Effects of dietary supplementation with selenium enriched yeast or sodium selenite on selenium tissue distribution and meat quality in lambs

Animal Feed Science and Technology 149 (2009) 228–239

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Effects of dietary supplementation with selenium enriched yeast or sodium selenite on selenium tissue distribution and meat quality in lambs D.T. Juniper a,∗, R.H. Phipps a, E. Ramos-Morales a, G. Bertin b a

Animal Science Research Group, School of Agriculture, Policy and Development University of Reading, Earley Gate, Reading RG6 6AR, UK ALLTECH France, EU Regulatory Affairs Department, 14 Place Marie-Jeanne Bassot, Levallois-Perret 92300, France

b

a r t i c l e

i n f o

Article history: Received 4 October 2007 Received in revised form 4 June 2008 Accepted 26 June 2008 Keywords: Lamb Meat quality Selenium Selenocysteine Selenomethionine Tissues

a b s t r a c t The objective was to determine the concentration of total selenium (Se) and the proportion of total Se comprised as selenomethionine (SeMet) and selenocysteine (SeCys), as well as meat quality in terms of oxidative stability in post-mortem tissues of lambs offered diets with an increasing dose rate of selenized enriched yeast (SY), or sodium selenite (SS). Fifty lambs were offered, for a period of 112 d, a total mixed ration which had either been supplemented with SY (0, 0.11, 0.21 or 0.31 mg/kg DM to give total Se contents of 0.19, 0.3, 0.4 and 0.5 mg Se/kg DM for treatments T1, T2, T3 and T4, respectively) or SS (0.11 mg/kg DM to give 0.3 mg Se/kg DM total Se [T5]). At enrolment and at 28, 56, 84 and 112 d following enrolment, blood samples were taken for Se and Se species determination, as well as glutathione peroxidase (GSH-Px) activity. At the end of the study lambs were euthanased and samples of heart, liver, kidney, and skeletal muscle were retained for Se and Se species determination. Tissue GSH-Px activity and thiobarbituric acid reactive substances (TBARS) were determined in Longissimus Thoracis. The incorporation into the diet of ascending concentrations of Se as SY increased whole blood total Se and the proportion of total Se comprised as SeMet, and erythrocyte GSH-Px activity. Comparable doses of SS supplementation did not result in significant differences between these parameters. With the exception of kidney tissue, all other

Abbreviations: BW, body weight; DM, dry matter; GHS-Px, glutathione peroxidase; MAP, modified atmosphere packs; Se, selenium; SeCys, selenocysteine; SeMet, selenomethionine; SS, sodium selenite; SY, selenized yeast; TBARS, thiobarbituric acid reactive substances; TMR, total mixed ration. ∗ Corresponding author. Tel.: +44 118 378 6566; fax: +44 118 378 6595. E-mail address: [email protected] (D.T. Juniper). 0377-8401/$ – see front matter © 2008 Elsevier B.V. All rights reserved. doi:10.1016/j.anifeedsci.2008.06.009

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tissues showed a dose dependant response to increasing concentrations of dietary SY, such that total Se and SeMet increased. Selenium content of Psoas Major was higher in animals fed SY when compared to a similar dose of SS, indicating improvements in Se availability and retention. There were no significant treatment effects on meat quality assessments GHS-Px and TBARS, reflecting the lack of difference in the proportion of total Se that was comprised as SeCys. However, oxidative stability improved marginally with ascending tissue Se content, providing an indication of a linear dose response whereby TBARS improved with ascending SY inclusion. © 2008 Elsevier B.V. All rights reserved.

