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Influence of dietary cobalt on performance, nutrient digestibility and plasma metabolites in lambs
Animal Feed Science and Technology 135 (2007) 346–352
Short communication
Influence of dietary cobalt on performance, nutrient digestibility and plasma metabolites in lambs R.L. Wang a,b , X.H. Kong a , Y.Z. Zhang a , X.P. Zhu a , Narenbatu a , Z.H. Jia a,∗ a
College of Animal Science and Technology, China Agricultural University, Beijing 100094, PR China b Department of Animal Science, College of Agriculture Science, Guangdong Ocean University, Zhanjiang 524088, PR China Received 30 November 2005; received in revised form 22 June 2006; accepted 2 August 2006
Abstract Two experiments were conducted to study the effects of different inclusion levels of dietary cobalt (Co) on performance, nutrient digestibility and plasma metabolites in lambs. Experiment 1: sixty, 12-week-old weaned lambs (Chinese Poll Dorset × Small Tailed Han) were randomly divided into five groups which were fed with the basal diet containing 0.086 mg Co/kg DM, the basal diet supplied, respectively, with 0.25, 0.50, 0.75 and 1.00 mg Co/kg DM as reagent grade CoSO4 ·7H2 O for 10 weeks. Experiment 2: four lambs from each group in Experiment 1 were randomly allocated to the individual metabolic crates to measure the effects of dietary Co on apparent nutrient digestibility. Average daily gain and average daily feed intake were highest at the intermediate level (total diet Co of 0.586 mg/kg DM) (P<0.05). However, there was no difference in gain/feed. Supplemental 0.50 mg Co/kg DM resulted in highest digestibility of dry matter, organic matter, crude protein and acid detergent fiber and metabolizable energy intake (P<0.05). Neutral detergent fiber digestibility was increased at an increasing rate (P<0.05). Co supplementation at the intermediate level showed highest plasma vitamin B12 concentration and lowest plasma methylmalonic acid concentration (P<0.05), whereas there were no differences in plasma concentrations of folate and homocysteine. Plasma glucose concentration
Abbreviations: Co, cobalt; DM, dry matter; OM, organic matter; ME, metabolizable energy; CP, crude protein; NDF, neutral detergent fiber; ADF, acid detergent fiber; Ca, calcium; P, phosphorus; NaCI, sodium chloride; BW, body weight; ADG, average daily gain; ADFI, average daily feed intake; G/F, gain/feed; EE, ether extract; MMA, methylmalonic acid; HCY, homocysteine ∗ Corresponding author. Tel.: +86 10 62732728; fax: +86 10 62732728. E-mail address: [email protected] (Z.H. Jia). 0377-8401/$ – see front matter © 2006 Elsevier B.V. All rights reserved. doi:10.1016/j.anifeedsci.2006.08.011
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was linearly increased (P<0.05). In conclusion, the growth of Chinese Poll Dorset × Small Tailed Han lambs is compromised when fed a Chinese wildrye hay based diet containing 0.086 mg Co/kg DM. The recommended level of dietary Co for such lambs is between 0.336 and 0.586 mg/kg DM. © 2006 Elsevier B.V. All rights reserved. Keywords: Lamb; Cobalt; Vitamin B12 ; Performance; Nutrient digestibility; Metabolites
1. Introduction Cobalt (Co) is required for the synthesis of vitamin B12 by rumen microorganisms. Vitamin B12 acts as a co-factor for methylmalonyl-CoA mutase and methionine synthase which are important for gluconeogenesis and methionine synthesis. It has been reported that low level of dietary Co can lead to vitamin B12 deficiencies clinically manifested as anemia, inappetence and poor production and biochemically characterized by decrease in the plasma concentration of vitamin B12 , elevations in the plasma concentrations of methylmalonic acid (MMA) and homocysteine (HCY) in ruminants (Kennedy et al., 1990). The currently recommended dietary levels of 0.1–0.2 mg Co/kg dry matter (DM) in sheep (NRC, 1985) are based on observations in grazing animals; recent findings however have demonstrated that 0.10 mg/kg DM of Co intake does not meet the rumen microbial requirements (Kisidayova et al., 2001; Johnson et al., 2004). It was reported that 0.3–0.5 mg Co/kg DM enhanced ruminal microbial activity, fermentation and vitamin B12 synthesis (Singh and Chhabra, 1995). In addition, a higher level of dietary Co has been suggested both in beef cattle (Stangl et al., 2000) and cows (Tomlinson and Socha, 2003) than the recommended data. In northern China, Co is supplied to sheep at 0.1–0.2 mg/kg DM, and clinical cases of vitamin B12 deficiency especially in lambs have been reported (Wang et al., 2003). The optimal dietary levels of Co in lambs have not been properly established, and the effects of dietary Co concentration on sheep nutrient digestibility and metabolic characteristics were not well understood. Therefore, the present study was designed to evaluate the effects of dietary Co level on performance and nutrient utilization in Chinese Poll Dorset × Small Tailed Han sheep. 