soluble vitamins. Excess accumulation of vitamin A in the liver can lead to toxicity which manifests as bone pain, hepatosplenomegaly, nausea and diarrhea.

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Vitamin D

Vitamin D is a steroid hormone that functions to regulate specific gene expression following interaction with its intracellular receptor (VDR: vitamin D receptor). The VDR is a member of the steroid and thyroid hormone superfamily of nuclear receptors. The biologically active form of the hormone is 1,25-dihydroxy vitamin D3 (1,25-(OH)2D3, also called 1,25-dihydroxycholecalciferol or calcitriol). Calcitriol functions primarily to regulate calcium and phosphorous homeostasis.

Structures of major forms of vitamin D

Structures of the major vitamin D compounds. Ergosterol is the plant-derived precursor to vitamin D2, whereas, 7-dehydrocholesterol is the naturally produced precursor to vitamin D3.

Active calcitriol is derived from ergosterol (produced in plants) and from 7-dehydrocholesterol. 7-dehydrocholesterol is an intermediate in the synthesis of cholesterol that accumulates in the skin. Upon exposure to ultraviolet (uv) light from the sun and following thermal isomerization, 7-dehydrocholesterol is non-enzymatically converted to pre-vitamin D3 which then enters the bloodstream and is taken up by the liver where it undergoes the first of two activating hydroxylation reactions. Ergocalciferol (vitamin D2) is formed by uv irradiation of ergosterol.

Vitamin D2 and D3 are processed to D2-calcitriol and D3-calcitriol, respectively, by the same enzymatic pathways in the body. Cholecalciferol (or ergocalciferol) are absorbed from the intestine and transported to the liver bound to a specific vitamin D-binding protein. In the liver cholecalciferol is hydroxylated at the 25 position by a specific vitamin D 25-hydroxylase generating 25-hydroxy-D3 [25-(OH)D3] which is the major circulating form of vitamin D. The vitamin D 25-hydroxylase enzyme is encoded by the CYP2R1 (cytochrome P450 2R1) gene. Conversion of 25-(OH)D3 to its biologically active form, calcitriol, occurs through the activity of a specific 25-hydroxyvitamin D3 1-α-hydroxylase present in the proximal convoluted tubules of the kidneys, and in bone, keratinocytes, immune cells, and placenta. The 25-hydroxyvitamin D3 1-α-hydroxylase enzyme is encoded by the CYP27B1 (cytochrome P450 27B1) gene. 25-(OH)D3 can also be hydroxylated at the 24 position by a specific vitamin D3-24-hydroxylase in the kidneys, intestine, placenta and cartilage.

Structures of the major hormonal forms of vitamin D

Structures of 25-hydroxyitamin D3 and 1,25-dihydroxyvitamin D3. Formation of 25-hydroxyvitamin D3 occurs within the liver. Coversion of 25-hydroxyvitamin D3 to the hormonally active compound, calcitriol (1,25-dihydroxyvitamin D3 occurs within the kidneys.

Calcitriol functions in concert with parathyroid hormone (PTH) to regulate serum calcium and phosphorous levels. PTH is released from the parathyroid gland in response to low serum calcium and induces the production of calcitriol by activating the expression of the renal CYP27B1 gene. In contrast, reduced levels of PTH stimulate synthesis of the inactive 24,25-(OH)2D3. Within cells, calcitriol functions as a steroid hormone inducing the expression of calcium-binding proteins called calbindins. These proteins include calbindin 1, 28kDa (encoded by the CALB1 gene) and calbindin-D9k (encoded by the CALB3 gene). The CALB1 encoded protein is expressed in neuronal and endocrine cells and acts as a calcium sensor and calcium buffer. The CALB1 encoded protein is a member of the calcium-binding superfamily of proteins that includes calmodulin and troponin C. The CALB3 encoded protein is involved in intestinal calcium absorption. The steroid hormone action of vitamin D occurs via the action of calcitriol binding to a specific intracellular receptor that is a member of the nuclear receptor family of hormone receptors called the vitamin D receptor (VDR).

The increased absorption of calcium ions requires concomitant absorption of a negatively charged counter ion to maintain electrical neutrality. The predominant counter ion is Pi. When plasma calcium levels fall the major sites of action of calcitriol and PTH are bone where they stimulate bone resorption and the kidneys where they inhibit calcium excretion by stimulating reabsorption by the distal tubules.

