Discovery of Vitamin K and It’s Targets and Antagonists

Henrik Dam and Edward Doisy were awarded the 1943 Nobel Prize for Physiology or Medicine. Dam discovered vitamin K and Doisy isolated two forms of vitamin K. Vitamin K was discovered by Henrik Dam while carrying out experiments to study cholesterol metabolism in chicken.  Evidence available in the early twentieth century suggested that chicken unlike rats, mice and dog could not synthesize cholesterol. Henrik Dam fed chickens with artificial cholesterol free diet to see if they could synthesize cholesterol. Chicks developed haemorrhage after 2-3 weeks of cholesterol free diet. Their blood did not clot well. Dam supplemented the diet to determine the component of cholesterol free diet responsible for the haemorrhage.

  1. supplementing the diet with pure cholesterol did not stop the haemorrhages indicating that the lack of cholesterol was not the cause of bleeding
  2. supplementing with  linseed oil or triolin did not prevent the symptoms indicating that low fats were not the cause of the symptoms
  3. Chicken does not need vitamin C indicating that the bleeding was not because of scurvy

Holst and Halbrook at the University of California corrected the bleeding caused by cholesterol free diet by feeding chicken cabbage. They questioned the claims that chicken do not need vitamin C and proposed that it was the vitamin C in the cabbage that corrected the bleeding. Dam was able to disprove this hypothesis by demonstrating the failure of injectable vitamin C to correct bleeding. In 1934 Dam proposed that the bleeding was because of an unrecognized dietary factor, in 1935 he called it vitamin K (K from “koagulation” the Scandinavian and German for clotting) an in 1939 along with Karrar and co-workers made the first pure preparation of vitamin K (Henrik Dam’s Noble Prize Lecture).

In 1920 Northern US and Canada has an epidemic of a bleeding disease in cattle. Cows were bleeding spontaneously or following minor trauma. Twelve of the twenty-five bulls subjected to castration died of bleeding. The following year the disease was found to be due to ingestion of fodder containing moldy sweet clover. Unspoiled sweet clover was not found to cause the disease. The disease was demonstrated to be due to the deficiency of prothrombin in 1929. The substance causing the bleeding disease, 3,3′-methylenebis-(4-hydroxycoumarin) or dicumarol,  was not isolated until 1940. Dicumarol was the first oral anticoagulant to be used.

The first four letters of Warfarin are an acronym for Wisconsin Alumni Research Foundation that funded the research into the molecule. Warfarin was not developed as an oral anticoagulant but as a rodenticide. It was an unsuccessful suicide attempt by an US army recruit that was responsible for introduction of warfarin as an oral anticoagulant.  Warfarin was found to be more potent than dicumarol and has completely replaced the latter as the preferred oral anticoagulant.  4-Hydroxycoumarin derivatives that have short half-lives are used as oral anticoagulants as the effects are easy to titrate. These include warfarin, acenocoumarol  and phenprocoumon. Superwarfarins,  which have a substantially longer (weeks to months) half-life than warfarin,  are used as rodenticides. The suerwarfarins include difenacoum, flocoumafen, bromadiolone, tioclomarol and brodifacoum.

It became apparent early in the development of oral anticoagulants that their actions were antagonized by vitamin K. The function of vitamin K was however not discovered until 1974. In this year three laboratories reported that prothrombin from cows treated with warfarin had 10 glutamic acid residues at the amino end that were replaced by an unusual amino acid, gamma carboxy glutamic acid. Vitamin K was found to be a cofactor in the gamma carboxylation of glutamate that was essential for the action of prothrombin. Prothrombin is not the only protein that undergoes a post-translational modification. Others include coagulations factors (factors VII, IX, X), inhibitors of coagulation (protein C, protein S), proteins involved in bone mineralization (osteocalcin, matrix Gla protein (MGP), and protein S) and cell growth regulation (Gas6). Many non-coagulation vitamin K dependent proteins have been identified by the presence of gamma-carboxyl glutamic acid.

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