Chapter 1. vitamin A

1 Vitamin A: Nutritional Aspects of Retinoids and Carotenoids A Catharine Ross and Earl H Harrison CONTENTS Introduction Nutritional Aspects of Vitamin A and Carotenoids Historical Definitions of Vitamin A, Retinoids, and Carotenoids Properties, Nutritional Equivalency, and Recommended Intakes Properties of Nutritionally Important Retinoids Properties of Nutritionally Important Carotenoids Nutritional Equivalency Transport and Metabolism 10 Transport and Binding Proteins 10 Retinol-Binding Protein 12 Albumin 13 Lipoproteins 13 Intracellular Retinoid-Binding Proteins 13 Nuclear Retinoid Receptors 14 Intestinal Metabolism 15 Conversion of Provitamin A Carotenoids to Retinoids 15 Intestinal Absorption of Vitamin A 17 Reesterification, Incorporation into Chylomicrons, and Lymphatic Secretion 18 Hepatic Uptake, Storage, and Release of Vitamin A 18 Hepatic Uptake 18 Extrahepatic Uptake 19 Storage 19 Release 20 Plasma Transport 20 Plasma Retinol 20 Conditions in Which Plasma Retinol May Be Low 22 Other Retinoids in Plasma 23 Plasma Carotenoids 23 Plasma Retinol Kinetics and Recycling 23 Intracellular Retinoid Metabolism 23 Hydrolysis 23 Oxidation–Reduction and Irreversible Oxidation Reactions 24 Formation of More Polar Retinoids 24 ß 2006 by Taylor & Francis Group, LLC Conjugation Isomerization Vitamin A and Public Health Prevention of Xerophthalmia Actions of Vitamin A in the Eye Morbidity and Mortality Subclinical Deficiency Immune System Changes Medical Uses of Retinoids Dermatology Acute Promyelocytic Leukemia Prevention of Hypervitaminosis A of Nutritional Origin Excessive Consumption of b-Carotene References 25 25 25 25 26 27 27 27 28 28 29 29 29 30 INTRODUCTION Vitamin A (retinol) is an essential micronutrient for all vertebrates It is required for normal vision, reproduction, embryonic development, cell and tissue differentiation, and immune function Many aspects of the transport and metabolism of vitamin A, as well as its functions, are well conserved among species Dietary vitamin A is ingested in two main forms—preformed vitamin A (retinyl esters and retinol) and provitamin A carotenoids (b-carotene, a-carotene, and b-cryptoxanthin)—although the proportion of vitamin A obtained from each of these form varies considerably among animal species and among individual human diets These precursors serve as substrates for the biosynthesis of two essential metabolites of vitamin A: 11-cis-retinal, required for vision, and all-trans-retinoic acid, required for cell differentiation and the regulation of gene transcription in nearly all tissues Research on vitamin A now spans nine decades Over 34,000 citations to vitamin A, 7,000 to b-carotene, and 20,000 to retinoic acid can be found in the National Library of Medicine’s PubMed database [1], covering topics related to nutrition, biochemistry, molecular and cell biology, physiology, toxicology, public health, and medical therapy Besides the naturally occurring forms of vitamin A indicated earlier, numerous structural analogs have been synthesized Some retinoids have become widely used as therapeutic agents, particularly in the treatment of dermatological diseases and certain cancers In this chapter, we focus first on vitamin A from a nutritional perspective, addressing its chemical forms and properties, the nutritional equivalency of compounds that provide vitamin A activity, and current dietary recommendations We then cover the metabolism of carotenoids and vitamin A Finally, we provide a brief discussion of the key uses of vitamin A and retinoids in public health and medicine, referring to their benefits as well as some of the adverse effects caused by ingesting excessive amounts of this highly potent group of compounds NUTRITIONAL ASPECTS OF VITAMIN A AND CAROTENOIDS HISTORICAL Vitamin A was discovered in the early 1900s by McCollum and colleagues at the University of Wisconsin and independently by Osborne and Mendel at Yale Both groups were studying the effects of diets made from purified protein and carbohydrate sources, such as casein and rice flour, on the growth and survival of young rats They observed that growth ceased and the animals died unless the diet was supplemented with butter, fish oils, or a quantitatively ß 2006 by Taylor & Francis Group, LLC minor ether-soluble fraction extracted from these substances, from milk, or from meats The unknown substance was then called ‘‘fat-soluble A.’’ Not long thereafter, it was recognized that the yellow carotenes present in plant extracts had similar nutritional properties, and it was postulated that this carotenoid fraction could give rise through metabolism to the bioactive form of fat-soluble A, now called vitamin A, in animal tissues This was shown to be correct after b-carotene and retinol were isolated and characterized, and it was shown that dietary b-carotene gives rise to retinol in animal tissues Within the first few decades of vitamin A research, vitamin A deficiency was shown to cause several specific disease conditions, including xerophthalmia; squamous metaplasia of epithelial and mucosal tissues; increased susceptibility to infections; and abnormalities of reproduction Each of these seminal discoveries paved the way for many subsequent investigations that have greatly expanded our knowledge about vitamin A Although the discoveries made in the early 1900s may now seem long ago, it is interesting to note, as reviewed by Wolf [2], that physicians in ancient Egypt, around 1500 BC, were already using the liver of ox, a very rich source of vitamin A, to cure what is now referred to as night blindness DEFINITIONS OF VITAMIN A, RETINOIDS, AND CAROTENOIDS Vitamin A is a generic term that refers to compounds with the biological activity of retinol These include the provitamin A carotenoids, principally b-carotene, a-carotene, and b-cryptoxanthin, which are provided in the diet by green and yellow or orange vegetables and some fruits and preformed vitamin A, namely retinyl esters and retinol itself, present in foods of animal origin, mainly in organ meats such as liver, other meats, eggs, and dairy products The term retinoid was coined to describe synthetically produced structural analogs of the naturally occurring vitamin A family, but the term is now used for natural as well as synthetic compounds [3] Retinoids and carotenoids are defined based on molecular structure According to the Joint Commission on Biochemical Nomenclature of the International Union of Pure and Applied Chemistry and International Union of Biochemistry and Molecular Biology (IUPAC–IUB), retinoids are ‘‘a class of compounds consisting of four isoprenoid units joined in a head-to-tail manner’’ [4] All-trans-retinol is the parent molecule of this family The retinoid molecule can be divided into three parts: a trimethylated cyclohexene ring, a conjugated tetraene side chain, and a polar carbon–oxygen functional group Additional examples of key retinoids and structural subgroups, a history of the naming of these compounds, and current nomenclature of retinoids are available online [4] The IUPAC–IUB defines carotenoids [5] as ‘‘a class of hydrocarbons (carotenes) and their oxygenated derivatives (xanthophylls) consisting of eight isoprenoid units joined in such a manner that the arrangement of isoprenoid units is reversed at the center of the molecule.’’ All carotenoids may be formally derived from the acyclic C40H56 structure that has a long central chain of conjugated double bonds, by (i) hydrogenation, (ii) dehydrogenation, (iii) cyclization, or (iv) oxidation, or any combination of these processes PROPERTIES, NUTRITIONAL EQUIVALENCY, AND RECOMMENDED INTAKES Properties of Nutritionally Important Retinoids Nutritionally important retinoids and some of their metabolites are illustrated in Figure 1.1 The conventional numbering of carbon atoms in the retinoid molecule is shown in the structure of all-trans-retinol in Figure 1.1a Due to the conjugated double-bond structure of retinoids and carotenoids, these molecules possess very characteristic UV or visible light absorption spectra that are useful in their identification and quantification [6,7] ß 2006 by Taylor & Francis Group, LLC 16 17 19 345 20 11 10 13 12 14 15 COOH CH2OH 18 (a) All-trans-retinol (e) CH2OH (b) All-trans-retinoic acid 9-cis-Retinoic acid 3,4-Didehydroretinol (f) COOH CHO O OH All-trans-retinal O (c) OH OH O COOH 11-cis-Retinal (d) (g) Retinoyl-β-glucuronide CHO FIGURE 1.1 Nutritionally important retinoids and major metabolites The conventional numbering system for retinoids is shown for all-trans-retinol, the parent molecule of the retinoid family Furr and colleagues have summarized the light absorption properties of over 50 retinoids [8] and nutritionally active carotenoids [9] Some of the properties of several retinoids related to dietary vitamin A are summarized in Table 1.1 Retinoids tend to be most stable in the all-trans configuration Retinol is most often present in tissues in esterified form, where the fatty acyl group is usually palmitate with lesser amounts of stearate and oleate esters Esterification protects the