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Roles of selenium in farms and in human health
Carlos André Prauchner
Lambert Academic Publishing 2015 1
Carlos André Prauchner
Roles of selenium in farms and in human health
Lambert Academic Publishing 2015 2
I dedicate this book for my father (in memorian) and for my mother, that are my inspirations to live. I want to dedicate it also for all relatives of victims of the tragedy in Boate Kiss of Santa Maria, happened in January/27/2013 and that finished prematurely 242 lifes. The author.
3
SUMARY PRESENTATION INTRODUCTION
6 8
CHAPTER 1 A brief history about selenium 1.1 Introduction 1.2 Historic on searches with selenium
10 11 11
CHAPTER 2 Roles of selenium in farms 2.1 Introduction 2.2 Selenium in agriculture 2.2.1Selenium in soil and its cycle from oceans to continents 2.2.2Selenium as environmental pollutant 2.2.3Selenium metabolism in plants 2.2.3.1 The selenium assimilation pathway 2.2.3.2 The selenium volatilization pathway 2.2.3.3 The selenium methylation pathway 2.2.3.4 The double edge of selenium metabolites 2.2.4Crops production by supplementing selenium 2.2.5The “agronomic biofortification” 2.2.6The “genetic biofortification” 2.3Selenium in animal production 2.3.1 Selenium levels in animal diet 2.3.2 Selenium metabolism in animal tissues 2.3.3 Selenium supplementation for diary cattle 2.3.4 Selenium supplementation for meat producing 2.3.5 Selenium supplementation and animal reproduction 2.3.6 Selenium in the Veterinary clinic 2.3.6.1 The selenium and imune surveillance of animals 2.3.6.2 The oxidative stress in the Veterinary clinic 4
19 20 21 22 38 43 52 58 60 62 64 71 88 90 91 97 101 112 123 135 135 140
CHAPTER 3 Roles of selenium in human health 147 3.1 Introduction 148 3.2 Selenium levels in human diet 148 3.3 Some animal products as selenium carriers 149 3.4 The true biological value of selenium in foods 156 3.4.1 The bioacessibility 159 3.4.2 The bioavailability 160 3.4.3 The bioactivity 160 3.4.4 The selenium metabolism in human body depends on its chemical speciation 161 3.5 The selenium bioacessibility, bioavailability and bioactivity of some foods 165 3.6 Major diseases related to low selenium status 169 3.7 Selenium in preventing human diseases 175 3.7.1 Selenium in mitigating heavy metals poisoning 176 3.7.2 Selenium in preventing cardiovascular diseases 184 3.7.3 Selenium in preventing cancer 188 CHAPTER 4 Some organic selenium compounds 4.1 Introduction 4.2 Ebselen 4.3 Selenium-enriched yeast extracts 4.4 Diphenyl diselenide [(PhSe)2]
195 196 196 209 210
CHAPTER 5 Selenium toxicity 5.1 Introduction 5.2 Selenium poisoning in animal and humans
216 217 217
CONCLUSIONS REFERENCES
232 241 5
PRESENTATION
The book “Roles of selenium in farms and in human health” was elaborated with the proporsal to discuss some aspects related to selenium biochemistry, as well as to describe some applications of this important microelement in agriculture and animal production. Additionally, the book attempts to correlate the soil composition, the cycle of selenium in nature and several functions exerted by it in farms, with food production and with the prevention of some diseases in human population. Initially, the book shows a brief hystoric about evolution in searchs with selenium, discusses some topics on selenium in nature, as well as its danger as environmental pollutant. Afterwards, it is showed texts about roles of selenim in farms, i.e., its importance for agriculture and animal production. In this chapter the reader will find a discussion related to selenium in soil, its absorption and metabolism in plants, the classification of vegetals in terms of selenium tolerance, the importance of selenium fertilization to crop plants and its relation with the production of foods from vegetal origin. Concerning to animal production, it is showed explantions about selenim absoption and metabolism in animals, its requirement in diet according animal species and its importance to the production of foods from animal origin (milk, meat and eggs). Long the chapter it is emphasized the relation among soil composition, plants and animal production with human health. Following, the next chapter describes exclusively roles of selenium in human population, including the nutritional requirement, major selenium sources to our diet, the bioacessibility, bioavailability and bioactivity of selenium species in some foods, as well as a brief discussion about selenium metabolism in human body. Furthermore, it is described functions of the micronutrient in preventing/mitiganting some human diseases. In this line, the fourth chapter is destinated to describe biological effects of some organo selenium compounds with efficacy already confirmed or that
6
show potential to be tested in the prevention/treatment of several animal and human diseases. Finally, the last chapter discusses the pathogenesis of selenium toxicity. Thus, it describes some observational evidences in animals and human reported centuries ago that today are recognized as possibly caused by selenium overload. Moreover, are described some episodes of selenium poisoning occurred more recently. It was attempted to do the differentiation among acute, subchronic and chronic intoxication, emphasizing that the diagnosis should be done not exclusively based on serum selenium concentrations, but considering clinic signals and patient anamnesis. Texts were elaborated from extensive literature review, which including several works carried out by foreing researchs, but some done by national ones. I want apologize in advancing to colleagues and friends whose works were not commented, or although were, did not receive the sufficient attention. By the other hand, I want to detach that all comments, critiques, corrections or sugestions are welcome and will be useful to enrich new wordings. Thus, I am certain that the knowledge will be never ready, but it will be forever being built, and each adds only a little contribution! In this context, the cientific universe surround the selenium element has changed a lot of in last decades. Although we cannot foresse it, we need to accompain, or even better, to participate of new advances. Then, with these words the reader is invited to know somewhat about the potential and roles of selenium in farms and in human health.
The author. Santa Maria (RS), Brazil, 2015.
7
INTRODUCTION
The natural richness of Brazil in terms of arable soils, forests, water sources and climate favorable to development of agriculture and animal reare has facilitated the advance of the agribusiness in our country. Parallelly, universities and search centers are contributing with this evolution through developing modern insumes, new cultivars, better fertilization techniques, animal genetic improvements, more advanced techniques for animal management, new and better veterinary products, among other innovations. Thus, the national agribusiness has reached a status of an important supplier of raw matters and foodstuffs to inner and extern merkats. Particularly, in relation to food production, it should be remembered that many diseases that threat human health could be prevented by the consumption of determinated types of foods with a nutritional characteristic composition. As farm’ animals as human population depend on products elaborated by plants in the composition of part or totality of diet. In last degree, the composition of vegetal material, although reflects the physiologic charactertistics of each plant species, depends primarily on soil composition. Thus, the prevalence of some types of diseases in human population that inhabit determinated region of world could be related to soil composition. Micronutrients, as selenium, can improve the production of crop plants. At the same time, by absorving and concentrating selenium in eatable parts, plants become importante source of the micronutrient for animal and human diets. In the case of animals, the selenium can play a series of effects that help in maintaining the sanity, optimizing reproductive performance, improving productivity and quality of foods from animal origin. In relation to human health, the consumption of products from vegetal and animal origin that were selenium-enriched can help in preventing against cancer. Inserted in this context, the book “Roles of selenium in farms and in human health” induces the reader to “travel” by the cientific universe that describes selenium functions in nature, as in agricultural activities as in 8
human health. It attempts to emphasize a relatively recent worry that is the relation among soil composition, nutritional value of foods from vegetal and animal origin with human health. The book, that some times describes basic aspects related to farms, in other moments comments complicated cientific findings in terms of biochemical and phisological roles of selenium in plants, animals and humans. In addition, at some occasions the author discusses his interpretation about determined topics and challenge the reader to image important issues that may represent new opportunities for searching. Thus, I think that the book will be a good reading as for graduation and post-graduation students of rural, biological and health sciences, especially related to Agronomy, Zootechny, Veterinary, Biology and Nutrition, as for non-academic population.
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CHAPTER 1 A brief historic about selenium
10
1.1Introduction
The natural element selenium was somewhat enigmatic and mysterious for researches themselves. The unknown on its real importance for animals and higher plants did of this micronutrient the center of many discussions and dubiousness. The essential toxin, expression that some people utilized to call the selenium, represented the doubt that remained during many decades in scientific community about essenciality or toxicity of the selenium. Only from 56 years ago this paradigm was being slowly broken. Today our understanding indicates that selenium can be both, essential micronutrient and toxin, which depends only on the dose or concentration in diet. Really, the hazard there is and that frightened so researches in the past is clearer today: the allowance between selenium deficiency and toxicity to humans fall within a range of only 10 times. Fortunately, scientific advances were decisives in the comprehension of both effects of the element, the nutritional and the toxic one.
1.2Historic on searches with selenium
The moon inspirited not only passionates or, maybe, researches in the past were more romantic that are actual ones. According history, the selenium element (Se34) was identified by Swedish chemist Jöns Jacob Berzelius at near 200 years ago (BERZELIUS, 1817). It was describeb as a chemical element related to sulphur and tellurium, showed chemical properties seem to the last. As tellurium was called “tellus”, that in latim means terra, Berzelius called the new element of “selene”, that in Greek means moon (Figure 1.1). For more than one century, scientists did not found any biologic function to selenium. Indeed, this lack of comprehension helped to maintain a spirit 11
of mystery related to the element, doing it somewhat mystic until for researches.
Figure 1.1 In the early the selenium was somewhat enigmatic and mysterious for researches themselves. But, insofar as that science was being developed, that old and mystic look was slowly substituted by a rational conception. Today, the selenium is recognized as a micronutrient essential to the perfect growth of plants and animals and by preventing health problems in human population.
