Vitamin E activity in food is derived from a series of compounds of plant origin, the tocopherols and tocotrienols. The term vitamin E, according to the International Union of Pure and Applied Chemistry-International Union of Biochemistry (IUPAC-IUB) Commission on Biochemical Nomenclature, is used as a generic descriptor for all tocol and tocotrienol derivatives that qualitatively exhibit the biologic activity of alpha-tocopherol (IUPAC-IUB, 1973). Both the tocols (tocopherols) and tocotrienols consist of a hydroquinone nucleus and an isoprenoid side chain. Characteristically, tocols have a saturated side chain, whereas the tocotrienols have an unsaturated side chain containing three double bonds. For a more detailed description and an early assessment of tocopherol chemistry, refer to Pennock et al. (1964). Four isomers of each of these two classes of vitamin E exist (alpha, beta, gamma, delta), differentiated by the presence of methyl-groups at positions 5, 7 or 8 of the chromanol ring (Figure 4-1). Early researchers (Evans et al., 1936) isolated alpha-tocopherol from wheat germ oil. Emerson et al. (1937) were subsequently able to isolate beta-tocopherol and gamma-tocopherol. Stern et al. (1947) isolated delta-tocopherol from soybean oil and described its properties in the 1940s. Alpha-tocopherol, the most biologically active of these compounds is the predominant vitamin E active compound in feedstuffs, while the biological activity of the other tocols is limited (Table 4-1). However, some new functions have recently been found for non alpha-tocopherol forms of vitamin E (Schaffer et al., 2005; Freiser and Jiang, 2009). For a comprehensive review comparing the vitamin E biopotency of various “natural-derived” and chemically synthesized alpha-tocopherols, see Scherf et al. (1996).
Adapted from Machlin (1984)
Alpha-tocopherol is a yellow oil that is insoluble in water but soluble in organic solvents. Tocopherols are extremely resistant to heat but readily oxidized. Natural vitamin E is subject to destruction by oxidation, which is accelerated by heat, moisture, rancid fat, copper and iron. Alpha-tocopherol is an excellent natural antioxidant that protects carotene and other oxidizable materials in feed and in the body. However, in the process of acting as an antioxidant it is destroyed. Commercially available sources of vitamin E activity are shown in Table 4-2. Because there are three centers of asymmetry in alpha-tocopherol, eight stereoisomers are possible (Ames, 1979). Differences in biopotency of the stereoisomers of alpha-tocopherol can be seen from the definition of International Unit (IU). According to The United States Pharmacopeia (1980), dl-alpha-tocopheryl acetate (Illus 4-1) also called dl-rac-tocopheryl acetate) is the International Standard of vitamin E activity with 1 IU equivalent to 1 mg of dl-alpha-tocopheryl acetate (Table 4-2). This is the most widely available source of vitamin E activity for the supplementation of animal feeds. The acetate ester of d- or dl-alpha-tocopherol is synthesized to stabilize the compound from oxidation and maintain vitamin E activity.
Synthetic alpha-tocopherol (dl-or all-rac-alpha-tocopherol) is not identical to the naturally derived tocopherol (d- or RRR-alpha tocopherol). On the basis principally of rat bioassay work and using dl-alpha-tocopheryl acetate as a standard (1 mg=1 IU), 1 mg, dl-alpha-tocopheol equals 1.1 IU, 1 mg d-alpha-tocopheryl acetate equals 1.36 IU, and 1 mg d-alpha-tocopherol equals 1.49 IU of vitamin E. However, for some studies in several species including swine the naturally derived RRR, vitamin E form compared to the synthetic dl-alpha-tocopherol has been shown to be more effective in elevating milk, colostrum, plasma and tissue alpha-tocopherol concentrations when administered on an equil IU basis (Jensen et al. 2006). Pigs discriminate between RRR-alpha-tocopheryl acetate and all-rac-alpha-tocopheryl acetate with a preference for RRR-alpha-tocopheryl acetate, thus the official bioequivalence ratio of 1.36:1 RRR- to all-rac-α-tocopheryl acetate is underestimated (Mahan et al., 2000; Lauridsen et al., 2002; Wilburn et al., 2008; Yang et al., 2009). Feeding a higher dietary level of dl-alpha-tocopheryl acetate could circumvent the lower bioavailability of the dl-form-tocopheryl acetate. Early researchers investigated the effect of dietary fat on the intestinal absorption of tocopherol (Duncan et al., 1960). Vitamin E absorption is related to fat digestion and is facilitated by bile and pancreatic lipase (Sitrin et al., 1987). In humans, bioavailibility of vitamin E is dependent on fat content of the meal. Hays et al. (2001) reported that plasma alpha-tocopherol concentrations doubled when alpha-tocopheryl acetate (100 to 200 mg/day) was fortified in milk compared to orange juice. Jensen et al. (1997) found that the ileal digestibilities of tocopherols vary based on the dietary fat source. Whether presented as free alcohol or as esters, most vitamin E is absorbed as the alcohol. Esters are largely hydrolyzed in the gut wall, and the free alcohol enters the intestinal lacteals and is transported via lymph to the general circulation. Balance studies indicate that much less vitamin E is absorbed, or at least retained, in the body than is vitamin A. Vitamin E recovered in feces from a test dose was found to range from 65% to 80% in the human, rabbit and hen, although in chicks it was reported at about 25%. The efficiency of digestion and absorption of vitamin E varies with dietary inclusion level. At 10 IU per kg (4.5 IU per lb), there is about 98% uptake of vitamin E, while at 100 and 1,000 IU per kg (45.45 and 454.34 IU per lb), efficiency declines to 80% and 70%, respectively (Leeson and Summers, 2001). The naturally occurring tocopherol form is subject to destruction in the digestive tract to some extent, whereas the acetate ester is not. Much of the acetate is readily split off in the intestinal wall and the alcohol is reformed and absorbed, thereby permitting the vitamin to function as a biological antioxidant. Any acetate form absorbed or injected into the body is converted to the alcohol form. Vitamin E in plasma is attached mainly to lipoproteins in the globulin fraction. An alpha-tocopherol transfer protein has been identified (Traber, 2006). Rates and amounts of absorption of the various tocopherols and tocotrienols are in the same general order of magnitude as their biological potencies. Alpha-tocopherol is absorbed best, with gamma-tocopherol absorption 85% that of alpha- forms and with a more rapid excretion. One can generally assume that most of the vitamin E activity within plasma and other animal tissues is alpha-tocopherol (Ullrey, 1981). Lindberg (1973) reported that average values for tocopherol in plasma from pigs are much lower than those measured in plasma from dogs and that tocopherol levels are higher in pigs approaching slaughter weight than in newly weaned pigs. Froseth (1979) reviewed a variety of studies designed to evaluate effects of dietary additions of selenium, vitamin E and various inorganic elements on selenium and vitamin E metabolism, including serum tocopherol levels and tissue selenium. The diet of the sow influences stores in the young pig at birth and the amount obtained from mother’s milk. However, vitamin E is not highly effective in passing through swine placental membranes (Mahan, 1990; Lauridsen et al., 2002). Thus, vitamin E reserves of pigs at farrowing are low (Urbanova and Toulova, 1975). As placental transfer of tocopherol from dam to fetus is minimal, the importance of milk for enhancing the vitamin E status of the newborn is greater (Lauridsen, 2002).
Vitamin E is stored throughout all body tissues. Data regarding vitamin E storage in porcine liver offer conflicting conclusions. In some cases, it would appear that the liver contains the greatest stored vitamin E compared to other tissues (Jensen et al., 1990). However, Hoppe et al. (1993) indicated that adipose tissue contained the highest concentrations of vitamin E followed by liver, cardiac muscle and M. longissimus in decreasing order. Hoppe et al. (1993) suggested the discrepancy might be due to whether or not the pigs were sacrificed with or without fasting. The authors noted in their study that the pigs were slaughtered after 24 hours of fasting, which likely caused partial depletion of liver vitamin E. The results by Jensen et al., (1990) would support this explanation. In this work, the liver depletion of vitamin E was relatively rapid, while vitamin E levels in fat and muscle remained unchanged during a one-week depletion period. Liver contains only a small fraction of total body stores, in contrast to vitamin A, for which about 95% of the body reserves are in the liver. Grela and Jakobsen (1994) reported that 0.05% dl-alpha-tocopherol resulted in a greater content of vitamin E in depot fat than did 6.1 mg alpha-tocopheryl acetate per 100 g of feed, when either was supplemented to a standard diet with 10% soybean oil. Small amounts of vitamin E will persist tenaciously in the body for a long time. However, stores are exhausted rapidly by polyunsaturated fatty acids (PUFA) in the tissues, the rate of disappearance being proportional to the intake of PUFA. A major excretory route of absorbed vitamin E is bile, in which tocopherol appears mostly in the free form (McDowell, 2000).