1. Introduction In the early 1970s a specific biological role for Selenium (Se) became apparent with the discovery of the first selenoprotein, glutathione peroxidase (GSH-Px) (Rotruck et al., 1973). Glutathione peroxidase catalyses the reduction of lipid and hydrogen peroxides to less harmful hydroxides via the oxidation and subsequent reduction of selenocysteine (SeCys) which is the active centre of this enzyme (Arteel and Sies, 2001). The antioxidant functions of Se, via GSH-Px activity, have been shown to persist post-mortem in poultry muscle tissue (DeVore et al., 1983), delaying the onset of oxidation reactions, which affects adversely both the nutritive value and flavor of meat products (Morrissey et al., 1998). Plant Se concentration can be extremely variable and dietary Se supplements are generally required for ruminant diets. Selenium supplements are in two forms, inorganic mineral salts, such as sodium selenite (SS; Na2 SeO3 ) or selenate (Na2 SeO4 ), or in organic forms such as Se enriched yeast (SY), in which selenomethionine (SeMet) is the predominant form of Se (Korhola et al., 1986). The distribution and accumulation of Se and Se species in animal tissues depends very much on the source of the Se supplement. Surai (2006) reported that SeMet is retained in tissue proteins to a greater extent than SeCys or Se derived from inorganic forms. Selenium absorption occurs within the small intestine and whilst SeMet is absorbed via the methionine transporter system, the absorption of inorganic Se, such as SS, is less efficient and occurs mainly by passive diffusion (Weiss, 2003). Following absorption SeMet can be incorporated nonspecifically into general body proteins in place of methionine and can act as a biological pool for Se (Suzuki and Ogra, 2002) which can be utilised during periods of suboptimal Se intake. Conversely inorganic sources that are taken up through the small intestine are either utilised or methylated and subsequently excreted. Furthermore, Seko et al. (1989) showed that SS may act as prooxidant, which has the potential to be toxic at high dietary levels, whereas SeMet does not possess these properties. Irrespective of source, Se must undergo a metabolic transformation to selenide prior to its assimilation into SeCys and subsequent incorporation into selenoproteins via the UGA codon (Suzuki and Ogra, 2002). However, no such intermediate step is necessary for the incorporation of SeMet into general proteins. Consequently the biological actions of Se depend on the amount and chemical form of Se consumed and then specific studies measuring the accumulation of Se forms in edible tissues are needed. In addition, a correlation exits between tissue Se concentration and GSH-Px activity in meat (DeVore and Greene, 1982) and the Se supplementation of livestock diets has the potential to improve the oxidative stability in meat. However, SeCys forms the functional core of the GSH-Px enzyme and increases in tissue Se in the form of SeMet, although improving the nutritional quality of meat, may not necessarily result in improvements in oxidative stability. Therefore, the objective of this study was to determine the distribution of total Se and the proportion of total Se as SeMet and SeCys, as well as meat quality in terms of oxidative stability, within the postmortem tissues of lambs that had been offered either comparable doses of SY or SS, or the dose response of graded levels of SY within the diet.

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2. Materials and methods 2.1. Animal, experimental design and diets The work was conducted under the authority of the UK Animals (Scientific procedures) Act 1986 (Home Office, 1986) and procedures were undertaken by staff holding appropriate license authorities under the Act. All lambs were housed on straw bedding with sufficient space as prescribed by Home Office guidelines. Fresh potable water was available at all times. Fifty castrated male North Country Mule × Suffolk lambs, with mean age of 4 months and an initial body weight (BW) of 29.5 ± 0.46 kg, were enrolled onto the study. Animals were blocked by BW and then randomly allocated to one of five experimental treatments, an unsupplemented control (T1), those supplemented with ascending concentrations of SY (T2, T3 and T4) or supplemented with SS (T5). Animals were offered, for a period of 112 d, the same total mixed ration (TMR), which contained 800, 138, 39 and 23 g/kg DM of maize silage, soybean meal, ground wheat and vitamin and mineral supplement, respectively, that differed in either the quantity of SY (Sel-Plex® [Saccharomyces cerevisiae CNCM, Collection Nationale de Culture de Micro-organism, I-3060], Alltech, Nicholasville, KY, USA) or Se source (SS), depending on treatment designation. Diets were prepared fresh daily and offered ad libitum (refusal maintained at 100 g/kg DMI). 2.2. Feed analyses Representative samples of each TMR were taken weekly and frozen (−20 ◦ C). At the end of the study feed samples were bulked, sub-sampled and then analyzed for nutritional composition and total Se content. Oven DM content was determined by drying samples in a forced draught oven at 100 ◦ C for 24 h. Ash was obtained by incineration in a muffle furnace at 550 ◦ C for 16 h. Neutral detergent fibre (aNDFom) analyses were performed by the sequential procedure of Van Soest et al. (1991) using the Ankom 200 Fiber Analyzer (Ankom® Technology, Fairpoint, New York, NY). The NDF was assayed with alpha amylase. The NDF values were expressed without residual ash. Starch was determined by polarimetry with random samples analyzed using the enzyme technique (MacRae and Armstrong, 1968) to verify the calibration of the polarimeter. Nitrogen content was measured by the Kjeldahl technique. Oil sugars, neutral detergent cellulose + gammanase (NCGD) were determined as well as estimation of metabolizable energy (ME) by AFRC (1993). 2.3. Feed preparation All TMR were prepared fresh daily using a self-propelled feeder wagon (Calan Data Ranger, American Calan INC., USA) fitted with a weighing device (Weightronix Model 1015, Fairmount, USA), which recorded the individual weights of the different feed ingredients added to the hopper. Selenized yeast and SS were incorporated into the vitamin and mineral supplements (Dairy Direct, Bury St. Edmunds, UK) at the point of manufacture and subsequently incorporated into the SY and SS designated TMR at the time of mixing. Selenized yeast was included in the TMR at 0, 0.11, 0.21 or 0.31 mg/kg DM, to achieve a total dietary Se concentration of 0.19 (T1), 0.30 (T2), 0.40 (T3) or 0.50 (T4) mg/kg DM, respectively, or SS was included at 0.11 mg/kg DM to achieve a total Se concentration of 0.30 (T5) mg/kg DM. 2.4. Blood and tissue sampling Blood samples were taken via jugular venepuncture at enrolment, and on days 28, 56, 84, and 112 of the study from each individual animal. Samples were taken aseptically using the Vacutainer system (BD Diagnostics, Oxford, UK) using a 21 gauge needle into a 5 ml lithium heparin pretreated tube, for subsequent determination of Se, Se species and GSH-Px activity. At the end of the treatment period (112 d), all lambs were euthanased and samples of heart, liver, kidney and skeletal muscle (Longissimus Thoracis and Psoas Major) were retained for determination