In the 1930 decade, the predominant concept about selenium was of a toxic element. In a curious episode, one farm located at American State of South Dakota was sold successively, and each new owner sold it again. The discontent of many owners resulted in judicial actions against old owners, which would have acted in a dishonest conduct when sold the farm by first
12
time. The farm fields were devaluated more and more. One of reclamations was that hen eggs had low hatchability rate and, when chicken were hatched many of them were deformed and dead. These monstrosities were more frequent in hens fed with wheat grains produced in the farm. The United State Department of Agriculture (USDA) identified selenium compounds in these grains. Afterwards, it was demonstrated that the injection of selenium compounds in the air sac of eggs would reply those abnormalities observed in the farm (FRANKE et al., 1934). Then, it was concluded that the wheat produced by own farm was richer in selenium because high concentration of the micronutrient in soil. After a lot of polemic, “the mysterious farm”, as became known, was bought by USDA and thansformed in an experimental station, put on ending to disagreements and problems from those businesses. Later, it was identified that only one part of the farm had high selenium levels in soil. Thus, pioneer studies related to the importance of selenium for live organisms indicated to one element of natural occurrence that would be toxic and carcinogenic. However, Schwarz and Foltz (1957) identified this element as essential to the good growth of bacteria, mammals and birds, and called it as “factor 3”. Afterwards, there was a period where were reported some empiric observations on syndromes in animals maintained on selenium-deficient diet. At the same time, it was observed a crescent number of studies demonstrating that selenium would be need to perfect development of broiler, turkey, bovine, sheep, goat and other animals. In addition, it was demonstrated that selenium would be more effective than vitamin E in preventing exudative diathesis in chicken (NESHEIM and SCOTT, 1958; SCOTT, 1962), white muscle disease in lambs (MUTH et al., 1958) and calves (MUTH, SCHUBERT and OLFIELD, 1961) and hepatic necrosis in pigs (EGGERT et al., 1957; GRANT e THAFVELIN, 1958). However, in the early of 1970 decade, there were still consistent discussions in United States of America (USA) about if approving or not selenium supplementation to animal diet (SCOTT, 1973). Although there were many reports indicating positive effects of selenium on growth rate and 13
sanitary status of animals, mainly to broiler, the “Food and Drug Administration” (FDA) was not self-satisfied to allow selenium incorporation to animals rations, with allegation that the element would be a potent carcinogen. Although illegally, many veterinaries recommended to supply selenium for animal lots, because they were satisfied with the good results reached. After several studies have showed, on the contrary than was imaged in the past, that selenium could protect animals from different types of cancer, that ancient paradigm was slowly being broken. Selenium biochemistry emerged really after 1973, when two bacterial enzymes, formate dehydrogenase (ANDREESEN e LJUNGDAHL, 1973) and glycine reductase (TURNER e STADTMAN, 1973) were identified as containing selenium. At the same time, the role of selenium in mammals was being revealed after the discovery of its participation in active site of the enzyme glutathione peroxidase (GSH-Px) (FLOHÉ, GUNZLER and SHOCK, 1973; ROTRUCK et al., 1973). During last years of 1960 decade, searches showed that functions played by selenium in animals would be resultant from its participation in several important proteins. It includes selenoenzymes that act as part of the antioxidant defense mechanism of animal cells. The number of selenoproteins identified has increased substantially ultimately. Selenium is incorporated in proteins mainly as selenocysteine (SeCys), that is considerate the 21º amino acid. Otherwise, in other proteins the element is, more frequently, as selenomethionine (SeMet). It is known, at least, five families of glutathione peroxidase isoenzymes (containing 4 atoms gram of Se/mol of protein): the cellular gluthathione peroxidase (GSH-Px 1); the intestinal epithelium glutathione peroxidase (GSH-Px 2); the plasma glutathione peroxidase (GSH-Px 3); the phospholipid hydroperoxides glutathione peroxidase (GSH-Px 4) and; the sperm-containing glutathione peroxidase (GSH-Px 5) (REDERSTORFF, KROL and LESCURE, 2006). The GSH-Px 1 is the most abundant selenoprotein in mammals. Its activity in liver is regulated by selenium status. GSH-Px 1 is a cytosolic enzyme expressed in all tissues and it has been considerated the most 14
important antioxidant enzyme in mammals (TAPIERO, TOWNSEND e TEW, 2003). The GSH-Px 2 is very similar to GSH-Px 1, but is found mainly in gastrointestinal tract (CHU, DOROSHOW e ESWORTHY, 1993). The GSH-Px 3 is the second more important selenoprotein in plasma, ranked only behind the selenoprotein P. This enzyme is chemically and immunologically different from GSH-Px 1 (TAKAHASHI et al., 1987). The GSH-Px 4 is also called phospholipid hydroperoxides glutathione peroxidase (URSINI, MAIORINO e GREGOLIN, 1985). Differently from other isoforms, it is a monomer of, approximately, 20 kilodaltons (kDa), located in both, cytosol and mitochondria. GSH-Px 4 and GSH-Px 1 seem to be regulated differently by selenium status (LEI et al., 1995). The major biologic role of GSH-Pxs is maintaining peroxides produced by cellular metabolism at low levels, decreasing thus, the potential dangerous of these reactive species. GSH-Pxs provide a second defense line against hydroperoxides, which may impair membrane and other cellular structures. Selenium is a component of GSH-Px and acts synergistically with tocopherol to mitigate lipid peroxidation in cells and tissues. In a coordinated action with catalase, GSH-Pxs decompose the hydrogen peroxide (H2O2) to water and, in turn, produce the oxidized form of glutathione (GSSH). The regeneration to the reduced form is done by a cycle that includes the catalytic action of glutathione reductase (GR). In this cycle electrons are provided from reduced form of nicotinamide adenine dinucleotide phosphate (NADPH). NADP+ (oxidized form) is normally reformed back to reduced state by pentose phosphate pathway (TAPIERO, TOWNSEND e TEW, 2003). Other two important selenoproteins are iodothyronin-5’-deiodinase (with 1 atom gram of Se/mol of protein), that catalyses the 5’ deiodination of L-thyroxine (T4) to triiodothyronine (T3) (GROSS, OERTEL and KOHRLE, 1995), and the thioredoxin reductase. This last enzyme catalyses the reduction of thioredoxin by consuming NADPH and is largely utilized by biologic systems to reduce ribonucleotides and deoxyribonucleotides (TAPIERO, TOWNSEND e TEW, 2003).
15
The thyroid gland synthezised two important hormones, the triiodothyronine (T3), the active form in liver and kidney; and the thyroxine (T4), the active form produced only by thyroid gland. Formation of T3 from T4 depends on the deiodination catalysed by iodothyronin-5’-deiodinase type I, that occurs in peripheral tissues (ARTHUR and BECKETT, 1999). In the nervous system and in the hypophysis gland, deiodination of T4 to T3, mainly for local use, depends on the deiodinase type 2 activity (KÖHRLE, 1994). The system thioredoxin/thioredoxin reductase acts in the DNA synthesis process, in mechanisms for repairing DNA damage, in maintaining the redox balance of cells, in regulating the activity of transcription factors and in reforming antioxidants systems (ALLAN et al., 1999). In this cycle, thioredoxin in reformed by thioredoxin reductase (TR) by spending NADPH. Compounds as selenide, selenodiglutathione and SeCys are efficiently reduced by thioredoxins, but also directly by NADPH and TR (BJÖRNSTEDT et al., 1997). Other important selenoproteins including the selenoprotein P (with 10 atoms gram of Se/mol of protein) (AKESSON et al., 1994), which is found in plasma; and the selenoprotein W (with 0.92 atoms gram of Se/mol of protein) (VANDELAND et al., 1993; WHANGER, 2000), isolated from several tissues, with higher concentrations in skeletal and cardiac muscles (YEH et al., 1997). It has been stablished that expression of these selenoproteins is selenium dependent (ALLAN et al., 1999). Selenium is component of these enzymes in the form of the seleno amino acid SeCys. The ability to catalyse oxydo-reduction reactions played by GSH-Pxs is related to higher capacity of their selenol group to undergo ionization at physiologic pH, which replaces cystein-thiol groups in the protein. Substituting SeCys by Cys reduces dramatically the capacity of these enzymes to catalyse oxydo-reduction reactions (DRISCOLL and COPELAND, 2003). The major compound recognized worldwide as selenium source in rations and mineral supplements for animals is sodium selenite (Na2SeO3), although some times it has been utilized sodium selenate (Na2SeO4) or barium selenate 16
(BaSeO4). The Na2SeO4 is the most common selenium species in formulations for soil fertilization or in foliar spraying fertilizers, whose utilization is recommended in selenium-deficient geo-ecosystems. It was imagined, until recently, that selenium played no function in plants. However, Sing, Sing and Bhandari (1980) demonstrated that supplying 0.5 mg of Se/kg of soil (in the form of selenide) it would stimulate the growth and dry matter production by Indian mustard (Bassica juncea L.). More searches confirmed this finding and now it has been accepted by scientific community that selenium may exert an antioxidant function in crops that remember its activity in animal tissues. A concept relatively new is that soil composition and crop nutrition walking together with animal and human health. This new perspective induced the development of strategies for plant biofortification as a way to produce eatable tissues of plants richer in selenium for animal and human consumption. Likewise, in Veterinary science, selenium supplementation to farm’ animals reared to food production (meat, milk, eggs, etc) resulted in improves of flock sanity, as well as in the enrichment of nutritional value of these foods. The enrichment of vegetal and animal foodstuffs by selenium supplementation has been encouraged due the possibility to prevent health problems in human populations. Then, it has changed the look on farms, now as a supplier of better nutritional foods for human consumption, whose perspective fall well into the concept of nutraceutical foods. Although there are several studies related to roles of the microelement in nutrition of farm’ animals, positive effects of supplying selenium in human diet were evaluated in a limited number of searches. Specially, in concerning to cancer prevention by selenium, that requires long-term studies, there are a few studies published. In addition, some are restricted to a population living in regions constituted by soil poorer in selenium. In China, two diseases (Keshan disease and Kashin-Beck disease) have been linked to lower levels of selenium in the local population, but other factors seem to compete to the deflagraton of these infirmities.
17
Thus, after admissing selenium importance to the perfect development of farm’animals, recognizing its benefites to human health and discoverying its role in higher plants, important progress has been done in looking to know better selenium functions in biologic systems. Mainly in the last decade, two important lines of searche has emerged, that are: i) studies about effects of selenium fertilization on crop physiology and productivity and; ii) searches evaluating effects of organoselenium compounds (natural or synthetic) on animal production and human health. In following texts these aspects will be described with more details.
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CHAPTER 2 Roles of selenium in farms
19
2.1 Introduction
Selenium enters in food chain through its absorption by plants from soil. Then, it is transfered to foods of vegetal and animals origin, which constitute the human diet. In true, the onset of all is in soil composition, because will are possible soils seleneium-deficient, moderate and enriched in the micronutrient. Consequently, will are possible geo-ecosystems seleniumdowned and overloaded. Selenium plays role as in plant growth as in animals development. In crop plants selenium improve their productivity by mitigating oxidative stress caused by abiotic stress, which damage the photosynthetic apparatus. A better yield in photosynthesis will result in higher grain production, as to cereals as to oleaginous. Then, in regions constituted by selenium-poor soils, fertilizing selenium to crops, directly on soil or as foliar fertilizer, is a way to increment the crops productivity as well as to increase selenium in foodstuffs. Animals that eat selenium-enriched diet may reach higher productivity, because selenium contributes in preventing mastitis and periparturient complications. Moreover, selenium supplementation ameliorates the meat quality and the shelf life of cuts. This benefit is evidenced as to beef, as to swine and broiler meat products. In addition, selenium may ameliorate the reproductive performance of farm’s animals, especially for ruminants. Calves and lambs born from mothers supplemented with selenium during gestation seem healthier and show a more viogorous growth. Even, laying hends may produce selenium-enriched eggs if supplemented with the micronutrient. Furthermore, selenium may modulate or stimulate the immune surveillance of animals, guaranting better general health. More recently, it has been identified that selenium may play medicinal role in Veterinary clinic, mainly in therapy of diseases with the involvement of oxidative stress in their pathogeny. Thus, as it will be described following, selenium seems be an important micronutrient in farms, i.e, for agriculture and animal production.
