PUBLICATION DATE:  03/05/2007

RATING

AUTHOR:  PETER F. SURAI and JULIA E. DVORSKA - Avian Science Research Centre/Sumy State Agrarian University (Courtesy of Alltech Inc.)

Animal health depends on many factors; and increasingly it is appreciated that diet plays a pivotal role in health maintenance and prevention of various diseases. Among dietary factors, antioxidants have a special place in the battle for survival, well-being and content. This is owing to the detrimental effects of free radicals and toxic products of their metabolism on various metabolic processes in the animal body.

Free radicals are compounds containing one or more unpaired electrons. Most biologically relevant free radicals are derived from oxygen and nitrogen.

Free radicals are highly reactive, and are capable of damaging molecules such as DNA, proteins, lipids or carbohydrates. The animal body is under constant attack from free radicals, formed as a normal part of oxidative metabolism and as part of the immune system’s strategy for destroying invading microorganisms, as well as from outside sources.

For example, under normal physiological conditions about 3-5% of the oxygen taken up by the cell undergoes univalent reduction leading to the formation of free radicals (Singal et al., 1998).

Furthermore, immune cells produce free radicals and use them to destroy pathogens (Schwarz, 1996; Kettle and Winterbourn, 1997). Some internal and external sources of free radicals are shown in Table 1.

During evolution living organisms developed specific antioxidant protective mechanisms to deal with free radicals, which are constantly produced in the cells. These mechanisms aided survival in an oxygenated atmosphere when oxygen concentration was rising. These mechanisms are described by the general term ‘antioxidant system’ (Halliwell and Gutteridge, 1999).

In nature there are hundreds and thousands of compounds possessing antioxidant properties that are able to react with free radicals. They are fatsoluble (vitamin E and carotenoids, etc.) and water-soluble (ascorbic acid, glutathione, bilirubin, etc.), they can be synthesised in the body (ascorbic acid or glutathione) and are delivered with food (vitamin E, carotenoids, selenium, etc.). More importantly, there is a range of antioxidant enzymes that can be synthesised in the body, but require the presence of diet-derived co-factors. For example, selenium (Se) in the form of selenocysteine is an essential part of a family of enzymes called glutathione peroxidases (GSH-Px). Zinc (Zn), copper (Cu) and manganese (Mn) are integral parts of another antioxidant enzyme family called superoxide dismutases (SOD), while iron (Fe) is an essential part of the antioxidant enzyme catalase (CAT). Therefore only when these metals are delivered with the diet in sufficient amounts can the body synthesise the antioxidant enzymes.

Deficiency of those elements causes oxidative stress and results in damage to biological molecules and membranes.

Biological antioxidants are substances capable of reacting with free radicals or products of their metabolism, converting them into less reactive molecules and preventing or delaying oxidation. The most important and well-characterised natural antioxidants in the animal body are vitamins E and C. Plant pigments known as carotenoids also have antioxidant properties. The protective antioxidant compounds are located in organelles, subcellular compartments or the extracellular spaces enabling maximum cellular protection.

All antioxidants in the body operate in concert to bring about antioxidant defence, with one member of the group helping another to do its job efficiently.

For example, vitamin C restores vitamin E after oxidation; and glutathione does the same for vitamin C (Figure 1). Therefore if relationships in this team of antioxidants are effective, which happens only in the case of a balanced diet and sufficient provision of dietary antioxidants, then even low doses of an antioxidant such as vitamin E can be effective. On the other hand, under high oxidative stress conditions, when free radical production dramatically increases, then without external help it will be difficult to prevent damage to major organs and systems. This external help takes the form of increased supplementation of the diet with natural antioxidants, especially vitamin E and Se. For the nutritionist or feed formulator, the challenge is to understand when the antioxidant system requires help, and how much of this help can be justified given the additional expense.

An understanding of the major steps in antioxidant defence helps the nutritionist in making decisions about antioxidant supplementation:

• A delicate balance exists between the amount of free radicals generated and the antioxidants needed for protection.

• Excess free radicals or insufficient antioxidant protection can shift this balance to produce oxidative stress.