2. Materials and methods 2.1. Experiment 1 Sixty, 12-week-old weaned lambs (Poll Dorset × Small Tailed Han sheep) were housed in individual wooden pens with slatted floors in an open-sided barn and fed a Chinese wildrye hay based diet which was formulated to meet all nutrient requirements with the exception of Co for lambs (NRC, 1985) (Table 1). The Chinese wildrye hay was offered once a day ad libitum, and the concentrate (500 g/animal) was supplied in two equal amounts at 06:00 h and 16:30 h. All animals had free access to water containing undetectable concentrations of Co (analyzed less than 0.01 mg/kg). After 2 weeks adjustment to the experimental feeding system, lambs were weighed and randomly assigned to 5 treatments with 12 each: control
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Table 1 The composition of the basal diet fed to the lambs Chemical compositiona (g/kg DM)
Feed ingredients (g/kg) hayb,c
Chinese wildrye Maize flourc Wheat branc Soybean mealc Cottonseed mealc Limestone Salt Premixd
500 279 120 70 20 2 6 3
Organic matter Metabolizable energy (MJ/kg DM) Crude protein Neutral detergent fiber Acid detergent fiber Calcium Phosphorus Sodium chloride Cobalt (mg/kg DM)
920.3 8.92 139.6 398.0 237.6 3.4 3.4 5.7 0.086
a
Analyzed values except metabolizable energy. Composition of Chinese wildrye hay: ME, 7.84 MJ/kg; CP, 73 g/kg; EE, 36 g/kg; NDF, 575 g/kg; ADF, 328 g/kg; Ca, 2.2 g/kg; P, 1.4 g/kg. c Co content: Chinese wildrye hay, 0.03 mg/kg; maize flour, 0.05 mg/kg; wheat bran, 0.11 mg/kg; soybean meal, 0.48 mg/kg; cottonseed meal, 0.52 mg/kg. d Provided per kilogram of the diet: 40 mg of Zn as ZnSO ·7H O; 25 mg of Mn as MnSO ·H O; 1.0 mg of I 4 2 4 2 as KI; 50 mg of Fe as FeSO4 ·7H2 O; 10 mg of Cu as CuSO4 ·5H2 O; 0.2 mg of Se as Na2 SeO3 ·5H2 O; 1500 IU of Vitamin A; 250 IU of Vitamin D and 16 IU of Vitamin E. b
fed with the basal diet, and 4 supplemental groups which were fed the basal diet and supplied with 0.25, 0.50, 0.75 and 1.00 mg Co/kg DM, respectively. Co was added as CoSO4 ·7H2 O to the premix using finely maize flour as a carrier and mixed with concentrate. The experiment lasted for 70 days. Daily feed offerings and refusals were measured to obtain net feed intake for each animal. At the end of the experiment, the final body weight was measured. The blood samples were collected from each lamb before morning feeding via jugular venipuncture into heparinized vacutainers, then centrifuged at 4 ◦ C for 10 min at 1100 × g and stored at 70 ◦ C pending analyses. 2.2. Experiment 2 Four lambs from each group in Experiment 1 were randomly selected at the end of the feeding period and allocated to individual metabolism crate to study the effects of dietary Co on apparent nutrient digestibility. The digestibility trial lasted for 10 days with 5-day adaptation period and subsequent 5-day collection period during which daily feed intake and output of faeces were collected and recorded. Animals were fed the same diet as in Experiment 1. Representative daily hay and concentrate samples were collected every morning. Faeces from each lamb were collected using a total collection method in plastic bags, then weighted and subsampled. Sulfuric acid was added to the sample to prevent nitrogen loss. Animal feeds and faeces was dried to a constant weight at 70 ◦ C, then ground to pass a 1 mm screen and preserved for further analyses. 2.3. Analytic methods The Co concentrations in feeds were determined using an atomic absorption spectrophotometer (Model 5100, HGA-600 Graphite Furnace; Perkin-Elmer, USA) by absorbance at
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240.7 nm. Feed and faecal samples were analyzed for dry matter (DM), organic matter (OM), crude protein (CP) and ether extract (EE) according to AOAC (1990); neutral detergent fiber (NDF) by Van Soest et al. (1991) without sodium sulfite or alpha amylase; and acid detergent fiber (ADF) by Robertson and Van Soest (1981). CP in feaces was analyzed in dry samples. NDF and ADF are expressed with residual ash. Plasma concentrations of vitamin B12 and folate were determined using a competitive binding radio-immunoassay kit, in which the non-specific vitamin B12 -binding R-protein was removed by affinity chromatography (ICN, Costa Mesa, CA, USA); Plasma MMA concentration by McMurray et al. (1986) through a modified GC method (Agilent Instruments Model 6890, Agilent Technologies, Wilmington, DE). Plasma levels of HCY by Cornwell et al. (1993); and glucose by a commercial kit (Sigma chemical Co., St. Louis, MO). 2.4. Statistical analysis The nature of response to incremental additions of Co was analyzed by polynomial contrasts (SAS Inst. Inc., Cary, NC). The model included linear and quadratic contrast for effects of supplemental Co.