Vitamin E has been shown to be essential for integrity and optimum function of reproductive, muscular, circulatory, nervous and immune systems (Hoekstra, 1975; Sheffy and Schultz, 1979; Bendich, 1987; McDowell, 2000). It is well established that some functions of vitamin E, however, can be fulfilled in part or entirely by traces of selenium or by certain synthetic antioxidants. Even sulfur-bearing amino acids, cystine and methionine affect certain vitamin E functions. The effects of methionine have been considered in vitamin E-selenium studies (Reid et al., 1968; Sharp et al., 1972a). Much evidence points to undiscovered metabolic roles for vitamin E that may be paralleled biologically by roles of selenium and possible other substances. The most widely accepted functions of vitamin E are discussed in this section.
Vitamin E reacts or functions as a membrane-bound antioxidant, trapping lipid peroxyl free radicals produced from unsaturated fatty acids under conditions of “oxidative stress”. Orientation of vitamin E within cell membranes appears to be critical to its functionality (Dunnett, 2003). Lipids, especially phopholids present in cell membranes are particularly susceptible to oxidative damage, being positively correlated with the degree of unsaturation of its fatty acids. This function is closely related to and synergistic with the role of selenium. Selenium has been found to be part of 25 selenoproteins with most of the functions unknown, although these selenoproteins generally participate in antioxidant and anabolic processes (Hatfield and Gladyshev, 2002). Selenium has been shown to act in aqueous cell media (cytosol and mitochondrial matrix) by destroying hydrogen peroxide and hydroperoxides via the enzyme glutathione peroxidase (GSH-Px) of which it is a co-factor. In this capacity, it prevents oxidation of unsaturated lipid materials within cells, thus protecting fats within the cell membrane from breaking down. If lipid hydroperoxides are allowed to form in the absence of adequate tocopherols, direct cellular tissue damage can result, in which peroxidation of lipids destroys structural integrity of the cell and causes metabolic derangements.The various GSH-Px enzymes are characterized by different tissue specificities and are expressed from different genes. In general, different forms of GSH-Px perform their protective functions in concert, with each providing antioxidant protection at different sites of the body.
Highly reactive oxygen species such as superoxide anion radical (O2-), hydroxyl radical (OH), hydrogen peroxide (H2O2) and singlet oxygen (O2) are continuously produced in the course of normal aerobic cellular metabolism. Also, phagocytic granulocytes undergo respiratory burst to produce oxygen radicals that destroy intracellular pathogens. However, these oxidative products can, in turn, damage healthy cells if they are not eliminated. Antioxidants serve to stabilize these highly reactive free radicals, thereby maintaining the structural and functional integrity of cells (Chew, 1995). Therefore, antioxidants are very important to immune defense and the health of humans and animals.
Vitamin E reacts or functions as a chain-breaking antioxidant, thereby neutralizing free radicals and preventing oxidation of lipids within membranes. At least one important function of vitamin E is to interrupt production of free radicals at the initial stage. Myodystrophic tissue is common in cases of vitamin E-selenium deficiency with leakage of cellular compounds such as creatinine and various transaminases through affected membranes into plasma. Fontaineet al. (1977a) reported that serum creatine phosphokinase activity increases are associated with the occurrence of subclinical muscular dystrophy and that vitamin E and selenium deficiencies have marked additive effects on the induction of skeletal muscular disease in pigs. The more active the cell (e.g., the cells of skeletal and involuntary muscles), the greater is the inflow of lipids for energy supply and the greater is the risk of tissue damage if vitamin E is limiting. This antioxidant property also ensures erythrocyte stability and maintenance of capillary blood vessel integrity.
Interruption of fat peroxidation by tocopherol explains the well-established observation that dietary tocopherols protect or spare body supplies of such oxidizable materials as vitamin A, vitamin C and the carotenes. Certain deficiency signs of vitamin E (i.e., muscular dystrophy) can be prevented by diet supplementation with other antioxidants. This tends to support the antioxidant role of tocopherols. Semen quality of boars was improved with selenium and vitamin E supplementation, with vitamin E playing a role in maintaining sperm integrity in combination with selenium (Marin-Guzman et al., 1989). In 1995, Brzezinska-Slebodzinska et al. evaluated the antioxidant effect of vitamin E and glutathione on lipid peroxidation in boar semen plasma. Seven weeks of oral dl-alpha-tocopheryl acetate (1,000 IU/day/boar) caused a decrease in the level of semen plasma thiobarbituric acid reactive substances, the measurement chosen to reflect lipid peroxidation. Furthermore, vitamin E supplementation significantly increased the number of spermatozoa per cubic centimeter of ejaculate. The authors indicated that vitamin E protects boar semen against fatty acid peroxidation and has a positive influence on semen quality. Marin-Guzman et al. (1997) confirmed that both dietary vitamin E and selenium can affect boar semen quality, but reported that the greater effect seemed to be from selenium. These authors also suggested that selenium and vitamin E might act in separate roles with vitamin E perhaps maintaining semen and sperm quality through its antioxidant properties.
Chemical antioxidants are only stored at very low levels, thus they are not as effective as tocopherol. It is clear that highly unsaturated fatty acids in the diet increase vitamin E requirements (McDowell, 2000). When acting as an antioxidant, vitamin E supplies become depleted, thus furnishing an explanation for the often observed fact that the presence of dietary unsaturated fats (susceptible to peroxidation) augments or precipitates a vitamin E deficiency.
In 1987, Duthie et al. proposed that stress-susceptibility syndrome may be associated with anomalous vitamin E metabolism. Subsequently, Duthie et al. (1989) reported that the results of their study indicated that stress-susceptible pigs have antioxidant abnormalities, which can be partially compensated for by increasing dietary vitamin E supplementation. Therefore, vitamin E supplementation may reduce the number of stress-related deaths among stress-susceptible pigs. In a related study, Hoppe et al. (1989) evaluated the effects of adding ascorbic acid in addition to vitamin E to increase the antioxidant content of the ration. These authors reported that in stress-susceptible pigs, antioxidant supplementation had a significant protective effect on cell membrane integrity.
In order to improve the oxidative stability, and thus increase the shelf life of meat, antioxidants have been successfully added to animal feeds. In the last few years, different compounds such as carotenoids, vitamin C, selenium and plant extracts have been tested in different experiments to verify their potential antioxidant effect on pork quality (Kerth et al., 2001; Hasty et al, 2002; Peeters et al., 2005; Guo et al., 2006a, 2006b; Morel et al., 2008). Of them all, alpha-tocopherol demonstrated the highest biological efficiency in preventing the lipid oxidation in vivo.
Alpha-tocopherol may be involved in the formation of structural components of biological membranes, thus exerting a unique influence on architecture of membrane phospholipids (Ullrey, 1981). It is reported that alpha-tocopherol stimulated the incorporation of 14C from linoleic acid into arachidonic acid in fibroblast phospholipids. Also, it was found that alpha-tocopherol exerted a pronounced stimulatory influence on formation of prostaglandin E from arachidonic acid, while a chemical antioxidant had no effect.
Vitamin E is an inhibitor of platelet aggregation in pigs (McIntosh et al., 1985), and may play a role by inhibiting peroxidation of arachidonic acid, which is required for formation of prostaglandins involved in platelet aggregation (Panganamala and Cornwell, 1982; Machlin, 1984).
Vitamin E is perhaps the most studied nutrient related to the immune response (Meydani and Han, 2006). Evidence accumulated over the years and in many species indicates that vitamin E is an essential nutrient for normal function of the immune system. Furthermore, studies suggest that beneficial effects of certain nutrients, such as vitamin E on reducing disease risk, can be through their effects on the immune response. Deficiency in vitamin E impairs B- and T-cell-mediated immunity. Vitamin E acts in part by reducing prostaglandin synthesis and by preventing the oxidation of PUFAs in cell membranes (Shankar, 2006).
Finch and Turner (1996) have reviewed the effects of selenium and vitamin E on the immune responses of domestic animals. The authors noted the significance of the basal nutritional status, type of supplements used and the route and timing of the different sources of selenium and vitamin E utilized. Considerable attention is presently being directed to the role vitamin E and selenium play in protecting leukocytes and macrophages during phagocytosis, the mechanism whereby animals immunologically kill invading bacteria. Both vitamin E and selenium may help these cells survive the toxic products produced to effectively kill ingested bacteria (Badwey and Karnovsky, 1980).
