Vitamin K



Vitamin K is a group of structurally similar, fat soluble vitamins that are needed for the posttranslational modification of certain proteins, mostly required for blood coagulation, but also involved in metabolic pathways in bone and other tissue. They are 2-methyl-1,4-naphthoquinone (3-)derivatives. This group of vitamins includes two natural vitamers: vitamin K1 and vitamin K2.

Vitamin K1 is also known as vitamin Kj, phylloquinone or phytomenadione (also called phytonadione). Vitamin K1 is synthesized by plants and is found in green leafy vegetables and can be found in soybean oil.

Vitamin K2 homologs (menaquinones) are characterized by the number of isoprenoid residues comprising the side chain. Menaquinones are abbreviated MK-n, where n represents the number of isoprenoid side chains. Thus, menaquinone-4 abbreviated MK-4, has 4 isoprene residues in the side chain. Bacteria can produce a range of vitamin K2 forms, including the conversion of K1 to K2 by bacteria in the small intestines. No known toxicity exists for vitamins K1 and K2.

Three synthetic types of vitamin K are known: vitamins K3, K4, and K5. Although the natural K1 and K2 forms are nontoxic, the synthetic form K3 (menadione) has shown toxicity.

Vitamin K was identified in 1929 by Danish scientist Henrik Dam when he investigated the role of cholesterol by feeding chickens a cholesterol-depleted diet. After several weeks, the animals developed hemorrhages and started bleeding. These defects could not be restored by adding purified cholesterol to the diet. It appeared that—together with the cholesterol—a second compound had been extracted from the food, and this compound was called the coagulation vitamin. The new vitamin received the letter K because the initial discoveries were reported in a German journal, in which it was designated as Koagulationsvitamin.

Subtypes of vitamin K2
Vitamin K2 (menaquinone), is itself a category of vitamin K that includes many types of vitamin K2. The two subtypes of vitamin K2 that have been most studied are menaquinone-4 (menatetrenone, MK4) and menaquinone-7 (MK7).

MK4 is produced via conversion of vitamin K1 in the body, in the testes, pancreas and arterial walls. While major questions still surround the biochemical pathway for the transformation of vitamin K1 to MK4, studies demonstrate that the conversion is not dependent on gut bacteria, occurring in germ-free rats and in parenterally-administered K1 in rats. In fact, tissues that accumulate high amounts of MK4 have a remarkable capacity to convert up to 90% of the available K1 into MK4.

In contrast to MK4, menaquinone-7 (MK7) is not produced by humans but is converted from phylloquinone in the intestines by gut bacteria. However, bacteria-derived menaquinones (MK7) appear to contribute minimally to overall vitamin K status. MK4 and MK7 are both found in the United States in dietary supplements for bone health.

The US FDA has not approved any form of vitamin K for the prevention or treatment of osteoporosis; however, MK4 has been shown to decrease fractures up to 87%. In the amount of 45 mg daily MK4 has been approved by the Ministry of Health in Japan since 1995 for the prevention and treatment of osteoporosis.

Vitamin K2 (MK4, but not MK7 or vitamin K1) has also been shown to prevent bone loss and/or fractures in the following circumstances:
 * caused by corticosteroids (e.g., prednisone, dexamethasone, prednisolone),
 * anorexia nervosa,
 * cirrhosis of the liver,
 * postmenopausal osteoporosis,
 * disuse from stroke,
 * Alzheimer’s disease,
 * Parkinson disease,
 * primary biliary cirrhosis
 * and leuprolide treatment (for prostate cancer).

Vitamin K absorption and dietary need
Previous theory held that dietary deficiency is extremely rare unless the intestine (small bowel) was heavily damaged, resulting in malabsorption of the molecule. The other at-risk group for deficiency were those subject to decreased production of K2 by normal flora, as seen in broad spectrum antibiotic use. Taking broad-spectrum antibiotics can reduce vitamin K production in the gut by nearly 74% in people compared to those not taking these antibiotics. Diets low in vitamin K also decrease the body's vitamin K concentration. Additionally, in the elderly there is a reduction in vitamin K2 production.