3. Results 3.1. Performance Average daily gain (ADG) and average daily feed intake (ADFI) were highest at the intermediate level (total diet Co of 0.586 mg/kg DM). However, there was no difference in gain/feed (F/G) (Table 2). 3.2. Digestibility Digestibility of DM, OM, CP and ADF and metabolizable energy intake were highest at the intermediate level (P<0.05) (Table 3), but there were no differences in EE digestibility. NDF digestibility was increased at an increasing rate (P<0.05). Table 2 Effects of dietary Co supplementation on performancea of lambs (DM basis) S.E.M.b
Co supplemental levels (mg/kg)
Initial BW (kg) Final BW (kg) ADG (kg) ADFI (kg) G/F (g/g) a
0
0.25
0.50
0.75
1.00
23.57 34.82 0.161 1.000 0.161
23.37 36.09 0.182 1.052 0.173
23.48 37.49 0.200 1.114 0.181
23.37 36.08 0.182 1.099 0.166
23.57 36.06 0.178 1.085 0.165
0.213 0.403 0.005 0.024 0.005
Co level responses Linear
Quadratic
0.978 0.491 0.057 0.137 0.952
0.379 0.250 0.018 0.049 0.300
Co, cobalt; BW, body weight; ADG, average daily gain; ADFI, average daily feed intake; G/F, average daily gain/average daily feed intake. b S.E.M.: standard error of mean, where n = 12 per treatment.
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Table 3 Effects of dietary Co supplementation on apparent nutrient digestibility coefficientsa in lambs (DM basis) S.E.M.b
Co supplemental levels (mg/kg)
DM OM CP EE NDF ADF ME (MJ/d)
0
0.25
0.50
0.75
1.00
0.692 0.715 0.703 0.685 0.502 0.484 8.924
0.704 0.727 0.712 0.688 0.512 0.493 9.381
0.712 0.731 0.715 0.702 0.520 0.501 9.935
0.711 0.730 0.715 0.703 0.525 0.503 9.805
0.711 0.731 0.715 0.694 0.522 0.507 9.678
0.004 0.004 0.003 0.003 0.002 0.002 0.213
Co level responses Linear
Quadratic
0.070 0.099 0.078 0.250 0.033 0.055 0.137
0.020 0.046 0.041 0.204 0.008 0.006 0.049
a Co, cobalt; DM, dry matter; OM, organic matter; CP, crude protein; EE, ether extract; NDF, neutral detergent fiber; ADF, acid detergent fiber; ME, metabolizable energy. b S.E.M.: standard error of mean, where n = 4 per treatment.
Table 4 Effects of dietary Co supplementation on plasma metabolitesa in lambs S.E.M.b
Co supplemental levels (mg/kg)
Vitamin B12 (pmol/L) Folate (nmol/L) MMA (mol/L) HCY (mol/L) Glucose (mmol/L) a b
0
0.25
0.50
0.75
1.00
202 17.34 5.93 11.62 3.59
1688 18.62 3.70 9.61 3.69
2225 19.00 3.15 8.92 3.77
2442 17.82 3.05 9.60 3.75
2386 18.26 3.03 9.24 3.83
66.699 0.529 0.141 0.308 0.060
Co level responses Linear
Quadratic
0.058 0.686 0.086 0.176 0.021
0.015 0.497 0.044 0.144 0.093
Co, cobalt; MMA, methylmalonic acid; HCY, homocysteine. S.E.M.: standard error of mean, where n = 12 per treatment.
3.3. Vitamin B12 , folate, MMA, HCY and glucose Co supplementation at the intermediate level showed highest plasma vitamin B12 concentration and lowest plasma MMA concentration (P<0.05) (Table 4). However, there were no differences in plasma concentrations of folate and HCY. Plasma glucose concentration was linearly increased with incremental additions of Co (P<0.05).
