The chemical structure of biotin includes a sulfur atom in its ring (like thiamin) and a transverse bond across the ring (Illus. 11-1). It is a monocarboxylic acid with the sulfur as a thioether linkage. Biotin with its rather unique structure contains three asymmetric carbon atoms; therefore, eight different isomers are possible. Of these isomers, only d-biotin has vitamin activity.
Biotin crystallizes from water as long, white needles. Its melting point is 232° to 233°C. Free biotin is soluble in dilute alkaline solutions and hot water and practically insoluble in fats and organic solvents. Biotin is quite stable under ordinary conditions. It is destroyed by nitric acid, other strong acids, strong bases and formaldehyde, and also is inactivated by oxidative rancidity reactions (Scott et al., 1982). It is gradually destroyed by ultraviolet (UV) radiation. Structurally related analogs of biotin can vary from no activity, to partial replacement of biotin activity, to anti-biotin activity. Mild oxidation converts biotin to the sulfoxide, and strong oxidation converts it to sulfone. Strong agents result in sulfur replacement by oxygen, resulting in oxybiotin and desthiobiotin. Oxybiotin has some biotin activity for chicks (one-third) but less for rats (one-tenth). A prominent nutrient-drug interaction is that biotin-dependent enzymes are reduced with the epilepsy drug carbamazepine (Rathman et al., 2002).
Biotin is present in feedstuffs in both bound and free forms, and much of the bound biotin is apparently unavailable to animal species. For poultry (and presumably other species), often less than one-half of the microbiologically determined biotin in a feedstuff is biologically available (Scott, 1981; Frigg, 1984, 1987). Naturally occurring biotin is found partly in the free state (fruit, milk, vegetables) and partly in the form bound to protein in animal tissues, plant seeds and yeast. Naturally occurring biotin is often bound to the amino acid lysine.
Biotinidase, present in pancreatic juice and the intestinal mucosa, releases biotin from biocytin (bound form of biotin) during the luminal phase of proteolysis. In most species that have been investigated, physiological concentrations of biotin are absorbed from the intestinal tract by a sodium-dependent active transport process, which is inhibited by dethiobiotin and biocytin (Said and Derweesh, 1991). Absorption of biotin by a Na+-dependent process was noted to be higher in the duodenum than the jejunum, which was in turn higher than that in the ileum, and it was concluded that the proximal part of the human small intestine was the site of maximum transport of biotin (Said et al., 1988). Biotin is absorbed intact in the first third to half of the small intestine (Bonjour, 1991). Also, biotin is absorbed from the hind gut of the pig; 50% to 61% of infused biotin disappears between the cecum and feces; this is accompanied by more than a four-fold increase of plasma biotin concentration and more than a six-fold increase of urinary biotin excretion (Barth et al., 1986). Scholtissek et al. (1990) suggested that under basal conditions 1.7% to 17% of the swine requirement for biotin is provided by colonic bacteria.
Biotin appears to circulate in the bloodstream both free and bound to a serum glycoprotein, which also has biotinidase activity, catalyzing the hydrolysis of biocytin. In humans, 81% of biotin in plasma was free and the remainder bound (Mock and Malik, 1992). Information is very limited on biotin transport, tissue deposition, and storage in animals and humans. Mock (1990) reported that biotin is transported as a free water-soluble component of plasma, is taken up by cells via active transport and is attached to its apoenzymes. Said et al. (1992) reported that biotin is transported into human liver via a specialized, carrier-mediated transport system. This system is Na+ gradient-dependent and transports biotin via an electroneutral process.
All cells contain some biotin, with larger quantities in the liver and kidney. Intracellular distribution of biotin corresponds to known locations of biotin-dependent enzymes (carboxylases). Investigations of biotin metabolism in animals are difficult to interpret, as biotin-producing microorganisms are present in the intestinal tract distal to the cecum. Often the amount of biotin excreted in urine and feces together exceeds the total dietary intake, whereas urinary biotin excretion is usually less than intake. Efficient conservation of biotin, together with the recycling of biocytin released from the catabolism of biotin-containing enzymes, may be as important as intestinal bacterial synthesis of the vitamin in meeting biotin requirements (Bender, 1992). 14C-labeled biotin showed the major portion of intraperitoneally injected radioactivity to be excreted in the urine and none in the feces or expired as CO2 (Lee et al., 1973). In rats and pigs, biliary excretion of biotin and metabolites is negligible (Zempleni et al., 1997).