20
2.2 Selenium in agriculture
Similar other micronutrients selenium shows a natural cycle that comprises its evaporation from ocean, from surface of soil and from leaves of plants, which produce selenium-volatile compounds. Afterwards, the microelement reachs the atmosphere where is transported by wind and goes back to surface of soil in the form of little particles that sediment or fell diluted in raining drops. Geo-ecosystems can be naturally selenium-overloaded, but what cause worrier to researches on this matter are environmental problems caused by selenium from anthropogenic mobilization. Fire of coal by thermoelectric powers and mining activity by shelters are examples of antropogenic sources to selenium environmental pollution. In the case of selenium-downed ecosystems, fundamentally, there are two manners to produce foods containing higher selenium levels, which are “agronomic fortification” and “genetic fortification”. The Finland case, maybe, is the better example of applying the agronomic fortification strategty to produce selenium-enriched foods, with positive repercussions on human health. With exception of Brassica and Allium genders, crops plants generally do not accumulate appreciable amounts of selenium in their eatable tissues. Selenium absorption and metabolism in plants share many carriers, enzymes and metabolic steps with sulphur. According metabolic pathways and fates, that are intrinsic to determined vegetal species, there are different plant catergories in terms of selenium tolerance. Some species may accumulate high levels of the micronutrient in tissues, being called hyperaccumulators. At general manner, plants that concentrate elevated amount of selenium in their tissues excluding it from protein synthesis by generating non-proteic amino acids. Other plants synthesized methylated compounds, including volatile ones, which are evaporated by leaves. Selenium fertilization to pasture plants, as roughages, may improve their dry matter production and selenium content, by this way benefiting grazing 21
animals. In addition to protect against oxidative stress to photosyntetic apparatus and the consequent increment in productivity, cereals as wheat and rice may be efficient carriers of selenium to human diet. An especial case is Brazilian nuts, a natural selenium-rich food produced by tall trees that grow in Amazon forest. Their fruits are collected by native habitants of the region and exported to several regions of the world. Then, in following lines the reader will find an explanation about these themes.
2.2.1 Selenium in soil and its cycle from oceans to continents
As other trace elements, selenium is a natural component of the terrestrial crust, and it is found in water too. Generally, selenium is introduced in food chain from its extraction from soil by plants, which determine the concentration of the micronutrient in vegetals, grains, forage and in foods from animal origin destinated to human diet. The micronutrient is normally measured in soil at concentrations between 0.01 – 2 mg/kg (mean of 0.4 mg/kg). Nevertheless, its distribution on terrestrial surface is not uniform, resulting in geo-ecosystems rich/excessive and other deficient in the microelement. Thus, soil containing elevated selenium levels (>2 – 1,250 mg/kg) are found in some regions around world, and they has been called seleniferous areas. Some of these regions include an area in USA (up to 28 mg/kg), Ireland (up to 1,200 mg/kg), China (up to 59 mg/kg) (HARTIKAINEN et al., 2005) and in South America, as Venezuela and Colombia (COMBS, 2001). Selenium-enriched soils are constituted by carboniferous shale (600 mg/kg), phosphoriferous rock (300 mg/kg) or coal (6,500 mg/kg) (FORDYCE, 2007). For example, soil samples from seleniferous regions as Niobara and Pierre shales (USA) possess up to 90 mg of Se/kg, while majority of nonseleniferous soil based on low-selenium granites and metamorphic sandstone contain levels significantly lesser than 2 mg of Se/kg (TRELEASE, 1945; 22
ERMAKOV, 1992). At Hubei province, in China, Yang et al. (1983) described that the selenium level in soil was approximately 790 ȝg/100 mg, of which 35 ȝg/100 mg were water-soluble selenium. Surprisingly, the level of selenium in samples of coal extracted from that region was 291 ȝg/g, but it was identified a sample with more than 84 mg/g, while, in USA, the level in coal normally is around 1 – 2 ȝg/g. By the other hand, regions where soils are poorer in selenium are found along an extensive band from South-West to North-East of China, with soils containing between 0.022 and 0.125 mg/kg (TAN et al., 2002), where is reported prevalence of both, Keshan and Kashin-Beck diseases. Based on selenium distribution in arable layer of Chinese soils and its relation with Keshan and Kashin-beck disease, Tan et al. (2002) categorized Chinese soil as deficient in the microelement, containing lesser than 0,125 mg of Se/kg (where were reported cases of two diseases); marginal soils, containing between 0.123 and 0.175 mg of Se/kg, and selenium-excessive soils, containing levels higher than 3 mg of the micronutrient/kg. In China, at municipality of Enshi, geo-ecosystems selenium-deficient and excessive may be identified as near as 20 km one from other due to natural variation in the geology between these regions (FORDYCE et al., 2000). Other regions poor in the microelement inclose Australian, north region of North Correa, Nepal, Tibet and the central region of Africa, particularly the Democratic Republic of Congo (TAPIERO, TOWNSEND and TEW, 2003). In USA, selenium deficient soils inclose a wide region that covers vales and mountains of the Beaverhead, Big Hole, basin of the Bitterrot River and hills at the South-East, as well as part of the basin of Clark Fork River, in Montana. In these locality plants harvested and evaluated showed lesser than 0.1 mg of Se/kg of dry matter, and the white muscle disease was a serious problem. Similarly, along a band at West from the Salt River until the Phoenix one also contain low selenium levels, as well as in a great part of States of Wisconsin, Illinois, Indiana, Michigan, Ohio and along all west of Virginia, Maryland, Pennsylvania, New York e New England. However, the region identified as containing lowest concentration of selenium in USA is 23
located at North-West of Pacific Ocean and along all South-East of the country, where the middle concentration of selenium in roughages was lower than 0.05 mg/kg of dry matter (KUBOTA et al., 1967). In Europe are found regions constituted by poor selenium soils in Finland (KOLJONEN, 1973a,b,c, 1974), Denmark, and Eastern and central regions of Siberia (Russia). Other regions certainly constituted by selenium-deficient geo-ecosystems inclose New Zealand (COMBS, 2001) and the Mexican Plateau, where were measured concentrations of selenium in soil as low as 0.047 mg/kg (RAMíREZ-BRIBIESCA, 2001a). In Brazil, at municipalities of Corumbá and Aquidauana (MS), it were evaluated selenium concentrations in animal tissues and identified low levels of the micronutrient ( organic matter joined > soluble > extractable. Probably, iron and aluminium oxides would be major responsible by selenium adsorption in these soils. Related to texture of soils, in a layer between 0 and 20 cm depth and, especially, in samples taken at 20 – 50 cm depth, the major portion of selenium was recovered from the fraction constituted by clay, what retained approximately 50% of all selenium in this soil. In a peculiar manner, in the samples of these three soils taken in Taiwan, at 0 – 20 cm depth layer, the major fraction of selenium were as selenite. Specifically, in samples of one of this soils (“Pichen”), at 0 – 20 cm depth layer, that showed clayish texture, whose pH was 4.59 and organic matter content was around 8.56 g/kg, approximated 70% of the recovered selenium was as selenite. This observation could be related to the low pH and, maybe, could be resultant from higher microbial activity of this soil. The chief transformation caused by microorganisms on selenium status in soils includes reduction process, as from Se(IV) to insoluble Se0 (ROUX et al., 2001), or production of volatile compounds (ZHANG and FRANKENBERGER, 1999). In addition, as selenium is an essential micronutrient, it may be incorporated to microorganisms in the form of selenoproteins (STOLZ and OREMLAND, 1999), contributing to the 29
micronutrient retention. Together, these transformations affect the selenium soluble fraction, which is the proportion available to absorption by plants. For example, in a soil that contained 787 ȝg of Se/100 g and with 2.5% of organic matter, the soluble selenium fraction was 35.4 ȝg/100 g (about 4.3% of the total). Meanwhile, in another soil containing 5.9 ȝg/100 g of total selenium, but with only 0.36% of organic matter, the level of soluble selenium was 0.44 ȝg/100 g (about 7.8% of the total) (YANG et al., 1983). It has been suggested that greater organic matter content in soil could provide reducer equivalents utilized by soil microorganisms in the reduction from Se(VI) to Se(IV). This last, as shows higher adsorption by forming inner sphere complexes with iron oxides than Se(VI) (HAYES et al., 1987), would be responsible by the reduction of selenium availability in this soil. Concerning to the influence of microbial activity on selenium distribution in soil, Février, Martin-Garin and Leclere (2007), by comparing variations of the distribution coefficient of selenium between a sterile and crude soil, observed that the Kd-Se increases from 22 L/kg to 50 L/kg after 21 days in contacting to crude soil (with microbial activity), while on sterile conditions the Kd-Se was not changed along the experiment (30 days). In addition, when crude soil was mixtured with glucose and nitrate (both at 5.6 mmol/L), i.e., a source of carbon and another of nitrogen, there was four times increase in microbial population in the soil. As result, this treatment caused a elevation from 30 to 50% in the proportion of Se(IV) and in an increment of kd-Se value from 22 L/kg to 108.3 L/kg. This observation demonstrate that microorganisms increase selenium imobilization in soil and that this change occurs concomitantly to the increment in proportion of Se(IV). Additionally, a fraction of selenium probably was reduced to elementary selenium. Thus, both, reduction of free selenium species to selenite and to elementary selenium, as well as the assimilation of selenium by microorganisms could explain the increase in imobilization of selenium provoked by microbial activity of soil (FÉVRIER, MARTIN-GARIN and LECLERE, 2007). Similarly, the microbial activity of soil also provokes volatilization of determined amount of selenium due forming volatile compounds, mainly
30
methylated species as dimethyl-selenide (DMSe) and dimethyl-diselenide (DMDSe). According demonstrated Bañuelos and Lin (2007), additioning carbon sources to soil increases the selenium volatilization rate by soil microorganisms. Originally, the soil studied contained approximately 4.8 mg of Se/kg. The addition of 71.4 mg of methionine to samples of this soil resulted in an approximated volatilization of 434 ȝg of Se/m2 in the first day of contact. When the carbon source was casein (572 mg/kg of soil), the volatilization rate reached about 346 ȝ of Se/m2 in the first day of contact. These results demonstrated that soils with high microbial activity and elevated levels of organic matter could retain more selenium by accelerating reduction of oxy-anions as a manner to render it less soluble, by immobilizing more selenium in microbial selenoproteins and due complexing more selenium by joining it to organic matter. Additionally, the conversion of inorganic selenium species into volatile metabolites by soil microorganisms allow transfers a significant fraction of the micronutrient to atmosphere. As it will discuss later, to soil highly polluted with selenium, the volatilization by microorganisms or by plants, that uptake elevated amounts of selenium from soil and eliminate volatile selenium species by leaves, has been studied as way to alleviate the selenium overload in soils. By the other hand, abiotic factors as the reduction potential and the oxygen concentration may affect the proportion among several chemical forms of selenium in reactive mediuns as soil and water. In this sense, Kumar and Riyazuddin (2011) evaluated samples of ground water taken from 65 dug wells in India before (May) and after (January) the rain station, between 2004 and 2007. Researchers observed that, before raining station, the level of Se(IV) of 64 samples evaluated varied between 0.15 and 0.43 ȝg/L. At the same period, the level of Se(VI) of these samples was between 0.16 and 4.73 ȝg/L. After raining station, it was observed increase in both, total selenium and selenate concentrations in relation to selenite in samples. In this period, the Se(IV) concentrations varied between 0.15 and 1.25 ȝg/L and the Se(VI) between 0.58 and 10.37 ȝg/L. In this water samples, pH was between 6.5 and 8.8, which characterize the water as lightly alkaline. The reduction 31