hydroxyl group from oxidation and significantly alters the molecule’s physical properties (Table 1.1) Retinyl esters in tissues are usually admixed with triglycerides and other neutral lipids, including the antioxidant a-tocopherol Retinyl esters are the major form of vitamin A in the body as a whole and the predominant form (often more than 95%) in chylomicrons, cellular lipid droplets, and milk fat globules Thus, they are also the major form in foods of animal origin Retinol contained in nutritional supplements and fortified foods is usually produced synthetically and is stabilized by formation of the acetate, propionate, or palmitate ester Minor forms of vitamin A may be present in the diet, such as vitamin A2 (3,4-didehydroretinol) (Figure 1.1b), which is present in the oils of fresh-water fish and serves as a visual pigment in these species [10] Several retinoids that are crucial for function are either absent or insignificant in the diet, but are generated metabolically from dietary precursors Due to the potential for the double bonds of the molecules in the vitamin A family to exist in either the trans- or cis-isomeric form, a large number of retinoid isomers are possible The terminal functional group can be in one of several oxidation states, varying from hydrocarbon, as in anhydroretinol, to alcohol, aldehyde, and carboxylic acid Many of these forms may be further modified through the addition of substituents to the ring, side chain, or end group These changes in molecular structure significantly alter the physical properties of the molecules in the vitamin A family and may markedly affect their biological activity While dozens of natural retinoids ß 2006 by Taylor & Francis Group, LLC ß 2006 by Taylor & Francis Group, LLC TABLE 1.1 Properties of Vitamin A Compounds and Their Metabolites Compound All-trans-retinol (vitamin A1) 3,4-Didehydroretinol (vitamin A2) Retinyl palmitate All-trans-retinal 11-cis-Retinal All-trans-retinoic acid 9-cis-Retinoic acid 13-cis-Retinoic acid Retinoyl-b-glucuronide 4-Oxo-all-trans-retinoic acid Formula and Molecular Mass C20H30O 286.44 C20H28O 284.44 C36H60O2 528 C20H28O; 284.44 C20H28O; 284.44 C20H28O2; 300.4 C20H28O2; 300.4 C20H28O2; 300.4 C26H36O8 476.1 C20H26O3; 314.4 Molar Extinction Coefficient, «, in Indicated Solvent Solvents in Which Soluble Physical State Wavelength of Maximum Absorption, lmax Absolute alcohol, methanol, chloroform, petroleum ether, fats, and oils Alcohols, ether Crystalline solid 324–325 Crystalline solid 350 52,770 in ethanol 51,770 in hexane 41,320 in ethanol Hexane, ether, dimethylsulfoxide Viscous oil 325 49,260 in ethanol Ethanol, chloroform, cyclohexane, petroleum ether, oils Ethanol, chloroform, cyclohexane, petroleum ether, oils Ethanol, methanol, isopropanol, dimethyl sulfoxide Ethanol, methanol, isopropanol, dimethyl sulfoxide Ethanol, methanol, isopropanol, dimethyl sulfoxide Aqueous methanol Crystalline solid Crystalline solid 383 368 380 365 350 42,880 48,000 24,935 26,360 45,300 Crystalline solid 345 36,900 in ethanol Crystalline solid 354 39,750 in ethanol Crystalline solid 360 50,700 in methanol Crystalline solid 360 58,220 in ethanol Ethanol, methanol, dimethyl sulfoxide Note: For additional absorption spectrum data, see Furr et al [8,9] Crystalline solid in ethanol in hexane in ethanol in hexane in ethanol Lycopene (a) BCO 15 15Ј BCO-2 10Ј 9Ј All-trans-β-carotene (b) α-Carotene (c) OH (d) HO (e) HO Lutein β-Cryptoxanthin FIGURE 1.2 Nutritionally important carotenoids (a) Lycopene, a nonprovitamin A carotene; (b) alltrans-bb0 -carotene; arrows indicate sites of cleavage by b-carotene monooxygenase, BCO, and BCO-2; (c) all-trans (a,b0 ) carotene; (d) lutein, a nonprovitamin A xanthophyll; (e) b-cryptoxanthin have been isolated, the molecules illustrated in Figure 1.1 and Figure 1.2 are the principal retinoids and carotenoids, respectively, of nutritional importance, and thus are the main focus of this chapter Nevertheless, it is important to recognize that numerous minor metabolites can be formed at several branch points as retinol and the provitamin A carotenoids are metabolized All-trans-retinal (Figure 1.1c) is the immediate product of the central cleavage of b-carotene as well as an intermediate in the oxidative metabolism of retinol to all-transretinoic acid The 11-cis isomer of retinal (Figure 1.1d) is formed in the retina and most of it is covalently bound to one of the visual pigments, rhodopsin in rods or iodopsin in cones The aldehyde functional group of 11-cis-retinal combines with specific lysine residues