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THE SELENIUM CONTENT OF MEATS, SEAFOODS, AND VEGETABLES by XIAOYUN ZHANG, B.Med., M.Phm. A THESIS IN FOOD AND NUTRITION Submitted to the Graduate Faculty of Texas Tech University in Partial Fulfillment of the Requirements for the Degree of MASTER OF SCIENCE Approved Accepted Nay, 1993 r- go5 ACKNOWLEDGEMENTS /3 fJcl' 4-93 ~ ~- J.+c- ,orq3 f/~ .03 I am deeply indebted to my advisor, Dr. Julian E. Spallholz, for his patient CJr.n, ;)- ~0' and helpful guidance during the research and preparation of this thesis. I thank the other members of my committee, Dr. Lee M. Boylan and Dr. Leslie D. Thompson for their helpful criticism. I would also like to thank the many faculty and staff members at the Department of Education Nutrition Restaurant Hotel Management who have helped me in these years. Finally, I would like to thank my husband and my family for their support during this time . .. 11 1 l ) j 1jf1>r TABLE OF CONTENTS ACKNOWLEDGEMENTS 11 LIST OF TABLES IV LIST OF FIGURES v CHAPTER I. INTRODUCTION 1 II. LITERATURE REVIEW 2.1 Biochemical functions of Se 2.2 Deficiency diseases of Se 2.3 Forms of Se 2.4 Bioavailability of Se in foods 2.5 RDA of Se 2.6 Distribution of Se 2. 7 Factors that affect the Se content of foods 3 3 5 I 8 9 9 10 III. METHODS AND MATERIALS 3.1 Selection of food samples 3.2 Preparation of food samples 3.3 Analytical methods 18 18 18 19 IV. RESULTS 4.1 Food Se concentration 4.2 Comparison of food Se content with other areas 4.3 Effect of drying methods on Se content 4.4 Effect of cooking methods on Se content 4.5 Contribution of 3 oz. of cooked meats to RDA for Se 22 22 22 23 24 24 DISCUSSION 29 V. VI. CONCLUSIONS 33 REFERENCES 35 Ill LIST OF TABLES 2.1 Bioavailability of selenium in foods 13 2.2 Summary of selenium content of foods in some areas 15 3.1 Contributions of selenium in the U.S.A. diet. 18 3.2 Procedure for selenium determination 19 4.1 Selenium content of seafood 21 4.2 Selenium content of ground beef and pork 21 4.3 Selenium content of poultry 22 4.4 Selenium content of vegetables 22 4.5 Selenium content of milk 22 4.6 Comparison of selenium content with other areas 24 4.7 Selenium content of raw, cooked and dried meats 25 4.8 Contribution of cooked meats to RDA for selenium 25 lV LIST OF FIGURES 2.1 Functions of glutathione peroxidase, superoxide dismutase and a-tocopherol in a cell. 12 2.2 Outline of selenium metabolism. 13 2.3 The regional distribution of forages and grain containing different levels of selenium in the U.S.A. 14 CHAPTER I INTRODUCTION Selenium (Se) is a very important trace nutrient in the human body. The Se-dependent enzymes, glutathione peroxidase (GSHPx) and phospholipidhydroperoxide glutathione peroxidase (PLGSHPx), are antioxidant enzymes which protect cells from oxidative damage. The major biochemical effect of selenium is to maintain GSHPx activity and PLGSHPx activity. A third Se-containing enzyme is type 1 iodothyronine 5'-deiodinase which catalyzes 1-thyroxin (T 4 ) to 3,3',5-triiodothyronine (T 3 ). A dietary selenium deficiency can cause heart disease which was first recognized in some areas of the People's Republic of China where the selenium content of diets is extremely low. After much research, the Food and Nutrition Board of the U.S. National Academy of Sciences set its 1989 Recommended Dietary Allowance (RDA) for selenium at 70 and 55 J.1g/day for adult men and women, respectively. Dietary selenium in foods varies from different countries and also from different areas of the same country mainly due to the concentration and biological availability of selenium in the soil. The interest in dietary selenium intake has increased because of selenium's important role in health. So the need for the assessment and improvement of selenium food composition data from different areas has also increased. A few studies of the selenium conteut in foods 1 2 from Maryland and Ohio have been reported but no information is available on the selenium content of foods in Lubbock, Texas. In this study, data is provided on the selenium content of some commonly consumed foods. CHAPTER II LITERATCRE REVIE\h/ Selenium (Se ), which is a group VI element under sulfur in the periodic table, was first discovered by Berzelius in 1817. Since then, selenium has become known as one of the most toxic elements ( cf. Shamberger [33]). In 1957, Schwarz and Foltz first reported that dietary selenium was important in metabolism because it could prevent liver necrosis in vitamin £-deficient rats (Schwarz and Foltz, [31]). Since then, some other animal diseases have been found to respond to selenium [24]. These diseases are white muscle disease in cattle, swine, and sheep; liver necrosis in swine and rats; and growth depression in cattle, rats. and sheep. Now, more than 40 species of animals are known to have a biological requirement for selenium. 