potential varied from 65 to 322 mV, indicating a character lightly reductive to moderately oxidant of this water. The diluted oxygen level was between 0.25 and 5.00 mg/L. Interestingly, after raining station (January) the increase on proportion of selenate in the water was accompanied by increment of the oxidant potential and elevation of the oxygen level diluted in water. For majority of samples, there was positive correlation between the increment in oxidant potential and in oxygen level diluted in water, accompanied by an elevation in selenate concentration. Other factors that, according authors, could impair the reduction from selenate to selenite and to elementary selenium in this water samples would be its lightly alkaline pH. In this context, it should be considerated the elevated bicarbonate concentrations in this samples (73 – 650 mg/L). As discussed above, selenium availability to plants is determined by factors related to soil, as its pH, its iron and aluminium oxides levels, its microbial activity, its reduction potential, its organic matter and clay content, in addition to competition with other elements contained in earth. Furthermore, many studies on availability of this microelement in soil have focused on its speciation. However, physical properties inherent to soil, as compaction level, erosion and water availability, may affect plant growing and root functions, ions transport process from surface of roots and lixiviation of solutes to far from the roots reach. Attempting to investigate the influence of these factors on selenium concentration in wheat grains, Zhao et al. (2007) carried out a field experiment utilizing samples of a loamy-sandy soil located in United Kingdom, which was submitted to four compaction levels and three irrigation regimes. This soil contained, approximately, 0.23 mg of Se/kg, 10.7 g/kg of organic matter, 1.2 g/kg of nitrogen and its pH was 6.2. Along the two years of study (2003 and 2004), selenium concentration in grains was in a range of 10 – 115 ȝg/kg, in 2003, and 11 – 54 ȝg/kg, in 2004. In 2003, as irrigation as soil compaction provoked reduction of selenium levels in wheat grains. In 2004, there was effect of irrigation, causing reduction between 30 and 75% of selenium concentration in wheat grains 32
produced by plots submitted to intermediate and high irrigation regimes, respectively. Authors commented that, in 2004, the compaction was less intense because the soil showed lower resistance to penetration of penetrometer. They suggested that this difference could be consequence of soil moisture (that was around 15% in 2003 and 5% in 2004) at the time when soil was compacted by successive crossing (0, 1, 4 or 8 times) of an 11-ton tractor. Hard soil compaction generally impairs root growing, which may restring nutrient uptake by plants, particularly of those in lesser concentration in soil, as is the case of selenium (ZHAO et al., 2007). The irrigation may affect selenium level in wheat grains by, at least, three mechanisms. The first and more simple is the rain effect. In the study carried out by Zhao et al. (2007), in 2003, intermediate and high irrigation regimes provoked increments in production about of 17 and 20%, respectively. In 2004, on same conditions, the increment reached 19 and 54%, respectively. This effect became more evident when Zhao et al. (2007) demonstrated there is negative correlation between production level and selenium concentration in wheat grains. However, as production increases approximately twice and selenium concentration diminished about 10 times, other factors could be influencing in this fall. The water utilized for plots irrigating contained high sulphur levels (49 mg/L), which could inhibit Se(IV) and Se(VI) uptake by wheat plants roots. In total, the intermediate irrigation regime would be carrying between 20 and 57 kg/ha of sulphur, while the highest one would add about 143 - 202 kg/ha of sulphur. Thus, by irrigating plots could accelerate Se(VI) lixiviation (ZHAO et al., 2007) causing reduction in microelement uptake by plants due carrying it to deeper layer of soil, too far to be reached by roots. In resume, it is observed that the interactions between plant and soil are very complex to the selenium, and several mechanisms inherent to soil and to climate may affect the micronutrient availability for plant uptaking. Similarly, practices of soil manegnament and fertilization schemes may also interfere on dynamic equilibrium of the plant-soil system. On top of food 33
chain, human nutrition and health in concerning to the selenium depends, at least instance, on soil characteristics where foods are produced. However, a new level of complexity needs to be considerate about selenium role in geoecosystems that are interactions among soil, climate and the effect of several life organisms on selenium recycling in nature. Selenium shows a cycle in nature that initiates by its evaporation by plants and microorganisms of soil, and following it is incorporated in tissue proteins and converted into volatile compounds (for example, dimethyl-selenide and dimethyl-diselenide). These metabolites are introduced in the atmosphere and brought back to soil by raining or attached to suspense particles that sediment on surface of earth (SHRIFT, 1964; STORK, JURY and FRANKENBERGER, 1999). Thus, the atmosphere represents the major source of selenium to earth surface, deposited by raining or through powder particles that sediment. However, despite activity of soil microorganisms and plants in converting inorganic selenium into volatile organic compounds, as methylated species, the major selenium input for atmosphere and for soil is from oceans surface slime. It was estimated that the marine environment would be responsible by 45 to 77% of total selenium emissions to atmosphere (MOSHER et al., 1987; AMOUROUX and DONARD, 1996). Like marine sulphur cycle, it has been thought that selenium volatile species are evaporated by oceans water slime and sequestered by air suspended particles. These aerosols are transported by wind from ocean to continents, where, then, will are deposited on surface of earth (MOSHER et al., 1987; ELLIS et al., 1993) (Figure 2.1).
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Figure 2.1 Oceans represent the major selenium source for life organisms. The evaporation of volatileselenium species from water slime is resultant from the phytoplankton activity. These volatile compounds are transported by wind from oceans to continents, where sediment attached to suspended particles or fall dissolved in raining drops (photography done by author: dawn at Imbé beach, RS, Brazil).
In this context, Amouroux et al. (2001) developed a study to investigate the cycle and distribution of selenium in marine environment. Researches taken samples at 1 meter depth in a region located at north of the Atlantic Ocean. In fractions of these samples it was evaluated the presence of selenium volatile species, which were determined by cryofocusing gas chromatography and inductively coupled plasma-mass spectrometric (ICP/MS) detection. Amouroux et al. (2001) identified three major volatile selenium species in their samples that were: dimethyl-selenide (DMSe), dimethyl-diselenide (DMDSe) and dimethyl-selenyl-sulfide (DMSeS). DMSe and DMSeS were
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detected at higher concentration in waters evaluated. Indeed, DMSe account to 50% of all selenium-volatile species determined in those samples. Concentrations of DMSe and DMSeS were compared with several species of phytoplankton found in ocean surface. Researches detected significant correlation between DMSe and the amount of seaweed of Coccolithophoridea family (expressed as mg of carbon/m3). In addition, Amouroux et al. (2001) identified dimethyl-sulfide (DMS) in water samples, and its concentration also showed correlation with the amount of seaweed of Coccolithophoridea family. These researches commented that, in another study done in this area and during bloom of phytoplankton it was reported correlation between of Coccolithophorideos, as the species Emiliania huxleyi, and the concentration of DMS (HOLLIGAN et al., 1993). Thus, Amouroux et al. (2001) suggested that DMSe also would be produced by Coccolithophorideos like DMS, by biosynthesis and cleavage of the metabolic dimethyl sulphonium propionate (ANDREAE, 1990). Furthermore, in a hyper-saline medium containing high concentrations of selenium, the dimethyl-selenonium ion (DMSe+-R) was identified in a proteic fraction from some seaweed tolerant to hypertonic medium (FAN, LANE and HIGASHI, 1997). These seaweed species produced high concentrations of selenium-volatile compounds, reinforcing evidences that the selenium volatilization process that occurs in the marine environment is really due to the phytoplankton activity. However, taking in account that there was none correlation between DMSeS concentrations and the amount of seaweed in samples, Amouroux et al. (2001) postulated that this metabolite would be produced by another pathway, because experiments from other laboratories already demonstrated a chemical/microbial process in aquatic sediments or marine waters doing the conversion of seleno amino acid into DMSeS (CHAU et al., 1976; AMOUROUX, PÉCHEYRAN and DONARD, 2000). It was postulated that DMSe would be produced by nucleophilic displacement from selenonio salt by water or halide, whereas DMSeS and DMSe would be formed by oxidation of methyl thiol and methyl-seleno-thiol moieties (AMOUROUX et al., 2001). 36
The molar ratio of Se to S in marine plankton (0.7 x 104) (CUTTER and BRULAND, 1984) is also quite correlated to the concentration ratio of volatile forms that were measured in seawater. This contrasts with the molar ratio of dissolved elements in seawater (where they are present largely as non-volatile species) of 3 x 108. Then, selenium may be apparently converted into volatile compounds faster than S. The formation of DMSe and DMS in marine environments occurs by uptaking inorganic species of Se and S dissolved in seawater by seaweeds and their biotransformation in the cell. In this process, many reduction and methylation steps are required to the formation of biogenic precursors of both volatile forms. Selenium is in seawater mainly as selenate (SeO42-) and selenite (SeO32-), and sulphur as sulphate (SO42-). The thermodynamic analyses revealed that the calculated free energy for SeO42- and SeO32- reduction into selenide (HSe;-22.2 and -22.9 kcal/eq., respectively) is four times higher than the reduction of SO42- to sulfide (HS-; -5.9 kcal/eq.). This important difference in terms of free energy demonstrates that reduction of selenium ions is more favorable than sulphate reduction, which could explain the relative abundance of DMSe in relation to DMS (AMOUROUX et al., 2001). In this study, researchers observed a ratio of volatile species of Se and S of, approximately, 1.5 x 10-4. In the past, the relation Se/S measured in ice cores was 1.6 x 10-4 (MILNE, 1998), for the period from 800 B.C. up to 1892, i.e., prior to major industrialisation. The relation diminished between 1892 to 1952, to reach a value about 0.7 x 10-4 after 1952, due, presumably to greater relative sulphur emission than selenium one. This tendency could be explained by the increment in sulphur emission by anthropogenic activities, as fire of excessive petroleum amounts, which shows a low relation Se/S (0.1 – 0.5 x 10-4). Thus, the similarity between volatile concentrations of Se/S found in sea waters and the relation between elements in ice cores indicates that marine emissions regulates the global cycle of both element in nature, which seems true, at least, to the preindustrial period (AMOUROUX et al., 2001).
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Taking in account the change model between sea and air developed by Liss and Merlivat (1986), Amouroux et al. (2001) estimated that selenium emissions from sea could reach the total of 35 x 109 grams/year. Considering food production, the recycle of selenium from sea represents an important economy of fertilizers. Unfortunately, the disastrous human hands have also perturbed this rich and wonderful equilibrium in nature. It is evident that we are able very more to pollute than to recycle, and the selenium would not escape from this lamentable rule.