• Oxidative stress plays a major role in many degenerative pathologies; and free radical formation is involved in the initiation or progression phase of various diseases. In general, it is widely believed that most animal diseases are associated with free radical production and metabolism at some stage.


Natural antioxidants in feed ingredients

It has been suggested that the antioxidant/prooxidant balance in the body is responsible for maintaining health, productive and reproductive performance of animals (Figure 2). This balance can be adversely affected by suboptimal nutrient intakes or positively affected by dietary supplementation. Therefore, feed components can modulate maintenance of this balance and may thereby influence the effects of aging as well as disease resistance. Thus, the most important step in preventing oxidative damage would be to enhance antioxidant capacity by optimising the dietary intake of antioxidants. Following is a brief summary of the activities of the major dietary antioxidants.

Figure 1. Vitamin E recycling (1 - thioredoxin reductase; 2 - glutathione reductase; 3 - glucose-6-phosphate dehydrogenase).

Figure 2. Antioxidant/pro-oxidant balance in the cell.

VITAMIN E

• Main chain-breaking antioxidant in biological systems.
• Located in biological membranes and lipid droplets.
• Absorbed in the small intestine with efficiency depending on diet composition, level of supplementation, age, sex and other individual characteristics.
• Accumulated to some extent in the liver and adipose tissues, however, deposition is not sufficient to meet long-term requirements.
• Crude plant oils are the richest source of vitamin E, however, oil refining decreases vitamin E concentration dramatically. Meat products are poor sources of vitamin E.
• Unstable and easily oxidized. Commercial preparations usually contain esterified forms of the vitamin (eg. tocopheryl acetate), which is comparatively stable during storage but does not possess antioxidant activity itself. Only after digestion in the intestine is it converted to activeα-tocopherol.
• Non-toxic. Even very high doses are not associated with hypervitaminosis.
• Comparatively expensive.
• Recent data show that increased vitamin E supplementation is beneficial for stressed animals.


CAROTENOIDS

• Pigments responsible for yellow, orange and sometimes red pigmentation in plants, insects, birds and marine animals.
• Have some health promoting properties, including immune system modulation.
• There are no established requirements for animals.
• Many plant-derived foods are rich in carotenoids, but animal-derived foods are poor sources.
• Unstable and oxidized during food storage.
• Reserves in the body are limited.


VITAMIN C

• Universal water-soluble antioxidant.
• Works in close relationship with vitamin E, recycling it from oxidized form.
• Important anti-stress agent.
• Synthesised in the body of most animals.
• In stress conditions dietary supplementation is beneficial.
• Easily oxidized.
• Meat-derived food is poor source of ascorbate.


GLUTATHIONE

• Cellular antioxidant.
• Synthesised in animals.
• Intracellular redox status sensor.
• Important in stress conditions.
• Dietary supplementation has not proven effective.


SELENIUM

• Dietary essential trace element
• Essential part of a range of selenoproteins, including GSH-Px, thioredoxin reductase (TrxR), iodothyronine deiodinase (ID) and others.
• Food ingredients contain variable concentrations of Se, but most are deficient.
• Physiological requirements are low, but if not met, antioxidant systems are compromised with detrimental consequences for animal health.
• Toxic in high doses.
• Two major sources in practical diets: a natural source in the form of selenomethionine from plants and inorganic selenium in the form of selenite or selenate.
• Natural Se (selenomethionine) has been proven to have a range of benefits as a nutritional supplement.


ZINC

• The second most abundant trace element in mammals.
• A component of >300 enzymes involved in:

– antioxidant defence as an integral part of SOD.
– hormone secretion.
– keratin generation and epithelial tissue integrity.
– nucleic acid synthesis.
– protein synthesis.
– sexual development and spermatogenesis.
– immune function.

• Organic Zn is characterised by improved availability in comparison to inorganic sources and is considered to be beneficial for animal health.


COPPER

• An essential component of a range of physiologically important metalloenzymes taking part in:

– antioxidant defence as an integral part of SOD.
– cellular respiration.
– cardiac function.
– bone formation.
– carbohydrate and lipid metabolism.
– immune function.
– connective tissue development.
– tissue keratinization.
– myelination of the spinal cord.