4. Discussion 4.1. Performance Lambs fed the control diet containing 0.086 mg Co/kg DM had a reduced appetite and gained less weight than the supplemented animals, suggesting that the inclusion level of 0.086 mg/kg DM Co was inadequate for normal growth of lambs. It was supported by Johnson et al. (2004) who reported that Omani goats fed with diet containing 0.12 mg Co/kg DM exhibited slower growth rate, dry ruffled hair coats and hepatic lipidosis. Clearly, the present study indicated that maximum growth rate in lambs was achieved at 0.50 mg/kg DM Co supplementation. On consideration of the unchanged F/G with Co levels, the addition of
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0.25–0.50 mg/kg DM Co seems to be optimal for improved performance of lambs, which agrees with recent work in cattle (Stangl et al., 2000). 4.2. Digestibility The results in nutrient digestibility were consistent with the findings of Kadim et al. (2003) who demonstrated that low levels of dietary Co in goats resulted in lower apparent nutrient digestibility compared to goats supplemented with parenteral injections of vitamin B12 . The increased digestibility of NDF and ADF indicate the possible role of Co in fiber digestion. This may largely be due to the increase in activity of fiber digesting bacteria (Tomlinson and Socha, 2003). In contrast, the elevated Co intake (2, 4 and 8 mg/kg DM) had no effects on degradability of DM, OM, NDF and ADF (P>0.05) (Kisidayova et al., 2001). This suggests that the optimum level of dietary Co is critical for improving nutrient utilization, and 0.50 mg supplemental Co/kg DM is recommended based on these results. 4.3. Vitamin B12 , folate, MMA, HCY and glucose Results in the present study showed that control lambs were deficient in Co because the plasma values for vitamin B12 and MMA were below normal level (220 pmol/L) and above the upper limit of normal (5 mol/L), respectively. Supplemental 0.50 mg Co/kg DM seems to be optimum level for enhanced plasma vitamin B12 or reduced MMA concentration, whereas, supplemental 0.25 mg Co/kg DM was enough to keep low plasma HCY concentration in lambs. These values are in agreement with Singh and Chhabra (1995). The result of plasma folate concentration was consistent with Stangl et al. (2000), but contrary to Tiffany and Spears (2005). This was not well understood. Apparently, Co supplementation was beneficial for glucose formation by gluconeogenesis. 5. Conclusion Control diet containing 0.086 mg Co/kg DM was inadequate for achieving optimal growth rate in meat-producing lambs. The level of dietary Co for Chinese Poll Dorset × Small Tailed Han lambs is recommended to be 0.336–0.586 mg Co/kg DM. References AOAC, 1990. Official Methods for Analysis, 15th ed. Association of Official Analytical Chemists, Arlington, VA, pp. 69–90. Cornwell, P.E., Morgan, S.L., Vaughn, W.H., 1993. Modification of a homocysteine in human plasma. J. Chromatogr. 617, 136–139. Johnson, E.H., AI-Habsi, K., Kaplan, E., Srikandakumar, A., Kadim, I.T., Annamalai, K., AI-Busaidy, R., Mahgoub, O., 2004. Caprine hepatic lipidosis induced through the intake of low level of dietary cobalt. Vet. J. 168, 174–179. Kadim, I.T., Johnson, E.H., Mahgoub, O., Srikandakumar, A., AI-Ajmi, D.S., Ritchie, A., Annamalai, K., AIAlhali, A.S., 2003. Effect of low levels of dietary cobalt on apparent nutrient digestibility in Omani goats. Anim. Feed Sci. Technol. 109, 209–216.