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of total Se and Se species. Glutathione peroxidase and thiobarbituric acid reactive substances (TBARS) were determined in Longissimus Thoracis muscle. 2.5. Selenium analyses Total Se in TMR, whole blood sample and tissues (heart, liver, kidney and skeletal muscle) was determined by mineralizing 1 g of each sample in 4 ml 16 M HNO3 and 2 ml 9.8 M H2 O2 within a closed-vessel heating block system. The solution was further diluted with water and Se subsequently determined using inductively coupled plasma mass spectrometry (ICP-MS) (PerkinElmer Elan 6100 ICPMS, Massachusetts, USA). Selenium speciation was determined using the method described by Palacios et al. (2005). Briefly, samples were initially incubated for 5 h with dl-dithiothreitol and iodoacetamide to reduce and alkylate SeCys. Samples were then spiked with SeMet77 and subsequently incubated for 24 h at 37 ◦ C with a mixture of protease and lipase maintained at a pH 7.5. Following incubation, the mixture was centrifuged and the supernatant separated and purified by cell exclusion liquid chromatography. Aliquots of the supernatant were analyzed by reversed-phase HPLC-ICPMS using an ICP MS equipped with a collision cell (PerkinElmer Elan 6100 ICPMS, Massachusetts, USA). 2.6. Meat quality assessment Samples of the Longissimus Thoracis muscle were used for analysis of lipid oxidation as TBARS and GSH-Px. The samples were packed into modified atmosphere packs (MAP, O2 :CO2 , 75:25) and subjected to simulated retail display (3 ◦ C, 700 lux for 16 h out of 24 h). Thiobarbituric acid reactive substances were determined on day 10 of display by the method of Tarladgis et al. (1960) modified by the use of a Büchi 321 distillation unit (BÜCHI Labortechnik AG, Postfach, Switzerland). Glutathione peroxidase activity was determined in samples of the Longissimus Thoracis muscle that were taken either immediately following euthanasia or in samples that had undergone 10 d aging in MAP, using a modified method of the coupled assay procedure of Paglia and Valentine (1967), modified by DeVore and Greene (1982), Daun et al. (2000) and Raes of Ghent University (unpublished). 2.7. Statistical analysis Statistically significant differences between the five treatment groups for whole blood Se content and erythrocyte GSH-Px activity were determined by ANOVA using the Mixed model procedure of SAS (v. 8.2; SAS Inst., Inc., Cary, NC). Sources of variation within the model included treatment (4 df), block (4 df) and time (4 df) where individual animal formed the repeated subject and time the repeated measure. Statistical tests were undertaken for main effects and treatment × time interactions. Statistical differences between the five treatment groups and linear and quadratic contrast between doses for tissue total Se content and meat quality assessments were determined by ANOVA using a general linear model. Sources of variation within the model included treatment. Results are presented as least square means with the S.E.M. and P value. Tukey simultaneous test were used to establish statistical differences (P<0.05) between individual treatment means. 3. Results 3.1. Feed analyses The nutritional composition and Se concentration of TMR for each treatment are shown in Table 1. Although the experimental diets were formulated to have the same nutritional composition, there was variation between treatments in a number of nutritional parameters. The concentrate blends for each treatment had Se contents of 0.4, 0.9, 1.1, 2.5 and 1.2 mg Se/kg DM for treatments T1, T2, T3, T4 and T5, respectively, when combined with Se content of the maize silage gave estimated TMR Se contents of 0.18, 0.28, 0.32, 0.3 and 0.6 mg/kg DM.

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Table 1 Laboratory determined nutrient composition (g/kg DM unless otherwise stated) of total mixed ration diets offered to lambs that were either unsupplemented (T1), supplemented with 0.11, 0.21, and 0.31 mg Se/kg DM as selenized yeast (T2, T3, and T4, respectively) or 0.11 mg Se/kg DM as sodium selenite (T5) for a period of 112 d Diet

DM (g/kg FW) Crude protein ME (MJ/kg DM) Starch Neutral detergent fibre Sugars NCGD Oil Ash Se (mg/kg DM)

S.E.M.