2.1 Biochemical functions of Se There are three Se-dependent enzymes, glutathione peroxidase (GSHPx), phospholipidhydroperoxide glutathione peroxidase (PLGSHPx) and type 1 iodothyronine 5'-deiodinase. In 1973, John Rotruck, a graduate student at the University of Wisconsin, discovered that the enzyme glutathione peroxidase (GSHPx) contained the element selenium [28]. Glutathione peroxidase is present in cells and in plasma and contains four atoms of selenium which is an integral part of the active site of glutathione peroxidase. Glutathione peroxidase can 3 4 reduce hydrogen peroxide (H2 0 2 ) to water and organic hydroperoxides (ROOH) to an alcohol (ROH), as shown in Equation (2.1). (2.1) Normal cellular reactions and the metabolism of toxic substances can produce hydrogen peroxide, hydroperoxides, superoxide, various free radicals including hydroxy radical and possibly singlet oxygen. These free radicals can initiate lipid peroxidation and cause membrane damage. Antioxidants in cells can prevent lipid peroxidation by breaking free-radical chain reactions or preventing the cellular accumulation of the free radicals. One of the important groups of antioxidants are the antioxidant enzymes synthesized by cells which includes glutathione peroxidase, catalase and superoxide dismutase (SOD). In a typical animal cell, glutathione peroxidase reacts with peroxides and superoxide dismutase reacts with superoxide in the cytosol and 0'-tocopherol scavenges free radicals in the membranes (Figure 2.1 ). Since glutathione peroxidase is a Se-dependent enzyme, selenium thus can serve as a biological antioxidant. The enzymic activity of glutathione peroxidase correlates well with the concentration of selenium in blood. Dietary Se-deficiency causes glutathione peroxidase activity to decline in tissues. According to Hafeman et al. [10], after four weeks of a Se-deficient diet, rat liver glutathione peroxidase activity falls to undetectable levels. This research indicates that one 5 of major functions of selenium is to maintain GSHPx activity that protects cells from oxidative damage. Another antioxidant Se-containing enzyme, phospholipidhydroperoxide glutathione peroxidase (PLGSHPx) discovered by Fulvio Ursini at the University of Padua in 1985 [45], protects membranes. PLGSHPx is present in phospholipids and contains one atom of selenium. PLGSHPx is similar to glutathione peroxidase in that selenium is also an integral part of the active site of PLGSHPx. PLGSHPx can protect liposomes and biomembranes from peroxidative damage. In 1990, Dietrich Behne in Germany found another selenoenzyme. type 1 iodothyronine 5 '-deiodinase [4). This Se-containing enzyme catalyzes the deiodination of the prohormone 1-thyroxine (T 4 ) to the biologically active thyroid hormone 3,3',5-triiodothyronine (T 3 ). Research has shown that in Se-deficient rats, the T 3 production is decreased in the lin'r. 2.2 Deficiency diseases of Se The first Se-deficiency disease in humans, named Keshan disease, was discovered in Keshan area of the People's Republic of China in 1979, an area where the selenium content in the soil is very low [48). Keshan disease affects the heart muscle. Children less than 15 years of age are most susceptible. The Keshan Disease Research Group of the Chinese Academy of Medical Sciences has treated 36,603 children with 1.0 mg of sodium selenite per week. Only 21 6 cases occurred during the four years of investigation. However, there were 107 cases in 9,642 children who were not treated with sodium selenite (14]. The mechanism by which Keshan disease develops is not totally clear. According to Yang et al. [48], Se-dependent GSHPx has a stabilizing effect on erythrocyte membranes and selenium may protect and stabilize membranes as PLGSHPx and lessen their susceptibility to pathogenic factors in the em·ironment. For example, in 1970 Shamberger found significantly lower death rate of heart