Recent research results also demonstrate that the small intestine and large intestine (colon) seem to be inefficient at absorbing vitamins K.  These results are reinforced by human cohort studies, where a majority of the subjects showed inadequate vitamins K amounts in the body. This was revealed by the presence of large amounts of incomplete gamma-carboxylated proteins in the blood, an indirect test for vitamins K deficiency. And in an animal model MK4 was shown to prevent arterial calcifications, pointing to its potential role in cardiovascular disease prevention. In this study vitamin K1 was also tested and shown to not prevent arterial calfications.

Chemical structure
All members of the vitamin K group of vitamins share a methylated naphthoquinone ring structure (menadione), and vary in the aliphatic side chain attached at the 3-position (see figure 1). Phylloquinone (also known as vitamin K1) invariably contains in its side chain four isoprenoid residues, one of which is unsaturated.

Menaquinones have side chains composed of a variable number of unsaturated isoprenoid residues; generally they are designated as MK-n, where n specifies the number of isoprenoids.

It is generally accepted that the naphthoquinone is the functional group, so that the mechanism of action is similar for all K-vitamins. Substantial differences may be expected, however, with respect to intestinal absorption, transport, tissue distribution, and bio-availability. These differences are caused by the different lipophilicity of the various side chains, and by the different food matrices in which they occur.

There are three synthetic forms of vitamin K, vitamins K3, K4, and K5, which are used in many areas including the pet food industry (vitamin K3) and to inhibit fungal growth (vitamin K5).

Physiology
Vitamin K is involved in the carboxylation of certain glutamate residues in proteins to form gamma-carboxyglutamate residues (abbreviated Gla residues). The modified residues are often (but not always) situated within specific protein domains called Gla domains. Gla residues are usually involved in binding calcium. The Gla residues are essential for the biological activity of all known Gla-proteins.

15 human proteins with Gla domains have been discovered, and they play key roles in the regulation of three physiological processes:
 * Blood coagulation: (prothrombin (factor II), factors VII, IX, X, protein C, protein S, and protein Z).
 * Bone metabolism: osteocalcin, also called bone Gla-protein (BGP), matrix Gla protein (MGP), and periostin.
 * Vascular biology: growth arrest-specific protein 6 (Gas6)
 * unknown function: proline-rich g-carboxy glutamyl proteins (PRGPs) 1 and 2, and transmembrane g-carboxy glutamyl proteins (TMGs) 3 and 4.

Like other liposoluble vitamins (A, D, E), vitamin K is stored in the fat tissue of the human body.

Recommended amounts
The U.S. Dietary Reference Intake (DRI) for an Adequate Intake (AI) of vitamin K for a 25-year old male is 120 micrograms/day. The Adequate Intake (AI) for adult women is 90 micrograms/day, for infants is 10–20 micrograms/day, for children and adolescents 15–100 micrograms/day. In 2002 it was found that to get maximum carboxylation of osteocalcin, one may have to take up to 1000 μg of vitamin K1.

Toxicity
Although allergic reaction from supplementation is possible, there is no known toxicity associated with high doses of the phylloquinone (vitamin K1) or menaquinone (vitamin K2) forms of vitamin K and therefore no tolerable upper intake level (UL) has been set.

Blood clotting (coagulation) studies in humans using 45 mg per day of vitamin K2 (as MK4) and even up to 135 mg/day (45 mg three times daily) of K2 (as MK4), showed no increase in blood clot risk. Even doses in rats as high as 250 mg/kg body weight did not alter the tendency for blood-clot formation to occur.

However, a synthetic form of vitamin K, vitamin K3 (menadione), is demonstrably toxic. In fact, the FDA has banned this synthetic form of the vitamin from over-the-counter supplements because large doses have been shown to cause allergic reactions, hemolytic anemia, and cytotoxicity in liver cells.

Drug interactions
Phylloquinone (K1) or menaquinone (K2) are capable of blocking the blood thinning action of anticoagulants like warfarin, which work by interfering with the action of vitamin K.  They also reverse the tendency of these drugs to cause arterial calcification in the long term.