• Inorganic copper is a strong pro-oxidant and can stimulate lipid peroxidation in feed or in the intestinal tract.
• Organic Cu does not possess pro-oxidant properties and can improve Cu status of animals.


IRON

• A vital role in many biochemical reactions taking part in:

– antioxidant defence as an essential component of catalase.
– energy and protein metabolism.
– heme respiratory carrier.
– oxidation/reduction reactions.
– electron transport system.

• A very strong pro-oxidant in oxide or sulphate form and can stimulate lipid peroxidation. This is especially relevant in the digestive tract where lipid peroxidation can cause enterocyte damage and decreased absorption of various nutrients (especially antioxidants). Organically complexed forms reduce oxidant potential.


MANGANESE

• Plays an important role in body metabolism as an essential part of a range of enzymes taking part in:

– antioxidant protection as an integral part of SOD.
– bone growth and egg shell formation.
– carbohydrate and lipid metabolism.
– immune and nervous function.
– reproduction.

Antioxidants and animal health

Dietary antioxidants play a crucial role in the health of companion animals by providing protection during exposure to elevated levels of toxins and free radicals. The results of recent in vivo assessments, clinical trials and research studies showed that oral supplementation with vitamin E, Se, glutathione, and taurine were beneficial for both maintaining natural antioxidant/pro-oxidant balance in the body and protecting against a number of degenerative diseases associated with free radical damage and toxin exposure (Scanlan, 2001). For example, pet dogs share the human environment and respond to many toxic insults in ways analogous to humans (Backer et al., 2001). In fact, a number of diseases affecting the heart are prevalent in canines. Various antioxidants including coenzyme Q10 and vitamin E have been evaluated in the prevention and treatment of many types of heart disease in dogs providing clear evidence for the benefit of these supplements (Dove, 2001). Therefore, optimal levels of certain dietary nutrients have been shown to increase life span, improve life quality, reduce symptoms and physical evidence of disease, and decrease mortality rates in these animals.


Food quality and antioxidants

Petfood quality control is very strict and ensures that only high quality ingredients are included in the diet. However, since environmental pollutants such as heavy metals or mycotoxins are very widely distributed, it is almost impossible to completely avoid their presence in food ingredients. For example, an analytical survey of 42 elements in 31 commercial canned pet foods noted that arsenic, bromine, cadmium, chromium, mercury and selenium were highest in fish-containing cat foods (Furr et al., 1976). Lead was most consistently high in those products containing chicken. Barium, nickel and tin also appeared high in some samples. An analytical survey of toxic elements including mutagens, nitrosamines and polychlorinated biphenyls as well as the toxicologically protective constituents Zn, Se and vitamin C, in 48 pet foods was conducted (Mumma et al., 1986). High concentrations of fluoride and iodide in some samples and increased concentrations of mercury and Se in certain cat foods containing fish were detected. Polychlorinated biphenyls were only detected in one cat food.

Ochratoxin A occurrence was investigated in canned (26 samples) as well as dry pet foods (17 samples) for cats and dogs (Razzazi et al., 2001).

Ochratoxin A was detected in 47% of the pet food samples at the levels of 0.1-0.8 ng/g food. Only two samples contained higher amounts (3.2 and 13.1 ng/g). Ochratoxin A was also detected in 62% of cat kidneys (0.35-1.5 ng/g tissue). Recently mycoflora of 21 dry pet foods (12 dog and 9 cat food) that corresponded to eight commercial brands imported from South America were analysed (Bueno et al., 2001). Ten genera and fungi classified as Mycelia sterilia were identified. The predominant genera were Aspergillus (62%), Rhizopus (48%), and Mucor (38%). The most prevalent among Aspergillus was Aspergillus flavus followed by Aspergillus niger and Aspergillus terreus, all of which are potential sources of mycotoxins.

Lipid peroxides are a major concern for petfood manufacturers. Since added vitamin E is usually added in esterified form, it does not possess antioxidant activity per se and only becomes active after de-esterification in the intestine. Therefore, various synthetic antioxidants are used for prevention of lipid peroxidation during pet food storage.