Biotin is an essential coenzyme in carbohydrate, fat and protein metabolism. It is involved in conversion of carbohydrate to protein and vice versa, as well as conversion of protein and carbohydrate to fat. Biotin also plays an important role in maintaining normal blood glucose levels from metabolism of protein and fat when the dietary intake of carbohydrate is low. As a component of several carboxylating enzymes, it has the capacity to transport carboxyl units and to fix carbon dioxide (as bicarbonate) in tissue (Camporeale and Zempleni, 2006). In carbohydrate metabolism, biotin functions in both carbon dioxide fixation and decarboxylation, with the energy-producing citric acid cycle dependent upon the presence of this vitamin. Specific biotin-dependent reactions in carbohydrate metabolism are:
In protein metabolism, biotin enzymes are important in protein synthesis, amino acid deamination, purine synthesis and nucleic acid metabolism. Biotin is required for transcarboxylation in degradation of various amino acids. Deficiency of the vitamin in mammals hinders the normal conversion of the deaminated chain of leucine to acetyl-CoA. Depletion of hepatic biotin results in reduction of hepatic activity of methylcrotonyl-CoA carboxylase, which is needed for leucine degradation (Mock and Mock, 1992). Likewise ability to synthesize citrulline from ornithine is reduced in liver homogenates from biotin-deficient rats. The urea cycle enzyme ornithine transcarbamylase was significantly lower in livers of biotin-deficient rats (Maeda et al., 1996). Acetylcoenzyme A (CoA)-carboxylase catalyzes addition of CO2 to acetyl CoA to form malonyl CoA. This is the first reaction in the synthesis of fatty acids. Biotin is required for normal long chain unsaturated fatty acid synthesis and is important for essential fatty acid metabolism. Deficiency in rats and chicks inhibited arachidonic acid (20:4) synthesis from linoleic acid (18:2) while increasing linolenic acid (18:3) and its metabolite (22:6) (Watkins and Kratzer, 1987a). Biotin deficiency resulted in reduced arachidonic acid (20:4) in chicks and, therefore, reduced plasma prostaglandin E2 (PGE2) since arachidonic acid is a precursor of prostaglandin (20:4) (Watkins and Kratzer, 1987b; Watkins, 1989).
More recently, evidence has emerged that biotin also plays unique roles in cell signaling, epigenetic control of gene expression, and chromatom structure (Rodríguez-Meléndez and Zempleni, 2003). Biotin regulates the genetic expression of holocarboxylase synthetase and mitochondrial carboxylases in rats (Rodríguez-Meléndez et al,, 2001). Manthey et al. (2002) reports that biotin affects expression of biotin transporters, biotinylations of carboxylases and metabolism of interleukin-2 in Jurat cells.
Dogs and cats have a metabolic requirement for biotin, but a dietary requirement has not been established when foods from natural ingredients are fed. Requirements for biotin are difficult to establish due to biotin variability in feed content and bioavailability. Likewise, it is difficult to obtain a quantitative requirement for biotin as the vitamin is synthesized by many different microorganisms and certain fungi. These microorganisms are found in the lower part of the intestinal tract, a region in which absorption of nutrients is generally reduced. There is evidence in the pig, however, that intestinal microflora make a significant contribution to the body pool of available biotin (Barth et al., 1986). Kopinski (1989) has reported that this microbially synthesized biotin is of little benefit to the pig, however. What is not known for the various species is the extent of microbial synthesis or the biotin availability to the host.It is concluded that microorganisms contribute to animal and human requirements, as the use of some sulfa drugs such as sulfathalidine can induce deficiency under some circumstances. Rate and extent of biotin synthesis may be dependent upon the level of other dietary components. In rats and poultry, it has been shown that polyunsaturated fatty acids (PUFA), ascorbic acid and other B vitamins may influence the demand for biotin. Addition of PUFA to fat-free, biotin-deficient diets increased severity of dermal lesions (Roland and Edwards, 1971). Biotin is rapidly destroyed as feeds become rancid. Pure biotin was inactivated to an extent of 96% in 12 hours when linoleic acid of a high peroxide number was added to the diet (Pavcek and Shull, 1942). In the presence of alpha-tocopherol, this destruction amounted to only 40% after 48 hours.
A definite biotin requirement for dogs has not been established. (NRC, 2006; AAFCO, 2007). However, diets containing raw egg white and/or antibiotics may need biotin supplementation. The NRC (1985) for dogs suggests 30 µg biotin per 1,000 kcal metabolizable energy (ME) as a safeguard against a possible deficiency.