Due to the critical role of vitamin D in the development of growing bone as well as the maintenance of healthy bone it has been added to milk since the 1930's. In the US all milk is fortified with 100 IU/cup. Fatty fishes such as salmon, sardines and tuna are also an excellent source of vitamin D. Cod liver oil is also very high in vitamin D. For vegetarians and vegans ultraviolet (uv) light irradiated mushrooms and yeast are the only sources of vitamin D that are not animal in origin. The irradiation of yeasts and mushrooms produces vitamin D2. Vitamin D content is listed as the amount in International Units (IU).

Food source

Vitamin D content (IU)

Cod liver oil, 1Tbs (15ml) 1360
Salmon, cooked, 3.5 oz 447
Sardines, canned in oil, 1.75 oz 250
Tuna, canned in water, 3 oz 154
Egg, 1 large 41
Swiss cheese 6
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Clinical Significances of Vitamin D

Vitamin D Deficiency

As a result of the addition of vitamin D to milk, deficiencies in this vitamin are rare in most developed countries. As a fat soluble vitamin, any lipid absorption disorder can be associated with deficiency in the vitamin. Patients with cystic fibrosis are particularly prone to deficiencies in the fat soluble vitamins due to defective pancreatic enzyme secretion and function. The main symptom of vitamin D deficiency in children is rickets and in adults is osteomalacia. Rickets is characterized by improper mineralization during the development of the bones resulting in soft bones. Osteomalacia is characterized by demineralization of previously formed bone leading to increased softness and susceptibility to fracture. The resultant decrease in Ca2+ uptake due to vitamin D deficiency leads to hyperparathyroidism as a compensatory mechanism. The excess release of parathyroid hormone (PTH) can cause metastatic calcification and peritrabecular fibrosis. The peritrabecular fibrosis, associated with hyperparathyroidism, is quite distinct and is clinically called osteitis fibrosa cystica (OFC). OFC is also called von Recklinghausen disease of bone. This latter name is distinct from von Recklinghausen disease which is neurofibromatosis type 1.

Vitamin D Toxicity

Due to the fat solubility of vitamin D it is possible to consume too much of the vitamin. Excess vitamin D intake results in a very characteristic toxicity profile. The excess vitamin results in and associated excess uptake of intestinal Ca2+ leading to hypercalcemia. This hypercalcemia is essentially indistinguishable from the hypercalcemia caused by secondary hyperparathyroidism in the context of vitamin D deficiency. The earliest tissue affected by the onset of the metastatic calcification with vitamin D toxicity is the kidney. The symptoms of the renal deficit are very similar to those experienced by type 1 diabetics: polydipsia and polyuria. Chronic vitamin D toxicity leads to metastatic calcification of numerous soft tissues and eventually results in permanent renal failure.

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Vitamin E

Structure of the tocopherols

Structures of the four tocopherols. The biologically active forms of vitamin E constitute a family of four related compounds called tocopherols. The most abundant tocopherol in non-hepatic tissues in humans is the α-tocopherol form.

Vitamin E is a mixture of several related compounds known as tocopherols and tocotrienols. The tocopherols are the major sources of vitamin E in the U.S. diet. The tocopherols differ by the number and position of methyl (–CH3–) groups present on the ring system of the chemical structure. The different tocopherols are designated α-, β-, γ-, and δ-tocopherol. Most vitamin E in U.S. diets is in the form of γ-tocopherol from soybean, canola, corn, and other vegetable oils. All four tocopherols are able to act as free radical scavengers thus they all have potent antioxidant properties. Vitamin E is absorbed from the intestines packaged in chylomicrons. It is delivered to the tissues via chylomicron transport and then to the liver through chylomicron remnant uptake. The liver can export vitamin E in very low density lipoproteins (VLDLs). Within the liver α-tocopherol transfer protein preferentially transfers α-tocopherol to VLDLs, thus α-tocopherol is the most abundant tocopherol in nonhepatic (liver) tissues. Due to its lipophilic nature, vitamin E accumulates in cellular membranes, fat deposits and other circulating lipoproteins. The major site of vitamin E storage is in adipose tissue.

The major function of vitamin E is to act as a natural antioxidant by scavenging free radicals and molecular oxygen. In particular vitamin E is important for preventing peroxidation of polyunsaturated membrane fatty acids. The vitamins E and C are interrelated in their antioxidant capabilities. Active α-tocopherol can be regenerated by interaction with vitamin C following scavenge of a peroxy free radical. Alternatively, α-tocopherol can scavenge two peroxy free radicals and then be conjugated to glucuronate for excretion in the bile.