2.2.2 Selenium as environmental pollutant
In countries that utilized coal as energy source to function industries or thermoelectric powers it has been reported environmental problems due selenium sedimentation, after attaching to fume particles derived from coal fire. On this concern, Reash et al. (2006) reported a case of selenium contamination by sedimentation of fume from a thermoelectric power at Ohio State, in USA. The power plant started to operate in 1974 and, since released fume from its chimneys, through tube systems on surface of a lake with, approximately, 100 ha of area. Components more dense in fume were deposited on water slime of lake and contaminated not only fishes, but also insect larva and other wild species that lived on surface of water, or that ate phyto and zooplankton. Along of several years, sediment of the bottom of lake also began to accumulate high selenium levels. In 1994, to check environmental concentrations of the micronutrient, researchers evaluated tissues of a species of fishe that, certainly, would be very sensitive to selenium and heavy metals contained in fume of coal. Samples were taken from a little river that drained the output of lake to the Ohio River, which was located below. Selenium amounts recovered from fishes body was 17.3 ȝg/g of dry weight, in ovaries the selenium level was 32.5 ȝg/g of dry weight and in testis the contamination reached 37.1 ȝg/g of 38
dry weight. The highest level recovered was from fishes liver, whose value achieved 57.33 ȝg of selenium/g of dry weight. The upper limit tolerated to selenium by fishes body and ovaries was determined as being 7.9 ȝg/g and 17 ȝg/g, respectively (US EPA, 2004). In another study, it was measured in liver a contamination of 29 ȝg/g of dry weight, which was sufficient to impairs fish reproduction, while at 34 ȝg of Se/g of dry weight it was observed fish mortality (LEMLY, 1993). Analyses of river sediment revealed contamination around of 20 ȝg of Se/g of dry weight. In addition, Reash et al. (2006) also evaluated tissues from fly living and eating on slime of water. This kind of flies showed selenium levels about 18.5 ȝg/g of dry weight. This value is very higher than that found in sediments and in flies colected on the reservoir of a sludge treatment company located near the power plant, which drained its output to Ohio River too. Samples from this lake contained 1,800 ng/g after fertilizing plants with 100 g/Se/ha. Similar variation was observed for bread, that showed a middle selenium concentration of 35 ng/g when made with flour from borders without selenate application (control-plots), but reaching concentrations >1,900 ng/g when produced with flour from borders fertilized with 100 g of Se/ha. Additionally, Hart et al. (2011) demonstrated that there is linear correlation between selenium content in fertilizer that was added to plots, selenium level in its respective flours and in bread produced with these flours. This observation indicates that there was an efficient incorporation and retention of selenium in wheat plants, as well as efficient transference of the micronutrient to grains, flours and breads. However, selenium amount carried by wheat grains to integral flour diminished insofar as fertilization level was increased: below 5 g of Se/ha, 100% of selenium was transferred, but this fraction decreased to 96% after adding 10g of Se/ha, 91% with 20g of Se/ha, but was maintained at 83% and 84% with 50g and 100g of Se/ha, respectively. Although it occurred variation in selenium concentration of soil and, hence, in grains produced by control plots during the two years along experiment (2006 and 2007), the enrichment rate of flours and breads was reproducible in both periods. For example, to the level of 15 g of Se/ha, the increment in selenium content of white flour in relation to control group was 78
254 ng/g in 2006 and 238 ng/g in 2007. In the case of integral flour produced with grains from border that were fertilized with 15g of Se/ha, the increase in 2006 was of 294 ng/g and, in 2007, of 287 ng/g (HART et al., 2011). The integral flour and the bread made from it showed higher selenium concentration for the same level of fertilization that white flour and its respective bread. The white bread showed, in mean, 13% less selenium that bread made with integral flour (HART et al., 2011). This difference may be due selenium distribution in wheat grains. Although selenium is more concentrated in embryo of seeds, it is distributed more uniform that other mineral of wheat grains, thereby a fraction should be deposited also in the husk (LYONS et al., 2005b). The predominant chemic specie of extractable selenium in both flours, white and integral, was SeMet. This seleno amino acid represented, approximately, 80% of total selenium measured in wheat products. After making bread, the content of SeMet fell somewhat, but was maintained near to 78% of the total extractable selenium. Other selenium species indentified in flour and breads were, approximately, SeCys (5%), MeSeCys (3%), selenito (5%) and selenate (4%) (HART et al., 2011). But, another study identified almost 90% of total selenium in wheat as SeMet (STADLOBER, SAGER and IRGOLIC, 2001). The selenium carried by wheat is well absorbed and retained in the organism, as demonstrated by another study with human healthy volunteers (FOX et al., 2005). Another study showed an increase of dietetic ingestion of the micronutrient by eating selenium-rich bread, providing 200 ȝg/day, causing an increase of plasmatic selenium from 65 ȝg/ml to 165 ȝg/ml (THOMSON, ONG and ROBINSON, 1985). Nevertheless selenium in the form of SeMet to be well absorbed and retained in organism, its bioavailability seems be not as high as that of selenito (THIRY et al., 2012). Over this limitation, diary bread consumption of British population is around 80 g for women and 122 g for man, corresponding to 2 and 3 slices, respectively (HENDERSON, GREGORY and SWAN, 2002). Taking in account selenium concentration in bread produced from grains whose plots were fertilized with 15 g of Se/ha and the amount consumed by local 79
population, the consumption of these bread would resulted in an ingestion of 11.6 – 13.5 ȝg for women and 17.4 – 20.2 ȝg for men, corresponding to 19 - 27% of the requirement fixed in UK. This estimated increment of selenium intake from bread made with this flour would increment the selenium consumption by British population to 60 ȝg for women and 75 ȝg for men. Because these differences of soil composition of UK and due variation on selenium status of British population, Hart et al. (2011) concluded that wheat biofortification through utilizing fertilizers could be reached by applying selenate to crops within a range from 5 to 15 g/ha. In Australia, a study revealed that medium selenium concentration of local population would be of 103 ȝg/L (LYONS et al., 2004a), which is lower than measured in many countries and only somewhat higher that 100 ȝg/L that is need to maximize the expression of important selenoenzymes, as GSH-Px (RAYMAN and CLARK, 2000). However, only 8% of the evaluated men showed levels higher than 120 ȝg/L, whose concentration is thought to protect against prostatic cancer (BROOKS et al., 2001), the most common solid tumor diagnosed in masculine Australian population (MEUILLET et al., 2004). Therefore, some strategies for increasing selenium levels of Australian population has been proposed, among them: i) consumption of natural selenium-enriched foods (Brazilian nuts, cereals, meats and fishes); ii) adds selenium to water; iii) enrichment of food at processing in industries; iv) biofortification, including supplementation of food sources, use of fertilizers containing selenium and; v) development of plants that concentrate selenium in eatable parts. Worldwide, wheat represents an important dietetic source of selenium, and to do the wheat to be selenium-enriched is relatively easier to be achieved in the practice. In Australia, it has been estimated that near to half of selenium intake by people is via wheat byproducts. Thus, it is rational to considerate this crop as a potential target for “agronomic biofortificaton” and for plant breeding to higher capacity to uptake from soil, translocate and concentrate selenium in eatable parts (“genetic biofortification”). It is likely that these two strategies are more promising and desirable methods for 80
increasing selenium status of human population and adequated to be applied in Australia. Then, in a study to evaluate levels and forms of selenium to be supplied to crops at South of Australia, Lyons et al. (2004b) observed that the application on soil was more effective than applying later, as foliar fertilizer (in the flowering). Soil fertilizing showed some advantages, as by not causing damage to plants, because the fertilization was carried out before the germination. In addition, wheat crops at South of Australia generally are on threat of drought and stress by heating, which result in lesser foliar area. Therefore, these researches recommended that the application of seleniumcontaining fertilizer on soil before seeding would be more adequate for that region. The highest conversion efficiency rate was obtained by the highest level of fertilization, i.e, 120 g/ha. Due this dosage it was possible to reach production of 2,700 kg/ha, whose grains contained 10 – 12 mg of Se/kg and with additional cost below $40/ha. Researchers commented, even, that wheat seems tolerate better high levels of selenium supplementation than other crops, as rice, tobacco, and soya (LYONS et al., 2005a). Field experiments has showed that fertilization with up to 120 g of Se/ha, as sodium selenate, provoked no reduction on productivity or any other adverse effect to wheat plants. Moreover, after applying 500 g/ha of sodium selenate on soil or 330 g/ha as foliar fertilizer, none toxic effect was observed. In an assay done in greenhouse, it was observed that selenium would be toxic for plants from concentration of 325 mg/kg, which would be achieved following a level of 2.6 mg of sodium selenate per kg of soil. In another study, where were evaluated selenium effects on germination process, it was demonstrated that up to concentration of 150 mg/L (as sodium selenate) or up to 23 – 39 mg/L (as sodium selenite) it was not observed undesired effects on germination. The inhibition of early roots grown was observed at concentration between 6 and 13 mg of Se/kg, to both selenium species, but the most pronounced effect was observed to selenite (LYONS et al., 2005a).
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One of worries that have emerged after the advent of fertilizing crops with selenium applied directly on soil is the possibility of ground water contamination. As debated in previous sections, only a little fraction of total selenium is found as water soluble micronutrient, which could be absorbed by plants (around 5 to 10% of the total selenium). This means that major portion of the microelement will is attached to metal hydroxides, joined to clay particle or to humus, whose proportion is on dependency to physical and chemical properties of the soil. But, along several years the excess retained could result in the micronutrient overload in soil. Insofar as buffering mechanisms are overtaken, the micronutrient could reach superficial waters, or even more grave, lixiviates and achieves ground water. Therefore, in countries as Finland, contamination levels in soil and water are on permanent and careful monitoring. Maybe, applying foliar fertilizers in the form of sodium selenate may represent an alternative to minimize this hazard. In resume, all data showed above indicate that, independently on selenium speciation or on manner to application, wheat may be utilized as a selenium carrier to human diet. In addition, it is probably that wheat tolerate well a wide range of selenium doses and toxicity seems not happen in field conditions. In this sense, searches evaluating selenium fertilization at levels between 10 to 200 g/ha, either applying on soil or by pulverization as foliar fertilizer, reported that it was no danger. Hence, this wide security allowance renders wheat crop a good alternative for carrying selenium to human diet (LYONS et al., 2005a). By the other hand, in other countries the cereal that has potential to be introduced into the “agronomic fortification” is the rice. For long it has been recognized that rice produced in determined regions of China and India is poorer in selenium than grains produced in USA (BIERI and AHMAD, 1976). These authors found that selenium concentration in rice produced in Bangladeshi varied between 0.06 and 0.17 mg/kg, while rice grains produced at American State of Louisiana contained about 0.46 mg/kg of the micronutrient. Chen et al. (2002) measured the level of selenium in rice produced in China and reported middle concentrations around 0.025 mg/kg. In another study, Liu et al. (2009) determined selenium concentration of 10 82
cultivars of cereal and reported medium values of 0.024 mg of Se/kg to integral rice. However, considerable portion of selenium is lost during the grain polish process (LIU et al., 2009). The integral rice is obtained by withdrawing husk from crude grains, what is constituted by a yellowish layer (6 – 7% of total weight of grain), by embryo (2 – 3% of total weight of grain) and by endosperm (about 90% of total weight of grain) (CHEN, SIEBENMORGEN and GRIFFIN, 1998). The integral rice shows more nutrients, as protein, lipids, fibers, vitamins and minerals than white rice. As nutrients of integral rice are concentrated mainly on the outer yellowish layer and in the germen, the polish process to produce the white rice cause a loss of, approximately, 28% and 84% of proteins and minerals, respectively (LAMBERTS et al., 2007). In this sense, Liu et al. (2009) demonstrated that the selenium loss during rice polish process is dependent of shape and consistency of grains. Therefore, grains more rounded and with higher proportion between thickness and length (shorter and wider) and with lesser porosity (more dense) showed lesser loss by forming bran than thiner, lengthy (with lesser proportion between thickness and lengthy) and more porous grains, to the same time of polish (10; 20; 30; 40; 50; 60; 70; 80; 90; and 100 seconds). Interestingly, according selenium concentration diminishes from outer layer to the endosperm, during the first 30 seconds of polish that occurs the major loss to the bran. Thus, there was linear correlation between the amount that is lost (as quantity of bran produced) and selenium concentration loss to the bran, mainly from 10 to 30 seconds of polish. The selenium loss increased from 6.51% to 34.28% at the same time that the percentage lost as bran increased from 2.71 to 17.96%. Hence, it was concluded that integral rice is richer in selenium that white one and, as more prolonged will be the polish time, higher will be the portion of the micronutrient removed from the surface of grain to into bran. The consumption of rice by Chinese population, mainly by rural people, varied between 300 and 500 g/person/day. As it is the major food and is ingested at high quantity, Chinese researches have looked for to develop
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strategies for enriching rice grains with selenium, because, in the majority of localities, concentration of the micronutrient in these grains is very low. On this concern, Chen et al. (2002) reported that selenium application by pulverization (as foliar fertilizer), in dose of 20 g/ha, resulted increase of the micronutrient concentration from 0.025 mg/kg (control group) to 0.47 and 0.64 mg/kg, when applied in the form of sodium selenite and sodium selenate, respectively. Authors concluded that selenate was more effective to increase selenium content in rice grains than selenite. At mean, selenium application as selenate resulted in concentrations of the micronutrient in grains about 35% higher than when application was done with selenite. Thus, Chen et al. (2002) finished concluding that if selenium content of rice grains could be maintained between 0.3 and 0.5 mg/kg and considering the diary cereal consumption by Chinese population of about 400 g, it would be sufficient to elevate the micronutrient ingestion from actual 7.5 - 12.5 ȝg to 100 – 200 ȝg/person/day. In relation to natural selenium-rich foods of vegetal origin, it needs to be remembered of Brazilian nuts, an important exportation product obtained by extractive activity of silkman and other native workers in Amazon forest (Figure 2.5A). Nuts are produced by a tall tree that, in Portuguese idiom, is called “castanheira” (Bertholletia excelsa). It grows in Amazon forest and may be identified among other trees (Figure 2.5B). Its fruit is involved by a hard shell and into there are some almonds, that is the eatable selenium-rich nut (Figure 2.5C and D). After collecting from forest, the shell is broken and nuts are withdrawn and polish, to then to be package and sent to market, inclusive, for overseas (Figure 2.5E and F). Nuts are rich in phenolic compounds, flavonoids, isoflavones and terpenes, which are responsible by its antioxidant capacity. These compounds are located, mainly, in outer layer of almond (“brown skin”) (JOHN and SHAHIDI, 2010). In addition, according declarated before, Brazilian nuts are an important source of selenium into vegetable kingdom, and its consumption can be recommended in face of its positive effects on human health.