Since vitamin E acts as an in vivo tissue antioxidant and aids in quenching free radicals produced in the body, any infection or other stress factor may exacerbate depletion of the limited vitamin E stores from various tissues. With respect to immunocompetence higher dietary levels have been shown to positively influence both cellular and humoral immune status of young pigs. These two responses are generally used as criteria for determining the requirement of a nutrient. Vitamin E is one of the few nutrients for which higher-than-recommended doses can further enhance certain immune functions, at least temporarily. The optimal dose of vitamin E for stimulating immune function has not been determined (Shankar, 2006). During stress and disease, there is an increase in production of glucocorticoids, epinephrine, eicosanoids, and phagocytic activity. Eicosanoid and corticoid synthesis and phagocytic respiratory bursts are prominent producers of free radicals, which challenge the animal’s antioxidant systems. Due to a number of factors, the antioxidant vitamin E status of an animal may be marginal or deficient prior to and after stress and disease.
Since vitamin E status is integral to an animal’s health and productivity, it is imperative to be able to determine the status. Vitamin E has effectively improved the humoral immune response of a variety of species when challenged with nonliving antigens, bacteria or a virus. Resistance to disease was significantly greater in chicks and turkeys challenged with Escherichia coli or sheep inoculated with Chlamydia (Nockels, 1979). Short term inoculation of pigs with E. coli led to a decreased liver alpha-tocopherol status (Lauridsen et al., 2011). Vitamin E and selenium supplementation in young pigs was beneficial in increasing whole cell agglutination against E. coli (Ellis and Vorhies, 1976). In this 56-day study, six- to eight-week-old pigs were supplemented with 0, 22 or 110 IU of vitamin E per kg (0, 10 or 50 IU per lb) of diet. Each pig was injected with E. coli bacteria intramuscularly on day 0 and 35. Supplementation with vitamin E increased antibody titers of pigs to E. coli bacteria 1.5 and 2.7 times for the 22 and 110 IU per kg (10 and 50 IU per lb) treatments, respectively, compared to unsupplemented controls. Teige et al. (1977) also found that the response to E. coli endotoxin was affected by the vitamin E supply available to pigs. Hidiroglou et al. (1995) reported that when piglets were injected intramuscularly with 500 IU of alpha-tocopherol at birth and 1,000 IU of alpha-tocopherol at 7 and 14 days of age, their antibody titers to Keyhole limpet hemocyanin were significantly higher than those of the control piglets.
Likewise, Morrow et al. (1987) reported that vitamin E improved the health status of pigs from sows housed in confinement and that it was possible to enhance the cellular and humoral immune responses of these young pigs with vitamin E as compared to pigs from sows that were kept on alfalfa pasture during gestation. These authors also indicated that pigs injected at birth and 10 days of age with 400 IU of vitamin E were heavier than controls at 28 days of age. Babinszky et al. (1991a) investigated the effects of dietary vitamin E in pregnant and lactating sow diets on serum vitamin E concentration and on cell-mediated and humoral immune response in suckling and weaned piglets. Not only did vitamin E in the sow’s diet increase serum vitamin E concentrations in the one-week-old piglets, but phagocytic measure of the piglets was also increased. Furthermore, the immune response against ovalbumin was increased at one week after immunization for weaned pigs from litters fed high levels (136 mg alpha-tocopherol per kg (62 mg/lb) of feed) of vitamin E. In a subsequent study, Babinszky et al. (1991b) indicated that the humoral immune system of lactating sows may be affected by the combinations of alpha-tocopherol and fat that are included in gestation and lactation diets. They did not find evidence that the cell-mediated immunity of sows was affected by either alpha-tocopherol or fat addition to their diets. Mudron et al. (1996) reported that T-lymphocyte percentages and the metabolic activity of phagocytes were greater after injection of vitamin E (20 mg tocopheryl acetate per kg (44 mg per lb) body weight).
Teige et al. (1977; 1978a) reported that when pigs were fed minced colon from swine with dysentery or with a pure culture of Treponema hyodysenteriae, vitamin E and selenium supplementation in a deficient diet increased resistance to the disease. In a separate study, Teige et al. (1982) indicated that the effect of selenium on T. hyodysenteriae was greater than the effect of vitamin E. Larsen and Tollersrud (1981) indicated that dietary vitamin E or selenium increased the phytohemagglutinin (PHA) response of peripheral pig lymphocytes. Furthermore, addition of vitamin E increased the PHA response regardless of the levels of selenium. Peplowski et al. (1981) found an additive effect of vitamin E and selenium, which increased in hemagglutination titers when these nutrients were provided either through dietary sources or injections. Jensen et al. (1988a) reported that lymphocyte response to pokeweed mitogen was at least twice as high in pigs with serum vitamin E values above 3 mg per liter compared with lymphocytes from animals with lower concentrations of serum vitamin E.
Hayek et al. (1989) reported that supplementation with vitamin E (1,000 IU) and (or) selenium (5 mg) as a single injection administered on day 100 of gestation augmented the transfer of immunity to pigs from sows fed levels of vitamin E and selenium that approximated the sow’s requirement (NRC, 1998). Colostral immunoglobulin transfer is essential to the pig’s immunity and survival, as the newborn pig is devoid of circulating antibodies (Bourne et al., 1978) and must attain maternal immunoglobulins via colostrum during the first few hours postpartum (Butler, 1984). Okere and Hacker (1997) evaluated whether prepartum injections of vitamin E and (or) selenium to sows were effective in enhancing reproductive performance and in augmenting colostral transfer of immunoglobulins to piglets. Piglet serum IgG was significantly higher in the treatment groups at 24 hours postpartum, seven days of age and at weaning. These authors hypothesized that immune deficient porcine neonates could benefit from the improved IgG transfer, which was attributed to vitamin E and (or) selenium supplementation of the sows. Nemec et al. (1994) were not able to consistently confirm the positive effects of vitamin E supplementation on immunoglobulin transfer in their study. Vitamin E-supplemented sows did have a significant dose-dependent increase in milk IgG levels but only on day 14 after parturition. However, for gilts that received 110 or 220 IU of vitamin E per kg (50 or 100 IU/lb) of diet during lactation, offspring required less time to acquire phytohemagglutinin response and displayed a greater concanavalin A response than did piglets from sows supplemented with 55 IU of vitamin E per kg (25 IU/lb). Thus, the maternal vitamin E supplementation improved the ability of newborn pigs to acquire cellular immunity as measured by these responses.
Data from Wuryastuti et al. (1993) implied that if gestating sows do not obtain sufficient vitamin E and (or) selenium, the sows and their piglets will be more susceptible to disease in the peripartum period. Selenium restriction impaired neutrophil function while vitamin E restriction affected both lymphocytes and neutrophils. The authors concluded that maintaining 0.3 mg of selenium per kg of diet for swine is justified and that the immune response should be considered when the NRC reevaluates vitamin E requirements. Bonnette et al. (1990) evaluated the influence of supplemental dietary vitamin E (11 or 220 IU per kg) (5 or 100 IU/lb) diet) on performance, humoral antibody production and serum cortisol levels of young pigs. The higher level of supplemental vitamin E increased serum vitamin E concentration, but performance, cortisol levels, immune response and antibody titers in red blood cells were not affected as measured under the conditions of this experiment. The authors concluded that pigs nursed by sows fed NRC-estimated nutrient requirements for vitamin E and selenium have sufficient stores of vitamin E at weaning so that providing the young pigs 220 versus 11 IU vitamin E per kg (100 versus 5 IU/lb) of diet offers no advantage. These results confirm the findings obtained in a separate study by Bonnette et al. (1990b). Goss and Bilkei (1994) were unable to detect a significant influence of dietary vitamin E supplementation to pregnant sows on perparturient hypogalactia syndrome (PHS) or on postnatal piglet losses. However, these authors did observe that supplementing the sow rations with vitamin E during gestation resulted in a significant weaning litter weight improvement. The authors suggested that possibly a subclinical manifestation of PHS in the sows fed the unsupplemented diet might have resulted in less milk production, which contributed to the lowered weight gain in these piglets.