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Kennedy, D.G., Cannavan, A., Molloy, A., Harty, F.O., Taylor, S.M., Kennedy, S., Blanchflower, W.J., 1990. Methylmalonyl-CoA mutase (EC5. 4. 99. 2) and methionine synthase (EC2. 1. 1. 13) in the tissues of cobalt–vitamin B12 deficient sheep. Br. J. Nutr. 64, 721–732. Kisidayova, S., Sviatko, P., Siroka, P., Jalc, D., 2001. Effect of elevated cobalt intake on fermentative parameters and protozoan population in RUSITEC. Anim. Feed Sci. Technol. 91, 223–232. McMurray, C.H., Blanchflower, W.J., Rice, D.A., Mcloughlin, M., 1986. Sensitive and specific gas chromatographic method for the determination of methylmalonic acid in the plasma and urine of ruminants. J. Chromatogr. 378, 201–207. NRC, 1985. Nutrient Requirements of Sheep, 6th ed. National Academy Press, Washington, DC, USA. Robertson, J.B., Van Soest, P.J., 1981. The Detergent System of Analysis and its Application to Human Foods. Cornell University, Ithaca, New York. Singh, K.K., Chhabra, A., 1995. Effect of dietary cobalt on ruminal vitamin B12 synthesis and rumen metabolites. J. Nucl. Agric. Biol. 24, 112–116. Stangl, G.I., Schwarz, F.J., Muller, H., Kirchgessner, M., 2000. Evaluation of the cobalt requirement of beef cattle based on vitamin B12 , folate, homocysteine and methylmalonic acid. Br. J. Nutr. 84, 645–653. Tiffany, M.E., Spears, J.W., 2005. Differential responses to dietary cobalt in finishing steers fed corn-versus barley-based diets. J. Anim. Sci. 83, 2580–2589. Tomlinson, D., Socha, M., 2003. More cobalt for mature cows? Feed Int. 8, 20–22. Van Soest, P.J., Roberston, J.B., Lewis, B.A., 1991. Methods for dietary fibre NDF and non-starch polysaccharides in relation to animal nutrition. J. Dairy Sci. 74, 3583–3597. Wang, G.Q., Wang, Y.H., Ren, Y., Wang, J.H., Wang, G., 2003. Investigation report of cobalt deficiency in grazing sheep. J. Jilin Agric. Univ. 25, 203–207.
Short communication
Influence of dietary cobalt on performance, nutrient digestibility and plasma metabolites in lambs R.L. Wang a,b , X.H. Kong a , Y.Z. Zhang a , X.P. Zhu a , Narenbatu a , Z.H. Jia a,∗ a
College of Animal Science and Technology, China Agricultural University, Beijing 100094, PR China b Department of Animal Science, College of Agriculture Science, Guangdong Ocean University, Zhanjiang 524088, PR China Received 30 November 2005; received in revised form 22 June 2006; accepted 2 August 2006
Abstract Two experiments were conducted to study the effects of different inclusion levels of dietary cobalt (Co) on performance, nutrient digestibility and plasma metabolites in lambs. Experiment 1: sixty, 12-week-old weaned lambs (Chinese Poll Dorset × Small Tailed Han) were randomly divided into five groups which were fed with the basal diet containing 0.086 mg Co/kg DM, the basal diet supplied, respectively, with 0.25, 0.50, 0.75 and 1.00 mg Co/kg DM as reagent grade CoSO4 ·7H2 O for 10 weeks. Experiment 2: four lambs from each group in Experiment 1 were randomly allocated to the individual metabolic crates to measure the effects of dietary Co on apparent nutrient digestibility. Average daily gain and average daily feed intake were highest at the intermediate level (total diet Co of 0.586 mg/kg DM) (P<0.05). However, there was no difference in gain/feed. Supplemental 0.50 mg Co/kg DM resulted in highest digestibility of dry matter, organic matter, crude protein and acid detergent fiber and metabolizable energy intake (P<0.05). Neutral detergent fiber digestibility was increased at an increasing rate (P<0.05). Co supplementation at the intermediate level showed highest plasma vitamin B12 concentration and lowest plasma methylmalonic acid concentration (P<0.05), whereas there were no differences in plasma concentrations of folate and homocysteine. Plasma glucose concentration
Abbreviations: Co, cobalt; DM, dry matter; OM, organic matter; ME, metabolizable energy; CP, crude protein; NDF, neutral detergent fiber; ADF, acid detergent fiber; Ca, calcium; P, phosphorus; NaCI, sodium chloride; BW, body weight; ADG, average daily gain; ADFI, average daily feed intake; G/F, gain/feed; EE, ether extract; MMA, methylmalonic acid; HCY, homocysteine ∗ Corresponding author. Tel.: +86 10 62732728; fax: +86 10 62732728. E-mail address: [email protected] (Z.H. Jia). 0377-8401/$ – see front matter © 2006 Elsevier B.V. All rights reserved. doi:10.1016/j.anifeedsci.2006.08.011