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A)
B)
C)
D)
E)
F)
Figure 2.5 Brazilian nuts collected from Amazon forest. A) Amazon forest is a very dense one where grow tall trees. Native habitants develop extractive activity by collecting natural products of commercial value (AMAZON FOREST, 2015). B) The tree called “castanheira” (Bertholletia excelsa) may be identified into the forest and produces a special kind of nut (CASTANHEIRA, 2015). C) Fruits are involved by a hard shell (FRUIT, 2015); D) and into there are some almonds, each is an eatable selenium-rich nut (NUT, 2015). E) Fruits fall on soil and are collected by native habitants (COLLECTING, 2015); F) and following processed to human consumption (PROCESSING, 2015).
Regarding to selenium content in Brazilian nuts, a study developed by Chang et al. (1995) reported that middle concentrations in almonds collected at a region of Acre State and at another of Rondônia State (Brazilian
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Amazon) would be about 3 mg of Se/kg, while for fruits collected at a region near Manaus city and Belém city (Brazilian Amazon) would be, approximately, of 36 mg of Se/kg. According theses authors, differences in selenium concentration among nuts collected from different sites in Brazilian Amazon could result from differential composition of soil at each of these regions and/or differences in patterns of selenium uptake from soil by trees. The variation linked to plant genetic variety would occur due physiological characteristics and adaptations of trees growing at different points of forest. On the medical potential of nuts, a study showed that the inclusion of it into rat diet caused a reduction of the number of mama tumors induced by carcinogen (IP and LISK, 1994a). Afterwards, another study demonstrated that the consumption of nuts would result in an increment of selenium plasmatic concentration in humans (LISK et al., 1998). In relation to human health, Cominetti et al. (2012) developed a study with 37 Brazilian women volunteers that were prone to undergo complications related to atherosclerosis, whose were oriented to eat one nut per day (containing total selenium content around 290 ȝg) during 8 weeks. Majority of parameters evaluated in the search (before and after the consumption of nuts) were selenium blood concentration (of plasma and of erythrocytes), GSH-Px activity of erythrocytes, lipid profile [total cholesterol (TC), triglycerides (TAG), the proportion among very low density lipoprotein (VLDL), low density lipoprotein (LDL), high density lipoprotein (HDL)] and determination of atherogenic risk [Castelli index I (proportion between TC and HDL) and Castelli index II (proportion of LDL and HDL). It is advised that before consuming nuts volunteers showed low plasmatic selenium levels (mean at 55.7 ȝg/L) and of erythrocytes (mean at 60.5 ȝg/L). Cominetti et al. (2012) demonstrated that the consumption of one nut per day, during 8 weeks, was sufficient to elevate selenium concentration of plasma and erythrocytes to 132.5 and 205 ȝg/L, respectively. At the same period, these researches observed that the elevation of blood selenium levels was accompanied by a significant increase in GSH-Px activity, i.e, from 36.6 to 53.6 UI/g of Hb. However, the most important result identified by group was the increment of serum concentrations of HDL caused by eating nut, 86
which was increased from 37.6 to 44.5 mg/dl. Consequently, there was an improvement in Castelli index I and II, demonstrating that the consumption of Brazilian nuts could help to prevent atherosclerosis. This result reinforces the role of selenium in preventing cardiovascular diseases and adds substantial value to Brazilian nuts, which may be considerated a nutraceutical food. Unfortunately, the consumption of nuts has been very restricted, inclusive, in Brazil. Taking in account this brief discussion about foods from vegetal origin as dietetic via to increase selenium status of human population it seems obvious that, in majority of cases, it will be need to fertilize crops with the micronutrient to achieve elevated levels of selenium in foods. With exception of some plants, as those belonging to Brassica and Allium genders, or Brazilian nuts and some species of mushroom, majority of crops does not accumulate selenium. Thus, to do of grains, as wheat, rice and their byproducts important sources of selenium to human diet, it will be need to utilize agricultural techniques as the “biofortification”. Among them, it is recommended to do crops fertilization with selenate at the time of seeding or, after, as foliar fertilizer applied at the time of flowering. Likely, due their wide consumption around world, wheat and rice are will the preferred selenium food carriers to be introduced into “ agronomic biofortification strategy”, with results economically more viable im terms of agricultural production. In relation to effects on public health (as increase of GSH-Px activity, protection against heavy metal toxicity, prevention of several types of cancers and cardiovascular diseases) easier to be achieved in the practice. Additionally, wheat and rice could be improved to absorbed more selenium from soil and to deposit it into grains.
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2.2.6 The “genetic biofortification”
The strategy based on development and selection of plants of agronomic interest that concentrate more selenium in their eatable parts is called “genetic biofortification”. In plants and microorganisms, selenium is generally associated to protein fractions as SeMet, the most abundant seleno amino acid. MeSeCys is the most common selenium source in seleniferous plants, but it is found in roots of onion and garlic too (WHANGER, 2002). Genetic differences among roughages also may modify the rate of the microelement accumulation in aerial parts, which are ingested by herbivorous animals (McQUINN et al., 1991). Thus, it was carried out many studies to identify wheat lines that would be able to concentrate selenium in grains, however majority differences were due to soil composition but not related to genetic variation. Nevertheless, some differences there are. For example, a study found difference on selenium deposition by diploid wheat and rye, which deposited higher selenium levels in grains, as well as other minerals (S, Fe, Zn, Mn) (LYONS et al., 2005c). Into leguminous group, as Astragalus gender, it may be found seleniumaccumulators and non-accumulators species, with concentrations between 1,322 and 289 ȝg/g of MS, respectively, measured in leaves (SORS et al., 2005). This and other studies have indicated that the capacity to accumulate selenium is dependent on plant metabolism adaptations and on tolerance mechanisms. In Astragalus species, the variation in amount of selenium deposited in leaves is due to metabolic pathways that lead up to the escape metabolites generation, as SeCys and MeSeCys (SORS et al., 2005). Selenium uptake by hyper-accumulators may depend on changes of uptake mechanisms by roots and selenium transport in tissues, in which selenium compete with sulphur transporters. By comparing molar proportion between selenium and sulphur of selenium-accumulator and nonaccumulator species revealed that accumulators prefer to uptake selenium, 88
while non-accumulators has more affinity to sulphur (BELL, PARKER and PAGE, 1992). It was observed a variation in uptaking and subsequently partition of selenium between roots and leaves of determined cultivars of tomato in relation to wild type plants, indicating that both, uptake and transport, may vary in terms of selectivity for different substrates (SHENNAN, SCHACHTMAN and CRAMER, 1990). Thus, the base for genetic manipulation objecting to obtain plants that uptake and deposit more selenium into eatable tissues, mainly grains, should be targeted on mechanisms as uptake, transport, partition and selenium metabolism. One via to obtain plants genetically biofortified could be achieved by selecting plant lines based on its transporter affinity, preferring those that showed more affinity to selenium instead of sulphur. Following in pathway of the micronutrient in plants, the second target would be to select plants based on their nutrient partition characteristics. Thus, more desirable plants would be those with capacity to accumulate more selenium in leaves and vegetative tissues and, afterwards, abler to re-mobilize high levels of the microelement to grains during the filling. The metabolic pathways of selenium also need to be considerate in an improvement program. It should be preferred plants that promote the conversion of inorganic selenium into organic metabolites, which, in these turn, may be sequestrated by plant tissues to, then, to serve as a dietetic source of selenium if deposited into eatable tissues. Hence, as both strategies (“agronomic biofortification” and “genetic biofortification”) are capable to increase selenium levels in foods, their importance may be relevant insofar as would reach all population and would be an efficient and security manner to supplement the micronutrient for human population. Soon, a paradigm change needs to occur in agriculture: in addition to achieve high productivity and to produce with sustainability, the agriculture needs provide foods with better nutritional value (LYONS et al., 2005a).
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2.3Selenium in animal production
The selenium was recognized as a micronutrient necessary to animal and human diet in the finale of 1950 decade, when was observed that it could substitute the vitamin E in diet of rats and chicken for preventing cardiovascular, muscular and hepatic lesions. In that occasion, it was no known the exact compound that was involved in this effect, therefore Schwarz and Foltz (1957) called that selenium-containing compound as “factor 3”. However, in the early of 1970 decade, it was identified participation of selenium as component of the enzyme GSH-Px. Because its incorporation in this enzyme, that acts as part of antioxidants system in animal organisms by protecting cells against hydroperoxides attack, this finding induced several studies to comprise biologic functions of the microelement in animals and humans. Selenium deficiency impairs severely animal productive performance and its health, provoking high mortality of newborns as result from degenerative lesions in myocardium (RAMÍREZ-BRIBIESCAet al., 2001b). Among effects caused by selenium deficiency on productive performance is low weight gain (SHEPPARD et al., 1984; OBLITAS et al., 2000), low production of milk, wool, reduced fertility and low number of pups by calving/hatching (SEGERSON and GANAPATHY, 1980; SHEPPARD, BLOM and GRANT, 1984) and low quality of sperm (BECKETT and ARTHUR, 2005). Although selenium deficiency may occur in all animal species, ruminants seem proner, with higher severity in sheep and goat. Lambs and kids are proner to develop degenerative alterations in myocardium, being that adult animals are on higher risk for developing muscular atrophy. However, calves may also show myocardium degeneration and to suffer sudden death. In resume, selenium may improve animal health and its productivity, ameliorates reproductive performance, elevates its resistance against diseases, as well as to refine the quality of foods produced, as milk and meat.
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Due all motives describe above, in last years it has increased the importance of selenium in animal diets.