Deficiency
Average diets are usually not lacking in vitamin K and primary vitamin K deficiency is rare in healthy adults. Newborn infants are at an increased risk of deficiency. Other populations with an increased prevalence of vitamin K deficiency include individuals who suffer from liver damage or disease (e.g. alcoholics), people with cystic fibrosis, inflammatory bowel diseases or those who have recently had abdominal surgeries. Groups that may suffer from secondary vitamin K deficiency include bulimics, those on stringent diets, and those taking anticoagulants. Other drugs that have been associated with vitamin K deficiency include salicylates, barbiturates, and cefamandole, although the mechanism is still unknown. There is no difference between the sexes as both males and females are affected equally. Symptoms of deficiency include heavy menstrual bleeding in women, anemia, bruising, and bleeding of the gums or nose. They could also have disorders such as coagulopathy.

Osteoporosis and coronary heart disease  are strongly associated with lower levels of K2 (menaquinone). Menaquinone is not inhibited by salicylates as happens with K1, so menaquinone supplementation can alleviate the chronic vitamin K deficiency caused by long term aspirin use.

Function
The function of vitamin K in the cell is to convert glutamate in proteins to gamma-carboxyglutamate (Gla).

Within the cell, vitamin K undergoes electron reduction to a reduced form of vitamin K (called vitamin K hydroquinone) by the enzyme vitamin K epoxide reductase (or VKOR). Another enzyme then oxidizes vitamin K hydroquinone to allow carboxylation of Glu to Gla; this enzyme is called the gamma-glutamyl carboxylase or the vitamin K-dependent carboxylase. The carboxylation reaction will only proceed if the carboxylase enzyme is able to oxidize vitamin K hydroquinone to vitamin K epoxide at the same time; the carboxylation and epoxidation reactions are said to be coupled reactions. Vitamin K epoxide is then re-converted to vitamin K by vitamin K epoxide reductase. The reduction and subsequent re-oxidation of vitamin K coupled with carboxylation of Glu is called the vitamin K cycle. One of the reasons humans are rarely deficient in vitamin K is that vitamin K is continually recycled in our cells.

Warfarin and other coumarin drugs block the action of the vitamin K epoxide reductase. This results in decreased concentrations of vitamin K and vitamin K hydroquinone in the tissues, such that the carboxylation reaction catalyzed by the glutamyl carboxylase is inefficient. This results in the production of clotting factors with inadequate Gla. Without Gla on the amino termini of these factors, they no longer bind stably to the blood vessel endothelium and cannot activate clotting to allow formation of a clot during tissue injury. As it is impossible to predict what dose of warfarin will give the desired degree of suppression of the clotting, warfarin treatment must be carefully monitored to avoid over-dosing. (See the warfarin article.)

Gamma-carboxyglutamate proteins, or Gla-proteins
At present, the following human Gla-containing proteins have been characterized to the level of primary structure: the blood coagulation factors II (prothrombin), VII, IX, and X, the anticoagulant proteins C and S, and the Factor X-targeting protein Z. The bone Gla-protein osteocalcin, the calcification inhibiting matrix Gla protein (MGP), the cell growth regulating growth arrest specific gene 6 protein (Gas6), and the four transmembrane Gla proteins (TMGPs) the function of which is at present unknown. Gas6 can function as a growth factor that activates the Axl receptor tyrosine kinase and stimulates cell proliferation or prevents apoptosis in some cells. In all cases in which their function was known, the presence of the Gla residues in these proteins turned out to be essential for functional activity.

Gla-proteins are known to occur in a wide variety of vertebrates: mammals, birds, reptiles, and fish. The venom of a number of Australian snakes acts by activating the human blood clotting system. Remarkably, in some cases activation is accomplished by snake Gla-containing enzymes that bind to the endothelium of human blood vessels and catalyze the conversion of procoagulant clotting factors into activated ones, leading to unwanted and potentially deadly clotting.

Another interesting class of invertebrate Gla-containing proteins is synthesized by the fish-hunting snail Conus geographus. These snails produce a venom containing hundreds of neuro-active peptides, or conotoxins, which is sufficiently toxic to kill an adult human. Several of the conotoxins contain 2–5 Gla residues.

