Biochemistry, Fat Soluble Vitamins

Fundamentals

Sources of Vitamin A

In animals, the body stores vitamin A as a molecule called retinol. Egg yolk, milk, liver, cheese, and butter are all rich in vitamin A. We derive vitamin A from plant sources in the form of plant carotenoids, which convert to retinol during digestion. Plants abundant in vitamin A include dark green leafy vegetables (spinach, amaranth, among others), carrots, squash, yellow maize, mangoes, and papayas.[1]

Sources of Vitamin D

Vitamin D is found primarily in 2 forms, D2 and D3. Vitamin D2 (ergocalciferol) is present in certain foods such as salmon, tuna, and mackerel. Smaller quantities are present in beef liver, cheese, and egg yolks. Many countries fortify natural milk with vitamin D. This practice is implemented to decrease the prevalence of rickets and osteomalacia. Vitamin D3 (cholecalciferol) is synthesized in the skin after exposure to sunlight, hence its nickname the "sunshine vitamin."

Sources of Vitamin E

The predominant form of vitamin E is a-tocopherol. However, other tocopherols and tocotrienols are also in circulation, such as the alpha, gamma, beta, and delta forms. Naturally occurring sources of vitamin E include vegetable oils, seeds, nuts, and whole grains.[2]

Sources of Vitamin K

Vitamin K has 2 primary forms, K1 and K2. Vitamin K1 (phylloquinone) is in green leafy vegetables, cabbage, and cauliflower. Lesser quantities are in fish, meat, and some fruits. The gut microflora synthesizes vitamin K2 (menaquinone).[3]

Cellular Level

Despite structural differences between fat-soluble vitamins, they are absorbed and transported similarly due to their low solubility in hydrophilic media. The body absorbs fat-soluble vitamins into newly forming micelles in the small intestine. Micelles are lipid clusters that contain hydrophobic groups internally and hydrophilic groups externally. This process relies on the secretion of bile and pancreatic enzymes. After absorption into enterocytes, fat-soluble vitamins become packaged into chylomicrons, which then get secreted into the lymphatic system before entering the bloodstream. Chylomicrons are metabolized by lipoprotein lipase, which causes the release of fat-soluble vitamins into tissues for use and storage.

Because they are stored in tissue, the fat-soluble vitamins are retained by the body for a longer time than the water-soluble vitamins. Remnants of the chylomicron are then taken back up by the liver and recycled. Alpha-tocopherol is targeted into lipoproteins in the liver by a specific tocopherol transfer protein (TTP), mutations of which can result in vitamin E deficiency.[4]

Function

Vitamin A plays an integral role in the differentiation and proliferation of epithelial cells in the eyes, salivary glands, and genitourinary tract. Vitamin A is a precursor to the nuclear hormone all-trans retinoic acid, which heterodimerizes with retinoic acid receptors (RAR) in the nucleus. RAR-retinoid X receptor heterodimers serve as transcription factors that bind certain elements in promoters of genes. These genes encode important structural proteins, extracellular matrix proteins, and enzymes throughout the body. Retinal, a component of vitamin A, derives its name from its ability to produce rhodopsin in the retina, thereby aiding in vision, especially in low-light settings. Additionally, vitamin A stimulates T-lymphocyte differentiation and B-lymphocyte activation in response to immune stimuli.[5]

The primary function of vitamin D is to raise plasma calcium and phosphate concentrations, which promotes the mineralization of osteoids in the bone. Elevating calcium levels is necessary for properly functioning the neuromuscular junction, nerve transmission, and secretion and actions of hormones. Vitamin D3 from the skin and vitamin D2 from the diet are prohormones that undergo hydroxylation to 25-hydroxycholecalciferol in the liver via the enzyme 25-hydroxylase. 25-hydroxycholecalciferol becomes further hydroxylated in the kidney to its biologically most active form, 1,25-dihydroxycholecalciferol. Hydroxylation in the kidney into a biologically active form occurs via 1-a-hydroxylase, an enzyme under tight regulation by the parathyroid hormone. The active form of vitamin D increases the duodenal absorption of calcium and phosphate and calcium reabsorption from the distal convoluted tubule by upregulating calcium transporters that move calcium across epithelial cells. Importantly, vitamin D activates osteoclasts, our body’s bone-resorbing cells. The human body maintains equilibrium with bone formation and resorption. To effectively mineralize bone, some level of bone resorption is necessary.[6] 

Vitamin E, exclusively acquired from the diet, is best known for its antioxidant activity. Vitamin E inhibits the generation of reactive oxygen species during fat oxidation. It protects polyunsaturated fatty acids in cell membranes from oxidative destruction, maintaining membrane fluidity and stability. While it inhibits lipid peroxidation, including oxidation of LDL, supplementation has not resulted in a reduction in cardiovascular events.[7]

Vitamin K is necessary to activate certain clotting factors in the liver responsible for coagulation. For activation to occur, the clotting proteins must bind calcium. Vitamin K-dependent gamma-carboxylation of certain glutamic acid residues allows the proteins to bind calcium and carry out the coagulation cascade. Specifically, vitamin K serves as a cofactor for gamma-glutamyl carboxylase and catalyzes the post-translational synthesis of gamma-carboxy-glutamyl residues. This process activates prothrombin and factors VII, IX, X, protein C, and S. Oxidation of vitamin K hydroquinone supplies energy for these carboxylation reactions. Regeneration of vitamin K hydroquinone relies on vitamin K epoxide reductase and vitamin K quinone reductase.