T1

T2

T3

T4

T5

412 148 10.9 216 320 24.2 719 35.5 61.1 0.08

445 166 10.8 272 344 13.0 713 32.7 56.0 0.12

442 161 11.0 285 330 12.7 558 34.4 53.2 0.18

435 165 11.0 271 337 15.0 589 33.4 55.3 0.23

414 170 11.1 276 323 10.8 725 36.2 54.4 0.15

11.3 14.9 0.15 24.6 16.8 2.56 84.1 1.22 1.58 0.01

3.2. Selenium concentration in whole blood and tissues There were significant treatment (P<0.001) and linear dose effects (P<0.001) to the graded addition of SY to the diet in both whole blood and plasma total Se concentrations, values increasing on average by 150 and 71 ng/g FW in whole blood and plasma, respectively, for each 1 mg increase in Se intake derived from SY. However, there were no differences between Se sources, both treatments having similar values (465.3 and 467.9 ng/g FW for T2 and T5, respectively). Whole blood total Se values, irrespective of treatment, increased over time but a Se steady state had not been achieved following 112 d exposure to the experimental diets. Estimations of time required to reach a Se steady state based on regression analysis (data not shown) indicated that asymptotic values were predictably higher and achieved faster at higher inclusion rates of SY (125 d for T4; 145 d for T3 and T5; over 145 d for T2 and T1 treatments). The Se concentration of heart, liver, kidney and skeletal muscle (Psoas Major and Longissimus Thoracis) (Table 2) differed between different tissue types. Total Se concentration was greatest in kidney, with values ranging between 5.6 and 6.5 mg/kg DM, followed by liver, heart and skeletal muscle. There were no effects of treatment or any dose responses on the concentration of total Se within kidney tissue. Conversely, there was a significant linear dose dependant response on cardiac (P=0.056), hepatic (P=0.002) and skeletal muscle (P<0.001, for both Psoas Major and Longissimus Thoracis) to the graded addition of SY to the diet, tissue Se concentrations increasing on average by 0.43, 1.20, 0.60 and 0.56 mg/kg DM for each 1 mg/kg DM increase in Se intake derived from SY. However, the concentration of total Se within liver, heart and kidney tissues appeared unaffected by Se source, although the Se content of the Longissimus Thoracis and Psoas Major were numerically (0.59 vs. 0.47 mg/kg DM) and Table 2 Total selenium concentration in heart, liver, kidney and muscle (Psoas Major and Longissimus Thoracis) (mg/kg DM) of lambs offered diets that were either unsupplemented (T1), supplemented with 0.11, 0.21, and 0.31 mg Se/kg DM as selenized yeast (T2, T3, and T4, respectively) or 0.11 mg Se/kg DM of sodium selenite (T5) for a period of 112 d Treatment

S.E.M.

P QuadraticA

T2

T3

T4

T5

Heart Liver Kidney

1.08 1.34a 5.67

1.24 1.63a 5.87

1.37 1.81ab 5.63

1.36 2.23b 6.47

1.12 1.70ab 5.67

0.095 0.128 0.409

0.187 0.008 0.586

0.056 0.002 0.248

0.452 0.574 0.435

Skeletal muscle tissue Psoas Major Longissimus Thoracis

0.42a 0.40a

0.61b 0.59a

0.71bc 0.67b

0.80c 0.78b

0.48a 0.47a

0.029 0.048

<0.001 0.002

<0.001 <0.001

0.179 0.500

a,b,c A

Treatment

LinearA

T1

Means within a row with different superscripts are significantly different (P<0.05). Linear and quadratic contrasts exclude treatment T5 (selenite 0.30 mg/kg total Se).

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Fig. 1. Whole blood total selenium and the proportion of total selenium comprised as selenomethionine, selenocysteine and other selenium species in lambs offered diets that were either unsupplemented (T1), supplemented with 0.11, 0.21, and 0.31 mg Se/kg DM as selenized yeast (T2, T3, and T4, respectively) or 0.11 mg Se/kg DM as sodium selenite (T5) for a period of 112 d.

significantly greater (P<0.05; 0.61 vs. 0.48 mg/kg DM), respectively, in those animals that had received SY when compared to those that had received comparable doses of SS. 3.3. Selenized amino acids contents of whole blood and tissues Total Se and the proportion of total Se comprised as the selenized amino acids SeCys and SeMet, as well as other unidentified Se fractions, for whole blood at enrolment and days 56 and 112 of the study, and those for visceral and muscle tissues at completion of the study are shown in Figs. 1–3, respectively.

Fig. 2. Total selenium and the proportion of total selenium comprised as selenomethionine, selenocysteine and other selenium species in liver and kidney tissues of lambs offered diets that were either unsupplemented (T1), supplemented with 0.11, 0.21, and 0.31 mg Se/kg DM as selenized yeast (T2, T3, and T4, respectively) or 0.11 mg Se/kg DM as sodium selenite (T5) for a period of 112 d.

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Fig. 3. Total selenium and the proportion of total selenium comprised as selenomethionine, selenocysteine and other selenium species in heart and muscle tissues (Psoas Major and Longissimus Thoracis) of lambs offered diets that were either unsupplemented (T1), supplemented with selenized yeast (T2, T3, and T4) or sodium selenite (T5) for a period of 112 d.