disease in the high selenium areas. Another disease linked to Se-deficiency in China is Kashin-Beck disease [48]. It is an endemic osteoarthritis that occurs in children or young people. Oral selenium supplementation seemed to prevent the disease [20]. A number of studies suggest that poor selenium status may be associated with an increased risk of developing cancer [18]. There are about fifty dietary experiments showing selenium to have anticancer actiYity in animals [34]. Some research has observed a reduction in skin tumors when mice were treated with 7, 12- dimethylbenzanthracene (D~IBA) [27,32,35]. Other research has show11 that certain amounts of selenium added to the diet or drinking water reduced the incidence of liver tumors, colon tumors and breast tumors in animals [34]. According to many experiments using animal models, it could be concluded that selenium may act as an anticarcinogenic agent when giYen either in the diet or in drinking water, but this protective action is not universal. A proper 7 amount of selenium (1-5 ppm Se) given in food or drinking water may or may not protect the organism against the effects of some carcinogens [14]. We already know that selenium is an essential part of the enzyme glutathione peroxidase which is an antioxidant and can keep the organism free of peroxide compounds. The function of selenium is to maintain glutathione peroxidase activity. Since free radicals, such as 02 or OH·, may initiate the process of carcinogenesis, selenium can increase glutathione peroxidase and reduce the concentrations of free radicals and inhibit the initiation of the carcinogenic process. Cancer patients are known to have lower serum activity of glutathione peroxidase than healthy persons [14]. 2.3 Forms of Se Selenium occurs naturally in foods in organic compounds which are selenomethionine, Se-methyl-selenomethionine, selenocystine, and selenocysteine [25,29]. Tlwse dietary forms of selenium are well absorbed by monogastrics. Absorbed selenium is transported through the body primarily bound with plasma proteins. There are two major forms of selenium in animal tissues, selenocysteine and selenomethionine. Selenocysteine is derived from animals [11] and selenomethionine from plants which cannot be synthesized in the body [25]. Selenomethionine is absorbed and transported via the same mechanisms as methionine [4 7], but the method of selenocysteine absorption is unclear. \rhen 8 dietary selenomethionine is absorbed by the body, it combines with a variety of proteins in tissues, but after dietary selenocysteine is absorbed, it can directly follow selenium metabolism in the body or combine with glutathione peroxidase (Figure 2.2). As dietary selenium intake decreases, the tissue form of selenomethionine is returned to the methionine pool and then transferred to the selenocysteine form. After about one year of a low dietary selenium intake, selenium levels in blood will be very low. Therefore, dietary selenium is a very important and only source of selenium in the body. 2.4 Bioavailability of Se in foods The determination of bioavailability of selenium in foods is as important as the selenium content determination because it is not certain whether all the different compounds that occur in foods can be metabolized to biologically active selenium, even if they are well absorbed. Since the major function of selenium is to maintain the activity of glutathione peroxidase, the best way to determine the bioavailability of selenium is to measure glutathione peroxidase activity in selenium-depleted animals. The bioavailability of selenium in some foods has been estimated in the rat model. In rats the increase in liver glutathione peroxidase activity is measured after feeding the foods to rats previously depleted of selenium {Table 2.1 ). According to same studies. the 9 availability of selenium in mushrooms was very low. Wheat, Brazil nuts, and beef kidney contained the most highly available selenium. 