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was linearly increased (P<0.05). In conclusion, the growth of Chinese Poll Dorset × Small Tailed Han lambs is compromised when fed a Chinese wildrye hay based diet containing 0.086 mg Co/kg DM. The recommended level of dietary Co for such lambs is between 0.336 and 0.586 mg/kg DM. © 2006 Elsevier B.V. All rights reserved. Keywords: Lamb; Cobalt; Vitamin B12 ; Performance; Nutrient digestibility; Metabolites
1. Introduction Cobalt (Co) is required for the synthesis of vitamin B12 by rumen microorganisms. Vitamin B12 acts as a co-factor for methylmalonyl-CoA mutase and methionine synthase which are important for gluconeogenesis and methionine synthesis. It has been reported that low level of dietary Co can lead to vitamin B12 deficiencies clinically manifested as anemia, inappetence and poor production and biochemically characterized by decrease in the plasma concentration of vitamin B12 , elevations in the plasma concentrations of methylmalonic acid (MMA) and homocysteine (HCY) in ruminants (Kennedy et al., 1990). The currently recommended dietary levels of 0.1–0.2 mg Co/kg dry matter (DM) in sheep (NRC, 1985) are based on observations in grazing animals; recent findings however have demonstrated that 0.10 mg/kg DM of Co intake does not meet the rumen microbial requirements (Kisidayova et al., 2001; Johnson et al., 2004). It was reported that 0.3–0.5 mg Co/kg DM enhanced ruminal microbial activity, fermentation and vitamin B12 synthesis (Singh and Chhabra, 1995). In addition, a higher level of dietary Co has been suggested both in beef cattle (Stangl et al., 2000) and cows (Tomlinson and Socha, 2003) than the recommended data. In northern China, Co is supplied to sheep at 0.1–0.2 mg/kg DM, and clinical cases of vitamin B12 deficiency especially in lambs have been reported (Wang et al., 2003). The optimal dietary levels of Co in lambs have not been properly established, and the effects of dietary Co concentration on sheep nutrient digestibility and metabolic characteristics were not well understood. Therefore, the present study was designed to evaluate the effects of dietary Co level on performance and nutrient utilization in Chinese Poll Dorset × Small Tailed Han sheep. 2. Materials and methods 2.1. Experiment 1 Sixty, 12-week-old weaned lambs (Poll Dorset × Small Tailed Han sheep) were housed in individual wooden pens with slatted floors in an open-sided barn and fed a Chinese wildrye hay based diet which was formulated to meet all nutrient requirements with the exception of Co for lambs (NRC, 1985) (Table 1). The Chinese wildrye hay was offered once a day ad libitum, and the concentrate (500 g/animal) was supplied in two equal amounts at 06:00 h and 16:30 h. All animals had free access to water containing undetectable concentrations of Co (analyzed less than 0.01 mg/kg). After 2 weeks adjustment to the experimental feeding system, lambs were weighed and randomly assigned to 5 treatments with 12 each: control
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Table 1 The composition of the basal diet fed to the lambs Chemical compositiona (g/kg DM)
Feed ingredients (g/kg) hayb,c
Chinese wildrye Maize flourc Wheat branc Soybean mealc Cottonseed mealc Limestone Salt Premixd
500 279 120 70 20 2 6 3
Organic matter Metabolizable energy (MJ/kg DM) Crude protein Neutral detergent fiber Acid detergent fiber Calcium Phosphorus Sodium chloride Cobalt (mg/kg DM)
920.3 8.92 139.6 398.0 237.6 3.4 3.4 5.7 0.086
a
Analyzed values except metabolizable energy. Composition of Chinese wildrye hay: ME, 7.84 MJ/kg; CP, 73 g/kg; EE, 36 g/kg; NDF, 575 g/kg; ADF, 328 g/kg; Ca, 2.2 g/kg; P, 1.4 g/kg. c Co content: Chinese wildrye hay, 0.03 mg/kg; maize flour, 0.05 mg/kg; wheat bran, 0.11 mg/kg; soybean meal, 0.48 mg/kg; cottonseed meal, 0.52 mg/kg. d Provided per kilogram of the diet: 40 mg of Zn as ZnSO ·7H O; 25 mg of Mn as MnSO ·H O; 1.0 mg of I 4 2 4 2 as KI; 50 mg of Fe as FeSO4 ·7H2 O; 10 mg of Cu as CuSO4 ·5H2 O; 0.2 mg of Se as Na2 SeO3 ·5H2 O; 1500 IU of Vitamin A; 250 IU of Vitamin D and 16 IU of Vitamin E. b