With respect to whole blood, SeCys was the predominant selenized amino acid, irrespective of treatment and time point accounting for between 70 and 80% of total Se. The SeMet content in whole blood was different between Se sources, values being greatest in treatment T2 when compared with T5, although proportionally there was little difference, as SeMet accounted for between 14 and 15% of total Se irrespective of source. Absolute SeMet contents were greatest in those animals that had received T4 and accounted for over 21% of total Se. When changes seen in the proportions of each of the selenized amino acids are expressed as a function of enrolment values then, with the exception of T1, SeMet values increased by 170% of enrolment values in treatments T2 and T5, whereas the greatest increase seen in enrolment values was seen in treatment T4 which increase by 280%. With regard to Se speciation in tissues, SeCys was the predominant selenized amino acid within the kidney, liver and heart, comprising 90, 65 and 55% of total Se, respectively, appearing unaffected by either Se source or dose. However, although SeMet was the smaller fraction of total Se, there was a small dose response in the quantity of SeMet within heart, liver, kidney and Psoas Major muscle. Furthermore, incorporation of SeMet into heart, liver and kidney tissues was greater when SY formed the Se source with SeMet comprising 9, 30 and 31%, respectively, of total Se when compared with 6, 22 and 23% when SS formed the Se source. 3.4. Whole blood glutathione peroxidase activity Glutathione peroxidase activity (Table 3) was greater (P=0.114) in those treatments that had been augmented with supplementary Se when compared to T1, although there was no significant effect of dietary Se source on GSH-Px activity. The plot of whole blood Se concentration and GSH-Px activity (Fig. 4) indicated a positive curvilinear response (R2 = 0.726) to ascending whole blood Se values. In addition, increases in GSH-Px activity were better correlated with the proportion of total Se comprised as SeCys (R2 = 0.932; P<0.001) than SeMet in whole blood. 3.5. Glutathione peroxidase activity and thiobarbituric acid reactive substances in Longissimus Thoracis Tissue GSH-Px activities determined at 0 and 10 d post-mortem and TBARS determined at 10 d post-mortem of the Longissimus Thoracis muscle is shown in Table 4.

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Table 3 Whole blood total selenium concentration and glutathione peroxidase activity of lambs offered diets that were either unsupplemented (T1), supplemented with 0.11, 0.21, and 0.31 mg Se/kg DM as selenized yeast (T2, T3, and T4, respectively) or 0.11 mg Se/kg DM as sodium selenite (T5) for a period of 112 d Treatment T1 Whole blood Glutathione peroxidase (GSH-Px/g Hb) Plasma

S.E.M.

T2 a

T3 a

T4 ab

T5 b

a

P Treatment

LinearA

QuadraticB

420.1 301.5

465.3 333.5

493.6 325.7

519.8 323.1

467.9 316.3

13.93 11.2

<0.001 0.355

<0.001 0.255

0.545 0.140

123.6a

145.6b

171.2c

168.3c

150.4b

4.54

<0.001

<0.001

0.023

a,b,c

Means within a row with different superscripts are significantly different (P<0.05). A Linear and quadratic contrasts exclude treatment T5 (selenite 0.30 mg/kg total Se).

Fig. 4. Trend line analysis of the relationship between total Se content and glutathione peroxidase activity in whole blood of lambs offered diets that were either unsupplemented (T1), supplemented with 0.11, 0.21, 0.31 mg Se/kg DM as selenized yeast (T2, T3, and T4, respectively) or 0.11 mg Se/kg DM as sodium selenite (T5) for a period of 112 d.

Thiobarbituric acid reactive substances values were greatest in the Longissimus Thoracis from T1, although statistical differences between dietary treatments were not established. However, there was an indication of a linear dose effect (P=0.102) to the graded addition of SY. When TBARS values are plotted against total Se content of the Longissimus Thoracis (Fig. 5) from treatments T1 to T4 it indicates Table 4 Thiobarbituric acid reactive substances measured at 10 d post-mortem and glutathione peroxidase measured followed euthanasia and after 10 d aging in modified atmosphere packs (O2 :CO2 , 75:25) in Longissimus Thoracis muscle of lambs offered diets that were either unsupplemented (T1), supplemented with 0.11, 0.21, and 0.31 mg Se/kg DM as SY (T2, T3, and T4, respectively) or 0.11 mg Se/kg DM as sodium selenite (T5) for a period of 112 d Treatment T1 TBARS 10 d post-mortem (mg MDAb /kg) Tissue GSH-Px 0 d post-mortem (GSH-Px/g) Tissue GSH-Px 10 d post-mortem (GSH Px/g) a b

S.E.M.

T2

T3

T4

T5

P Treatment

Lineara

Quadratica

1.47

1.31

0.80

0.64

0.42

0.34

0.186

0.102

0.995

14.15

15.38

14.86

13.59

13.05

1.22

0.690

0.697

0.323

14.68

14.80

14.43

13.73

13.04

0.96

0.690

0.496

0.696

Linear and quadratic contrasts exclude treatment T5 (selenite 0.3 mg/kg DM total Se). MDA = malondialdehyde.

