Vitamin B6 What it is Research work in 1926 on nutritionally-induced skin problems in rats produced the first identifiable deficiency of vitamin B6. Its effects were not separated from others of the B-complex until 1934. Extensive experiments during the later 1930’s showed that this ”new” vitamin was essential both for micro-organisms and all higher forms of life. The pure substance was finally isolated in 1938 and given the name adermin because it was thought to be involved only in the skin. This name was abandoned in 1944 when it was synthesised for the first time and found to affect the central nervous system and produce changes in blood. The chemical synthesised in 1944 was named pyridoxine because it was found to be an alcohol of pyridine. Two other compounds similar in structure to pyridoxine also have vitamin B6 activity. These are pyridoxal and pyridoxamine. They differ from each other only in the chemical group attached to the pyridine ring. Only pyridoxine is found in plants whereas animal products may contain pyridoxal and pyridoxamine as well. The vitamin B6 activity of a feed is the total content of all three substituted pyridine compounds. What it does After phosphorylation, pyridoxal and pyridoxamine are incorporated into various enzyme systems as pyridoxal 5’-phosphate (PALP). These enzymes are involved in a large number of metabolic processes. Although research has been very extensive the multiple functions of vitamin B6 through PALP are still not yet fully understood; over fifty enzymes are already known which depend on it. One of the functions of PALP which has been investigated is its activity as a coenzyme factor in a series of enzymes involved with the metabolism of amino acids. These include transaminases, decarboxylases, desulphydrases and dehydrogenases. The transaminases, for example, function by transferring amino groups in the form of pyridoxamine. One of the products of these reactions is aspartic acid. Vitamin B6, as a precursor of PALP, is an essential factor in energy production, (supplying metabolites to the Kreb’s cycle), fat metabolism, protein breakdown and build-up, central nervous system activity and haemoglobin production. Another involvement of vitamin B6 is in the synthesis of globulin which carry antibodies for disease resistance. Active horses in competitive events utilise considerable amounts of energy which they mobilise from available carbohydrate in the blood, stored carbohydrate (glycogen) and fats. Since all these metabolic energy-producing reactions are dependent on vitamin B6 it is essential to ensure that the supply is more than adequate for performance horses. Similarly young growing horses require adequate vitamin B6 in order to metabolise protein so that they can grow normally. If too much is given Vitamin B6 is a water soluble vitamin and any excess is normally excreted. Exceedingly high doses (greater than 4g/kg liveweight) have been shown to produce convulsions in dogs and rats. Biosynthesis Micro-organisms in the caeca of horses synthesise pyridoxine but, as with other B-vitamins, it may not be absorbed efficiently. There is little evidence that horses are able to benefit directly from the caecal synthesis because the colon does not appear to be able to absorb vitamins. It follows that horses are dependent on a regular dietary supply, since there are no body stores and any excess vitamin B6 is excreted within 48 hours of intake. How it is measured Vitamin B6 occurs in three different forms so that any analytical procedure designed to determine accurately the total vitamin B6 activity must include all three. Unfortunately, no chemical method has yet been found which can measure all three compounds without first converting them to a single product. Such methods exist but the recoveries are low. A biochemical method using Neurospora sitophila seems to respond equally well to all three vitamin B6 forms and is generally used for vitamin B6 determinations in animal and vegetable products. Most pelleted feeds consist mainly of vegetable products (containing only pyridoxine as the vitamin B6 component) supplemented with pyridoxine hydrochloride, so manufactured compounds may be assayed chemically for pyridoxine, and the minor contributions of pyridoxal and pyridoxamine from the small amounts of animal products ignored. Chromatographic separation (preferably HPLC) followed by fluorimetric determination gives reproducible results for pyridoxine contents of feeds. These are usually expressed in weight units of pyridoxine hydrochloride because there are no International Units of vitamin B6. Research papers from the late 1930’s occasionally refer to ”Rat Units” which are equivalent to about 7.5mg pyridoxine hydrochloride. Pyridoxine hydrochloride contains 82% pyridoxine. Assessment of status It is usually impractical to attempt to measure the vitamin B6 status of an animal from the quantity of pyridoxine in a particular organ or in blood. Vitamin B6 components would have to be determined individually and they are so widely spread around the body that no one site can be considered representative. Similarly, no single enzyme can be regarded as an indicator of status. Research has shown that there are two possible ways of measuring total B6 status. The first relates to the presence of xanthurenic acid in the urine. This is normally not present but appears when the animal has insufficient vitamin B6. The second involves the measurement of nitrogen retention. Optimum nitrogen retention is only possible in the presence of adequate vitamin B6, so nitrogen balance experiments can indicate vitamin B6 status. It is worth noting from this that increased dietary protein intake leads to increased vitamin B6 requirement. Antagonists A number of chemically-prepared pyridoxine derivatives are powerful antagonists of vitamin B6. However, these do not occur naturally. Some medicinal products such as diethylstilboestrol and tyroxin appear to inhibit vitamin B6 activity. Some of the antibiotics have a similar effect. It has also been found that a hydrazic acid derivative with antibiotic properties in linseed is antagonistic to vitamin B6. Sulphonamides also appear to interfere with vitamin B6 activity. Relationships with other ingredients Vitamin B6 is closely involved in such a large number of metabolic pathways that there are direct relationships with many other dietary components. The linear relationship between protein and vitamin B6 has already been noted. The first amino acid to suffer during a shortage of the vitamin is tryptophan and a disturbance of the normal metabolism of tryptophan leads to the production and excretion of xanthurenic acid. Disordered fat metabolism also results from vitamin B6 deficiency because of its requirement for the biosynthesis of coenzyme A. Mineral retention, particularly of calcium, phosphorus, sodium, potassium and zinc is also dependent on vitamin B6 adequacy. Vitamin B6 is synergistic with other vitamins such as thiamine, niacin, riboflavin, biotin, ascorbic acid and vitamin E, and deficiencies of B6 also affect the modes of action of pantothenic acid and vitamin B12. It is also worth noting that the formation of PALP is dependent on an oxidase enzyme which contains riboflavin so any shortage of riboflavin induces a vitamin B6 deficiency as well. Requirements and allowances Horses’ needs for vitamin B6 depend on very many factors - age, performance, protein uptake and the presence of feed additives such as sulphonamides. Although deficiency symptoms have not been identified all the evidence suggests that the amount of vitamin B6 in feeds may not be sufficient for optimum performance at any age. Active horses appear to require a minimum dietary level of pyridoxine greater than 2.5 mg/kg. However levels above this minimum have been found to produce increases in growth rate and performance. The dose response curve appears to plateau when the diet contains a total of about 10mg pyridoxine per kg. The aim should be a supplement of about 3 mg per kg feed which therefore supplies an active, performance horse with 30 mg/day, a resting adult with 18 mg/day, mares and stallions with 12 mg/day and foals and yearlings with 3 -- 10 mg/day. Stability The natural components of vitamin B6 are stable and relatively unaffected by moisture, heat or oxygen. However they are affected by light and by alkaline pH, and minor losses are found in feed processing by pelleting or extrusion. The supplementation recommendations include an allowance of about 10% for losses during processing and storage. Livestock conditions suggesting further needs If growth and appetite are below normal and the horse appears dull and listless it is worth increasing the pyridoxine supplementation to see if this produces any response.