2.5 RDA of Se In 1980, because few data about selenium in human nutrition were available, the Food and Nutrition Board of the U.S. used the results of animal experiments and proposed a safe and adequate daily dietary intake of selenium of 50 - 200 J.Lg for adults [24]. Since more research and evidence showed the important roles of selenium in health, the Food and Nutrition Board in 1989 set a Recommended Dietary Allowance (RDA) for selenium of 70 and 55 J.Lg/ day for adult men and women, respectively. For pregnant and lactating women the RDA for selenium is 65 and 75 J.Lg/ day, respectively. For infants age 0 - 0.5 years, the RDA is 10 J.Lg/day and to age one it is 15 J.Lg/day. 2.6 Distribution of Se Selenium is widely distributed in the earth and can be found in soil, rocks, water, air, plants, and animal and human tissues [12,26,43,46]. According to Combs [6] the selenium content in soils varies in different areas and the amount of selenium in food plants reflects the soil selenium. Jenkins [15] indicated that the regional distribution of forages and grain containing different levels of selenium in the United States and Canada (Figure 2.3). In adequate selenium regions, about 80% of all forage and grain contained > 0.10 ppm selenium. In 10 intermediate regions, about 50% of all forage and grain contained > 0.10 ppm selenium. Regions considered low selenium have about 80% of all forage and grain containing < 0.10 ppm selenium. 2. 7 Factors that affect the Se content of foods A number of factors influence the selenium concentration in foods. 1. different geographic locations or the natural selenium content of soils [44]. 2. the soil concentration of biologically available plant selenium [44]. 3. the treatment of soil by topdressing [22]. 4. supplementation of animal feed [22]. 5. the processing of food products [9) 6. cooking methods [13]. 7. the protein content of the food [16). Of all these factors, the most important factor is the geographic region of production. Plants have no known selenium requirement. If no selenium is available for uptake, the plant still thrives. However, animals have a physiological requirement for selenium. If the selenium concentration in feedstuffs is very low, the selenium level in these animal tissues are also very low. Certain selenium compounds are easily destroyed, so the cooking methods may affect the selenium content in foods. According to Higgs et al. [13), broiling 11 chops and steaks caused little change in the selenium content of the meat and baking had little effect on the selenium level of chicken or fish. Many studies of the selenium content of foods have been published. Table 2.2 is data summarized from a number of studies in the United States (21,37,30], in England (42], in France (36], in New Zealand (41], and in Canada (7). As we have already indicated, current research suggests that selenium is a very important element in the human body and dietary selenium is a major source. Since the selenium content of foods \'aries according to geographic location and since interest in dietary selenium intake has increased, due to the important role of selenium in health, there is an increased need for the assessment and improvement of the selenium food composition data acquisition from a wide variety of areas. There are some reports of the selenium content in foods from the Maryland and Ohio, and also a report of selenium content in canned foods or food products from Houston, Texas (16). Ko information is available on the selenium content of foods in the Lubbock, vVest Texas area. In fact, this provided part. of the motivation for the present study which provides data on the selenium content of some commonly consumed foods in Lubbock. Texas, and examines whether the selenium content is influenced during the cooking or drying processes. 12 KEMBRANES CYTOSOL soluble enzymes drug metabolism mitochondrial membrane and E. R enzymes Se-GSHPx H2 o2 ~ 2GSH 2H 20 esse USFA-~ (O) 2 (FA . ) (HO ·) HO. ~ R-H USFA - OOH Se-eSHPx ROOH -~--=--o::::---~> ROH 2GSH Figure 2.1: Functions of glutathione peroxidase, superoxide dismutase and a-tocopherol in a cell. ( ) = species scavenged or quenched by a-tocopherol (modified from Sunde and Hoekstra, [19]). 13 TISSUE FORMS DIETARY FORMS (follow methionine metabolism pathways) SELENOMETHIONINE ~ METHIONINE :;:::: COMBINE (derived from plants) POOL WITH PROTEINS ! SELENOCYSTEINE (derived from animals) :;:::: SELENIUM METABOLISM COMBINE \VITH GSHPx ! EXCRETION IN URINE, BREATH Figure 2.2: Outline of selenium metabolism (modified from Levander and Burk. [19]) Table 2.1: Bioavailability of selenium in foodsa. Foods Whole wheat bread 6 Brazil nutsc Beef kidney, cookedd Baltic herring, bakede Wheat, boiled d Shrimp, boilede Crab, boilede Tuna, canned, water-packedd Oysters, boilede Tuna, canned, oil-packedb Mushroom, freeze-driedc a. Levander, [18] b. Alexander et al., [1] c. Chansler et al., [5] d. Douglass et al., [8] e. Mutanen et al., [23] Availability (%) (selenite = 100) 142 117 97 90 83 73 58 57 38 32 4 14 ~ L:.:.