Methods of assessment
Prothrombin time test:
 * Measures the time required for blood to clot
 * Blood sample mixed with citric acid and put in a fibrometer.
 * Delayed clot formation indicates a deficiency.

Unfortunately insensitive to mild deficiency as the values do not change until the concentration of prothrombin in the blood has declined by at least 50 percent

Plasma Phylloquinone:
 * Was found to be positively correlated with phylloquinone intake in elderly British women, but not men

However an article by Schurges et al. reported no correlation between FFQ and plasma phylloquinone

Urinary γ-carboxyglutamic acid:
 * Urinary Gla responds to changes in dietary vitamin K intake.
 * Several days are required before any change can be observed.

In a study by Booth et al. increases of phylloquinone intakes from 100 μg to between 377–417 μg for 5 days did not induce a significant change Response may be age-specific

Function in bacteria
Many bacteria, such as Escherichia coli found in the large intestine, can synthesize vitamin K2 (menaquinone-7), but not vitamin K1 (phylloquinone). In these bacteria, menaquinone will transfer two electrons between two different small molecules, in a process called anaerobic respiration. For example, a small molecule with an excess of electrons (also called an electron donor) such as lactate, formate, or NADH, with the help of an enzyme, will pass two electrons to a menaquinone. The menaquinone, with the help of another enzyme, will in turn transfer these 2 electrons to a suitable oxidant, such fumarate or nitrate (also called an electron acceptor). Adding two electrons to fumarate or nitrate will convert the molecule to succinate or nitrite + water, respectively. Some of these reactions generate a cellular energy source, ATP, in a manner similar to eukaryotic cell aerobic respiration, except that the final electron acceptor is not molecular oxygen, but say fumarate or nitrate (In aerobic respiration, the final oxidant is molecular oxygen (O2), which accepts four electrons from an electron donor such as NADH to be converted to water.)  Escherichia coli can carry out aerobic respiration and menaquinone-mediated anaerobic respiration.

Vitamin K injection in newborns
The blood clotting factors of newborn babies are roughly 30 to 60 percent that of adult values; this may be due to the reduced synthesis of precursor proteins and the sterility of their guts. Human milk contains between 1 and 4 micrograms/litre of vitamin K1, while formula derived milk can contain up to 100 micrograms/litre in supplemented formulas. Vitamin K2 concentrations in human milk appear to be much lower than those of vitamin K1. It is estimated that there is a 0.25 to 1.7 percent occurrence of vitamin K deficiency bleeding in the first week of the infant's life with a prevalence of 2-10 cases per 100,000 births. Premature babies have even lower levels of the vitamin and are at a higher risk from this deficiency.

Bleeding in infants due to vitamin K deficiency can be severe, leading to hospitalizations, blood transfusions, brain damage and death. Supplementation with vitamin K can prevent most cases of vitamin K deficiency bleeding in the newborn. Intramuscular administration of vitamin K is more effective in preventing late vitamin K deficiency bleeding than oral administration.

USA
As a result of the occurrences of vitamin K deficiency bleeding, the Committee on Nutrition of the American Academy of Pediatrics has recommended that 0.5 to 1.0 mg vitamin K1 be administered to all newborns shortly after birth.

UK
In the UK, vitamin K is administered to newborns as either a single injection at birth or three orally administered doses given at birth and then over the baby's first month.

Controversy
Controversy arose in the early 1990s regarding this practice when two studies were shown suggesting a relationship between parenteral administration of vitamin K and childhood cancer (14). However, poor methods and small sample sizes led to the discrediting of these studies and a review of the evidence published in 2000 by Ross and Davies found no link between the two.

Vitamin K and bone health
There is physiological and observational evidence that vitamin K plays a role in bone growth and the maintenance of bone density, but efforts to delay the onset of osteoporosis by vitamin K supplementation have proven ineffective.