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Fig. 5. Trend line analysis of the relationship between total Se content and thiobarbituric acid reactive substances of the Logissimus Thoracis muscle of lambs offered diets that were either unsupplemented (T1), supplemented with 0.11, 0.21, and 0.31 mg Se/kg DM as selenized yeast (T2, T3, and T4, respectively) or 0.11 mg Se/kg DM as sodium selenite (T5) for a period of 112 d.

a reduction in TBARS values with ascending total tissue Se content (R2 = 0.302), although it does suggest that further increases in tissue Se contents would not necessarily result in further reductions in TBARS values. Although there were no statistical differences between Se sources in tissue GSH-Px activity, these values were greater for SY supplemented animals when compared with SS, both at 0 (15.38 vs. 13.05) and 10 d (14.80 vs. 13.04) post-mortem, which is consistent with total selenium content of the Longissimus Thoracis.Subsequent regression analysis failed to establish any relationship between tissue total Se content and tissue GSH-Px activity at 0 and 10 d post-mortem (R2 < 0.001). Similarly, regression analysis of TBARS against GSH-Px activity at 0 and 10 d post-mortem failed to establish a relationship between these two parameters. 4. Discussion 4.1. Feed analyses The variation showed between the different dietary treatments in a number of measured nutritional parameters was probably the result of variation found within each sub-sample as a consequence of the coarse nature of the compositional ingredients. A number of samples for each of the constituent ingredients had been analyzed for total Se prior to the commencement of the study and required doses calculated to meet target levels. However the variation seen between expected and laboratory Se concentrations may once again be the result of the coarse nature of the compositional ingredients. 4.2. Selenium concentration in whole blood and tissues There were no differences between sources in whole blood total Se. However, other authors (Van Ryssen et al., 1989) reported higher whole blood Se content in sheep fed organic Se when compared to those fed comparable amounts of SS, although dietary Se concentrations used by these authors were notably greater than those used in this present study (1 mg/kg DM). In this study a linear dose response was shown on whole blood Se concentration to the graded addition of SY to the diet. Similar linear dose dependant responses on whole blood Se content to the graded addition of organic SY to dairy cow diets (Juniper et al., 2006) and SS to wether sheep diet (Cristaldi et al., 2005) have been reported. Whole blood Se values, irrespective of treatment, increased over time although a Se steady state was not achieved after 112 d continuous exposure to the experimental diets. This is consistent with the findings of Van Ryssen et al. (1989) who reported that the Se content of whole blood Se

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of sheep failed to achieve a plateau following a similar experimental period during which sheep were offered diets containing up to 1 mg/kg DM whether administered in organic or inorganic forms. Extrapolation data from this current study indicates that Se steady state may be achieved between 125 and 150 d, reflecting the turnover rate of erythrocytes (Mahan et al., 1999). Changes in whole blood total Se in T1 was unexpected and could be due to the background Se content of the basal TMR being probably higher than that of the pasture animals had grazed prior to study enrolment. The Se concentrations of tissues differed between different tissues types, although the hierarchy of tissue Se contents were consistent with those of wether lambs offered diets containing up to 2.9 mg Se/kg DM reported by Taylor (2005). The lack of treatment effect on the concentration of total Se within kidney tissue, are consistent with the results reported previously by Van Ryssen et al. (1989). The greater Se concentration of Longissimus Thoracis and Poas Major, when animals received SY supplementation in comparison with SS was not unexpected, as organic Se supplementation has been shown to result in greater Se accumulation in muscle in comparison with inorganic Se (Van Ryssen et al., 1989; Lawler et al., 2004). Other authors (Qin et al., 2007) reported significantly greater total Se concentrations in the kidney, liver and muscle tissues of lambs offered SY supplementation (0.10 mg/kg diet) when compared with those offered comparable amounts of SS. 4.3. Selenized amino acids contents of whole blood and tissues The increases seen in whole blood total Se concentration over time, irrespective of treatment, were more attributable to increases in the proportion of total Se comprised as SeCys than SeMet or other unidentified Se fractions. The proportion of Se comprised as SeMet was greater in animals receiving SY supplements when compared to those offered comparable doses of SS, which is most likely ascribed to the high concentration of SeMet in the SY product (SeMet comprised 54–74% of total Se; Rayman, 2004). With regard to the Se speciation of tissues, SeCys was the predominant selenized amino acid, irrespective of Se source or dose, within the kidney, liver and heart. Given the greater metabolic activity of these tissues these results would suggest the incorporation of Se into functional seleno-enzymes rather than the non-specific incorporation into tissue protein. Conversely, the proportion of total Se that was comprised as SeMet was greater in skeletal muscle tissue, especially when SY was the Se source, reflecting better uptake and the non-specific incorporation of SeMet into tissue protein (Schrauzer, 2003). Similar results have been reported in sheep (Van Ryssen et al., 1989) and steers (Lawler et al., 2004) when organic Se was compared with inorganic sources. 4.4. Whole blood glutathione peroxidase activity Although it has been shown that whole blood Se concentration is closely correlated with GSHPx activity (Koller et al., 1984), the percentage of Se in erythrocytes associated with GSH-Px may vary depending on Se source (Beilstein and Whanger, 1986; Van Ryssen et al., 1989). The lack of effect of the source of Se on erythrocyte GSH-Px activity showed in this study supports the review of 11 studies cited by Weiss (2003). This author reported that in nine of those studies, there was no significant difference in GSH-Px activity when comparing SY with SS, even though the values were numerically higher for SY supplemented animals. According to Weiss (2003) these results indicated that SY provided equal to or slightly higher amounts of selenide for SeCys synthesis compared with SS. However, other authors (Qin et al., 2007) reported significantly higher whole blood Se concentrations and GSH-Px activities in lambs fed with SY enriched diets (0.10 mg/kg diet, during 8 weeks) when compared with lambs fed similar concentrations of SS. The increases in GSH-Px activity were better correlated with the proportion of total Se comprised as SeCys than SeMet of whole blood, which would tend to indicate that the larger proportion of whole blood total Se was incorporated into GSH-Px.