Cats probably do not require a dietary source of biotin unless antimicrobial agents or antivitamins are present in the diet. The NRC (1986) for cats reported that a purified diet containing 60 µg biotin per kg (27.3 µg per lb) supported pregnancy and lactation in queens and normal growth in kittens. Therefore, with adjustment for dietary caloric density, a requirement of 70 µg per kg (31.8 µg per lb) of diet is recommended.The latest NRC (2006) for cats suggests a biotin requirement of 75 µg per kg (34 µg per lb) of diet for all classes of cats. The Association of American Feed Control Officials (AAFCO, 2007) recommends 70 µg biotin per kg (31.8 µg per lb) of diet. The AAFCO recommendation is qualified by the statement that biotin does not need to be added unless the diets contain antimicrobial or antivitamin compounds.
Biotin is present in many feedstuffs; however, corn, wheat, other cereals, meat and fish are relatively poor sources (Table 11-1). There is considerable variation in biotin content within individual sources. For example, 59 samples of corn analyzed for biotin varied between 56 and 115 µg per kg (26 and 52 µg per lb) and 62 samples of meat meal ranged from 17 to 323 µg per kg (8 to 146 µg per lb) (Frigg, 1987). In comparison to cereal grains, oilseed meals are better sources of total biotin. Soybean meal, for instance, contains a mean biotin content of 270 µg per kg (122 µg per lb) with a range of 200 to 387 µg per kg (91 to 176 µg per lb) (Frigg, 1976; 1984; 1994).
Less than one-half of the biotin in various feedstuffs, as determined by microbiological assay, is biologically available (Frigg, 1976; 1984; 1987). Thus, it is important to know the form of biotin (e.g., bound or unbound) as well as its overall content in feed.
In alfalfa meal, corn, cottonseed meal and soybean meal, bioavailability of biotin is estimated at 100%. However, biotin availability is variable for other feedstuffs, for example, 20% to 50% in barley, 62% in corn gluten meal, 30% in fish meal, 20% to 60% in sorghum, 32% in oats and 0% to 62% in wheat (McDowell, 2004).Frigg (1976) reported that the biotin content of wheat and barley may be totally unavailable. Buenrostro and Kratzer (1984) found biotin in sorghum to have an availability of 10% to 20%, while the bioavailability of corn ranged from 75% to 100%.
Information is lacking on the availability of feedstuff sources of biotin to ruminants. It seems likely, however, that much of the dietary biotin would also be unavailable to rumen microorganisms. The primary source of biotin to ruminants is that synthesized by microorganisms in the rumen, and quantities are produced regardless of dietary composition (Hayes et al., 1966). Milk produced by ruminants is and excellent source of biotin; cow colostrum contained from 1.0 to 2.7 µg biotin per 100 ml (Foley and Otterby, 1978). Biotin in feedstuffs may be destroyed by heat curing, solvent extraction, and improper storage conditions, while pelleting has little effect on biotin content of feed (McGinnis, 1986). Milling of wheat or corn and canning of corn, carrots, spinach or tomatoes reduced biotin concentrations (Bonjour, 1991). Biotin is unstable to oxidizing conditions and, therefore, is destroyed by heat, especially under conditions that support simultaneous lipid peroxidation.
Biotin is commercially available as a 100% crystalline product or as various dilutions, premixed and low potency, spray-dried preparations. The d form of biotin is the biologically active form. A 2% spray-dried biotin product is also commercially available for use either in feed or drinking water.
Biotin is important for normal function of the thyroid and adrenal glands, the reproductive tract and the nervous system. Its effect on the cutaneous system is most dramatic, since a severe dermatitis is the major obvious clinical sign of biotin deficiency in animals, including dogs and cats. Biotin is active in skin formation and maintenance, and a deficiency causes abnormal keratinization and cornification of the epidermis, leading to low tensile strength and more skin lacerations. Perhaps more importantly, biotin deficiency can slow the process of wound healing.
Biotin deficiency results in reduced growth rate and impaired feed conversion and produces a wide variety of clinical signs, particularly dermal lesions. There is a hyperkeratosis of the superficial and follicular epithelia, giving the animal a scruffy appearance due to exfoliation of the skin. These lesions begin on the limbs and face and spread over the body. Naturally occurring biotin deficiency is very rare in dogs and cats (NRC, 2006). Apparently biotin deficiency in dogs and cats has occurred only when animals have been fed a biotin antagonist (e.g., avidin) and/or have been treated with antibiotics. Because gut microbial synthesis may meet half the biotin requirement, antimicrobials that decrease the population of the intestinal microflora may also result in signs of biotin deficiency (Hand et al., 2010).