fed with the basal diet, and 4 supplemental groups which were fed the basal diet and supplied with 0.25, 0.50, 0.75 and 1.00 mg Co/kg DM, respectively. Co was added as CoSO4 ·7H2 O to the premix using finely maize flour as a carrier and mixed with concentrate. The experiment lasted for 70 days. Daily feed offerings and refusals were measured to obtain net feed intake for each animal. At the end of the experiment, the final body weight was measured. The blood samples were collected from each lamb before morning feeding via jugular venipuncture into heparinized vacutainers, then centrifuged at 4 ◦ C for 10 min at 1100 × g and stored at 70 ◦ C pending analyses. 2.2. Experiment 2 Four lambs from each group in Experiment 1 were randomly selected at the end of the feeding period and allocated to individual metabolism crate to study the effects of dietary Co on apparent nutrient digestibility. The digestibility trial lasted for 10 days with 5-day adaptation period and subsequent 5-day collection period during which daily feed intake and output of faeces were collected and recorded. Animals were fed the same diet as in Experiment 1. Representative daily hay and concentrate samples were collected every morning. Faeces from each lamb were collected using a total collection method in plastic bags, then weighted and subsampled. Sulfuric acid was added to the sample to prevent nitrogen loss. Animal feeds and faeces was dried to a constant weight at 70 ◦ C, then ground to pass a 1 mm screen and preserved for further analyses. 2.3. Analytic methods The Co concentrations in feeds were determined using an atomic absorption spectrophotometer (Model 5100, HGA-600 Graphite Furnace; Perkin-Elmer, USA) by absorbance at
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240.7 nm. Feed and faecal samples were analyzed for dry matter (DM), organic matter (OM), crude protein (CP) and ether extract (EE) according to AOAC (1990); neutral detergent fiber (NDF) by Van Soest et al. (1991) without sodium sulfite or alpha amylase; and acid detergent fiber (ADF) by Robertson and Van Soest (1981). CP in feaces was analyzed in dry samples. NDF and ADF are expressed with residual ash. Plasma concentrations of vitamin B12 and folate were determined using a competitive binding radio-immunoassay kit, in which the non-specific vitamin B12 -binding R-protein was removed by affinity chromatography (ICN, Costa Mesa, CA, USA); Plasma MMA concentration by McMurray et al. (1986) through a modified GC method (Agilent Instruments Model 6890, Agilent Technologies, Wilmington, DE). Plasma levels of HCY by Cornwell et al. (1993); and glucose by a commercial kit (Sigma chemical Co., St. Louis, MO). 2.4. Statistical analysis The nature of response to incremental additions of Co was analyzed by polynomial contrasts (SAS Inst. Inc., Cary, NC). The model included linear and quadratic contrast for effects of supplemental Co.
3. Results 3.1. Performance Average daily gain (ADG) and average daily feed intake (ADFI) were highest at the intermediate level (total diet Co of 0.586 mg/kg DM). However, there was no difference in gain/feed (F/G) (Table 2). 3.2. Digestibility Digestibility of DM, OM, CP and ADF and metabolizable energy intake were highest at the intermediate level (P<0.05) (Table 3), but there were no differences in EE digestibility. NDF digestibility was increased at an increasing rate (P<0.05). Table 2 Effects of dietary Co supplementation on performancea of lambs (DM basis) S.E.M.b
Co supplemental levels (mg/kg)
Initial BW (kg) Final BW (kg) ADG (kg) ADFI (kg) G/F (g/g) a
0
0.25
0.50
0.75
1.00
23.57 34.82 0.161 1.000 0.161
23.37 36.09 0.182 1.052 0.173
23.48 37.49 0.200 1.114 0.181
23.37 36.08 0.182 1.099 0.166
23.57 36.06 0.178 1.085 0.165
0.213 0.403 0.005 0.024 0.005
Co level responses Linear
Quadratic
0.978 0.491 0.057 0.137 0.952
0.379 0.250 0.018 0.049 0.300
Co, cobalt; BW, body weight; ADG, average daily gain; ADFI, average daily feed intake; G/F, average daily gain/average daily feed intake. b S.E.M.: standard error of mean, where n = 12 per treatment.
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Table 3 Effects of dietary Co supplementation on apparent nutrient digestibility coefficientsa in lambs (DM basis) S.E.M.b
Co supplemental levels (mg/kg)
DM OM CP EE NDF ADF ME (MJ/d)
0
0.25
0.50
0.75
1.00
0.692 0.715 0.703 0.685 0.502 0.484 8.924
0.704 0.727 0.712 0.688 0.512 0.493 9.381
0.712 0.731 0.715 0.702 0.520 0.501 9.935
0.711 0.730 0.715 0.703 0.525 0.503 9.805
0.711 0.731 0.715 0.694 0.522 0.507 9.678
0.004 0.004 0.003 0.003 0.002 0.002 0.213
Co level responses Linear
Quadratic
0.070 0.099 0.078 0.250 0.033 0.055 0.137
0.020 0.046 0.041 0.204 0.008 0.006 0.049
a Co, cobalt; DM, dry matter; OM, organic matter; CP, crude protein; EE, ether extract; NDF, neutral detergent fiber; ADF, acid detergent fiber; ME, metabolizable energy. b S.E.M.: standard error of mean, where n = 4 per treatment.