The effects of different preservative treatments, ethoxyquin plus butylated hydroxyanisole (BHA), mixed tocopherols and ascorbyl palmitate and mixed tocopherols, applied to extruded dog food were studied (Gross et al., 1994). After processing the dog foods were placed in bags and stored for 16 weeks at 48.8 °C or for 12 months at 22.2 °C. Best results were observed with the combination of ethoxyquin and BHA. Interestingly, in both the high and ambient temperature tests dogs consumed more of the foods with the lowest peroxide value when given a two-bowl choice.


Anorexia in companion animals and its connection to antioxidants

One of the most frequent motivations for seeking veterinary attention for a cat is loss of normal appetite (anorexia) (Michel, 2001). The mechanisms underlying decreased food intake are of course multifactorial, however one aspect of appetite regulation involves interaction of external stimuli with signals from the gastrointestinal tract and central nervous system (Michel, 2001). It is likely that lipid peroxidation in the intestine could be a triggering factor for anorexia given the willingness to select foods with lower levels of peroxides (Gross et al., 1994). Insufficient erythrocyte tocopherol, together with altered antioxidant enzyme activities, suggests a certain degree of oxidative damage in anorexia nervosa since micronutrient intake is insufficient and oxidative stress a probability (Moyano et al., 1999).

Cytokines likely have a specific role in anorexia development. In fact, the pro-inflammatory cytokines and oxidant molecules produced during the inflammatory response may be beneficial or detrimental to the animal, depending on the amounts and contexts in which they are produced (Grimble, 1998). In particular, aberrant or excessive production of those molecules has been implicated in inflammatory disease and sepsis. Complex systems exist for the control of cytokine production and cellular redox status. Cytokine activities influence use of antioxidant nutrients including influence of the hypothalamo-pituitary-adrenal hormones, acute phase proteins, and endogenous inhibitors of interleukin-1 and tumor necrosis factor as well as endogenously synthesized antioxidants such as glutathione and dietary antioxidants such as tocopherols, ascorbates and cachectins (Grimble, 1998). At the same time, nutrients change cytokine production and potency by influencing tissue concentrations of many of the molecules involved in cytokine biology.

It is well recognised, for example, that low antioxidant intake results in enhanced cytokine production and effects. The anorexia that in many cases follows infection and injury may be purposeful to permit release of substrate from endogenous sources to support and control the inflammatory process (Grimble, 1998). Therefore, prior as well as concurrent antioxidant intake are of great importance in determining the outcome of the inflammatory response. It is interesting that anorexia was one of the major clinical signs of toxicosis in aflatoxin B₁- treated calves (Brucato et al., 1986), and Se supplementation improved feed intake in animals exposed to mycotoxins. Vitamin E-selenium deficiency was also associated with the development of anorexia and diarrhoea in wild ducks (Dhillon and Winterfield, 1983). Similarly, silver toxicity in weanling pigs was associated with lesions typical of Se-vitamin E deficiency including anorexia and diarrhoea (Van Vleet, 1976). A relationship between antioxidant supplementation and intake was also indicated when administration of seleno-DL-methionine increased voluntary feed consumption of Se-deficient chickens within 2-3 hrs (Bunk and Combs, 1980). Dietary antioxidants could be the supplements of choice when dealing with animal anorexia.


Antioxidant deficiencies in companion animals

Signs of antioxidant deficiency in companion animals are usually very similar to those described for laboratory and farm animals. For example, a case of nutritional myopathy primarily caused by vitamin E deficiency was described in a cat that had been fed a diet consisting almost entirely of boiled Norwegian coley (Dennis and Alexander, 1982). The animal had swollen muscles in both the hind and forelegs. Analyses of biopsy material revealed chronic, severe myositis, with normal muscle tissue undergoing a series of degenerative changes.