Detection of biotin deficiency in various species includes decrease in urine and plasma biotin and reduced activity of carboxylase enzymes. Feeding raw egg white significantly reduced urinary biotin and the activity of various biotin-dependent enzymes (pyruvate and propionyl CoA carboxylases) in the liver and kidneys of dogs (Shen et al., 1977). Biotin-deficient cats had a decrease in the activity of liver propionyl CoA carboxylase (Carey and Morris, 1977).
Biotin deficiency was observed in dogs fed a diet containing spray-dried egg white and sulfaguanidine (Greve, 1963). Dogs exhibited scurfy skin (due to hyperkeratosis of the superficial and follicular epithelia) and a marked decline in urinary biotin concentration. Biotin deficiency also includes alopecia and eventually diarrhea and anorexia (Case et al., 1995). Biotin responsive disease conditions in dogs include dull coat, brittle hair, loss of hair, scaly skin, pruritis and dermatitis (Frigg et al., 1989; NRC, 2006).
Biotin deficiency was produced in cats by feeding diets containing 18.5% to 32.0% raw egg white in a semi-purified diet to kittens (Carey and Morris, 1975; 1977). Growth was normal up to about 150 days. At this time, dried secretions were evident around the eyes and nose and at the angles of the mouth. Also seen were scaly dermatitis of the nasomaxilla-mandibular region, general alopecia and hypersalivation. Alopecia that began on the face has been reported in cats (Carey and Morris, 1977), but not in dogs (Greve, 1963). These signs increased in severity and were later accompanied by bloody diarrhea and marked anorexia and emaciation in the terminal stages of the deficiency (NRC, 2006). After 11 weeks of consuming a semi-purified diet containing egg white, young female cats developed signs of alopecia, dried secretion around the eyes, nose, mouth and feet, focal dermatitis of the lips near the eyeteeth and a brownish appearance of the skin (Pastoor et al., 1991).
Biotin deficiencies are not generally a problem. However, the treatment of dogs and cats with antibiotics that decrease the bacterial population of the large intestine will cause an increase in the dietary requirement for biotin. Likewise, including such antagonists in the diet as avidin and rancid fats will inactivate biotin in feeds of biological origin. Carey and Morris (1977) developed biotin deficiency in cats with a semi-purified diet containing dried egg white and sulfathiazole. The deficiency was reversed by administering 250 µg of supplemental biotin by injection every other day. In addition, due to the extreme variation of the biotin content and bioavailability in feedstuffs, pet foods can be unreliable as consistent sources of biotin. Biotin deficiency for dogs is apparently more prevalent than once thought. In a study with small animal practitioners, 60% of clinical signs were cured and a further 31% improved for dogs (119 cases) with hair and skin conditions (Frigg et al., 1989). Losses of biotin during storage can be considerable. Biotin is readily destroyed by feed rancidity (Pavcek and Shull, 1942). Preparing fresh feeds, storing them only for short periods and keeping them dry and in a well-ventilated storage area will minimize rancidity problems. Also, the diet should be low in feedstuffs containing high levels of pro-oxidants and/or properly protected by an effective antioxidant to avoid destruction of biotin. Biotin is relatively stable in multivitamin premixes, as well as in naturally occurring sources in feeds, and is fairly stable during processing. However, Gadient (1986) reported a 10% to 30% reduction in biotin after three months of storage when the vitamin supplement contained choline and trace minerals. Chlorine has been shown to inactivate biotin. This may be of importance for pets receiving chlorinated drinking water.
Paul et al. (1973) reported that an acute dose of biotin at the level of 5 mg per 100 g (5 mg per 6.25 oz) of body weight caused irregularities of the estrus cycle with heavy infiltration of leukocytes in the vagina of the rat up to 14 days after treatment. In additional rat studies, a dose of biotin at least 5,000 or 10,000 times the daily requirement had no deleterious effects (Mittelholzer, 1976). Studies with poultry and swine indicate that those species can safely tolerate dietary biotinlevels four to 10 times their nutritional requirements (NRC, 1987). In view of the poor retention of biotin for most species, higher levels may be tolerated. There is a lack of data available to estimate dietary tolerance levels of biotin for dogs and cats.
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