Lipopolysaccharide (LPS) administration is a well-documented model for evaluating disease stress, and the response to LPS can be attributed to events that include cytokine synthesis and release (Dinarello, 1996; Webel et al., 1997). Cytokines are released from activated macrophages in response to immunologic challenge and are primarily responsible for the subsequent metabolic effects (Johnson, 1997). Webel et al. (1998) investigated whether or not vitamin E might affect pro-inflammatory cytokine production. In their in vivo study, effects of intramuscular injection of vitamin E (d-alpha-tocopherol) on plasma interleukin-6 (IL-6) and cortisol in pigs subjected to a challenge dose of LPS were evaluated. Pigs that received vitamin E prior to the LPS challenge had substantially lower peak levels of IL-6 and cortisol than pigs not provided with vitamin E injections. The authors suggested that the improved survival and growth performance of pigs after weaning noted by other authors, who provided pigs with vitamin E injections, might be partially due to the reduction of excess pro-inflammatory cytokines during the stressful postweaning period.
Antioxidants, including vitamin E, play a role in resistance to viral infection. Vitamin E deficiency allows a normally benign virus to cause disease (Beck et al., 1994). In mice, enhanced virulence of a virus resulted in myocardial injury that was prevented with adequate vitamin E. A selenium or vitamin E deficiency leads to a change in viral phenotype, such that a non-virulent strain of a virus becomes virulent and a virulent strain becomes more virulent (Beck, 1997, 2007; Sheridan and Beck, 2008). Thus, host nutritional status should be considered a driving force for the emerengence of new viral strains or newly pathogenic strains of known viruses.
There is limited evidence that vitamin E is involved in biological oxidation-reduction reactions (Hoffmann-La Roche, 1972). Vitamin E also appears to regulate the biosynthesis of DNA within cells. Vitamin E appears to be of particular importance in cellular respiration in heart and skeletal muscles (Leeson and Summers, 2001).
Both vitamin E and selenium provide protection against toxicity of various heavy metals (Whanger, 1981). Vitamin E is highly effective in reducing toxicity of metals such as silver, arsenic and lead and shows slight effects against cadmium and mercury toxicity. Vitamin E but not selenium, was found to be effective in preventing the development of lesions induced by silver supplementation in weanling pigs’ diets (Van Vleet, 1976). Miller (1971) reported that supplemental vitamin E partially prevented copper toxicity. Based on results of their study, Van Vleet et al. (1981) suggested that increased amounts of selenium and vitamin E might be needed to prevent the development of selenium-vitamin E deficiency in animals fed rations containing large concentrations of several trace elements (silver, tellurium, cobalt, zinc, cadmium or vanadium). Tollerz (1973) indicated that certain pigs have an abnormally high sensitivity to iron and that vitamin E or selenium supplementation raised the level of tolerance to iron in piglets and other species. Nielsen et al. (1979) reported that iron toxicity was observed in piglets that were not supplied with adequate dietary vitamin E. Vitamin E can be effective against other toxic substances. For example, treatment with vitamin E gave protection to weanling pigs against monensin-induced skeletal muscle damage (Van Vleet et al., 1987).
There is a close working relationship between vitamin E and selenium within tissues. Selenium has a sparing effect on vitamin E and delays onset of deficiency signs. Likewise, vitamin E and sulfur amino acids partially protect against or delay onset of several forms of selenium deficiency syndromes. Tissue breakdown occurs in most species receiving diets deficient in both vitamin E and selenium, mainly through peroxidation. Peroxides and hydroperoxides are highly destructive to tissue integrity and lead to disease development. It now appears that vitamin E in cellular and subcellular membranes is the first line of defense against peroxidation of vital phospholipids (Figure 4-2), but even with adequate vitamin E, some peroxides are formed. Berlin et al. (1994) suggested that vitamin E may be involved in a homeostatic mechanism that preferentially protects certain organs from peroxidation. In their study, alpha-tocopherol levels were reduced in the liver but not in the heart of pigs fed diets containing 15% fat from menhaden oil. Selenium, as part of the enzyme glutathione peroxidase (GSH-Px), is a second line of defense that destroys these peroxides before they have an opportunity to cause damage to membranes. Therefore, selenium, vitamin E and sulfur-containing amino acids, through different biochemical mechanisms, are capable of preventing some of the same nutritional diseases. Vitamin E prevents fatty acid hydroperoxide formation, sulfur amino acids are precursors of GSH-Px and selenium is a component of GSH-Px (Smith et al., 1974).
Although alpha-tocopherol has been the most widely studied form of vitamin E, other tocopherols and tocotrienols have recently been shown to have biological significance (Qureshi et al., 2001; Eder et al., 2002; McCormick and Parker, 2004; Schaffer et al., 2005; Nakagawa et al., 2007; Sun et al., 2008; Freiser and Jiang, 2009). The greater emphasis on alpha-tocopherol undoubtedly arises from observations that gamma-tocopherol and delta-tocopherol are only 10% and 1% as effective as alpha-tocopherol, respectively, in experimental animal models of vitamin E deficiency. Tocotrienols have been shown to possess excellent antioxidant activity in vitro and have been suggested to suppress reactive oxygen substances more efficiently than tocopherols (Schaffer et al., 2005). Studies have shown that tocotrienols exert more significant neuroprotective, anti-cancer and cholesterol-lowering properties than do tocopherols (Qureshi et al., 2001; Sun et al., 2008). Gamma-tocopherol has beneficial properties as an anti-inflammatory and possibly anti-atherogenic and anti-cancer agent (Wolf, 2006). Research with tocotrienols and non-alpha-tocopherol, has been carried out with laboratory animals and in vitro studies; the significance for farm animals is unknown.
Additional functions of vitamin E that have been reported (Scott et al., 1982) include (1) normal phosphorylation reactions, especially of high-energy phosphate compounds, such as creatine phosphate and adenosine triphosphate (ATP), (2) a role in the synthesis of vitamin C (ascorbic acid), (3) a role in synthesis of ubiquinone, and (4) a role in sulfur amino acid metabolism. Pappu et al. (1978) have reported that vitamin E plays a role in vitamin B12 metabolism. A deficiency of vitamin E interfered with conversion of vitamin B12to its coenzyme 5′-deoxyandenosylcobalamin and concomitantly metabolism of methylmalonyl-CoA to succinyl-CoA.