Multivitamin and mineral supplementation led to a complete clinical recovery with the cat regaining full use of its legs within 14 days. A dog fed a purified vitamin E-deficient diet developed retinopathy, which was ophthalmoscopically visible as early as 3 months (Riis et al., 1981). Typical lesions developed first in the central and outer layers and progressed slowly toward the periphery with the tapetal retina being more extensively involved, showing color change and mottling. Microscopic examination revealed large amounts of yellow autofluorescent pigment accumulated within the retinal epithelium. Later stages of retinopathy showed this pigment was present in migrating cells in all the retinal layers. Supplementation of vitamin E protected the retina from changes, but addition of Se alone was not effective.

Experimentally induced vitamin E-Se deficiency in the growing dog was also characterised (Van Vleet, 1975). Beagle pups (5-8 weeks old) were fed a semisynthetic basal diet deficient in vitamin E and Se for 55 to 70 days. Clinical signs of vitamin E-Se deficiency developed after 40 to 60 days and was characterised by muscular weakness, subcutaneous edema, anorexia, depression, dyspnea, and eventual coma. Gross lesions on necropsy included ventral subcutaneous edema, generalized skeletal muscular pallor and edema with scattered white longitudinal streaking, prominent brownish yellow discoloration of the intestinal musculature, and a layer of white chalky material at the renal corticomedullary junction. Microscopic examination revealed extensive skeletal muscular degeneration and regeneration, focal subendocardial necrosis in the ventricular myocardium, intestinal lipofuscinosis and renal mineralization. A case of cardiac myopathy resembling Se-vitamin E deficiency myopathy was described in a dog by van Rensburg and Venning (1979).


Practical applications of antioxidants in companion animal nutrition

IMMUNE SYSTEM

One of the most important applications of antioxidants in companion animal nutrition is modulation of immune response. Among many different nutrients involved in regulation of the immune system, natural antioxidants have a special role (for review see Surai, 2002). Phagocytic cells produce free radicals (superoxide radical, hydroxyl radical) and toxic reactive oxygen species (ROS, hydrogen peroxide, hypochlorous acid) and use them to kill pathogens. However the same ROS can also damage biological molecules including various membrane structures as well as phagocytes themselves unless they are neutralized by antioxidants.

Therefore, even phagocyte function is compromised when antioxidant defences are low. Furthermore, efficiency of immune response depends on the communication among various cells including neutrophils, macrophages, B- and T-lymphocytes, NK cells etc., which is due to presence of very sensitive receptors on the surface of those immune cells. These receptors are sensitive to damage by ROS. In addition, production of communicating molecules (cytokines, eicosanoids, etc.) is also antioxidant-dependent. Therefore, antioxidant protection of the immune cell receptors is of great importance for the maintenance of their communications and ultimately for immunocompetence. The consequences of a compromised antioxidant system in relation to immunocompetence are shown in Figure 3.

Antioxidant compounds such as vitamin E, selenium and carotenoids are well-known to be involved in immunomodulation. However, the dietary levels of antioxidants that bring about these effects are usually several times higher than those necessary for animal growth and development.

Furthermore, other trace minerals, including Zn, Cu and Fe are also involved in immune system regulation as cofactors for antioxidant enzymes and important biosynthetic pathways.

Figure 3. Oxidative stress and the immune system (adapted from Surai, 2002).

Data on effects of antioxidants on companion animal immunity are restricted to several recent publications. For example, dietary ß-carotene increased cell-mediated and humoral immune responses in female Beagle dogs (Chew et al., 2000). Compared with unsupplemented dogs, those fed 20 or 50 mg of ß-carotene had higher CD4+ cell numbers, CD4:CD8 ratio and plasma IgG concentration. Furthermore, the delayed-type hypersensitivity response to phytohemagglutinin (PHA) and vaccination was heightened in ßcarotene- supplemented dogs. On the other hand, immune response was impaired in dogs classified as low ß-carotene absorbers. Dietary lutein stimulated immune responses in the domestic canine in a similar fashion (Kim et al., 2000). Female Beagle dogs (17-18 months old) were supplemented daily with 0, 5, 10 or 20 mg lutein for 12 weeks.

Delayed-type hypersensitivity (DTH) response to saline, PHA and a polyvalent vaccine was assessed on weeks 0, 6 and 12. Lutein-supplemented dogs had stimulated DTH response to PHA and vaccine by week 6. Dietary lutein also increased lymphocyte proliferative response to mitogens and increased the percentages of cells expressing CD5, CD4, CD8 and major histocompatibility complex class II molecules. The production of IgG also increased in lutein-fed dogs after the second antigenic challenge (Kim et al., 2000).