Testing

Vitamin A

Vitamin A levels are tested by measuring serum retinol. Testing is indicated in patients who exhibit signs of vitamin-A deficiency, such as night blindness, xerophthalmia, and Bitot spots. The World Health Organization (WHO) states that serum retinol accurately estimates hepatic vitamin A stores in extreme deficiency and excess states. Low serum retinol levels are considered to be less than 0.70 micromole/L and can evaluate the extent of vitamin A deficiency. Plasma retinol levels in vitamin A-toxicity are generally greater than 3.5 micromole/L.[8] The recommended dietary allowance (RDA) of vitamin A is 900 microgram retinol activity equivalents in men and 700 microgram retinol activity equivalents in women. 

Vitamin D

Vitamin D is measurable in the serum in 25-hydroxyvitamin D and 1,25-dihydroxy vitamin D. 25-hydroxyvitamin D is the principal circulating form (levels in ng/ml). It has a half-life of 2 weeks and is the best measure of vitamin D status. Testing is indicated in populations at high risk for fractures, including those with osteoporosis, osteopenia, and the elderly. Treatment initiation depends on the extent of the deficiency. The Endocrine Society states that levels of 25-hydroxyvitamin D below 20 ng/ml are considered insufficient. According to The US Institute of Medicine, a 25-hydroxyvitamin D level of 20 ng/mL or greater is optimal for bone strength. Levels exceeding 100 ng/ml are considered toxic and put the patient at risk for hypercalcemia, calculi, and renal damage. Hence, optimum levels appear to be between 20 to 30 ng/ml, with levels over 50 ng/ml avoided. Testing levels of 1,25-hydroxyvitamin D may also be indicated in patients with kidney failure or suspected hypercalcemia from granulomas in sarcoidosis or suspected hyperparathyroidism. Low levels of the active metabolite are commonly observed in early renal failure, while increased levels may present in sarcoidosis and primary hyperparathyroidism. The RDA for vitamin D is 600 IU per day. In individuals older than 70 years, the RDA is 800 IU per day.

Vitamin E

The best indicator of the vitamin E level is serum alpha-tocopherol. The recommended level of vitamin E ranges from 5 to 17 micrograms/mL in adults and 3 to 18.4 micrograms/mL in children. In patients with hyperlipidemia, it is preferable to report standardized lipid levels. Testing may be necessary for patients suffering from sensorimotor neuropathy or a fat malabsorption disorder such as cystic fibrosis. The RDA for vitamin E is 15 mg per day. 

Vitamin K

According to the Food and Nutrition Board, a healthy intake of Vitamin K ranges from 70 to 140 micrograms daily. Vitamin-K deficiency and toxicity are rare in the US population; thus, dietary vitamin-K testing is not generally indicated. The principal test utilized to evaluate bleeding due to a possible vitamin-K deficiency is prothrombin time (PT). In the case of prolonged PT, vitamin K injections or oral supplements may be necessary. Vitamin K deficiency can be confirmed as the cause of bleeding if PT/INR normalizes in response to the injection or oral supplementation of vitamin K. The RDA for vitamin K in individuals under 6 months is 2 mcg daily. The RDA in adult males and females is 120 mcg per day and 90 mcg per day, respectively.

Pathophysiology

Deficiency

Vitamin A

Although rare in developed nations, vitamin A deficiency is a significant health concern in non-industrialized countries. It is responsible for over 500,000 cases of corneal lesions in children per year. In the United States, vitamin A deficiency most commonly results from fat malabsorption syndromes, alcoholism, and liver disease. Vitamin A uptake can also be impaired by iron deficiency, pancreatic insufficiency, and inflammatory bowel disease. Severe deficiency can lead to various ocular signs, notably night blindness (nyctalopia) and xerophthalmia. Keratin accumulation in the conjunctiva causing Bitot’s spots is a pathognomic physical finding. Other ocular manifestations include conjunctival xerosis, corneal drying and ulceration, and follicular hyperkeratosis.[1][9] Due to the role of vitamin A in T-lymphocyte proliferation and differentiation, deficiency also increases the risk of infections. Vitamin A is an effective treatment for measles, decreasing mortality in children and hospitalized patients.[10] Additionally, by inducing differentiation of acute promyelocytic anemia cells, all-trans retinoic acid (ATRA) is an effective treatment for acute promyelocytic leukemia.[11] In outpatient settings, isotretinoin is a common prescription for treating severe acne vulgaris.

