Exhaustive Study Notes on Lipid Soluble Vitamins: Vitamin A and Vitamin D

General Introduction to Vitamins

• Definition: Vitamins are organic compounds occurring in natural foods that are required in trace amounts for normal health and development.

• Synthesis: Generally, vitamins cannot be synthesized by the human body and must therefore be supplied through the diet.

• Historical Context:     * In 1912, Funk isolated a crystalline substance from rice polishings that could prevent or cure polyneuritis in pigeons.     * Chemically, this substance was found to be an "amine" and was essential to life, leading to the name "vitamine."     * Throughout the 20th century, 13 vitamins were identified.

Vitamers

• Definition: Vitamers are several closely related compounds that possess the activity of a particular vitamin.

• Examples:     * Vitamin B6: Refers to pyridoxine, pyridoxal, and pyridoxamine.     * Vitamin A: Refers to retinol, retinal, and retinoic acid.

Classification and Characteristics of Vitamins

• Water-Soluble Vitamins:     * Non-B-Complex: Ascorbic acid (Vitamin C).     * B-Complex (Energy-releasing): Thiamine ($B_1$), Riboflavin ($B_2$), Niacin ($B_3$), Biotin, Pantothenic acid.     * B-Complex (Hematopoietic): Folic acid, Vitamin $B_{12}$.     * B-Complex (Other): Pyridoxine ($B_6$), Pyridoxal, Pyridoxamine.

• Fat-Soluble Vitamins:     * Vitamin A (retinol, $\beta$-carotenes).     * Vitamin D (cholecalciferol).     * Vitamin K (phylloquinones, menaquinones).     * Vitamin E (tocopherols).

Comparative Characteristics

Fat-Soluble Vitamins: General Principles

• Nature: Hydrophobic compounds.

• Absorption Requirement: Can only be absorbed efficiently when there is normal fat absorption.

• Transport: Transported in the blood within lipoproteins or attached to specific binding proteins.

• Clinical Significance: Conditions affecting lipid digestion and absorption (e.g., very low-fat diets, steatorrhea, biliary system disorders) can lead to vitamin deficiency syndromes.

Vitamin A: Chemistry and Forms

• Distribution:     * Active forms are present only in animal tissues.     * The pro-vitamin, $\beta$-carotene, is found in plant tissues.

• Important Forms (Retinoids): Includes all molecules chemically related to retinol (natural and synthetic).     * Retinol: An alcohol ($-CH_2OH$).     * Retinal (Retinaldehyde): An aldehyde ($-CHO$).     * Retinoic Acid: A carboxylic acid ($-COOH$).

• Provitamin A (Carotenoids): $\alpha$, $\beta$, $\gamma$-carotenes and cryptoxanthin are quantitatively most important. Over 500 carotenoids exist, but less than $10\%$ have provitamin A activity.

• Molecular Structure ($C_{20}$):     * Consists of a trimethyl cyclohexenyl ring ($\beta$-ionone ring).     * Contains an all-trans isoprenoid chain with four double bonds.     * $\beta$-carotenes: A symmetrical molecule with two terminal $\beta$-ionone rings connected by an $18$-Carbon hydrocarbon chain with 4 conjugated double bonds. Theoretically, one molecule of $\beta$-carotene can yield two molecules of vitamin A.

Absorption and Metabolism of Vitamin A

• Conversion and Equivalency:     * $6\,\mu g$ of $\beta$-carotene is equivalent to $1\,\mu g$ of preformed retinol.     * Retinol is the key player and can be converted to other forms.     * Pathway: Retinol $\rightleftharpoons$ Retinal $\rightarrow$ Retinoic acid. (Note: Conversion of retinal to retinoic acid is irreversible).     * Reductase: Retinal is reduced to retinol by an $NADH$ or $NADPH$ dependent retinal reductase.

• Absorption Process:     * $\beta$-carotene is cleaved by a di-oxygenase to form retinal.     * Absorption requires bile salts.     * Within mucosal cells, retinol is re-esterified with fatty acids.     * It is then incorporated into chylomicrons and transported to the liver.     * It is transported from the liver by Retinol Binding Protein (RBP).

Dietary Sources and Requirements of Vitamin A

• Animal Sources: Fish, liver, eggs (excellent sources), milk, egg yolk.

• Plant Sources: Dark green, deeply colored fruits and vegetables (tomatoes, carrots, green-yellow vegetables, spinach, mangoes, papayas, corn, sweet potatoes) and alfalfa.

• Cooking: Moderate cooking enhances carotenoid release for gut uptake.

• Unit of Activity: $1\text{ RAE}$ (Retinol Activity Equivalent) = $3.33\text{ IU}$ of retinol and $20\text{ IU}$ of $\beta$-carotene.