The NRC (1998) requirement for vitamin E in growing swine varies from 11 to 16 IU per kg (5 to 7 IU per lb) of diet. Limited information is available on the vitamin E requirements for reproduction. The NRC (1998) has estimated 44 IU per kg (20 IU per lb) of diet is required for breeding and lactating swine. After conducting a survey of the Australian pig industry, Cargill et al. (1994) concluded that levels above 35 mg alpha-tocopherol per kg (15.9 mg/lb) of diet are required to maintain adequate plasma tocopherol levels in sows and piglets at weaning. These authors also concluded that tocopherol levels in sow diets appear to have a larger influence on the alpha-tocopherol status of weanling piglets than do weanling pig diets. Thus, feeding diets adequate in alpha-tocopherol to sows during gestation and lactation to prevent problems for the piglets after weaning, was strongly recommended. Mahan (1991) found that supplemental vitamin E levels less than 16 IU per kg (7.3 IU/lb) of diet were inadequate for sows. If sows were not provided adequate dietary vitamin E, the author indicated that smaller litter size, sow agalactia and pig mortality during the first week after birth might be exacerbated. Increasing the sows’ dietary vitamin E level was found to improve the alpha-tocopherol status of the nursing piglets throughout lactation in addition to proving beneficial during the postweaning period. Mahan concluded that even if some tissue tocopherol might be mobilized, the main source of vitamin E for reproducing sows, nursing pigs and weaned pigs should be the diet. In a subsequent study, Mahan (1994) indicated that vitamin E requirements of reproducing sows are higher than the 1988 NRC recommendations of 10 to 22 IU per kg of diet. In his 1994 report, Mahan indicated that litter size improved and the incidence of mastitis, metritis and agalactia was reduced when 44 or 66 IU dl-alpha-tocopheryl acetate per kg (20 or 30 IU per lb) of diet was provided during gestation and lactation. These higher dietary vitamin E levels resulted in a better vitamin E status of piglets at weaning compared to the piglets from the sows fed the lower dietary level. Grandhi et al. (1993) did not detect a significant response in growth rate, feed intake or feed efficiency when gilts were fed vitamin E at above 1988 NRC-recommended levels during prepubertal development. Grandhi et al.(1993) did suggest, ,however, that gilts may require a higher supplementation of vitamin E during prepubertal development (50 or 100 mg per kg of diet) (22.7 or 45.4 per lb) for improved estrus and a minimum of 50 IU per kg during prebreeding and gestation. The effect of vitamin E levels in sow diets on postweaning progeny and vitamin E deficiency lesions postweaning was studied by Mahan (1985). Sows were fed a corn-soybean meal diet with no supplemented selenium. Supplemental vitamin E was added at the rate of either 0, 44 or 220 IU per kg (0, 20 or 100 IU per lb) of diet. Results suggest that sows deplete tissue tocopherol from the first to second parity. Progeny from sows not supplemented with vitamin E had a higher incidence of cardiac muscle degeneration, liver necrosis and ceroid pigmentation of body fat than those of supplemented sows at four weeks postweaning. All pigs from unsupplemented sows in the second parity exhibited gut edema; no offspring of supplemented sows were affected in this manner. Also, sows from the second parity supplemented with 220 IU per kg (100 IU per lb) of diet had no vitamin E deficiency signs at necropsy. Young et al. (1977) and Whitehair et al. (1983a) studied the effects of supplemental vitamin E and (or) selenium in diets containing high moisture corn and soybean meal. Young et al. (1977) found that supplementation of gilt diets with selenium or vitamin E led to increased levels of these nutrients in the gilts’ serum and colostrum as well as in their offspring’s serum. The vitamin E supplementation of the sows had the greatest effect on the piglets through the colostrum versus the placental transfer of vitamin E. In the Whitehair et al.(1983a) study, sows were fed high-moisture corn with or without 50 IU per kg (23 IU per lb) of supplemental vitamin E and 0.10 ppm selenium. Supplemented sows had a greater number of pigs born and weaned, as well as increased litter weight gains compared to unsupplemented sows. Also, 50% of the unsupplemented sows had clinical signs of mastitis-metritis-agalactia (MMA). Vitamin E-supplemented sows were not observed with clinical MMA. Bertsch et al. (1975) investigated the effect of dietary vitamin E on the subsequent uptake of radioactive selenium by swine erythrocytes in vitro and also on reproductive performance. The authors concluded that dietary vitamin E partially alleviated the increased need for dietary selenium due to lactation stress. They also reported that hypogalactia was less pronounced in supplemented sows although the incidence of MMA and reproductive performance were not significantly influenced by dietary vitamin E (150 mg of dl-alpha tocopherol per day during gestation and lactation plus an additional 50 mg per pig nursing).Sow colostrum and milk vitamin E concentrations are affected by dietary vitamin E. Mahan (1985) reported that, regardless of dietary vitamin E supplementation level, colostrum has a high alpha-tocopherol content that decreases rapidly as lactation proceeds. Pregnant gilts and sows supplied with adequate dietary vitamin E will produce healthy, normal baby pigs with limited amounts of vitamin E (Mahan, 1986). Nielsen et al. (1973) and Loudenslager et al. (1986) reported that swine colostrum has high concentrations of alpha-tocopherol compared to that of milk and this has been confirmed by others (e.g., Hidiroglou et al., 1993; Mahan, 1994). Higher concentrations of beta- and gamma-tocopherol were present in milk than in colostrum. As sows age, vitamin E content of colostrum decreases (Mahan, 1985). Consequently, the amount of vitamin E activity available to the pig decreases, resulting in poor vitamin E tissue status in the weaning pig. Mahan (1985) has also indicated that vitamin E status in the breeding female can be compromised with continued reproduction. These observations, coupled with the fact that non-alpha-tocopherols in milk at or near weaning have low vitamin E biological activity (10% to 40%), result in young pigs consuming low quantities prior to weaning. Meyer et al.(1981) reported that plasma tocopherol concentrations decreased in pigs postweaning. These data suggest that weaned pigs are highly susceptible to vitamin E deficiency, which may increase mortality at weaning or shortly afterward until consumption of a well-fortified vitamin E diet is adequate. Carrion et al. (1994) evaluated the effect of injectable (600 IU of d-alpha-tocopherol injected I.M. at breeding and at day 110 of gestation) versus dietary (23 IU of d-alpha-tocopherol acetate per lb) of vitamin E on tocopherol status of sows and their progeny over three parities. The researchers found that both dietary and injectable tocopherol supplementation enhanced tocopheryl status of sows (based on serum and milk tocopherol concentrations) and their offspring.Additionally, reproductive performance was improved for sows consuming a diet of 15 IU per kg (7 IU per lb) of vitamin E and 0.1 ppm selenium when injectable vitamin E and selenium were administered (Chavez and Patten, 1986). Litter size, total litter weight at birth and litter size at weaning were significantly greater for supplemented animals. Additionally, the effect of vitamin E and selenium on reproductive performance was more pronounced in older sows with three or more parities than in gilts. Piatkowski et al. (1979) reported that 0.1 ppm of added selenium and 22 IU of added vitamin E per kg (10 IU per lb) of diet appear necessary to maintain tissue vitamin E levels. The Agricultural Research Council (ARC, 1981) has suggested a higher requirement than the NRC (1998) for growing pigs: 20 to 50 IU per kg (9 to 23 IU per lb) of diet. In order to obtain a certain safety margin for prevention of vitamin E and selenium deficiency syndromes, Jensen et al. (1988b) suggested growing swine receive 30 IU of supplemental vitamin E per kg (14 IU per lb) of diet.
Myer et al. (1992) found no effect on farrowing percentage or the number of pigs born alive when sows were injected with 500 to 600 IU vitamin E and 10 to 12 mg selenium prior to breeding. Anderson et al. (1997) investigated the effect of vitamin A and (or) beta-carotene injections just before and (or) shortly after breeding and of dietary supplementation of vitamin E on blood and tissue concentrations of alpha-tocopherol in gestating gilts. Although previous authors had presented evidence with chicks and laboratory rats that very high dietary vitamin A may interfere with vitamin E absorption and blood alpha-tocopherol concentrations, Anderson et al. (1997) did not find any detrimental effect of three 350,000-IU injections of vitamin A shortly before, at and shortly after breeding on vitamin E status of gilts during early gestation. Increases in alpha-tocopherol concentration in serum and tissues were noted following dietary vitamin E supplementation as expected. Anderson et al. (1995a) also found no consistent evidence that high levels of dietary vitamin A interfere with performance or serum and tissue concentrations of alpha-tocopherol in growing-finishing swine. Hoppe et al. (1992) reported the effects of dietary retinol on plasma and tissue alpha-tocopherol in pigs. While no effects on M. longissimus and backfat alpha-tocopherol levels were evident, alpha-tocopherol levels in the heart and liver showed an inverse relationship with dietary retinol. Dietary retinol up to 10,000 IU per kg did not affect tocopherol concentrations except in cardiac muscle.
In boars, it was reported that vitamin E enhanced fertility even when supplementation was raised from 40 to 80 mg per kg (18 to 36 lb) feed (Westendorf and Richter, 1977). Not only was production improved, but semen volume and sperm concentration were increased. Marin-Guzman et al. (1997) reported that when diets low in selenium and vitamin E were fed, the percentage of abnormal sperm increased and sperm motility declined. Boars that were fed low-selenium diets had a higher percentage of abnormal sperm primarily due to abnormal tail morphology. In these experiments, Marin-Guzman and co-workers evaluated the effects of dietary selenium at 0 or 0.5 ppm and vitamin E at 0 or 220 IU per kg (0 or 100 IU/lb). They indicated that boars with a low selenium status had fewer sperm that reached and penetrated the zona pellucida, which could lower the fertilization rate. The authors stated that vitamin E had a less dramatic effect than did selenium. This led them to suggest that although both selenium and vitamin E have important roles in maintaining semen and sperm quality in boars, their effects are unique with vitamin E, perhaps acting through its antioxidant properties. Brzezinska-Slebodzinska et al. (1995) observed that supplementation with vitamin E increased the concentration of spermatozoa in semen, an effect possibly linked to the antioxidant properties of this vitamin
Vitamin E requirements are exceedingly difficult to determine because of the interrelationships with other dietary factors. The requirement may be increased with increasing levels of PUFA, oxidizing agents, vitamin A, carotenoids, gossypol, or trace minerals and decreased with increasing levels of fat-soluble antioxidants, sulfur amino acids or selenium (McDowell et al., 1996; Franklin et al., 1998). In otherwise adequate diets containing sufficient sulfur amino acids, selenium and a minimum of PUFA, vitamin E requirements appear to be low.