Vitamin K takes part in the post translational modification as a cofactor in γ-carboxylation of vitamin K dependant proteins (VKDPs). VKDPs have glutamate residues (Glu). Biophysical studies have suggested that supplemental vitamin K promotes osteotrophic processes and slows osteoclastic processes via calcium bonding. Study of Atkins et al. revealed that phylloquinone, menatetrenone (MK4) and menadione promote in vitro mineralisaton by human primary osteoblasts. Other studies have shown that vitamin K antagonists (usually a class of anticoagulants) lead to early calcification of the epiphysis and epiphysial line in mice and other animals, causing seriously decreased bone growth. This is due to defects in osteocalcin and matrix Gla protein. Their primary function is to prevent overcalcification of the bone and cartilage. Vitamin K is important in the process of carboxylating glutamic acid (Glu) in these proteins to gamma-carboxyglutamic acid (Gla), which is necessary for their function. Vitamin D is reported to regulate the OC transcription by osteoblast thereby showing that vitamin K and vitamin D work in tandem for the bone metabolism and development. Lian and his group discovered two nucleotide substitution regions which they named "osteocalcin box" in the rat and human osteocalcin genes. They found a region 600 nucleotides immediately upstream from the transcription start site that support a 10-fold stimulated transcription of the gene by 1,25-dihydroxy vitamin D.

Vitamin K1 and bone health
Data from the 1998 Nurses Health Study, an observational study, indicated an inverse relationship between dietary vitamin K1 and the risk of hip fracture. After being given 110 micrograms/day of vitamin K, women who consumed lettuce one or more times per day had a significantly lower risk of hip fracture than women who consumed lettuce one or fewer times per week. In addition to this, high intakes of vitamin D but low intakes of vitamin K were suggested to pose an increased risk of hip fracture. The Framingham Heart Study is another study that showed the similar result. Subjects in the highest quartile of vitamin K1 intake (median K1 intake of 254 μg/ day) has 35% lower risk of hip fracture than those in the lowest quartile. Comparing with the daily recommended intake (DRI) of 90 and 120 μg/ day, both the above intakes are higher than existing DRI.

In the face of this evidence, a large multicentre randomized placebo-controlled trial was performed to test the supplementation of vitamin K in post-menopausal women with osteopenia. Despite heavy doses of vitamin K1, no differences were found in bone density between the supplemented and placebo groups.

Vitamin K2 (MK4) and bone health
In contrast, MK4 has been shown in numerous studies to reduce fracture risk, stop and reverse bone loss. In Japan, MK4 in the dose of 45 mg daily is recognized as a treatment for osteoporosis. MK4 has been shown to decrease fractures up to 87%. In the amount of 45 mg daily MK4 has been approved by the Ministry of Health in Japan since 1995 for the prevention and treatment of osteoporosis.

MK4 (but not MK7 or vitamin K1) prevented bone loss and/or fractures in the following circumstances:
 * caused by corticosteroids (e.g., prednisone, dexamethasone, prednisolone),
 * anorexia nervosa,
 * cirrhosis of the liver,
 * postmenopausal osteoporosis,
 * disuse from stroke,
 * Alzheimer’s disease,
 * Parkinson disease,
 * primary biliary cirrhosis and
 * leuprolide treatment (for prostate cancer).

Vitamin K2 (MK7) and bone health
Menaquinone-7 (MK7), which is abundant in fermented soybeans (natto), has been demonstrated to stimulate osteoblastic bone formation and to inhibit osteoclastic bone resorption. In another study, use of MK-7 caused significant elevations of serum Y-carboxylated osteocalcin concentration, a biomarker of bone formation. MK-7 also completely inhibited a decrease in the calcium content of bone tissue by inhibiting the bone-resorbing factors parathyroid hormone and prostaglandin E2. On 19 February 2011, HSA (Singapore) approved a health supplement which contains vitamin K2 (MK7) and vitamin D3 for increasing bone mineral density.

Vitamin K and Alzheimer's disease
Research into the antioxidant properties of vitamin K indicates that the concentration of vitamin K is lower in the circulation of carriers of the APOE4 gene, and recent studies have shown its ability to inhibit nerve cell death due to oxidative stress. It has been hypothesized that vitamin K may reduce neuronal damage and that supplementation may hold benefits to treating Alzheimer's disease, although more research is necessary in this area.