• Daily Requirements:     * Men (>19 yrs): $900\,\mu g\text{ RAE}$.     * Women (>19 yrs): $700\,\mu g\text{ RAE}$.     * Pregnancy: $770\,\mu g\text{ RAE/d}$.     * Lactating: $1300\,\mu g\text{ RAE/d}$.     * Children (1-18 yrs): $300-900\,\mu g\text{ RAE}$.

• Blood Levels:     * Normal blood level: $18-60\,\mu g/dl$.     * Carotenoids: $100-300\,\mu g/dl$.

Roles and Functions of Vitamin A

• Vision: Retinal is essential for vision and maintenance of the cornea.

• Protein Synthesis and Cell Differentiation: Involved in the differentiation of epithelial and goblet cells (Retinoic acid).

• Reproduction and Growth: Retinol supports these processes.

• Immunity: Supported by retinoic acid and carotenoids.

• Bone Health: Involved in bone growth and remodeling.

• Glycoprotein Synthesis: Retinol phosphate acts as a carrier of oligosaccharide chains to glycoprotein molecules.

• Mucopolysaccharide Synthesis: Retinoic acid is essential for the sulfation of mucopolysaccharides in matrix collagen and bones.

• Collagen Preservation: Inhibits collagenase, preventing collagen breakdown.

• Skin Health: Prevents keratinization of epithelial cells, keeping mucous membranes moist and healthy. Used in treating cystic acne by inhibiting keratinization and decreasing sebum secretion.

• Antioxidant: Specifically associated with $\beta$-carotene activity.

Wald’s Visual Cycle (The Rhodopsin Cycle)

• Initial Isomerization: All-trans retinol from blood is isomerized to $11$-cis retinol, then oxidized to $11$-cis retinal by retinol dehydrogenase in the presence of $NAD^+$.

• Rhodopsin Formation: The aldehyde group of $11$-cis-retinal (prosthetic group) covalently attaches to opsin (apoprotein) via a Schiff base to the $\epsilon$-amino group of a Lysine ($Lys$) residue, forming Rhodopsin (holoprotein).

• Light Activation:     * Absorption of light by rhodopsin causes the isomerization of $11$-cis-retinal.     * Sequence of intermediates:         1. Photorhodopsin ($10\text{ picoseconds}$).         2. Bathorhodopsin ($45\text{ picoseconds}$).         3. Lumirhodopsin ($30\text{ nanoseconds}$).         4. Metarhodopsin I ($75\text{ microseconds}$).         5. Metarhodopsin II ($10\text{ milliseconds}$; active signaling state).         6. Metarhodopsin III (minutes).     * Result: Final breakdown into Opsin and all-trans retinal. At this stage, the eye becomes less sensitive to light.

• Signaling Mechanism:     * In the dark: Rod cells contain enough $cGMP$ to keep voltage-gated $Ca^{2+}$ and $Na^+$ channels open.     * Light exposure: Decreases $cGMP$ levels via phosphodiesterase activation (triggered by Transducin-$GTP$).     * Lowered $cGMP$ causes ion channels to close, hyperpolarizing the plasma membrane and creating a nerve impulse.

Vitamin A and Gene Expression

• Retinoic acid is transported into the nucleus by a binding protein.

• It combines with receptor proteins $RAR$ and $RXR$.

• This complex binds to regulatory regions of the $DNA$ strand and interacts with other transcription factors.

• This "switches" genes on or off, increasing or decreasing the $mRNA$ of specific proteins.

Vitamin A Deficiency

• Night Blindness (Nyctalopia): The earliest sign; characterized by impairment in dark adaptation. Deficiency depresses the re-synthesis of rhodopsin.

• Eye Changes (Xerophthalmia):     * Dry conjunctiva.     * Bitot’s spots (white patches on the conjunctiva).     * Keratomalacia (softening of the cornea).     * Corneal ulceration.

• Skin Changes:     * Dryness and roughness.     * Papular eruptions and follicular hyperkeratosis (Phrynoderma or "toad skin").

• Systemic Effects:     * Increased susceptibility to respiratory tract infections.     * Keratinization of the urinary tract, leading to calculi formation.

Vitamin A Toxicity

• Carotenemia: Yellowing of the skin (but strictly not the sclerae) following ingestion of >30\,mg/d. Often seen in hypothyroid patients due to impaired carotene breakdown.

• Hypervitaminosis A (Acute): Raised intracranial pressure ($ICP$), vertigo, diplopia, bulging fontanels in children, seizures, exfoliative dermatitis, and death.

• Hypervitaminosis A (Chronic): Dry skin, cheilosis, glossitis, alopecia, bone demineralization and pain, hypercalcemia, liver fibrosis with portal hypertension, lymph node enlargement, and pseudotumor cerebri (raised $ICP$ and papilledema).