The immuno-modulatory action of lutein was also shown in domestic cats. Female Tabby cats (10 months old) were supplemented daily for 12 weeks with 0, 1, 5 or 10 mg lutein (Kim et al., 2000a). The DTH response to vaccine increased in a dosedependent manner on week 6. Compared to controls, cats given lutein also showed enhanced response to concanavalin A and proliferation of pokeweed mitogen-stimulated peripheral blood mononuclear cells. Dietary lutein also increased the percentages of CD4+ and CD21+ lymphocytes on week 12. Plasma IgG was higher in cats fed 10 mg lutein on weeks 8 and 12 (Kim et al., 2000a).

Sera from dogs maintained on a vitamin E- and Se-deficient diet had a severe suppressive effect on the in vitro response of lymphocytes to mitogenic stimulation (Lessard et al., 1993). Added vitamin E enhanced the responsiveness in serum from the dogs with vitamin E deficiency and it seems likely that the depressed lymphocyte response in sera from vitamin E-deficient dogs may be due in part to a loss of antioxidant activity (Langweiler et al., 1983).

In earlier experiments, depressed lymphocyte responsiveness in pups fed a vitamin E-deficient diet was concluded to be the result of a suppressive factor in the serum. When effects of vitamin E deficiency on dog lymphocytes was examined using the lymphocyte blastogenesis assay with PHA, concanavalin A, and pokeweed as mitogenic stimulants, lymphocytes from pups fed deficient diets were poorly responsive compared with the controls. (Langweiler et al., 1981). At the same time lymphocytes from controls, which had good responses when cultured in bovine fetal serum or normal canine serum, were poorly responsive when cultured in serum from pups fed the vitamin E-deficient diet. The capability of sera from vitamin E-deficient pups to support blastogenesis of lymphocytes from control dogs was restored following dietary supplementation with vitamin E.

It seems likely that antioxidant compounds, including Se, vitamin E and carotenoids in a combination with other immunomodulators such as omega-3 fatty acids, could be beneficial for the improvement of immune response in companion animals.


ANTIOXIDANTS AND EXERCISE IN DOGS

It is well known that exercise is essential to health for humans or pets, including dogs. However, where exercise is associated with overproduction of ROS, antioxidants have protective effects. Dogs in the 1998 Iditarod Race with pre-race plasma vitamin E concentrations >40.7 μg/ml were 1.9 times more likely to finish and had 1.8 times less risk of being withdrawn for every mile run than dogs with plasma vitamin E concentrations between 16.3 and 40.7μg/ml (Piercy et al., 2001). Antioxidant supplementation of sled dogs may attenuate exercise-induced oxidative damage. For example, effects of dietary antioxidant supplementation on plasma concentrations of antioxidants, exerciseinduced oxidative damage, and resistance to oxidative damage during exercise were studied in Alaskan sled dogs given 400 units of α-tocopherol acetate, 3 mg of ß-carotene and 20 mg of lutein orally per day or no supplement.

After one month, all dogs completed three days of exercise (Baskin et al., 2000). Dogs given the antioxidant supplements had increased plasma concentrations of antioxidants, decreased DNA oxidation and increased resistance of lipoprotein particles to in vitro oxidation. Twenty-four lightly trained sled dogs were used in an experiment to determine the effects of repetitive endurance exercise on antioxidant defences (Hinchcliff et al., 2000). Results demonstrated that repetitive endurance exercise in dogs was associated with lipid peroxidation and a reduction in plasma antioxidant concentrations, suggesting that the antioxidant mechanisms of minimally trained dogs may be inadequate to meet the antioxidant requirements of exercise. These reports suggest that increased dietary antioxidant levels are needed in exercise-stressed dogs.