=; LOW·A"AOXIMAHLY 10' OF ALL 'ORACl ANO CRAIN CONTAIN (O 10 "'M SH(,.IUM c:J D 1/ARIAIL£-A''ROXIMAT(L Y SO'.; CONTAIN )o 10 ""' S(L(NIUM II,.ClVD£5 AlASII.AI AOIOUATl-10 .. OF All '0RAGlS AND CRAIN CONTAIN )c IC ; , ... SHE,.IVM IIHCLVDI S HAWAIII Figure 2.3: The regional distribution of forages and grain containing different levels of selenium in the U.S.A. (modified from Jenkins, [15]) 15 Table 2.2: A summary of the selenium content of foods from a number of studies in the United States and other countries. Foods Beef & Pork beef,raw beef,ckd pork,raw pork,ckd Seafoods tuna fish cod,fillet shrimp,raw shrimp,ckd salmon Poultry chicken,raw chicken,ckd turkey,raw turkey,ckd Vegetables lettuce,raw potatoes,raw tomatoes,raw Dairy prod. milk, whole milk,2% fat milk,skim ckd=cooked MD (1970) Can. (1972) Se concentration (Jlg/ g) Eng. OH MD (1978) (1987) (1987) 0.195 0.127 0.030 0.239 0.296 0.140 0.428 0.588 0.116 0.210 0.068 0.068 0.131 0.720 0.531 0.530 0.640 0.600 0.160 0.200 0.180 0.34 0.250 0.008 0.023 <0.010 0.012 0.150 <0.010 0.060 0.080 0.100 0.010 0.005 0.005 0.048 I 0.863 0.220 0.260 0.330 0.350 Fr. (1988) 0.531 0.178 N. Z. (1990) 0.019 0.023 0.120 0.170 0.120 0.200 0.230 0.340 0.200 0.280 0.050 0.160 0.120 0.479 0.015 0.013 0.007 0.015 <0.001 0.006 <0.001 0.003 0.004 0.002 0.020 0.030 0.030 0.012 0.008 0.014 0.013 0.006 0.006 0.008 CHAPTER III METHODS AND MATERIALS 3.1 Selection of food samples Following the guidelines in Schubert et al. [30], a list of dietary selenium contributors from foods was developed (see Table 3.1). The food selection process was intended to include those frequently consumed foods that contribute to the bulk of selenium in the diets of the United States population. These foods are "core foods 1' for selenium which contribute about 41.39( of the selenium in the U.S. diet. From these "core foods," selections were made from commonly consumed meats, seafood, poultry, dairy products, and vegetables. All these food samples were purchased from a commercial packer or local supermarket; some of the vegetables were also purchased from a road-side market. 3.2 Preparation of food samples All meats, seafoods, and poultry were prepared as raw, cooked, freeze-dried or air-dried meats. The vegetables were analyzed on a fresh. dried. or cooked basis. We used ground beef, pork, chicken and turkey as samples. Fish was deboned and shrimp was shelled and deheaded. Samples were homogenized in a glass food blender. Meat, poultry and seafood samples were cooked in Teflon pans at a med-high temperature until well-done. They also were dried in a drying oven at 100°C for 24 hours or freeze-dried (Labconco freeze dryer). The 16 11 vegetables were peeled, outer leaves removed and dirt rinsed off using deionized distilled water. They were dried in an oven at 100°C for 24 hours or boiled and steamed for 10 minutes and then homogenized in a glass food blender. 3.3 Analytical methods The selenium content was determined using the fluorometric technique of Spallholz et al. [38]. This method involves: ( 1) preparation of samples and standards, (2) digesting 0.10-0.50 g of samples in a molybdate sulfuric perchloric acid mixture, (3) masking potential mineral interference with 0.008 M disodium ethylenediamine tetraacetate (EDTA). EDTA can chelate Ca, Zn, Fe, Mn and Mg, (4) adjusting the pH to 2 using either NH 4 0H or H 2 S04, (5) complexing selenium with freshly prepared 3, 3'-diaminonaphthalene (DAN) solution, (6) adding cyclohexane, (7) shaking each tube for 5 minutes and extracting the Se-DAN complex into cyclohexane, (8) samples were then analyzed in a spectrophotofluorometer with the excitation at 363 nm and the emission at 525 nm. 18 (9) a standard curve was run with each set of analyses. (10) the selenium content of the foods and standard solution were compared with a National Bureau of Standards (NBS) liver standard 1577a containing 0. 