2.3.1 Selenium levels in animal diet
Ruminants are naturally more susceptible to selenium deficiency due peculiar characteristic of fermentative process. The activity of microorganism may generate insoluble substances, especially selenures, and significant loss of the micronutrient because its assimilation by own microorganisms. The microbial population converts inorganic selenium species into organo selenocompounds, mainly seleno amino acids. There are reports according selenium concentrations in ruminal flora of adult sheep may achieve, approximately, 46 times the concentration found in the diet ingested. The microbial selenium possess high digestibility. However, the major compound is the seleno amino acid SeMet, that, although shows high bioacessibility and bioavailability, its bioactivity is somewhat limited. The NRC (1996) there has recommended that diet for bovines would need to contain between 0.1 and 0.4 ppm of selenium. In the case of diary cattle, for cows at early of lactation and calves in both stages, weaned and young, the inclusion levels would be between 0.1 and 0.3 mg/kg of diet (NRC, 1996). But, the NRC (2001) fixed selenium requirement for bovine at 0.3 mg/kg of dry matter. For broiler, the NRC (1994) recommended the inclusion of, at the least, 0.15 mg of Se/kg of diet, warning that levels equal or higher than 10 mg/kg of diet would be toxic for these animals. In the case of swine, the recommendation is that diet needs contain 0.3 mg of /kg for all developmental phases (NRC, 2001). For sheep, the inclusion levels would be of 0.2 mg/kg of diet (NRC, 2001). However, nor all selenium content of diet will digested and absorbed by animals; one fraction will metabolized and excreted. For diary cattle intaking approximately 2.5 mg of Se/day, it is estimated that the urinary loss would achieve 0.5 mg of Se/day (IVANCIC and WEISS, 2001). Similar values 91
were reported to dry cow fed with similar quantities of selenium (HARRISON and CONRAD, 1984a). Thus, for cow at the last trimester of gestation eating 10 kg of DM/day, the selenium requirement would be of 0.7 mg/day. Taking in account the absorption coefficient, which is estimated in 40%, the diary amount that diet should be carrier would increase to about 1.75 mg of Se/kg of DM. For diary cattle producing around of 30 kg of milk/day, the selenium requirement would be of 1.7 mg of Se/day, which means that diet would need to supply an amount of 4 mg of Se/day (HARRISON, CONRAD, 1984a). According this estimative, dry cow ingesting 1.4 mg of Se/day (HARRISON and CONRAD, 1984a) and lactation cow eating a total of 4.2 mg of Se/day (IVANCIC and WEISS, 2001) showed a lightly positive selenium balance. Many studies have demonstrated that selenium organic species are more effective to increase concentration of the micronutrient in blood and in milk compared with inorganic ones. However, a higher selenium concentration in milk means an increase of selenium excretion via this fluid, mainly by high production cows. In the mammary gland, the selenium excretion occurs afterwards its random incorporation as seleno amino acids (especially SeMet from diet) to casein and other milk proteins (MUÑIZ-NAVEIRO et al., 2005). In this situation, the proportion that remains for the animal to supply its selenium requirement, i.e., to maintain on normal activity several selenoenzymes, is not known yet (GIERUS, 2007). High production cows regulates their metabolism to maximize the partition of nutrient for milk secreting (BAUMAN, 2000) and, consequently, organic selenium species may be excreted at elevated levels in milk. Hence, the probability of high production cows showed signals of selenium deficiency is greater (GIERUS, 2007), justifying the importance of the micronutrient supplementation for these animal lots. A problem related to selenium absorption and digestion in ruminants is that the micronutrient present in foods could cause alterations in ruminal fermentation, at the same time that the fermentative process could provoke reduction in selenium availability to ruminant absortion.
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In this sense, Kim et al. (1997) simulated conditions of ruminal fermentation by incubating in vitro ruminal samples containing crescent concentrations of SeMet (0; 0,2 and 2 ppm) during a period of 12 hours. These authors observed an increment of acetate molar proportion in relation to other volatile fatty acids by incubating material containing 2 ppm of SeMet. In a more recent study, Wang et al. (2009) investigated effects of supplementating diary cattle with selenium-enriched yeast extracts on ruminal fermentation, diet digestibility, lactation performance and milk composition. In this study, animals were divided into 4 groups: basal diet (control), containing only 0.07 mg of Se/kg; 0.15 mg/kg (group 2); 0.3 mg of Se/kg (group 3) and; 0.45 mg of Se/kg of diet (group 4). The experimental period was of 45 days, of which 30 days were for physiological adaptation of animals and 15 days for observations. Wang et al. (2009) reported that the ruminal pH was lower in the group 3. Total production of volatile fatty acids was higher in all experimental groups supplemented in relation to control, and this increase was more expressive for groups that received 0.3 mg of Se/kg of diet. However, on the contrary than was observed by Kim et al. (1997), Wang et al. (2009) reported elevation in molar proportion of propionate, none alteration in proportion of acetate and reduction in butirate generation. Nitrogen content in the form of ammonia was lesser in groups 3 and 4 in relation to the control, and diminished linearly according the supplementation level. Dry matter intake and proportion of mainly milk components (lactose, fat and protein) was not affected by treatments. Milk total production and milk corrected to 4% of fat was higher in groups 2 and 3 than in groups 4 and control. Selenium secretion in milk increases linearly according the increment in supplementation level. The digestibility of dry matter, organic matter, crude protein, ether extract, neuter detergent fiber and acid detergent fiber in total digestive tract was higher in groups 2 and 3 compared with groups 4 and control. Authors concluded that selenium supplementation as yeast extract improved the ruminal fermentation pattern, milk production, selenium secretion in milk and diet digestibility. They suggested that these effects 93
could be due a stimulator effect played by selenium on intra-ruminal microorganisms or on their enzymes, which would be translated in an improvement of fermentation pattern, with positive reflexes being observed in lactation performance. Numerically, milk production was higher when cows were supplemented with 0.3 mg of Se/kg of diet. Considering dry matter consumption by this group, these cows ingested, approximately, 5 mg of Se/animal/day, which caused an increment in molar proportion of propionate, higher total production of volatile fatty acids and more significant fall of ruminal fluid pH, results that could be closely associated. Although authors did not measured bacterial biomass production in ruminal fluid, the reduction in ammonia level observed to groups 2 and 3 could be an evidence of the increment in ruminal microbial population, with elevation in nitrogen consumption in the rumen. Results also suggest that there was an improvement in digestibility of the main diet components due selenium supplementation observed in groups 2 and 3, which could be a consequence of the increment in ruminal fermentation rate. Selenium availability is related to chemicaç species, therefore, as more reduced is the microelement status, lesser will be its availability for animals. Ruminants absorbed selenium at low magnitude (about 54%) compared with monogastric animals (about 80%), because rumen is an environment that facilitates the reduction (ORTOLANI et al., 2002). In addition, the rumen pH exerts influence on selenium availability. As more acid is the pH of ruminal fluid, lesser will be the quantity of oxidized species of selenium and, consequently, lesser is to be its availability to absoption. Considering higher inclusion of concentrated foods in ruminant nutrition, the resultant fall of ruminal fluid pH would reduce the micronutrient absorption rate. The proportion between voluminosus and concentrate in diet may also affect availability and chemical species of selenium by alterations on microbial population, resulting in different patterns of fermentation and of metabolites that are generated. Finally, selenium availability depends also on digestibility of different foods containing selenium and on profile of selenium compounds into each of these sources.
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Selenide (SeO3-2) and selenate (SeO4-2) may be reduced to elementary selenium (Se0) by ruminal microorganisms as an alternative to detoxification (KIM et al., 1997). Microorganisms may also to incorporate Se in microbial protein (HIDIROGLOU and LESSARD, 1976; KIM et al., 1997). By this via, inorganic selenium of ration may be converted to organic selenium, in the form of seleno amino acids SeMet and SeCys, which are components of microorganism cells (GIERUS, 2007). The apparent digestibility of selenium in roughages and concentrates is about 30 and 60% for sheep, goat and dry cow (HARRISON and CONRAD, 1984a,b). The microelement in forms as sodium selenate, sodium selenide and yeast extract shows digestibility between 40 and 50% (HARRISON and CONRAD, 1984a). It seems there is an antagonistic or competitor effect among selenium and other minerals by digestion and absorption process. Thus, it has been demonstrated that apparent digestibility of selenium was reduced when cows were feeding with diet containing high (1.3%) or low (0.5%) calcium concentrations (HARRISON and CONRAD, 1984b). The increase of sulphur intake may elevate selenium requirement. For sheep it has been demonstrated that increments in sulphur level (in the form of sulphate) from 0.05% to 0.24% diminished selenium absorption (POPE et al., 1979). Additionally, true digestibility of the microelement in diet of lactating cows fell from 56% to 46% due increasing in sulphur concentration from 0.2% to 0.7% (in the form of calcium sulphate or magnesium sulphate) (IVANCIC and WEISS, 2001). However, it is not a rule because sulphur supplementation as anionic salts during last 3 weeks of gestation did not affect selenium levels in diary cattle (GANT, SANCHEZ and KINCAID, 1998). Comparatively, in beef cattle fed during long period with diet containing 0.2 and 0.5% of sulphur neither affected selenium blood levels nor GSH-Px activity (KHAN et al., 1987). It may occur, even, interaction between selenium and copper. For sheep fed with diet containing 0.3 mg of selenium/kg of DM, the increase in cooper concentration from 7 to 21 mg/kg of DM provoked an increment of selenium
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concentration in liver, but diminished it in muscle (HATMANN and Van RYSSEN, 1997). In relation to zinc, the increment in concentration of this microelement in diet of rats, from 5 to 20 mg/kg, reduced selenium absorption in 25% (HOUSE and WELCH, 1989). Selenium absorption in ruminants is similar to monogastrics. The selenide absorption occurs by diffusion and, hence, is proportional to quantity of the element in intestinal lumen (GIERUS, 2007). The selenate absorption occurs by active transport, being helped by transporters (Na+ and OH-) and the mechanism is the same utilized by sulphate. The SeMet is also absorbed by active transport, similarly to methionine (VENDELAND, 1994). It should be emphasized that one part of selenate in diet will be absorbed by diffusion. Only a little of elementary selenium is absorbed in intestine due it low solubility and, hence, it will be excreted in feces (GIERUS, 2007). In the case of ruminants, there are studies demonstrating that organic selenium compounds (as yeast extracts enriched with selenium and seleno organic compounds in diet) could provoke more increment in plasmatic concentration of the microelement and in GSH-Px activity than the same quantity of the selenium as inorganic salts (CONRAD and MOXON, 1979; JOHANSSON et al., 1990). However, it should be remebered that comparative studies among different sources of selenium in relation its absorption and bioavailability needs consider the microelement concentrations in blood and tissues, as well as evaluate the activity of GSH-Px, that shows one of biologic effects of selenium in animal organism. Thus, the micronutrient deposited in the form of muscular proteins (as SeMet) does not exert functional role, except by representing a source of selenium due the protein turnover, since that seleno amino acids will be recycled into cells. Organic forms of selenium that are incorporated as non-functional proteins, as wool proteins, hair, hull or milk, are irreversible lost.