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4.5. Glutathione peroxidase activity and thiobarbituric acid reactive substances in Longissimus Thoracis The greatest TBARS values which were indicative of greater lipid oxidation were seen in the Longissimus Thoracis from T1 animals, although statistical differences between dietary treatments were not established. Other authors (Skˇrivanová et al., 2007) have failed to establish significant effects of SY supplementation on the oxidative stability of the Longissimus Thoracis in calves. Several authors reported a correlation between GSH-Px activity and Se content of tissues (DeVore and Greene, 1982; Gatellier et al., 2004). Although there were no statistical differences between Se sources in tissue GSH-Px activity, these values were greater for SY supplemented animals when compared with SS at 0 and 10 d, which is consistent with total selenium content of Longissimus Thoracis. However, Van Ryssen et al. (1989), in sheep, and Zhan et al. (2007), in pigs, reported similar tissue GSHPx activities in animals that were offered organic Se supplementation when compared to inorganic Se although organic supplemented animals had greater tissue total Se content. These authors attributed this fact to the different metabolic pathways that differing sources of Se follow for incorporation into GSH-Px (Matte, 2007). There was no relationship between TBARS values and GSH-Px activity in Longissimus Thoracis, the improvements seen in oxidative stability with ascending tissue Se concentrations may have been attributable to other Se dependant antioxidant mechanisms. 5. Conclusions The incorporation into growing lambs diet of ascending concentrations of Se in the form of SY significantly increased total Se in whole blood and tissues, except kidney. Comparable doses of SS supplementation did not result in significant differences in these parameters. However, Se content of muscle was higher in animals offered SY when compared with SS, indicating improvement in Se uptake and retention. There were no significant effects of treatment on meat quality assessments, although oxidative stability was seen to improve marginally with ascending tissue Se content. Acknowledgments This research was funded by Alltech EU Regulatory Affairs Department (trial ref. SEL/LAM/DOS/01/0405/UK). The authors would also like to give special thanks to Ryszard Lobinski (UT2A laboratories) for performing the selenium analyses. References AFRC, 1993. Energy and Protein Requirements of Ruminants. An advisory manual prepared by the AFRC Technical Committee on Responses to Nutrients. CABI International, Wallingford, UK. Arteel, G.E., Sies, H., 2001. The biochemistry of selenium and the glutathione system. Environ. Toxicol. Pharmacol. 10, 153–158. Beilstein, M.A., Whanger, P.D., 1986. Deposition of dietary organic and inorganic selenium in rat erythrocyte proteins. J. Nutr. 116, 1701–1710. Cristaldi, L.A., McDowell, L.R., Buergelt, C.D., Davis, P.A., Wilkinson, N.S., Martin, F.G., 2005. Tolerance of inorganic selenium in wether sheep. Small Rumin. Res. 56, 205–213. ´˚ Daun, C., Johansson, M., Önning, G., Akesson, B., 2000. Glutathione peroxidase activity, tissue and soluble selenium content in beef and pork in relation to meat ageing and pig RN phenotype. Food Chem. 73, 313–319. DeVore, V.R., Greene, B.E., 1982. Glutathione peroxidase in post-rigor bovine semitendinosus muscle. J. Food Sci. 47, 1406–1409. DeVore, V.R., Colnago, G.L., Jensen, L.S., Greene, B.E., 1983. Thiobarbituric acid values and glutathione peroxidase activity in meat from chickens fed a selenium supplemented diet. J. Food Sci. 48, 300–301. Gatellier, P., Mercier, Y., Renerre, M., 2004. Effect of diet finishing mode (pasture or mixed diet) on antioxidant status of Charolais bovine meat. Meat Sci. 67, 385–394. Home Office, 1986. Animal Scientific Procedures Act 1986. Her Majesty’s Stationary Office, London. Juniper, D.T., Phipps, R.H., Jones, A.K., Bertin, G., 2006. Selenium supplementation of lactating dairy cows: effect on selenium concentration in blood, milk, urine and feces. J. Dairy Sci. 89, 3544–3551. Koller, L.D., South, P.J., Exon, J.H., Whitbeck, G.A., Maas, J., 1984. Comparison of selenium levels and glutathione peroxidase activity in bovine whole blood. Can. J. Comp. Med. 48, 431–433. Korhola, M., Vainio, A., Edelmann, K., 1986. Selenium yeast. Ann. Clin. Res. 18, 65–68. Lawler, T.L., Taylor, J.B., Finley, J.W., Caton, J.S., 2004. Effect of supranutritional and organically bound selenium on performance carcass characteristics and selenium distribution in finishing beef steers. J. Anim. Sci. 82, 1488–1493.