71 ± 0.07 ppm selenium. Table 3.1: Contributions of seleniwn in the U.S.A. dieta. Foods Mean(J.LgSe/g) % daily intake Beef, cooked 17 0.26 Pork, cooked 0.35 9 Chicken, cooked 0.21 6 0.016 Whole milk 3 Tuna, canned 2 0.72 1.8 2% milk 0.026 Turkey, cooked 0.25 1 0.6 Shrimp, cooked 0.64 0.5 Skim milk 0.027 0.2 White potato 0.009 0.1 Lettuce, raw 0.015 0.1 Tomato, raw 0.007 a. Adapted from Schubert et al., [30) 19 Table 3.2: Procedure for selenium determination a. Step 1 2 3 4 5 6 7 8 9 10 11 12 13 Operation Prepare samples or standards Add digestion mixture (2 ml) Digestion Dilute digest with 0.008 M EDTA Add 2.1- 2.2 NH 4 0H (concentrated) Adjust pH to 2 (NH 4 0H or H 2 S0 4 ) Prepare DAN solutionb Add DAN solution ( 4 ml) Incubate at 50°C in water bath Cool tubes Add cyclohexane ( 5 ml) Extract by shaking Measure fluorescence intensity: excitation 363 nm; emission 525 run 14 Plot relative fluorescence against ng Se/5 ml cydohexane a. Spallholz et al., [38] b. Prepare fresh; wash well with cyclohexane Total Volume (ml) 1 3 7 9.2 9.2 13.2 18.2 Approx. Time (Min) 15 5 45 5 5 20 45 5 20 5 5 5 15 CHAPTER IV RESULTS 4.1 Food Se concentration The concentration of selenium of different foods is presented in the Tables 4.1 to 4.5. Foods with the highest average selenium concentrations (0.64 j.lg Se/g wet weight) were the seafood group: tuna fish (1.09 J.lg/g), shrimp (0.58 J.lg/g) and cod fillet (0.26 J.lg/g); followed by the beef and pork group: beef with 20% fat (0.14 J.lg/g), beef with 15% fat (0.19 J.lg/g) and pork (0.26 J.lg/g). In poultry, turkey (0.30 J.lg/g) had a higher selenium content than chicken (0.13 J.lg/g) which was even higher than cod, pork and beef. Animal fat contained very low selenium content. In the vegetable group, most vegetables were found to contain less than 0.03 j.lg Se/g except mushrooms which contained 0.16 J.lg/g. According to Allaway [2], a poisonous mushroom, Amanita, took up significant amounts of selenium even when grown in soils containing low levels of selenium. Milk was also low in selenium content. Skim milk (0.043 J.lg/ g) was higher than 2% low fat milk (0.038 J.lg/ g) which was higher than whole homogenized milk (0.029 J.lg/ g). The order of selenium content of all these foods was: tuna fish > shrimp > turkey > cod fillet = pork > beef> mushrooms > chicken > milk > vegetables. 20 21 Table 4.1: Selenium content of seafood (J-Lg/g mean± SD). Samples Shrimp (no shell) Raw Cooked Air-dried Cod Filet Raw Cooked Air-dried Tuna Filet Raw Cooked Freeze-dried Number of Samples Selenium 5 5 5 0.585±0.021 0.735±0.007 2.810±0.056 5 5 5 0.265±0.035 0.350±0.056 1.435±0.021 6 5 5 1.090±0.071 1.420±0.212 3.330±0.665 Table 4.2: Selenium content of ground beef and pork (J-Lg/ g mean ± SD ). Samples Ground Beef (15% fat) Raw Cooked Air-dried Ground Beef (20% fat) Raw Cooked Freeze-dried Air-dried Fat Ground Pork (extra lean) Raw Cooked Air-dried Number of Samples Selenium 5 6 5 0.190±0.014 0.250±0.056 0.500±0.085 6 5 5 5 5 0.145±0.007 0.240±0.071 0.335±0.007 0.420±0.085 0.030±0.000 5 6 5 0.265±0.035 0.370±0.056 0. 735±0.120 22 Table 4.3: Selenium content of poultry (p,g/ g mean ± SD ). Samples Ground Chicken Raw Cooked Freeze-dried Air-dried Fat Ground Turkey Raw Cooked Air-dried Number of Samples Selenium 5 6 6 5 5 0.130±0.028 0.275±0.049 0.385±0.021 0.365±0.007 0.015±0.007 5 5 5 0.300±0.056 0.445±0.021 1.169±0.269 Table 4.4: Selenium content of vegetables (p,g/ g mean ± SD ), n = 5. Sample Broccoli Mushroom Lettuce Onion Tomato Potato Fresh 0.032±0.003 0.165±0.035 0.014±0.002 0.012±0.001 0.002±0.001 0.019±0.001 Dry 0.180±0.020 2.050±0.230 0.210±0.020 0.054±0.008 0.022±0.007 0.067±0.006 Boil 0.205±0.007 1.335±0.130 Steam 0.255±0.007 Table 4.5: Selenium content of milk (p,g/ g mean±SD ), n = 5. Samples Skim milk 2% milk Whole milk Selenium 0.0430±0.001 0.0385±0.001 0.0295±0.001 23 4.2 Comparison of food Se content with other areas Some areas of England, France, New Zealand and Canada have been known as low selenium areas with low levels of selenium in soils and continuing up the food chain resulting in low selenium status of its residents. The data listed in Table 4.6 were reported to be from low selenium areas in these countries. Maryland and Ohio are also considered as low selenium areas. In comparison with England, France, New Zealand and Canada, the selenium content of beef, pork and chicken in Lubbock was higher than that of meats in those countries. But the selenium content in cod fillet from New Zealand and Canada was higher. The selenium content in beef, pork and poultry from Lubbock was lower than those from Maryland area, but the beef and pork contained higher content of selenium than that from the Ohio area. Shrimp from these three areas contained a higher and similar amounts of selenium. Vegetables were generally low for all countries or states and did not differ greatly from the Lubbock values. The selenium concentration in milk from Lubbock was similar to that from the Maryland area which was higher than those from Ohio and other countries except Canada. The whole milk in Canada contained 0.15 j.lg Se/ g which was much higher than the other areas. 