Although α-tocopherol is the most abundant tocopherol in tissues outside of the liver, it is not the most potent antioxidant form of the vitamin. Because of the unmethylated carbons in the ring structure of γ- and δ-tocopherol, these two forms of vitamin E are much more active at trapping free radicals, in particular reactive nitrogen species. In addition, research has recently shown that the anticancer effects of vitamin E are due to the γ- and δ-tocopherol forms and is not associated with α-tocopherol.

Recommended daily intake amounts for vitamin E (as α-tocopherol) are listed in milligram (mg) amounts and also in International Units (IU). To convert from mg to IU: 1 mg of α-tocopherol is equivalent to 1.49 IU of the natural form or 2.22 IU of the synthetic form. Nuts, seeds, and vegetable oils are among the best sources of α-tocopherol, and significant amounts are available in green leafy vegetables and fortified cereals. Most vitamin E in US diets is in the form of γ-tocopherol from soybean, canola, corn, and other vegetable oils.

Food source

Vitamin E content (mg)

Wheat germ oil, 1 tablespoon 20.3
Almonds, dry roasted, 1 ounce 7.4
Sunflower oil, 1 tablespoon 5.6
Safflower oil, 1 tablespoon 4.6
Peanut butter, 2 tablespoons 2.9
Peanuts, dry roasted, 1 ounce 2.2
Corn oil, 1 tablespoon 1.9
Broccoli, chopped, boiled, ½ cup 1.2
Soybean oil, 1 tablespoon 1.1
Kiwifruit, 1 medium 1.1
Spinach, raw, 1 cup 0.6
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Clinical Significances of Vitamin E Deficiency

Due to adequate vitamin E intake in the average American diet, no major deficiency syndromes are common. However, conditions that result from vitamin E deficiency are related to disturbances in nerve cell membrane lipid homeostasis. These conditions include cerebellar ataxias, myopathies, retinopathy, loss of deep tendon reflexes, and dysarthria (speech problem associated with difficulty articulating due to defects in the motor component of speech). Another major symptom of vitamin E deficiency in humans is an increase in red blood cell fragility due to the increased level of erythrocyte membrane lipid peroxidation in the absence of tocopherols. Since vitamin E is absorbed from the intestines in chylomicrons, any fat absorption disorder can lead to deficiencies in vitamin E intake. Patients with cystic fibrosis are particularly prone to deficiencies in the fat soluble vitamins due to the defective pancreatic enzyme secretion and function. Increased intake of vitamin E is recommended in premature infants fed formulas that are low in the vitamin as well as in persons consuming a diet high in polyunsaturated fatty acids. Polyunsaturated fatty acids tend to form free radicals upon exposure to oxygen and this may lead to an increased risk of certain cancers.

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Vitamin K

The K vitamins exist naturally as K1 (phylloquinone) in green vegetables and K2 (menaquinone) produced by intestinal bacteria and K3 is synthetic menadione. When administered, vitamin K3 is alkylated to one of the vitamin K2 forms of menaquinone.

Structures of vitamin K molecules

Structures of the three major vitamin K molecules. The biologically active forms of vitamin K constitute a family of three related compounds, two of which are naturally produced (K1 and K2) and one of which is manufactured as a food additive (K3). The "n" after the isoprene structure in vitamin K2 represents the fact that 6, 7, or 9 isoprene groups are found in the most commonly derived forms of the vitamin.

The major function of the K vitamins is in the maintenance of normal levels of the blood clotting proteins, factors II, VII, IX, X and protein C and protein S, which are synthesized in the liver as inactive precursor proteins. Conversion from inactive to active clotting factor requires a posttranslational modification of specific glutamate (E) residues. This modification is a carboxylation and the enzyme responsible requires vitamin K as a cofactor. The resultant modified E residues are γ-carboxyglutamate (gla). This process is most clearly understood for factor II, also called preprothrombin. Prothrombin is modified preprothrombin. The gla residues are effective calcium ion chelators. Upon chelation of calcium, prothrombin interacts with phospholipids in membranes and is proteolysed to thrombin through the action of activated factor X (Xa).

During the carboxylation reaction reduced hydroquinone form of vitamin K is converted to a 2,3-epoxide form. The regeneration of the hydroquinone form requires the action of the enzyme vitamin K epoxide reductase (VKORC1) which involves a two-step reaction. These latter reactions are the site of action of the coumarin based anticoagulants such as warfarin (trade name: Coumadin®).