Although various vegetable oils (e.g., corn oil, cotton seed oil, peanut oil) contain PUFA they are less likely to result in a vitamin E deficiency because these oils are short-chained fatty acids (e.g., linoleic and linolenic) with only 2 or 3 double bonds. Fish oils, however, are very destructive to vitamin E as these long chained PUFA, containing 4 to 6 double bonds, can really destroy the vitamin (McDowell, 2000). The fish-oil PUFA which contain arachidonic acid (20:4), docosapentaenoic acid (22:5), and docosahexaenoic acid (22:6), require much more vitamin E for stabilization than typical plant PUFA, such as linoleic acid (18:2) and linolenic acid (18:3).
Simesen et al. (1979) reported that oxidized herring meal increased the requirement for vitamin E while oxidized lard had very little effect on the requirement. Gebert et al. (1999a) evaluated the requirements for vitamin E supplementation when dietary phytase was included in pig diets susceptible to lipid oxidation. They concluded that an extra dietary supplementation with alpha-tocopheryl acetate might be recommended when dietary phytase is included to avoid detrimental effects on fat digestibility. In a separate report, Gebert et al. (1999b) determined that the combined addition of vitamin E and phytase increased the cecal digestibility of dietary fat, oleic acid and linoleic acid.
The levels of PUFA found in plant and fish oils can increase vitamin E requirements, especially if these oils are allowed to undergo oxidative rancidity in the diet or are in the process of peroxidation when consumed by the animal. If they become completely rancid before ingestion, the only damage is the destruction of the vitamin E present in the oil and in the feed containing the rancidifying oil. But if they are undergoing active oxidative rancidity at the time of consumption, they apparently cause destruction of body stores of vitamin E as well (Scott et al., 1982). A combination of the stress of infection and the presence of oxidized fats in swine diets was reported to exaggerate vitamin needs still further (Teige et al., 1978a). These researchers reported that supplements of 100 IU of vitamin E per kg (45 IU per lb) of diet and 0.1 ppm selenium did not entirely prevent deficiency lesions in weanling pigs afflicted with dysentery and fed 3% cod liver oil. Under the conditions of certain experiments, the effect of including dietary fat in gilt rations on plasma alpha-tocopherol concentrations or in colostrum or milk is not always observed (Hidiroglou et al., 1993, 1995). However, Wang et al. (1996) reported that vitamin E and lipid peroxidation were both affected by dietary vitamin E and dietary PUFA. In their study, the requirement for vitamin E in young pigs increased as PUFA levels in the diet increased. Furthermore, the authors indicated that lipid peroxidation response of pigs was a suitable index for evaluating vitamin E requirements. The lipid peroxidation response was assessed by thiobarbituric acid-reactive substances in erythrocytes and ethane and pentane levels in exhaled gases.
Sauer et al. (1997) indicated that when milk replacer diets contain canola oil as the sole fat source, additional vitamin E supplementation is required. This finding did not appear to be related to PUFA concentration. The authors indicated that the factor or factors present in canola oil that increase vitamin E requirements for the young pigs remain to be determined. However, they referred to the fact that the higher phytosterol content in canola oil could interfere with vitamin E absorption or accelerate its rate of depletion in tissues.
The amount of vitamin E needed to maintain adequate growth and reproduction would not necessarily be enough to ensure optimal immune function as noted previously. For example, in the rat, Bendich et al. (1986) reported that 15 IU of vitamin E per kg (7 IU per lb) of diet prevented muscle abnormalities and 50 IU per kg (23 IU per lb) of diet was necessary to prevent red blood cell breakdown, but for maximum lymphocyte stimulation 200 IU of vitamin E per kg (91 IU per lb) of diet was required. For pigs, 150 IU of supplemental vitamin E per kg (68 IU per lb) of diet resulted in a lysozyme with higher activity and a higher rate of yeast lysis (Riedel-Caspari et al., 1986).
Requirements of both vitamin E and selenium are greatly dependent on the dietary concentrations of each other. Hitchcock et al. (1978) investigated the effects of dietary vitamin E on utilization of selenium by swine. Hakkarainen et al. (1978b) determined that either selenium or vitamin E supplementation individually was inadequate, but together, in appropriate amounts they did prevent the development of vitamin E-selenium deficiency. In reports by Van Vleet and Ruth (1977) and Van Vleet (1982), the efficacy of supplements in the prevention or control of selenium-vitamin E deficiency was evaluated. In 1982, Van Vleet suggested that approximately 0.2 mg of selenium per kg of feed and 30 IU of vitamin E per kg (13.6 IU/lb) of feed be included in the diets of growing swine to prevent selenium-vitamin E deficiency. Bengtsson et al. (1978a) were able to prevent the occurrence of vitamin E and selenium-deficiency syndrome in pigs fed a selenium-deficient diet by supplementing with 45 mg of dl-alpha-tocopheryl acetate per kg of diet beyond that present in the basal diet. In a related study, Bengtsson et al. (1978b) reported that selenium supplementation in the diet of weaned pigs could not overcome the effects of vitamin E-deficient basal rations. Wastellet al. (1972) investigated the effects of supplemental vitamin E and (or) selenium on performance, deficiency symptoms and blood components of growing-finishing pigs fed diets low in vitamin E and selenium.
Regional dietary considerations arise as well. Ku et al (1972) noted significant regional differences in dietary selenium content after surveying dietary selenium and vitamin E concentrations in diets fed to growing swine in 13 different states. Piper et al. (1975) indicated that when cull pea rations are utilized, as is common in the Pacific Northwest regions of the United States, selenium-vitamin E deficiency can develop unless a minimum selenium requirement for growing pigs of 0.06 to 0.07 ppm is provided. Determination of vitamin E requirements is further complicated because the body has a fairly large ability to store both vitamin E and selenium. Sows maintained on a diet deficient in vitamin E and selenium produced normal piglets during the first reproductive cycle of the deficiency and clinical deficiency signs occurred only after five such cycles (Glienke and Ewan, 1974). A number of studies to establish requirements for both nutrients have underestimated the requirements by failing to account for their augmentation from both body stores as well as experimental dietary concentrations. Regarding the relationship between selenium and vitamin E, Malm et al.(1976a) suggested that selenium may play a role in the transport of vitamin E across the placenta in swine. In this study, efficient placental transfer of vitamin E was observed even in dams with low serum concentrations of alpha-tocopherol.
A unique consideration arises for pigs fattened under certain organic farming systems. Kienzle et al. (1993) reported that a variety of deficiencies including vitamin E and selenium were observed in one such herd with resulting deaths.
Many vitamin E analyses of foods and feedstuffs have been reported using a variety of analytical techniques; however, there is a lack of characterization of individual tocopherols in the majority of analyses. Total tocopherol analysis of a food or feedstuff is of limited value in providing a reliable estimate of the biological vitamin E value.
Because alpha-tocopherol is the most active form of vitamin E, many nutritionists prefer listing this form in feeds versus the unreliable total tocopherol values. Some of these less active tocopherols, particularly gamma-tocopherol, are present in mixed diets in amounts two to four times greater than that of alpha-tocopherol. If only alpha-tocopherol in a mixed diet is reported, the value in mg can be increased by 20% to account for other tocopherols that are present, thus giving an approximation of total vitamin E activity as mg of alpha-tocopherol equivalents.
Vitamin E is widespread in nature with the richest sources being vegetable oils, cereal products containing the oils, eggs, liver, legumes, and, in general, green plants. In nature the synthesis of vitamin E is a function of plants and thus their products are by far the principal sources. It is abundant in whole cereal grains, particularly in germ, and thus in byproducts containing the germ (McDowell, 2000; Traber, 2006). There is a wide variation in vitamin content of particular feeds with many feeds having a three- to ten-fold range in reported alpha-tocopherol values. Feed table averages are often of little value in predicting individual content of feedstuffs or bioavailability of vitamins. Vitamin E content of 42 varieties of corn varied from 11.1 to 36.4 IU per kg, (5.0 to 16.5 IU per lb) a 3.3-fold difference (McDowell and Ward, 2008). The vitamin E levels found in various feedstuffs are shown in Table 4-3, which illustrates the high variability of vitamin E in the same feed component. Alpha-tocopherol is especially high in wheat germ oil, safflower oil and sunflower oil. Corn and soybean oils contain predominantly gamma-tocopherol, as well as some tocotrienols (McDowell, 2000; Traber, 2006). Cottonseed oil contains both alpha- and gamma-tocopherols in equal proportions.
Animal by-products supply only small amounts of vitamin E, and milk and dairy products are poor sources. Eggs, particularly the yolk, make a significant contribution depending on the diet of the hen. Wheat germ oil is the most concentrated natural source, and various other oils such as soybean and peanut, and particularly cottonseed are also rich sources of vitamin E. Unfortunately, most of the oilseed meals now marketed are almost devoid of these oils because of their removal by solvent extraction (McDowell, 2000). Although fish oil and rapeseed oil are rich sources of tocopherol, coconut oil contains relatively little tocopherol (Jensen et al., 1997). Green forage and other leafy materials, including good quality leaf meals, are very good sources, with alfalfa being especially rich.
