Vitamin K What it is Early this century research workers investigating various haemorrhagic problems discovered that many affected animals had deficiencies of prothrombin essential for blood coagulation. Then, in 1929, Henrik Dam discovered a dietary anti-haemorrhagic factor which was responsible for maintaining plasma prothrombin levels. He went on to show that a deficiency of this nutritional factor, vitamin K, was responsible for the reduction of prothrombin levels and the occurrence of haemorrhagic problems. In 1939 a highly purified form of the vitamin was isolated from vegetable sources and identified as phylloquinone, one of the naphthoquinones. Subsequent investigations established that other naphthoquinones have vitamin K activity. In particular, substances containing methyl naphthoquinone (or menadione) are vitamin K-active. The natural form of the vitamin, phylloquinone, is now designated vitamin K1 and the menadione compounds are all forms of vitamin K3. Soon after the discovery of the biological activity of menadione it was suggested that menadione is a provitamin which is alkylated after absorption into a prenyl menaquinone. Such prenyl menaquinones had previously been isolated from putrefying plant and animal products and had been labelled vitamin K2. What it does There are three stages in blood coagulation. First is the blood platelet adhesion reaction to form a platelet plug. Then follows the platelet release reaction which produces substances to constrict the blood vessel and trigger coagulation. The third, and final, phase is the formation of a fibrin clot from the conversion of soluble fibrinogen. Prothrombin in the blood is activated by the products of tissue injury to become thrombin which converts fibrinogen to fibrin. There are also some thirteen additional factors which speed up the process. The synthesis of prothrombin and factors VII, IX and X are all vitamin K-dependent. An unusual feature common to all these four proteins is the presence of g-carboxyglutamic acid, a very specific amino acid capable of binding Ca++ ions. So far no other function of vitamin K has been discovered. If too much is given Injections containing more than 2mg vitamin K3 per kg bodyweight induced renal toxicosis in healthy racehorses. They had been given these extremely large amounts to try to control exercise-induced pulmonary haemorrhage. Vitamin K3 given as a feed supplement could induce similar problems but more than 1 g/kg feed would be needed. Clinical signs in affected horses include renal colic, blood in the urine and electrolyte abnormalities. Biosynthesis Many unicellular organisms appear to be able to manufacture vitamin K, usually as K1 or K2. While synthesis in the caecum enhance the amount of vitamin K available for intestinal absorption there is considerable doubt whether caecal synthesis in horses leads to any increased uptake. How it is measured The only real measurement of vitamin K activity is the biological response in prothrombin production. Chicks are used for this time-consuming estimation. Most chick assay results suggest that the vitamin K activities of menadione and phylloquinone compounds are reasonably similar when compared on a gram molecular basis. Compound Weight of 1 gram molecule Equivalence (Phylloquinone = 1) Phylloquinone 451 1,00 Menadione 172 0,38 Menadione sodium bisulphite (MSB) 330 0,73 Menadione sodium bisulphite complex (MSBC) 538 1,19 Menadione dimethyl pyrimidinol bisulphite (MPB) 379 0,84 This table shows that 0.73 g MSB or 0.84 g MPB should have similar vitamin K activity to 1 g vitamin K1. Assessment of status It is practically impossible to assess the vitamin K status of a horse from determinations of vitamin K1, K2 or K3 in blood or liver tissue samples. The only method of determining the existence of a possible vitamin K deficiency is to measure the prothrombin clotting time (PCT). Any significant increase in PCT suggests vitamin K deficiency. Antagonists Some of the early research work on vitamin K was inspired by finding haemorrhagic problems after cattle were fed hay containing sweet clover. Later it was found that sweet clover contains a derivative of coumarin, the organic material responsible for the smell of new mown hay. Several coumarin derivatives are now known to antagonise the activity of vitamin K. High concentrations are found in sweet clover and sweet vernal grass. There is a risk, therefore, that horses fed hay containing these grasses could develop vitamin K-responsive haemorrhagic conditions. This antagonistic effect is used commercially in the production of rodent poisons such as warfarin which cause death by internal haemorrhage by interfering with vitamin K-dependent prothrombin production. Several of the sulpha drugs such as sulphaquinoxaline also antagonise vitamin K activity and prothrombin production. In the event of such antagonism, the menadione compounds (vitamin K3) are almost ineffective. Phylloquinone (vitamin K1) is the only effective antidote to coumarin poisoning. Requirements and allowances Since it is difficult to assess vitamin K status it is exceptionally difficult to determine accurate requirements and allowances for this vitamin. It is generally assumed that horses benefit from vitamin K supplements. The true requirement level is probably about 0.5-1 mg vitamin K3 per kg feed but the inherent instability of these products makes it essential to increase allowances substantially. If feeds are likely to contain any vitamin K antagonists a source of vitamin K1 is required. Young horses, mares and stallions generally need more vitamin K than adults and it is always advisable to ensure that performance horses have adequate vitamin K to ensure maximum healing ability. Recommendations for feed supplementation cannot be regarded as absolute because of the variations in processing conditions. They allow for losses, and any source of vitamin K (related to actual menadione content) can be used. Product Menadione Content MSB 52% MSBC 33% MPB 46% Generally, feeds for horses of all ages should be supplemented with 1 mg menadione per kg using one of the above compounds or a stabilised commercial derivative. Contents of feed ingredients A number of leafy materials contain relatively large amounts of K1 so that grazing horses should be adequately supplied. Animal products, with the exceptions of liver and egg yolks, are generally very poor sources. Dried or processed vegetables lose much of their vitamin K-activity during processing. Most compound feeds contain less than 1 mg vitamin K per kg. Stability Vitamin K1, K2 and menadione are stable in dry heat but are labile to oxidation, alkali, strong acids, light, humidity and irradiation. All forms of manufactured vitamin K3 are affected by feed processing. Pelleting with steam is particularly aggressive. Cubed or pelleted feed requires 50% more vitamin K than meal mixes. Some forms of vitamin K3 are said to be more resistant to pelleting than others. Research results are very contradictory and there is no clear choice for ag- gressive situations. Therefore it is best to provide 50% more menadione (in the form of one of the vitamin K3 analogues) than specified for the supplement to allow for losses.