Vitamin D

Vitamin D deficiency has become a global concern with dire health consequences. Common risk factors include old age, exclusively breastfed infants, immobility, reduced kidney function, dark skin, malabsorption syndromes, decreased sunlight exposure, and obesity.[12] Manifestations of deficiency include muscle aches and weakness with bone pain in the back, extremities, and pelvis. In children, vitamin D deficiency leads to impaired cartilage mineralization at growth plates, leading to rickets. Patients with rickets may have a bow-leg deformity, rachitic rosary, stunted growth with short stature, dental deformities, abnormal spinal curvature, craniotabes, and frequent fractures. In adults, low vitamin D levels lead to impaired mineralization of osteoid, leading to osteomalacia. Osteomalacia characteristically demonstrates diffuse bone and joint pain, myopathy, hypocalcemic tetany, and a waddling gait.

Vitamin E

Vitamin-E deficiency is extremely rare and principally occurs in individuals with fat malabsorption, abetalipoproteinemia (defect in microsomal transfer protein), and hypobetalipoproteinemia (mutation in apolipoprotein B) disorders. Deficiency symptoms include limb and truncal ataxia, hyporeflexia, and upward gaze limitations. Rarer manifestations are muscle weakness and constriction of visual fields. If left untreated, deficiency can result in blindness, memory impairment, and arrhythmias.[13] Multiple clinical trials have shown that vitamin-E supplementation decreases histological and biochemical evidence of liver dysfunction in patients with nonalcoholic fatty liver disease (NAFLD) and nonalcoholic steatohepatitis (NASH). The increased inflammation and oxidative stress observed in obesity theoretically heighten vitamin E requirements in this subset of patients. Additionally, these findings raise whether vitamin E deficiency could exacerbate liver dysfunction.[4]

Vitamin K

Vitamin K deficiency is clinically significant due to its prevalence in various patient populations. Risk factors include antibiotic use, which interferes with vitamin K production in the gut, nutritional deficiency, and high ingestion of vitamins A and E. Newborns are also at risk for deficiency due to immature gut flora, poor placental transfer, and low content in breast milk. The risk in newborns becomes further increased with a maternal history of anticonvulsant and anticoagulant use. A common clinical syndrome that results from vitamin-K deficiency is a hemorrhagic disease of the newborn, a life-threatening bleeding condition in neonates. Neonates with this condition present with failure to thrive, low birth weight, and excessive bleeding from the umbilical stump and mucous membranes. They are at higher risk for intracranial hemorrhage. This condition is treated prophylactically through vitamin K injections at birth. In adults, deficiency can also cause easy bleeding and bruising with an elevated PT.

Toxicity

Vitamin A

Vitamin A toxicity is most commonly the result of over-supplementation, wild game liver consumption, and isotretinoin therapy. Hypervitaminosis A leads to intracranial swelling, which manifests as headaches, papilledema, and seizures. Other findings include arthralgias, alopecia, dry mucous membranes, skin desquamation, hypercalcemia, and liver damage. Isoretinoic acid, an acne treatment, is contraindicated in women who are pregnant or may become pregnant due to a risk of spontaneous abortion and birth defects in the fetus.[14]

Vitamin D

Vitamin D toxicity, although rare, can occur in individuals taking large doses of vitamin D supplements with a heavy intake of fortified foods. Most symptoms of hypervitaminosis D stem from hypercalcemia caused by excessive calcium absorption in the duodenum and distal convoluted tubule. Clinical manifestations include gastrointestinal issues such as decreased appetite, diarrhea, nausea, vomiting, and constipation. Hypercalcemia can result in polyuria, polydipsia, pruritus, and the development of kidney stones. Bone, muscle, and joint pain are also common manifestations.[12]

Vitamin E

Hypervitaminosis E is most commonly a result of over-supplementation and is otherwise very rare. Since high doses of Vitamin E (800 mg per day) inhibit platelet aggregation, it is contraindicated in patients on anticoagulants.[15]

Vitamin K

Vitamin K toxicity is uncommon but is more prevalent in formula-fed infants and those receiving menadione injections, a water-soluble synthetic vitamin K precursor. Symptoms of hypervitaminosis K include hemolytic anemia, jaundice in newborns with hyperbilirubinemia, and liver damage.

Clinical Significance

Fat-soluble vitamin deficiencies and toxicities prevention rely on a diverse team of healthcare professionals. The interprofessional team involves physicians, medical assistants, dieticians, pharmacists, and patients. For example, a pediatrician must recognize risk factors for vitamin D deficiency in a neonate, including being exclusively breastfed and having darker skin. Clinicians, who often spend the most time with patients, are in a position to recognize abnormal signs and symptoms in high-risk patients and report their findings to the physician. Their watchful eye is vital in catching alarming symptoms early, such as headaches and seizures caused by vitamin-A toxicity. Dieticians and nutritionists carry out primary and tertiary prevention of fat-soluble vitamin excess or deficiency and play an essential role in modulating a patient’s diet and ensuring the meeting of their dietary needs.[16] 

Pharmacists are responsible for ensuring that other interprofessional team members are aware of the potential risks of medications that may lead to vitamin excess or deficiency. Finally, the physician's role is to follow proper screening protocols, recognize signs and symptoms of fat-soluble vitamin abnormalities early, order the correct labs, and work to coordinate the other members of the patient’s team optimally.