Concentration of tocopherols per unit dry matter in fresh herbage is between five and 10 times greater than in some cereals or their by-products (Hardy and Frape, 1983). A more available form of vitamin E is present in pastures and green forages. They contain ample quantities of alpha-tocopherol versus less bioavailable forms in grains (McDowell, 2004).
Mutetikka and Mahan (1993) reported that pasture lots consisting of predominately grass species had lower alpha-tocopherol contents than pastures containing alfalfa. These authors found that both vitamin E and selenium levels of gilts and litters were influenced by pasture, but that vitamin E was influenced more than was selenium.
Stability of all naturally occurring tocopherols is poor and substantial losses of vitamin E activity occur in feedstuffs when processed and stored, as well as in manufacturing and storage of finished feeds. Vitamin E sources in these ingredients are unstable under conditions that promote oxidation of feedstuffs— heat, oxygen, moisture, oxidizing fats, and trace minerals. For concentrates, oxidation increases following grinding, mixing with mineral, the addition of fat and pelleting. Oxidation, which is accelerated by heat, moisture, unsaturated fat, rancid fat and trace minerals, rapidly destroys natural vitamin E. Therefore, predicting the amount of vitamin E activity in feed ingredients is difficult. When feeds are pelleted, destruction of both vitamins E and A may occur if the diet does not contain sufficient antioxidants to prevent their accelerated oxidation under conditions of moisture and high temperature. Iron salts (i.e., ferric chloride) can completely destroy vitamin E. Dove and Ewan (1987, 1991) investigated the effect of numerous trace minerals (18% crude protein soybean meal diets with no trace minerals, standard trace mineral levels, or with standard trace minerals plus either 250 ppm copper, 1,000 ppm iron, 1,000 ppm zinc or 100 ppm manganese) on stability of vitamin E in swine grower diets. They concluded that the rate of oxidation of natural tocopherols is increased in diets containing increased levels of copper, iron, zinc or manganese. The authors also indicated that corn-soybean meal diets containing high levels of copper may require alpha-tocopheryl acetate to maintain recommended levels of vitamin E diets during storage, especially when added unsaturated fat is included. Dove and Ewan (1990) had previously reported that addition of growth-promoting levels of copper increased the loss of natural tocopherols from feed and that supplemental tocopherol may be required to maintain serum tocopherol levels of pigs fed diets containing 250 ppm copper. In alfalfa stored at 33°C for 12 weeks, vitamin E losses of 54% to 73% have been observed and 5% to 33% losses have been obtained with commercial dehydration of alfalfa.
In a study testing vitamin E stability, Orstadius et al. (1963) reported that vitamin E content of corn was reduced from 50 to 30 mg per kg (23 to 14 mg per lb) to about 5 mg per kg (2.2 mg per lb) of dry weight as a result of artificial drying at 100°C for 24 hours under a continuous flow of air. Similarly, it has been reported that artificial drying of corn for 40 minutes at 190°F produced an average 19% loss of alpha-tocopherol and a 12% loss of other tocopherols (Adams, 1973). When corn was dried for 54 minutes at 93°C, the alpha-tocopherol loss averaged 41%. Artificial drying of corn results in a much lower vitamin E content. Young et al. (1975) reported a concentration of 9.3 or 20 mg per kg (4 or 9 mg per lb) alpha-tocopherol in artificial dried corn versus undried, respectively. Preservation of moist grains by ensiling caused almost complete loss of vitamin E activity. Corn stored as acid-treated (propionic or acetic-propionic mixture) high-moisture corn contained approximately 1 mg of alpha-tocopherol per kg (0.45 mg per lb) of dry matter, whereas similar corn artificially dried following harvesting contained approximately 5.7 mg per kg (2.6 mg per lb) alpha-tocopherol (Young et al., 1978). Madsen et al. (1973) observed considerably lower levels of vitamin E in barley that had a high moisture content and was treated with propionic acid than in barley that had been conventionally dried. Apparently damage is not due to moisture alone, but to the combined propionic acid/moisture effect (McMurrayet al., 1980). Further decomposition of alpha-tocopherol occurs over a more extended period of time until the grain eventually has alpha-tocopherol levels of less than 1 mg per kg (0.45 mg per lb) of dry matter, which are commonly found in propionic acid-treated barley.
The most active form of natural vitamin E found in feed ingredients is d-alpha-tocopherol. For many years the primary source of vitamin E in animal feed was the natural tocopherols found in green plant materials and seeds. The principal commercially available forms of vitamin E used in the feed industry are the ester of RRR- and all-rac-alpha-tocopherol (Table 4-2). During commercial synthesis of dl-alpha-tocopheryl acetate, all-rac-alpha-tocopherol is esterified to the acetate form to stabilize it, with the ester extremely resistant to oxidation. Thus, dl-alpha-tocopheryl acetate does not act as an antioxidant in the feed. However, it does have antioxidant activity after it is hydrolyzed in the intestine and free dl-alpha-tocopherol is released and absorbed.
The acetate forms of alpha-tocopherol are commercially available from two basic sources: (1) d-alpha-tocopheryl acetate is made by extraction of natural tocopherols from vegetable oil refining by-products—molecular distillation to obtain the alpha form, and then acetylation to form the acetate ester, and (2) dl-alpha-tocopheryl acetate is made by complete chemical synthesis, producing a racemic mixture of equal proportions of the d and l isomers. Numerous authors have evaluated the relative potencies of these various forms (for a recent comprehensive review, see Scherf et al., 1996). Ames (1979) utilized a rat fetal-resorption bioassay and concluded that the commonly accepted conversion value of 1.36 may be too low. Anderson et al. (1995) evaluated the bioavailability of various vitamin E compounds by supplementing the diets of finishing swine for 28 days and comparing the concentration of alpha-tocopherol in blood serum and tissue. These authors indicated that both the acetate ester and alcohol forms of vitamin E resulted in rapid increases in serum concentration of vitamin E in finishing pigs. However, the serum concentrations remained high longer and the tissue concentrations of alpha-tocopherol were high when supplementation was with the acetate ester forms. This difference was likely due to the greater stability of acetate forms in mixed feed. The greatest relative bioavailability in this swine trial, based on serum and tissue tocopherol concentrations, was found when d-alpha-tocopheryl acetate was supplemented. Furthermore, the authors concluded that the bioavailability of this form seems to be greater for finishing swine than the predicted values from traditional rat bioassays. The RRR-alpha-tocopherol potency has been found to be underestimated compared to all rac-alpha-tocopherol in bioavailability for weanling pigs (Wilburn et al., 2008), finishing swine (Yang et al., 2009), and sows and suckling piglets (Mahan et al., 2000; Lauridsenet al., 2002).
Huang et al. (1995) evaluated effectiveness of individual tocopherols and mixtures of these tocopherols in inhibiting the formation and decomposition of hydroperoxides in bulk corn oil, which had been stripped of natural tocopherols. On the basis of hydroperoxide formation, the concentrations required for maximum antioxidant activity of 1:1 mixtures of alpha- and gamma-tocopherol or natural soybean tocopherol mixtures (alpha:gamma:delta=13.64:21) were 250 and 500 ppm, respectively.