D.T. Juniper et al. / Animal Feed Science and Technology 149 (2009) 228–239

239

MacRae, J.C., Armstrong, D.G., 1968. Enzyme method for determination of ␣-linked glucose polymers in biological materials. J. Sci. Food Agr. 19, 578–581. Mahan, D.C., Cline, T.R., Richert, B., 1999. Effects of dietary levels of selenium-enriched yeast and sodium selenite as selenium sources fed to growing-finishing pigs on performance, tissue selenium, serum glutathione peroxidase activity, carcass characteristics, and loin quality. J. Anim. Sci. 77, 2172–2179. Matte, J.J., 2007. Selenium metabolism, the glutathione peroxidase system and their interaction with some B vitamins in pigs. In: Lyons, T.P., Jaques, K.A., Hower, J.M. (Eds.), Proceedings of Alltechs 23rd Annual Symposium. Nottingham University Press, Nottingham, UK, pp. 87–96. Morrissey, P.A., Sheehy, P.J.A., Galvin, K., Kerry, J.P., Buckley, D.J., 1998. Lipid stability in meat and meat products. Meat Sci. 49, 73–86. Paglia, D.E., Valentine, W.N., 1967. Studies on the quantitative and qualitative characterization of erythrocyte glutathione peroxidase. J. Lab. Clin. Med. 70, 158–169. Palacios, O., Encinar, J.R., Bertin, G., 2005. Analysis of the selenium species distribution in cow blood by size exclusion liquid chromatography-inductively coupled plasma collision cell mass spectrometry (SEC-ICPccMS). Anal. Bioanal. Chem. 383, 516–522. Qin, S., Gao, J., Huang, K., 2007. Effects of different selenium sources on tissue selenium concentrations, blood GSH-Px activities and plasma interleukin levels in finishing lambs. Biol. Trace Elem. Res. 116, 91–102. Rayman, M.P., 2004. The use of high-selenium yeast to raise selenium status: how does it measure up? Br. J. Nutr. 92, 557–573. Rotruck, J.T., Pope, A.L., Ganther, H.E., Swanson, A.B., Hafeman, D.G., Hoekestra, W.G., 1973. Selenium: biochemical role as a component of glutathione peroxidase. Science 179, 588–590. Schrauzer, G.N., 2003. The nutritional significance, metabolism and toxicology of selenomethionine. Adv. Food Nutr. Res. 47, 73–112. Seko, Y., Saiti, Y., kitahara, J., Imura, N., 1989. Active oxygen generation by the reaction of selenite with reduced glutathione in vitro. In: Wendel, A. (Ed.), Selenium in Biology and Medicine. Springer-Verlag, Berlin, pp. 33–70. Skˇrivanová, E., Marounek, M., De Smet, S., Raes, K., 2007. Influence of dietary selenium and vitamin E on quality of veal. Meat Sci. 76, 495–500. Surai, P.F., 2006. Selenium distribution and reserves in human body. In: Selenium in Nutrition and Health. Nottingham University Press, Nottingham, UK, pp. 161–171. Suzuki, K.T., Ogra, Y., 2002. Metabolic pathway for selenium in the body: speciation by HPLC-ICP MS with enriched Se. Food Addit. Contamin. 19, 974–983. Tarladgis, B.G., Watts, B.M., Younathan, M.T., 1960. A distillation method for the quantitative determination of malonaldehyde in rancid foods. J. Am. Oil Chem. Soc. 37, 44–48. Taylor, J.B., 2005. Time-dependant influence of supranutritional organically bound selenium on selenium accumulation in whether lambs. J. Anim. Sci. 83, 1186–1193. Van Ryssen, J.B.J., Deagen, J.T., Beilstein, M.A., Whanger, P.D., 1989. Comparative metabolism of organic and inorganic selenium by sheep. J. Agric. Food Chem. 37, 1358–1363. Van Soest, P.J., Robertson, J.B., Lewis, B.A., 1991. Methods for dietary fibre, neutral detergent fibre and non-starch polysaccharides in relation to animal nutrition. J. Dairy Sci. 74, 3583. Weiss, W.P., 2003. Selenium nutrition of dairy cows: comparing responses to organic and inorganic selenium forms. In: Lyons, P., Jaques, K.A. (Eds.), Proceedings of the 19th Alltech Annual Symposium on Nutritional Biotechnology in the Feed and Food Industries. Nottingham University Press, Nottingham, UK, pp. 333–334. Zhan, X., Wang, M., Zhao, R., Li, W., Xu, Z., 2007. Effects of different selenium source on selenium distribution, loin quality and antioxidant status in finishing pigs. Anim. Feed Sci. Technol. 132, 202–211.