24 Table 4.6: Comparison of the selenium concentration in food samples in Lubbock, Texas with those from other areas. Foods Beef Pork Chicken Turkey Shrimp Cod Tuna Lettuce Potato Tomato Whole milk 2% milk Skim milk a. b. c. d. e. f. g. Lbka 0.190 0.265 0.130 0.300 0.585 0.265 1.090 0.014 0.019 0.002 0.029 0.038 0.043 Se concentration (J-Lg/ g, wet weight bases) MDb OHC Fr.e Eng.d N.Z? 0.220 0.068 0.030 0.060 0.019 0.330 0.131 0.140 0.080 0.120 0.200 0.178 0.050 0.120 0.340 0.530 0.531 0.230 0.100 0.200 0.340 0.720 0.531 0.120 0.015 0.015 <0.001 0.003 0.013 0.004 <0.010 0.006 0.007 <0.001 0.002 0.020 0.012 <0.010 0.013 0.006 0.030 0.008 0.030 0.014 0.006 0.008 Can. 9 0.127 0.296 0.160 0.863 0.008 0.023 0.150 This study, Lbk Lubbock Schubert et al., (30], MD Maryland Snook et al., (37), OH Ohio Thorn et al., [41], Eng. England Simon off, (36), Fr. France Thomason and Robinson, [41), N.Z. New Zealand Arthur, [3], Can. Cananda 4.3 Effect of drying methods on Se content According to our study (Table 4. 7), the selenium concentration in foods increased in the dried samples which may be due to loss of the moisture. Selenium was not lost during drying. There were no real differences between freeze-dried or air-dried samples of foods. 25 4.4 Effect of cooking methods on Se content According to the results in Table 4. 7, all cooked food samples contained higher selenium concentrations than did raw samples, but contained lower selenium concentrations than dried food samples. One of the reasons may be that part of the moisture and fat was lost during the cooking process. Table 4.7: Selenium content of raw, cooked and dried meats, poultry and seafood (J-Lg/ g, mean ± SD ), n=5. Foods Beef (20% fat) Beef (15% fat) Pork Chicken Turkey Shrimp Cod Fillet Tuna Fish Raw 0.145±0.007 0.190±0.014 0.265±0.035 0.130±0.028 0.300±0.056 0.585±0.021 0.265±0.035 1.090±0.071 Cooked 0.240±0.071 0.250±0.056 0.370±0.056 0.275±0.049 0.445±0.021 0. 735±0.007 0.350±0.056 1.420±0.212 Dried 0.420±0.085 0.500±0.085 0.735±0.120 0.365±0.007 1.160±0.269 2.810±0.056 1.435±0.021 3.330±0.665 Table 4.8: Contribution of 3 ounces of cooked meat, poultry and seafood to RDA for selenium (3 oz. = 85.05g). Food Beef (15% fat) Pork Chicken Turkey Shrimp Cod, fillet Tuna J-Lg/Se 16.2 22.5 11.1 25.5 49.8 22.5 92.7 % RDA Women 55 1-Lg/day 29.5 40.9 20.2 46.4 90.5 40.9 168.5 % RDA Men 70 J-lg/day 23.1 32.1 15.9 36.4 71.1 32.2 132.4 26 4.5 Contribution of 3 oz. of cooked meats to RDA for Se In Table 4.8, we estimated how three ounces of each meat, poultry and seafood would contribute to the percent of RDA of selenium for women and men in Lubbock, Texas. Seafoods, meats and poultry, along with cereal grains, are the major contributors to the RDA of selenium. Unfortunately, analytical quantitation alone does not insure an adequate bioavailable intake of selenium since the bioavailability of selenium in foods is thought to vary considerably. CHAPTER V DISCUSSION According to Schubert et al. [30], there is a set of criteria specifically for evaluating the quality of selenium data. There are five categories: number of samples, analytical methods, sample handling, sampling plan, and analytical quality control. Based on these criteria, at least five samples for each food were assayed. We used a modification of one official fluorimetric method (Spallholz et al., [38]) as the analytical method for selenium which has been used for long time and is satisfactory in precision, accuracy and recovery of selenium. Samples were selected from " core foods " except cereal grains for selenium (Schubert et al. [30]). All selected foods are commonly consumed and contributed to the RDA for selenium or significant sources of dietary selenium. Food samples were analyzt>d in duplicate or were stored in plastic containers at -4°C until analysis, typically within two weeks after procurement. The analyses were always performed in one identical manner, in the same laboratory and using the same