Nafstad and Tollersrud (1970) and Nafstad (1973) reviewed some of the experimental work and reports available at that time concerning vitamin E-selenium deficiency. In 1976, MacDonald et al. provided a comprehensive review of various aspects of the etiology, pathogenesis and biochemistry of vitamin E-selenium responsive diseases. Van Vleet and Kennedy (1989) provided a review of selenium-vitamin E deficiency in swine, including its effects on the liver, heart and skeletal muscle, diagnosis and differential diagnosis and control and prophylactic measures. Mortimer (1983) discussed a variety of factors that may contribute to the deficiency.Numerous authors have studied relationships between levels of alanine aminotransferase and aspartate aminotransferase activities and vitamin E- deficiency. The findings have not been of consistent diagnostic value (Simesen et al., 1979; Tollersrud and Nafstad, 1970). Young et al. (1976) found correlations between percent peroxide hemolysis and vitamin E intake or serum vitamin E. Fontaine and Valli (1977) concluded that red cell lipid peroxide measurement was a reliable test for vitamin E deficiency in swine. Red cell lipid peroxides were found to be increased in vitamin E-deficient pigs but not in selenium-deficient pigs. Thode-Jensen et al. (1979) reported that both glutathione peroxidase activity and resistance against erythrocyte lipid peroxidation (ELP) measurement were valuable methods for assessing vitamin E or selenium status in growing pigs. Simesen et al. (1982) stated that although blood and liver selenium and blood vitamin E were regarded as the best indicators for selenium-vitamin E deficiencies, determination of glutathione peroxidase activity and ELP may be suitable alternatives. Thode-Jensen et al. (1983) determined that ELP was a more sensitive index of vitamin E status than were vitamin E levels in young pigs. Malm et al.(1976b) reported that providing 0.5 ppm of supplemental selenium to diets of sows prevented the serum enzyme changes commonly associated with vitamin E-selenium deficiency. Rammell et al. (1988) suggested that rather than analyzing for one analyte, monitoring multiple factors such as feed and tissue vitamin E, selenium and PUFAs would provide a more accurate diagnosis. These authors indicated that liver vitamin E concentration of greater than 10 mmol per kg appeared to be adequate, while less than 2.5 mmol per kg could indicate vitamin E deficiency. The selenium and PUFA measurements would help determine deficiencies in the case that vitamin E concentrations fell to intermediate levels. Ewan (1971) reported that calcium levels were increased in livers of pigs fed diets deficient in selenium and vitamin E and that sodium levels were increased in muscle tissue from this same group. Vitamin E displays the greatest range of deficiency signs of all vitamins. Deficiency signs differ among species and even within the same species. Blaxter (1962) reported that muscular dystrophy seemed to be the one syndrome commonly encountered in vitamin E deficiency in all species. Fundamentally, this is Zenker’s degeneration of both skeletal and cardiac muscle fibers (Illus. 4-2). Connective tissue replacement that follows is observed grossly as white striations in the muscle bundles (Smith, 1970).
Occurrence of muscular dystrophy is worldwide, but its incidence or at least diagnosis, particularly in a mild or subclinical form, varies widely in different countries and regions within countries (McDowell et al., 1983). Considerable research has revealed the positive relationship between selenium content in soil and geographical occurrence of vitamin E-selenium responsive muscular dystrophy. Numerous authors have described ultrastructural alterations in the cardiac tissue (Sweeney and Brown, 1972; Piper et al., 1975; Van Vleet et al., 1977a, b) and skeletal muscle (Piper et al., 1975; Van Vleet et al., 1976) of pigs with selenium-vitamin E deficiency that had been experimentally induced. Ruth and Van Vleet (1974) reported that a selective destruction of type I skeletal muscle fibers and a lack of phosphorylase activity in type II fibers were evident in experimentally induced selenium-vitamin E deficiency in growing swine. Fontaine et al. (1977b) summarized their studies concerning vitamin E-selenium deficiency with regard to hematological and biochemical changes in young pigs. They reported that serum creatine phosphokinase activities were increased in association with vitamin E deficiency and selenium deficiency and that the interaction was also significant. They concluded that these increases reflect the occurrence of subclinical muscular dystrophy and that selenium and vitamin E deficiencies have additive effects in the induction of skeletal muscular dystrophy. Nafstad (1965) and Nafstad and Nafstad (1968) reported that vitamin E-deficient pigs had nuclear abnormalities in erythroid precursors within their bone marrow, inadequate erythroid production, increased destruction of erythroid cells and abnormalities in the formation of myeloid cells. In 1972, Baustad and Nafstad indicated that piglets born of vitamin E-deficient mothers had morphological abnormalities of blood and bone-marrow cells similar to those reported in growing pigs following vitamin E deficiency. The authors suggested that vitamin E might be regarded as a hemopoietic factor in the newborn piglet. However, results by Fontaine et al.(1977b) suggested that vitamin E is not a limiting factor for normal erythropoiesis in young growing pigs. Most vitamin E deficiency signs for the pig have been associated with selenium deficiency, and scientists usually refer to a vitamin E and (or) selenium deficiency since it is not clear which is involved. Generally, dietary levels of both must be low to bring about deficiency signs and lesions. Since the early 1950s, reports in the European literature have revealed tissue degeneration signs in swine under field conditions associated with vitamin E deficiency, with the significance of selenium deficiency not realized until 1957. Muscular dystrophy and hepatosis dietetica (toxic liver dystrophy) were particularly widespread in the swine industry in Sweden. Obel (1953) reported that records from the State Veterinary Medical Institute of Stockholm from 1947 to 1952 revealed that of a total of 4,382 pigs autopsied, more than 10% suffered from hepatosis dietetica. The description of and experimental induction of hepatosis dietetica reported by Obel (1953) was confirmed shortly thereafter by Hove and Seibold (1955). Hepatosis dietetica is characterized by extensive necrosis of the liver. Selenium-vitamin E deficiencies have been readily produced in swine diets through use of both highly unsaturated fats (i.e., cod-liver oil as in Lannek et al., 1961) and rancid fats. However, naturally occurring vitamin E-selenium deficiencies were not reported in the United States until the late 1960s (Michel et al., 1969), and in the 1970s they became widespread. High incidence of vitamin E-selenium deficiencies in swine was believed to be due to a number of factors (Trapp et al., 1970), including (1) swine raised in complete confinement, without access to pasture, (2) low selenium content in midwestern U.S. feeds, (3) solvent-extracted protein supplements low in vitamin E, (4) limited feeding programs for sows, (5) loss of vitamin E and selenium from corn due to oxidation as a result of air and heat drying or storing high-moisture grains, and (6) selection for leaner pigs that require more selenium. Evidence also suggests that moldy feed in bulk-holding bins may produce mycotoxins that either inhibit the uptake of vitamin E in the small intestine or affect the antioxidant balance of cells. An increase in confinement rearing of swine on concrete floors or slats has been accompanied by a decrease in the utilization of pasture and forages. Such crops are not only excellent sources of vitamin E but also provide the more highly available form of the vitamin, alpha- versus gamma-tocopherol. Vitamin E-selenium deficiency in swine is often associated with sudden death (Piper et al., 1975; Rice and Kennedy, 1989). In most cases clinical signs of the condition were not observed prior to death (Michel et al., 1969; Trapp et al., 1970), although occasionally pigs were observed with clinical signs of icterus, difficult locomotion, reluctance to move and weakness. Clinical signs also include peripheral cyanosis (particularly the ears), dyspnea (abdominal respiration) and a weak pulse, all occurring shortly before death. In many cases the faster-growing, more thrifty-appearing pigs died suddenly.
The most common pathologic lesions include massive hepatic necrosis (hepatosis dietetica) (Illus. 4-3, Illus. 4-4 and Illus. 4-5), degenerative myopathy of cardiac (Illus. 4-6) and skeletal (Illus. 4-7) muscles, edema, esophagogastric ulceration, icterus, nephrosis, hemoglobinuria, acute congestion, hemorrhaging in various tissues (Illus. 4-8) (Ewan et al., 1969; Trapp et al., 1970; Piper et al., 1975) and yellowish discoloration of adipose tissue (“yellow fat”). Many pathological reports of vitamin E-selenium deficiency note that the most striking lesion was liver necrosis (Trapp et al., 1970), but bilateral paleness of skeletal muscle was the gross lesion most commonly found. In some pigs, microscopic lesions in the liver were either absent or minimal, whereas changes in skeletal muscles were extensive. In other cases, the reverse was true. Bengtsson et al. (1978a,b) reported a variety of symptoms of vitamin E and selenium deficiency including lesions such as hepatosis dietetica, mulberry heart, muscular degeneration and microangiopathy in weaned pigs fed a selenium-deficient diet. Only the highest vitamin E supplementation level utilized in their study could overcome the symptoms. Dietary induction of mulberry heart and hepatosis dietetica has been studied by numerous authors including Sharp et al. (1970, 1972a). Sharp et al. (1972a) reported that dietary selenium (0.5 ppm) and (or) vitamin E (25 IU per kg of diet) prevented skeletal muscular dystrophy and exudative diathesis. Hakkarainen et al. (1978a) investigated whether providing dietary vitamin E and selenium could reverse the vitamin E-selenium deficiency (VESD) once the syndrome had been allowed to develop. By using the same combination (5 mg alpha-tocopheryl acetate and 135 mg selenium per kg diet) that was shown previously to prevent the onset of VESD, the therapy was found to result in recovery from VESD syndrome. However, with less supplementation (5 mg alpha-tocopherol acetate and 45 mg selenium per kg diet), the VESD syndrome was not reversed.
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