Structure of a γ-carboxyglutamamte (gla) residue and mechanism of incorporation into protein

Incorporation of a gla-residue into prothrombin: The incorporation of a gla-residue into a protein such as prothrombin requires the hydroquinone (KH2) form of vitamin K (either K1, K2, or synthetic K3). The utilization and regeneration of the KH2 form in the overall process of the γ-glutamyl carboxylase (GGCX) reaction is referred to as the vitamin K cycle. Either following the carboxylation, or directly from dietary quinone forms of vitamin K, the action of vitamin K epoxide reductase (VKORC1) is to provide a continuous source of the KH2 form. An addition enzyme, vitamin K quinone reductase (VKQR) can catalyze the conversion of the quinone form to the hydroquinone form using NADH as a cofactor.

In the average U.S. diet, meats and eggs are the most common food sources of the menaquinone form of vitamin K. Excellent sources of vitamin K include spinach, Brussels sprouts, Swiss chard, green beans, asparagus, broccoli, kale, mustard greens, green peas and carrots. Fermentation of foods can increase their vitamin K content. Fermented soy foods play a unique role in supplying vitamin K in certain traditional cuisines (like that of Japan). Sometimes you will see the word "natto" being used to refer to these fermented soy foods since Bacillus natto are bacteria that can convert vitamin K1 into K2 and are often used in the production of fermented soy products. Some cheeses are also fermented in a way that optimizes their vitamin K content.

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Clinical Significances of Vitamin K Deficiency

Naturally occurring vitamin K is absorbed from the intestines only in the presence of bile salts and other lipids through interaction with chylomicrons. Therefore, fat absorption disorders can result in vitamin K deficiency. Patients with cystic fibrosis are particularly prone to deficiencies in the fat soluble vitamins due to defective pancreatic enzyme secretion and function. The synthetic vitamin K3 is water soluble and absorbed irrespective of the presence of intestinal lipids and bile. Since the vitamin K2 form is synthesized by intestinal bacteria, deficiency of the vitamin in adults is rare. However, long term antibiotic treatment can lead to deficiency in adults. The intestine of newborn infants is sterile, therefore, vitamin K deficiency in infants is possible if lacking from the early diet. The primary symptom of a deficiency in infants is a hemorrhagic syndrome. Since vitamin K is critical for the generation of gla residues in clotting factors, as described above, deficiency in this vitamin results in dysfunction in the processes of blood coagulation. Since the defect will be at the level of factors II, VII, IX, X and protein C and protein S, the effects of vitamin K deficiency can be seen in the context of the prothrombin time (PT) assay. Whereas, vitamin C deficiency is associated with bleeding dysfunction at the level of platelet adhesion to collagen and is, therefore, measurable with an associated increase in bleeding time, vitamin K deficiency has no effect on bleeding time.

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Alpha-Lipoic Acid, LA

Alpha-lipoic acid, LA, (chemical name: 1,2-dithiolane-3-pentanoic acid; also known as thioctic acid), is a naturally occurring dithiol compound synthesized enzymatically in the mitochondrion from the medium-chain fatty acid octanoic acid. Because LA can be synthesized in the body it is not technically considered a vitamin but because of its vital role in overall cellular metabolism it is considered as an important, but not necessary,  dietary supplement. Lipoic acid has one chiral center and therefore exists in both R- and S-enantiomeric forms (see the Figure below). However, only R-LA is conjugated to conserved lysine residues in an amide linkage (forming a lipoamide), thus making this isoform essential as a cofactor in biological systems. Enzymes containing lipoamide are typically mitochondrial multi-enzyme complexes that catalyze the oxidative decarboxylation of α-keto acids such as the pyruvate dehydrogenase complex (PDHc), 2-oxoglutarate dehydrogenase (also known as α-ketoglutarate dehydrogenase), and branched-chain keto acid dehydrogenase (BCKD), and the glycine cleavage complex (GCC).

Given that LA is a necessary cofactor for mitochondrial α-ketoacid dehydrogenases, it thus serves a critical role in mitochondrial energy metabolism. In addition to de novo synthesis, LA is also absorbed intact from dietary sources, and it transiently accumulates in many tissues. LA has been described as a potent biological antioxidant, a detoxification agent, and a diabetes medicine. LA has been used to improve age-associated cardiovascular, cognitive, and neuromuscular deficits, and has been implicated as a modulator of various inflammatory signaling pathways. Accumulating evidence indicates that LA, supplied as a supplement in the diet, may not be used as a metabolic cofactor but instead, elicits a unique set of biochemical activities with potential therapeutic value against a host of pathophysiological insults.