in these proteins as a Schiff’s base All-trans-retinoic acid (Figure 1.1e) is the most bioactive form of vitamin A When fed to vitamin A-deficient animals, retinoic acid restores growth and tissue differentiation and prevents mortality, indicating that this form alone, or metabolites made from it, is able to support nearly all of the functions attributed to vitamin A A notable exception is vision, which is not restored by retinoic acid because retinoic acid cannot be reduced to retinal in vivo Retinoic acid is also the most potent natural ligand of the retinoid receptors, RAR and RXR (described later), as demonstrated in transactivation assays Several cis isomers of retinoic acid have been studied rather extensively, but they are still somewhat enigmatic as to origin and function 9-cis-Retinoic acid (Figure 1.1f ) is capable of binding to the nuclear receptors and may be a principal ligand of the RXR 13-cis-Retinoic acid is present in plasma, often at a concentration similar to all-trans-retinoic acid, and its therapeutic effects are well demonstrated (see the section Dermatology), but it is not known to be a high-affinity ligand for the nuclear retinoid receptors It is possible that 13-cis-retinoic acid acts as a relatively stable precursor or prodrug that can be metabolized to all-trans-retinoic acid or perhaps ß 2006 by Taylor & Francis Group, LLC another bioactive metabolite Di-cis isomers of retinoic acid also have been detected in plasma, further illustrating the complex mix of retinoids in biological systems Retinoids that are more polar than retinol or retinoic acid are formed through oxidative metabolism of the ionone ring and side chain These include 4-hydroxy, 4-oxo, 18-hydroxy, and 5,6-epoxy derivatives of retinoic acid, and similar modifications of other retinoids Conjugation of the lipophilic retinoids with very polar molecules such as glucuronic acid renders them water-soluble As an example, retinoyl-b-glucuronide (Figure 1.1g) is present as a significant metabolite in the plasma and bile Although some of these polar retinoids are active in some assays, most of the more polar and water-soluble retinoids appear to result from phase I and phase II metabolic or detoxification reactions They may, however, be deconjugated to some extent and recycled as the free compound Many retinoids have been chemically synthesized A large number of structural analogs have been synthesized and tested for their potential as drugs that may be able to induce cell differentiation In the field of dermatology, 13-cis-retinoic acid (isotretinoin) and the 1,2,4-trimethyl-3-methoxyphenyl analog of retinoic acid (acetretin) are prominent differentiation-promoting and keratinolytic compounds Other retinoids have been developed as agents able to selectively bind to and activate only a subset of retinoid receptors Some synthetic retinoids show none of the biological activities of vitamin A, but still are related in terms of structure Retinoids that show selectivity in binding to the RXR receptors rather than RAR, sometimes referred to as rexinoids, also have been synthesized [11,12] As analytical methods have improved, additional retinoids have been discovered Retinol metabolites have been identified in which the terminal group is dehydrated (anhydroretinol); the 13,14 position is saturated or hydroxylated; or the double bonds of the retinoid side chain are flipped back into a form known as a retro retinoid [4] These retinoids tend to be quantitatively minor or limited in their distribution, and their significance is still uncertain Properties of Nutritionally Important Carotenoids Carotenoids are synthesized by photosynthetic plants and some algae and bacteria, but not by animal tissues The initial stage of biosynthesis results in the formation of the basic polyisoprenoid structure of the hydrocarbon lycopene (Figure 1.2a), a 40-carbon linear structure with an extended system of 13 conjugated double bonds Further biosynthetic reactions result in the cyclization of the ends of this linear molecule to form either a- or b-ionone rings The carotene group of carotenoids comprises hydrocarbon carotenoids in which the ionone rings bear no other substituents The addition of oxygen to the carotene structure results in the formation of the xanthophyll group of carotenoids The double bonds in most carotenoids are present in the more stable all-trans configuration, although cis isomers can exist Carotenoids are widespread in nature and are responsible for the yellow, orange, red, and purple colors of many