• Teratogenicity: Excess Vitamin A in pregnant women can cause congenital malformations, spontaneous abortions, craniofacial abnormalities, and valvular heart disease ($VHD$). Daily doses should not exceed $3\,mg$.

• Note for Smokers: High doses of $\beta$-carotene should be avoided as they may increase the risk of lung cancer.

Vitamin D (Calciferol)

• Forms:     * Vitamin $D_2$ (Ergocalciferol): Synthetic form produced by irradiation of plant steroid ergosterol (plant source).     * Vitamin $D_3$ (Cholecalciferol): Produced photochemically in the skin from $7$-dehydrocholesterol by sunlight/$UV$ light (animal source).

• Classification Debate: Vitamin D is not strictly a vitamin because the body can synthesize it from cholesterol in the skin if exposed to enough sunlight; it also functions as a hormone.

Synthesis and Activation of Vitamin D

1. Skin: $7$-dehydrocholesterol $\xrightarrow{UV\,rays}$ Photolysis $\rightarrow$ Secosterol (cis) $\rightarrow$ Cholecalciferol ($D_3$) (trans).

2. Liver: Cholecalciferol $\xrightarrow{25-hydroxylase}$ $25$-hydroxy cholecalciferol ($25(OH)D$). This is the main storage form and binds to Vitamin D Binding Globulin.

3. Kidney: $25$-hydroxy cholecalciferol $\xrightarrow{1-alpha-hydroxylase}$ Calcitriol ($1,25$-Dihydroxy cholecalciferol), which is the active form.

Vitamin D Requirements and Regulation

• RDA:     * Standard: $400\text{ IU}$ ($10\,\mu g$).     * >60\text{ yrs}: $800\text{ IU}$ ($20\,\mu g$).     * Therapeutic: Up to $2000\text{ IU}$ for treating osteoporosis.

• Regulation of Active Form ($1,25(OH)_2D$):     * Stimulants: Parathyroid Hormone ($PTH$) and hypophosphatemia increase $1\alpha$-hydroxylase activity. Hypocalcemia acts indirectly by stimulating $PTH$ secretion.     * Inhibitors: Hypercalcemia, hyperphosphatemia, and $1,25(OH)_2D$ itself (negative feedback).     * Metabolism: $1,25(OH)_2D$ induces $24$-hydroxylase, converting $25(OH)D$ to $24,25(OH)_2D$ (the most prevalent dihydroxylated form in serum but biologically less active).

Functions of Vitamin D (Calcitriol)

• Mineral Homeostasis: Maintains calcium and phosphate in serum by acting on:     * Small Intestine: Increases absorption of $Ca^{2+}$ and phosphate. Involves $CaT1$ (epithelial transporter), Calbindin-$D_{9k}$ (diffusion), and $CaATPase$ (exit across basolateral membrane).     * Bone: Dual role in remodeling. High concentrations enhance bone resorption by increasing osteoclast activity; low to normal levels promote bone formation/mineralization by stimulating osteoblasts.     * Kidney: Stimulates calcium resorption in the Distal Convoluted Tubule ($DCT$).

• Hormonal Actions:     * Regulation of gene expression.     * Inhibits synthesis and secretion of $PTH$ (ultra-short feedback loop).     * Increases calcium-sensing receptor concentration in the parathyroid gland.

• Emerging Roles:     * Regulation of immune response and epithelial differentiation.     * Correlation with lower cancer incidence.

Clinical Aspects and Deficiency of Vitamin D

• Medical Correlations:     * Heart Health: Levels <15\,ng/ml double the risk of heart problems; may alleviate hypertension.     * Diabetes: Type 1 DM is more common in areas with less sunlight.     * Cancer: Breast cancer cells have Vitamin D Receptors ($VDR$); activation can slow growth or kill cancer cells.     * PAD: Protection against peripheral artery disease; low levels increase risk of leg pain, numbness, and amputation.

• Deficiency Syndromes:     * Rickets (Children): Poor $Ca^{2+}$ absorption leads to undermineralized bone. Symptoms include bow legs, knock knees, swollen joints (epiphyses), "hot-cross bun" head appearance (delayed fontanelle closure), rachitic rosary (beaded ribs), pigeon breast (pectus carinatum), and late teeth eruption.     * Osteomalacia (Adults): Bone demineralization. Common in women with low sunlight exposure, multiple pregnancies, or those observing purdah.

Vitamin D Toxicity

• Soft Tissue Calcification: Occurs in lungs, heart, and blood vessels.

• Arterial Hardening: Calcification of the arteries.

• Hypercalcemia: Elevation of blood calcium above normal (~$10\,mg/dl$).

• Renal Stones: Excess calcium leads to kidney stone formation.

• Systemic Symptoms: Lack of appetite, excessive thirst, excessive urination, and constipation.