equipment. From this study, the best sources of dietary selenium based on concentration (on a wet method basis) were: fish and seafoods, beef and pork, and poultry. These foods had moderate to high levels of selenium ( vegetables contained > 0.13 f-Lg/ g). Milk and < than 0.03 ILg/g selenium. Mushroom, one exception, contained 0.165 f-Lg/g selenium. _, •)~ 28 According to previous studies (Table 2.2) and this study, animal products, like muscle meats, usually contain higher concentrations of selenium than vegetables which have a very low selenium content. This is because selenium is an essential trace element for animal species but not for plants. If animal tissue selenium levels are too low, the animals will not survive. Therefore, they will try to increase the retention of selenium at a low selenium intake to maintain selenium concentrations in muscle and liver. Plant species as noted do not require selenium for growth. They still thrive even if no soil selenium is available for uptake. In contrast with the results stated in [6], our work suggests that plants may be very low in selenium no matter how high or low the soil selenium content. Many factors influence the levels of selenium in foods. These include the different geographic location, the soil concentration of biologically available selenium, the treatment of soil, the supplementation of animal feeds, the processing of food products, the cooking methods, and the protein content of the food where much selenium is formed. In all studies on selenium in foods, data for cooked foods is lacking. There is no separation of data on foods by cooking methods and no specific conclusions on changes in selenium concentration as a result of different cooking methods [30]. Table 4. 7 shows that selenium concentrations in cooked meats, poultry and seafoods were higher than their raw counterparts, which is probably due to lost moisture or part of the animal fat 29 during the cooking processes. This is very limited cooking method but it does suggest that most selenium is retention in cooked foods. Importation of foods from other geographical locations is another factor that influences the selenium content of foods in any area. Lubbock, Texas, is in an adequate-selenium area and Maryland is considered as low selenium region. Table 4.6 shows that some foods purchased from the local Lubbock supermarket, i.e., beef and pork, had a lower selenium content than those purchased from Maryland. Therefore, food importation may cause wide variability in the selenium content of some foods. Selenium present in food appears to be associated with protein, so the protein content of food affects the selenium concentration in the food. There is no doubt that animal foods usually contain higher selenium content. Much of the selenium in food appears to be selenocysteine in the animaL which is the active form of selenium in glutathione peroxidase [40]. The major plant form of dietary selenium is selenomethionine. A large proportion of the selenium in wheat and about 60% of the selenium in soy beans has been identified as selenomethionine [25,49], which may also be present in animals eating grains. As discussed in the literature review, it is not certain whether the different selenium compounds that occur in foods can be metabolized to biologically active selenium. Therefore, the determination of bioavailability of selenium in foods is also important. For example, seafoods usually contain higher concentrations of 30 selenium, but the bioavailability of selenium in tuna fish and certain other seafoods like crab and oysters is reported to be relatively low, perhaps because of heavy metals, particularly mercury, complexing with selenium [8]. CHAPTER VI CONCLUSIONS According to the literature and this experimental study, the following conclusions may be made. 1. The best sources of selenium are: fish and other seafoods, beef, pork, and poultry. The selenium level in these foods is typically higher than 0.13 J-Lg/ g. The selenium concentration in milk and vegetables is usually very low ( < 0.03 J-Lg/ g ) no matter how high or low the selenium level for that area. 2. Animal products like muscle meats usually contain a higher selenium content and vegetables contain a very low selenium content because animals require dietary selenium but vegetables do not require soil selenium. 3. 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