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2.3.2 Selenium metabolism in animal tissues
In the post-absorption metabolism of inorganic species, the selenide is uptaken by erythrocytes, immediately reduced to seleneto (HSe-), coupled to albumin and transported by plasma until the liver. The selenate, in this turn, is incorporated directly by hepatocytes, utilizing the same transport system for phosphates of the blood current (GIERUS, 2007). The selenide is also produced directly by ȕ-liase activity, while SeMet is converted in SeCys through a way similar to the transsulphuration, that occurs between methionine and cysteine (GIERUS, 2007), that is called transselenation (COMBS, 2001). It has been demonstrated that animals, on the contrary to plants, are not capable to synthesize SeMet (TAPIERO, TOWNSEND and TEW, 2003). After intestinal absorption, organic compounds containing selenium in foods, as seleno amino acids SeMet and SeCys, are brought by portal vein to the liver. In this organ both, SeMet and SeCys, may be metabolized by animals as amino acids, which may be degraded or shunted to general circulation, entering to amino acids pool. This metabolic fate occurs mainly to SeMet, which, similarly to methionine, may be incorporated into proteins as those of skeletal muscle and to serve as a stock of the micronutrient in animal organism. Consequently, about 40 – 50% of corporal selenium may be in the form of SeMet in muscular tissue, which is called non-functional selenoprotein (GIERUS, 2007). However, the SeMet may no substitute SeCys in the expression of specifics seleno enzymes that contains this amino acid (GSH-Px, for example). Then, in the first it needs to be uptaken from the amino acid pool and converted, by transselenation, to SeCys. At the same form, for doing that SeMet could serve as a selenium source to supply a reduction of plasmatic concentrations of the microelement due its deficiency in diet, as consequence of a deviation to fetus or because an increment of colostrum or milk secretion, it will depend on the tissue protein 97
turnover. Only after proteolysis is that SeMet will enter to amino acids pool, and then will be possible to generate SeCys or to be degraded to hydrogen selenide (H2Se), that is the primary source of selenium for different biochemical pathways in animal organism. There is a homeostatic control in the metabolism of majority of microminerals in animals, according the organism regulates situations of deficiency, by mobilizing reserves, and of excess, through excretion of the overplus or by reducing intestinal absorption (KIRCHGESSNER, 1993; SCHWARZ, KIRCHGESSNER and STANGL, 2000). However, differently that occurs with other microelements, it seems there is no regulation on absorption of different selenium forms (VENDELAND et al., 1994). The excretion of selenium excess, in this turn, occurs by conversion of the microelement into methylated species, that are lesser toxic than selenite, selenate and seleno amino acids, as methyl-selenol, DMSe, trimethylselenonium ion (trimethyl selenol) or seleno sugars (FRANCESCONI and PANNIER, 2004). Briefly, in the metabolism of selenium in animal cells, especially in hepatocytes, oxidized forms of selenium (selenate – SeO4-2; and selenide – SeO3-2) suffer reductive metabolism to produce H2Se. This species is the central selenium compound in animal tissues, which may follow by anabolic or catabolic pathways, according momentary necessities. In anabolic pathway, the H2Se may be utilized in formation of seleno amino acid SeCys, which is analogue of sulphured amino acid cysteine. Afterwards, this seleno amino acid may be incorporated into selenoenzymes. In the SeCys synthesis process, its formation occurs through incorporation of selenium atom, in the form of selenophosphate, in place of oxygen into O-phosphoseryl-tRNA ([Ser]), generating the group selenocysteyl-tRNA ([Ser]Sec). It is no known because the synthesis of SeCys requires a hydroxyl group of serine instead of the thiol group of cysteine. It is hypothesized that this characteristic could reduce the probability of mistakes in the process of selenoprotein synthesis by avoiding that cells “confuse” SeCys with cysteine (NOGUEIRA and ROCHA, 2011).
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The incorporation of SeCys in selenoenzymes occurs by modifying cotranslational residues of serine linked to tRNA in determined loci codified by UGA codon. This codon possesses the sequence to insertion of SeCys, in region 3’non-translated of mRNA, and utilizes the anti-codon ACA (BERRY et al., 1993; ALLAN, LACOURCIERE and STADTMAN, 1999; STADTMAN, 1996). The codon UGA generally signalizes to finish protein synthesis. To avoid that after reading it the synthesis will be interrupted prematurely, cells need to utilize some signaling factors. Thus, the incorporation of SeCys depends on assistance of a specific elongation factor (EFSec), of the SeCys insertion factor (SECIS) and of the SECIS binding protein 2 (SBP2). In the absence of these factors, the read of UGA codon determine the finalization of synthesis by entrying the ribosome release factor 2 (RF2). The allosteric competition or relative proportion between elongation factor and RF2 will define between the synthesis of selenoenzyme or interruption of the process (DRISCOLL and COPELAND, 2003). Alternatively, in the catabolical pathway, the SeMet may be metabolized to generate SeCys. Then, it may be degraded to composit the H2Se pool by action of a ȕ-liase. Another liase is necessary to convert MeSeCys, from some foods (as garlic), to methyl-selenol (CH3SeH). Successive methylation of H2Se detoxified the excess of Se, producing methyl-selenol (CH3SeH), dimethyl-selenide [(CH3)2Se] and trimethylselenonium ion [(CH3)3Se+]. Metabolites dimethyl-selenol and trimethylselenol are excreted by breath and urine, respectively. In the light overload of selenium, the preferential via is urinary excretion (trimethyl-selenol). However, in the case of higher selenium overload, the respiratory elimination will be a complementary via to excrete the selenium excess (as methylselenol and dimethyl-selenol) (COMBS, 2001; GIERUS, 2007). The oxidation of the excess of H2Se as selenium dioxide (SeO2) results in generation of superoxide anion and other ROS (COMBS, 2001), which may provoke oxidative damage in hepatocytes. In resume, selenium incorporation in tissue proteins, as those of skeletal muscle, requires the microelement in the form of SeMet, that will be directly 99
incorporated in substitution to methionine. The SeMet may be no incorporated directly in proteins without to be converted in the first to SeCys. Thus, SeMet needs to be converted to SeCys through a metabolic pathway called transselenation. This via is similar to transsulphuration, that occurs between methionine and cysteine. By the other hand, SeCys is not preferentially incorporated into tissue proteins, but deviated to synthesis of specific selenoenzymes. Inorganic selenium species are more quickly converted to selenide than organic ones. SeMet and SeCys may, even, to be oxidized to selenite and selenate (GIERUS, 2007). In addition its function as antioxidants, there are several organo selenocompounds (synthetic) that are capable to play many other functions in animal organism as anti-viral, anti-bacterial, anti-fungus, immunemodulator, anti-sepsis, anti-carcinogenic, anti-inflammatory and antiangiogenic effects. Inclusive, it should be commented that the selenium metabolism and some of its biologic effects described to animals, as well as some toxic effects, are shared by human body. However, in Veterinary the major selenium compound utilized in rations and mineral supplements is the sodium selenite (Na2SeO3). Although lesser dynamic in its effects than organic compounds, there are a series of benefites associated to Na2SeO3 consumption by animals, especially in relation to udder health, animal reproduction, prevention of nutritional myopathy (white muscle disease), improvement of the quality of meat products and prevention against cellular damage by oxidative damage. But, as all microelement, the super-dosage or prolonged exposition to supra-nutritional levels may deflagrate signals of intoxication in animals and humans. According described above, selenium metabolism in liver, at the same time that allow the micronutrient incorporation by animal cells due its conversion into chemic species that are easly assimilated by them, generates ROS. Then, as it will be discussed later, ROS produced by metabolizing the microelement overload participates of the selenium intoxication pathogenesis.
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2.3.3 Selenium supplementation for diary cattle
Mineral concentrations in roughates are affected directly by its level in soils. Generally, grasses of heat station containing lower mineral concentration than winter grass and leguminous. Bovines require minerals to maintain their health and to allow the perfect operation of vital functions. Mineral deficiencies, although light, may cause reduction of feed conversion rate, reduction in milk production, low fertility and lesser resistance to diseases, resulting in economic loss to the farmer and depreciation of final animal products (Figure 2.6). Mastitis is a great problem for milk production because impairments that causes. Elevation of expenditure with mastitis is not only related to treatment itself, but due economic loss caused by reduction in milk production, low quality of milk, necessity of specific strategies of management and milk disposal. It was estimated that economic expenditure due mastitis would overtake US$126,00/cow/lactation. Of these, about 60% would be loss due production fall provoked by subclinical mastitis, 15% resulting from clinical mastitis losses, 12% caused by non-functional quarters, 6% due extra labor, 2% resulting from animal reposition and 2% due veterinary services (HOLANDA et al., 2005). In a study involving 4,710 Holstein and Girolando cows objecting to identify and quantify causes for animals disposal, it was concluded that approximately 17% of discards were due alterations in mammary gland (SILVA et al., 2004). Hence, strategies for reducing mastitis prevalence in cow lots will have an important positive effect on profitability of farms.
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Figure 2.6 Selenium supplementation for diary cattle herds. Supplementation with the micronutrient may reduce mastitis prevalence, diminishs tank SCC, ameliorates reproductive performance of herds and improves milk and derivatives quality. Selenium-enriched foods, as milk and derivatives, are differentiated products that may achieve higher aggregated value (photography merely illustrative).
In this sense, after publishing the work of Maus et al. (1980), it there was an greater interest in relation to the importance of selenium for dairy cattle. In that opportunity, it was demonstrated that ingestion of selenium at very low dose, as 6 mg/cow/day (as Na2SeO3) would result in plasmatic concentrations of 112 ȝg Se/L already at 7 initial days after the onset of supplementation. At the same period, the secretion of selenium in milk would reach 56 ȝg/L. Thus, results showed by Maus et al. (1980) prompted to a search line on possible benefites of selenium supplementation to cows and subsequent positive effects to mammary gland health. Before, Arthur and Boyne (1979) already had demonstrated that neutrophils isolated from selenium-deficient bulls showed lesser capacity to kill cells of Candida albicans phagocyted, as well as were lesser able to reduce tetrazolium blue than neutrophils from bulls with normal selenium plasmatic levels. 102
Afterwards, Harrison et al. (1984c) demonstrated that diary cattle supplemented with selenium (0.1 mg/kg of body weight, i.m) at 21 days before calving showed reduction of 12% in prevalence of clinical mastitis, 22% less cases of metritis and reduction of 62% in the prevalence of ovarian cysts. When selenium was supplemented to diary cattle diet associated with 0.74 g of vitamin E/day, the reduction of metritis prevalence achieved 31% and the diminution in prevalence of placenta retention reached 100% (none case) (Table 2.1).
Table 2.1 Selenium supplementation levels and/or vitamin E and their effects on mastitis prevalence and reproductive problems of diary cattle (as % of reduction). Seleniuma Vitamin Eb Clinical mastitis Placenta retention 0.1 mg ---12 w/e ---1g/day 37 w/e 0.1 mg 1g/day s/e 100 Adapted from HARRISON et al. (1984c). w/e: without effect. a Intramuscular injection (0.1 mg/kg of body weight). b Oral supplementation (1g/day).
Metritis Ovarian cysts 22 62 w/e w/e 31 62
In a similar study, Smith et al. (1984) reported that selenium reduced by 46% the duration and symptoms of clinical mastitis, being that this reduction achieved 62% when were associated 0.1 mg of Se/kg of body weight (i.m, at 21 days before calving) + 0.74 g of vitamin E/cow/day. By reinforcing these results, Harrison et al. (1986) demonstrated that this treatment accelerated the uterus involution of fresh-calved cow suffering metritis. Selenium participation in prevention/cure of mastitis was better understood after results presented by Erskine et al. (1987). They carried out a search with two groups of diary cattle, one with low somatic cells count (SCC) in milk (