structure of alpha-lipoic acid

Structure of α-Lipoic Acid

Lipoic acid, either as a dietary supplement or a therapeutic agent, modulates distinct redox circuits because of its ability to equilibrate between different subcellular compartments as well as extracellularly. Because of its role in regulating redox states, LA is a critical component of the antioxidant network. It is important in the regeneration of other antioxidants, such as vitamins E and C, it increases the intracellular levels of glutathione (GSH), and provides redox regulation of numerous proteins and transcription factors. The extracellular thiol/disulfide red-ox environment (determined by the interconversion between cysteine and cystine) is able to modulate cell proliferation, apoptosis, cell adhesion molecules, and proinflammatory signaling. Lipoic acid may play an important role in the modulation of the extracellular red-ox state via the involvement of dihydrolipoic acid (DHLA) in the reduction of cystine to cysteine. This reduction facilitates a rapid uptake of the cysteine into the cell making it available to stimulate GSH synthesis. Cellular uptake of LA has been shown to occur via several systems including the medium chain fatty acid transporter, a Na+-dependent vitamin transport system, and a H+-linked monocarboxylate transporter for intestinal uptake. The cellular reduction of LA to DHLA is accomplished by NAD(P)H-driven enzymes, thioredoxin reductase, lipoamide dehydrogenase, and glutathione reductase.

interconversions of dihydrolipoic acid (DHLA) and lipoic acid (LA)

Interconversions of α-lipoic acid (LA) and dihydrolipoic acid (DHLA)

Lipoic acid is an essential cofactor for the E2 component of α-ketoacid dehydrogenase complexes that are exclusively located in mitochondria. These complexes include the pyruvate dehydrogenase complex (PDHc), 2-oxoglutarate (α-ketoglutarate) dehydrogenase (OGDH), and branched chain α-ketoacid dehydrogenase (BCKD) complexes. The PDHc catalyzes the oxidative carboxylation of pyruvate which serves as the entry point for carbohydrates into the TCA cycle as acetyl-CoA. OGDH catalyzes the conversion of 2-oxoglutarate (α-ketoglutarate) to succinyl-CoA in the TCA cycle. Evidence demonstrates that the activities of both PDH and OGDH are substantially decreased during aging and in neurodegenerative disorders.

Extensive evidence suggests that LA may have therapeutic usefulness in lowering blood glucose levels in diabetic conditions and that the intracellular redox status plays a role in the modulation of insulin resistance. Lipoic acid has been shown to stimulate glucose uptake by affecting components of the insulin signaling pathway. The signaling networks of insulin receptor activation include the insulin receptor substrates (IRS1, IRS2, and IRS4), PI3K, and PKB/AKT. There are three members of the PKB/AKT family of serine/threonine kinases identified as AKT1 (PKB, also PKBα), AKT2 (PKBβ), and AKT3 (PKBγ). Insulin-mediated activation of PI3K and PKB/AKT is necessary for the translocation of GLUT4 from an intracellular pool to the plasma membrane to allow for uptake of plasma glucose. Lipoic acid has been shown to augment tyrosine phosphorylation and the activity of components of insulin signaling including the insulin receptor, IRS1, PI3K, PKB/AKT, and p38MAPK. In addition, LA stimulated glucose uptake upon translocation and regulation of the intrinsic activity of GLUT4, an effect that is likely mediated by p38 MAPK. R-LA, as well as oxidized isoforms have been shown to stimulate glucose transport in differentiated 3T3-L1 adipocytes by a mechanism entailing changes in the intracellular redox status. These effects of LA are in agreement with an alteration of the thiol reactivity of redox components of the insulin signaling pathway caused by a thiol/disulfide exchange mechanism.

The stress-activated MAPK and JNK pathways play a central role in the progression of insulin resistance and diabetic neuropathies. The activation of these kinases results in the phosphorylation of IRS1 on serine 307 resulting in inhibition of the insulin-stimulated tyrosine phosphorylation of IRS1 leading to inhibition of insulin signaling. Lipoic acid has been shown to inhibit the JNK pathway and IRS1 serine phosphorylation, resulting in improved insulin sensitivity. The precise mechanism by which LA inhibits the JNK pathway remains unclear. Nevertheless, these effects place LA at the cross-road of insulin- and JNK signaling favoring glucose uptake and metabolism, thus ameliorating insulin resistance and improving the status of diabetes.

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RDA Values for Vitamins and Minerals

Click the following link to open a PDF file containing the recommended daily intake values for various vitamins and minerals:

RDA Tables for Vitamins and Minerals

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Last modified: January 10, 2017