NUTRITIONAL STRATEGIES TO PREVENT DAMAGING EFFECTS OF FREE RADICALS: SELENIUM SUPPLEMENTATION

While the body synthesises antioxidant enzymes, dietary availability of Se, Mn, Zn and Cu are among major restrictions for this synthesis. For example, the availability and the efficiency with which dietary Se is used, and therefore its ability to perform its physiological functions, greatly depends on whether the source is organic or inorganic. The basic difference is that the organic form, which is predominately selenomethionine in plant protein, can be stored in animal tissues (primarily muscle) as well as transferred across the placenta and into colostrum and milk. Other forms, such as inorganic Se from selenate or selenite as well as selenocysteine, are not retained in animal tissues to a significant extent.

The differences among mineral forms must be taken into account when formulating companion animal diets with the goal of optimising nutrition.

Animal diets include Se in order to prevent deficiency and to maintain health and reproductive performance. The reproductive system of companion animals, for example, depends very much on antioxidant status in both the male and female. Spermatozoa are rich in polyunsaturated fatty acids and require adequate antioxidant protection. Therefore, if antioxidant protection is compromised, male fertility will be compromised.

In the female, increased amounts of Se are needed to supply the developing foetus during gestation and for colostrum and milk formation without compromising the antioxidant nutrient needs of the dam.

Tissue reserves during periods of increased demand such as gestation/lactation are critical.

Unlike selenite Se, dietary selenomethionine is actively absorbed from the intestine following the same pathways as methionine. However, also like methionine, animals cannot synthesise the selenium analogue. Only plants, yeast and marine algae can form either compound, which is the reason that they are essential in the diet. The differences in metabolism of organic and inorganic Se forms underscore the fact that animals evolved to digest and metabolise the organic form. This does much to explain why the organic form is much more effective, particularly in animals subjected to stress or other conditions that increase Se demand.

In dogs, similar to other animals, the absorption of selenomethionine is accelerated by the specific amino acid active transport mechanisms in the gut mucosa. In contrast, sodium selenite is absorbed more slowly, by simple diffusion through the intestinal mucosa. For example, the triple-lumen perfusion method was used to measure the rate of absorption of trace quantities of Se from the jejunum when given as D,L-selenomethionine, D,Lselenocystine, or sodium selenite to healthy dogs (Reasbeck et al., 1985). Selenium absorption from the test segment (expressed as percent administered dose per centimeter) was 1.97 from D,Lselenomethionine, 1.15 from D,L-selenocystine, and 0.51 from sodium selenite.

It is interesting that Se concentration in serum of cats and dogs was substantially higher than that in human or farm animals (Table 2). The main conclusion from the Forrer et al. (1991) work was that up to 20% of the geriatric donors in Switzerland had inadequate selenium levels.

We can consider two different scenarios for antioxidant defence in companion animals relative to the form of selenium used. The first, and most common, situation exists when inorganic selenium is added to the diet (Figure 4). In response to oxidative stress, the body draws on nutrient reserves to synthesise additional selenoproteins such as GSHPx.

In this scenario, use of selenite to provide selenium has limited the establishment of selenium reserves. In consequence, the ability to synthesise selenoproteins and expand antioxidant protection is restricted. The antioxidant/pro-oxidant balance shifts toward overproduction of free radicals.

Therefore, in this scenario we would expect immunity and general health to be compromised and reproductive success decreased.

It should be noted that we are not speaking about dramatic differences, but after several stresses animal behaviour and health can be dramatically affected. This is especially important for newborn puppies or kittens, since antioxidant defence in neonates depends on antioxidant transfer from mothers through colostrum and milk. Since inorganic selenium is not transferred to the milk in any substantial amount, we would not expect an antioxidant system improvement through this route in this scenario.

The antioxidant defence scenario changes when organic selenium is used (Figure 5). The major benefit comes from the existence of selenium reserves in the form of selenomethionine in muscle.

Under stress conditions, cellular organelles called proteasomes catabolise tissue protein to provide amino acids for synthesis of specialized proteins including antibodies and selenoproteins.

Selenomethionine is released during catabolism, thereby becoming available for the synthesis of additional selenoproteins to prevent damaging effects of free radical overproduction. This is especially important since many stresses are associated with decreased food consumption. The ability of newly synthesised selenoproteins to prevent lipid peroxidation limits the impact of oxidative stressors and allows the animal to maintain immunocompetence and reproductive performance.