fruits, flowers, birds, insects, and marine animals In photosynthetic plants, carotenoids improve the efficiency of photosynthesis, while they are important to insects, birds, animals, and humans for their colorful and attractive sensory properties Although some 600 carotenoids have been isolated from natural sources, only about one-tenth of them are present in human diets [13], and only about 20 have been detected in blood and tissues b-carotene (Figure 1.2b), a-carotene (Figure 1.2c), lycopene, lutein (Figure 1.2d), and b-cryptoxanthin (Figure 1.2e) are the five most prominent carotenoids in the human body However, only b-carotene, a-carotene, and b-cryptoxanthin possess significant vitamin A activity To be active as vitamin A, a carotenoid must have an unsubstituted b-ionone ring and an unsaturated hydrocarbon chain The bioactivity of all-trans-b-carotene, with two symmetrical halves, is about twice that of an equal amount of a-carotene and b-cryptoxanthin, in which only one unsubstituted b-ionone ring is present Even though lycopene, lutein, and zeaxanthin can be relatively abundant in the diet and ß 2006 by Taylor & Francis Group, LLC humans can absorb them across the intestine into plasma, they lack vitamin A activity because of the absence of a closed unsubstituted ring In plants, provitamin A carotenoids are embedded in complex cellular structures such as the cellulose-containing matrix of chloroplasts or pigment-containing chromoplasts Their association with these matrices of plants is a significant factor affecting the efficiency of their digestion, release, and bioavailability [14,15] Nutritional Equivalency Units of Activity Different forms of vitamin A differ in their biological activity per unit of mass For this reason, the bioactivity of vitamin A in the diet is expressed in equivalents (with respect to all-transretinol) rather than in mass units Several different units have been adopted over time and most of them are still used in some capacity In 1967, the World Health Organization (WHO)=FAO recommended replacing the international unit (IU), a bioactivity unit, with the retinol equivalent (RE); RE was defined as mg of all-trans-retinol or mg of b-carotene in foods [16] In 2001, the U.S Institute of Medicine recommended replacing the RE with the retinol activity equivalent (RAE) and redefining the average equivalency values for carotenoids in foods in comparison with retinol [15] These sequential changes in units were in large part a response to better knowledge of the efficiency of utilization of carotenoids [15,16]; mg RAE is defined as mg of all-trans-retinol, and therefore is the same as mg RE Both are equal to 3.3 IU of retinol The equivalency of provitamin A carotenoids and retinol in the RAE system is illustrated in Figure 1.3 These currently adopted conversion factors are necessarily approximations Because the RAE terminology is not yet fully used, the vitamin A values in some food tables, food labels, and supplements are still expressed in RE or IU Another term, daily value (% DV), is used in food labeling It is not a true unit of activity, but provides an indication of the percentage of the recommended dietary allowance (RDA)* present in one serving of a given food Form consumed Equivalency after bioconversion Dietary or supplemental vitamin A (1 µg) Retinol (1 µg) Supplemental β-carotene (pure, in oily solution) (2 µg) Retinol (1 µg) Dietary β-carotene (in food matrix) (12 µg) Retinol (1 µg) Dietary α-carotene or β-cryptoxanthin (in food matrix) (24 µg) Retinol (1 µg) FIGURE 1.3 Approximate nutritional equivalency of dietary provitamin A carotenoids and retinol, as revised in 2001 The values shown are used to convert the contents of carotenoids in supplements and foods to equivalent amounts of dietary retinol (From Institute of Medicine, Dietary Reference Intakes for Vitamin A, Vitamin K, Arsenic, Boron, Chromium, Copper, Iodine, Iron, Manganese, Molybdenum, Nickel, Silicon, Vanadium, and Zinc, National Academy Press, Washington, 2002, pp 8–9.) * Based on the percentage of the RDA of a nutrient, for a person consuming a 2000 kcal diet ß 2006 by Taylor & Francis Group, LLC TABLE 1.2 Recommended Dietary Allowances (RDA) and Upper Level (UL) Values for Vitamin A by Life Stage Group Life Stage Group RDA (mg=day)a UL (mg=day)b Infants 0–12 months 400c 600 Children 1–3 years 4–8 years 300 400 600 900 Adolescent and adult males 9–13 years 14–18 years 19 to 70 years 600 900 900 1700 2800 3000 Adolescent and adult females 9–13 years 14–18 years 19 to 70 years 600 700 700 1700 2800 3000 Pregnancy