Vitamin K used topically
Vitamin K may be applied topically, typically as a 5% cream, to diminish postoperative bruising from cosmetic surgery and injections, broken capillaries (spider veins), to treat rosacea and to aid in the fading of hyperpigmentation and dark under-eye circles.

Vitamin K and cancer
While researchers in Japan were studying the role of vitamin K2 as the menaqunone-4 (MK-4) form in the prevention of bone loss in females with liver disease, they discovered another possible effect. This two year study which involved 21 women with viral liver cirrhosis found that women in the supplement group were 90 percent less likely to develop liver cancer. A German study performed on men with prostate cancer found a significant inverse relationship between vitamin K2 consumption and advanced prostate cancer.

Vitamin K as antidote for poisoning by 4-hydroxcoumarin drugs
Vitamin K is a true antidote for poisoning by 4-hydroxycoumarin anticoagulant drugs (sometimes loosely referred to as coumarins). These include the pharmaceutical warfarin, and also anticoagulant-mechanism poisons such as bromadiolone, which are commonly found in rodenticides. 4-Hydroxycoumarin drugs possess anticoagulatory and rodenticidal properties because they inhibit vitamin K-dependent synthesis of some clotting factors by the liver. Death is usually a result of internal hemorrhage. Treatment usually consists of repeated intravenous doses of vitamin K, followed by doses in pill form for a period of at least two weeks, though possibly up to 2 months, afterwards (in the case of the more potent 4-hydoxycoumarins used as rodenticides). If caught early, prognosis is good, even when great amounts of the drug or poison are ingested.

History of discovery
In 1929, Danish scientist Henrik Dam investigated the role of cholesterol by feeding chickens a cholesterol-depleted diet. After several weeks, the animals developed hemorrhages and started bleeding. These defects could not be restored by adding purified cholesterol to the diet. It appeared that—together with the cholesterol—a second compound had been extracted from the food, and this compound was called the coagulation vitamin. The new vitamin received the letter K because the initial discoveries were reported in a German journal, in which it was designated as Koagulationsvitamin. Edward Adelbert Doisy of Saint Louis University did much of the research that led to the discovery of the structure and chemical nature of vitamin K. Dam and Doisy shared the 1943 Nobel Prize for medicine for their work on vitamin K (K1 and K2) published in 1939. Several laboratories synthesized the compound(s) in 1939.

For several decades the vitamin K-deficient chick model was the only method of quantifying vitamin K in various foods: the chicks were made vitamin K-deficient and subsequently fed with known amounts of vitamin K-containing food. The extent to which blood coagulation was restored by the diet was taken as a measure for its vitamin K content. Three groups of physicians independently found this: Biochemical Institute, University of Copenhagen (Dam and Johannes Glavind), University of Iowa Department of Pathology (Emory Warner, Kenneth Brinkhous, and Harry Pratt Smith), and the Mayo Clinic (Hugh Butt, Albert Snell, and Arnold Osterberg).

The first published report of successful treatment with vitamin K of life-threatening hemorrhage in a jaundiced patient with prothrombin deficiency was made in 1938 by Smith, Warner, and Brinkhous.

The precise function of vitamin K was not discovered until 1974, when three laboratories (Stenflo et al., Nelsestuen et al., and Magnusson et al. ) isolated the vitamin K-dependent coagulation factor prothrombin (Factor II) from cows that received a high dose of a vitamin K antagonist, warfarin. It was shown that while warfarin-treated cows had a form of prothrombin that contained 10 glutamate amino acid residues near the amino terminus of this protein, the normal (untreated) cows contained 10 unusual residues that were chemically identified as gamma-carboxyglutamate, or Gla. The extra carboxyl group in Gla made clear that vitamin K plays a role in a carboxylation reaction during which Glu is converted into Gla.

The biochemistry of how vitamin K is used to convert Glu to Gla has been elucidated over the past thirty years in academic laboratories throughout the world.