Figure 4. Inorganic selenium scenario.

Figure 5. Organic selenium scenario.

Since selenomethionine is transferred to the milk and colostrum, newborn animals are better able to withstand oxidative stress and disease challenge.

Furthermore, the selenoenzyme iodothyronine deiodinase is involved in thyroid hormone activation and body thermoregulation in general. Therefore better Se status in the second scenario would be associated with better thermoregulation in neonates.

Though this scenario affords better antioxidant protection than when inorganic Se is added to feeds, there is a limitation to the extent to which stresses can be ameliorated. For example, when very high levels of toxins are present in the diet or environmental stresses are too high, the body cannot respond sufficiently to prevent pathobiological changes. On the other hand, this scenario is very effective over a wide range of everyday stress conditions. Common stresses that can compromise health if antioxidant protection is inadequate include:

Oxidized fat in the diet. Oxidation of fats during storage produces toxic peroxides and further stimulates lipid peroxidation.

Mycotoxins or any other toxic compounds. Mycotoxins have pro-oxidant effects, which can stimulate lipid peroxidation in the digestive tract as well as in stored feed.

Lack of exercise or over-exercising. Both can cause oxidative stress for companion animals.

Disease challenge. This is the most important stress. Immune cells themselves produce free radicals as a weapon to kill pathogens. Therefore, disease challenge is associated with increased free radical production which can damage cellular machinery if antioxidant enzymes are not sufficiently active. In addition, Se has a specific role in immune system regulation, which could be independent of its antioxidant functions.

Vaccination. Vaccination is an immune challenge, and is also a substantial stress. In some cases using vitamin E, for example, as a vaccine adjuvant can help improve vaccination efficiency.

Various medications in the diet. Some compounds can interfere with antioxidant absorption or assimilation.

The types or sources of stressors can vary with the species of animal, however overproduction of free radicals and the critical need for antioxidant protection are common to all. The overall relationship of antioxidants with fertility and immunity of companion animals is shown in Figure 6. Indeed, antioxidant defence and antioxidant/pro-oxidant balance in the body are major determinants of many physiological functions and more attention should be paid to this important issue. New forms of trace minerals in conjunction with stabilised forms of vitamin E available on the market today could substantially improve antioxidant defence, with consequent effects on health and reproduction.


Conclusions

The studies reviewed in this paper make it clear that effects of antioxidants on companion animals are very similar to those described for humans and other animals, since they share common environments, stresses and in some cases diseases.

Therefore, interaction between antioxidants and pro-oxidants begins at the level of the stomach and small intestine as described for humans (see Surai, 2002 in this volume). Therefore, dietary supplementation with various antioxidants is an effective strategy to prevent damaging effects of free radicals and toxic products of their metabolism in companion animals. In particular, specific attention should be paid to the form of dietary antioxidant used. For example, organic Se (selenomethionine) can improve the antioxidant capacity of the digesta while selenite in the same conditions could promote peroxidation. In addition, organic Se allows tissue reserves to accumulate against periods of increased Se demand. Furthermore, inorganic Cu and Fe are the major stimulators of lipid peroxidation in the digestive tract; and use of organic forms of those elements could avoid or mimimize detrimental actions.

Use of various plant-derived antioxidants such as flavonoids, could also help to prevent oxidative damage in the gastrointestinal tract owing to their antioxidant properties and ability to chelate Fe and Cu.

Of particular interest to the companion animal nutritionist are data showing protective effects of natural antioxidants during animal and human exercise as well as their immunomodulating properties. It is well known that vitamin E and carotenoids are nontoxic for animals even at very high leves of supplementation. However, more research is needed to establish optimal antioxidant supplementation of the companion animals.

Figure 6. Antioxidants in relation to fertility and immunity.

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PUBLICATION DATE:  03/05/2007

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AUTHOR:  PETER F. SURAI and JULIA E. DVORSKA - Avian Science Research Centre/Sumy State Agrarian University (Courtesy of Alltech Inc.)

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Estados Unidos de América - Dunboyne

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