Table 4 Effects of dietary Co supplementation on plasma metabolitesa in lambs S.E.M.b
Co supplemental levels (mg/kg)
Vitamin B12 (pmol/L) Folate (nmol/L) MMA (mol/L) HCY (mol/L) Glucose (mmol/L) a b
0
0.25
0.50
0.75
1.00
202 17.34 5.93 11.62 3.59
1688 18.62 3.70 9.61 3.69
2225 19.00 3.15 8.92 3.77
2442 17.82 3.05 9.60 3.75
2386 18.26 3.03 9.24 3.83
66.699 0.529 0.141 0.308 0.060
Co level responses Linear
Quadratic
0.058 0.686 0.086 0.176 0.021
0.015 0.497 0.044 0.144 0.093
Co, cobalt; MMA, methylmalonic acid; HCY, homocysteine. S.E.M.: standard error of mean, where n = 12 per treatment.
3.3. Vitamin B12 , folate, MMA, HCY and glucose Co supplementation at the intermediate level showed highest plasma vitamin B12 concentration and lowest plasma MMA concentration (P<0.05) (Table 4). However, there were no differences in plasma concentrations of folate and HCY. Plasma glucose concentration was linearly increased with incremental additions of Co (P<0.05).
4. Discussion 4.1. Performance Lambs fed the control diet containing 0.086 mg Co/kg DM had a reduced appetite and gained less weight than the supplemented animals, suggesting that the inclusion level of 0.086 mg/kg DM Co was inadequate for normal growth of lambs. It was supported by Johnson et al. (2004) who reported that Omani goats fed with diet containing 0.12 mg Co/kg DM exhibited slower growth rate, dry ruffled hair coats and hepatic lipidosis. Clearly, the present study indicated that maximum growth rate in lambs was achieved at 0.50 mg/kg DM Co supplementation. On consideration of the unchanged F/G with Co levels, the addition of
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0.25–0.50 mg/kg DM Co seems to be optimal for improved performance of lambs, which agrees with recent work in cattle (Stangl et al., 2000). 4.2. Digestibility The results in nutrient digestibility were consistent with the findings of Kadim et al. (2003) who demonstrated that low levels of dietary Co in goats resulted in lower apparent nutrient digestibility compared to goats supplemented with parenteral injections of vitamin B12 . The increased digestibility of NDF and ADF indicate the possible role of Co in fiber digestion. This may largely be due to the increase in activity of fiber digesting bacteria (Tomlinson and Socha, 2003). In contrast, the elevated Co intake (2, 4 and 8 mg/kg DM) had no effects on degradability of DM, OM, NDF and ADF (P>0.05) (Kisidayova et al., 2001). This suggests that the optimum level of dietary Co is critical for improving nutrient utilization, and 0.50 mg supplemental Co/kg DM is recommended based on these results. 4.3. Vitamin B12 , folate, MMA, HCY and glucose Results in the present study showed that control lambs were deficient in Co because the plasma values for vitamin B12 and MMA were below normal level (220 pmol/L) and above the upper limit of normal (5 mol/L), respectively. Supplemental 0.50 mg Co/kg DM seems to be optimum level for enhanced plasma vitamin B12 or reduced MMA concentration, whereas, supplemental 0.25 mg Co/kg DM was enough to keep low plasma HCY concentration in lambs. These values are in agreement with Singh and Chhabra (1995). The result of plasma folate concentration was consistent with Stangl et al. (2000), but contrary to Tiffany and Spears (2005). This was not well understood. Apparently, Co supplementation was beneficial for glucose formation by gluconeogenesis. 5. Conclusion Control diet containing 0.086 mg Co/kg DM was inadequate for achieving optimal growth rate in meat-producing lambs. The level of dietary Co for Chinese Poll Dorset × Small Tailed Han lambs is recommended to be 0.336–0.586 mg Co/kg DM. References AOAC, 1990. Official Methods for Analysis, 15th ed. Association of Official Analytical Chemists, Arlington, VA, pp. 69–90. Cornwell, P.E., Morgan, S.L., Vaughn, W.H., 1993. Modification of a homocysteine in human plasma. J. Chromatogr. 617, 136–139. Johnson, E.H., AI-Habsi, K., Kaplan, E., Srikandakumar, A., Kadim, I.T., Annamalai, K., AI-Busaidy, R., Mahgoub, O., 2004. Caprine hepatic lipidosis induced through the intake of low level of dietary cobalt. Vet. J. 168, 174–179. Kadim, I.T., Johnson, E.H., Mahgoub, O., Srikandakumar, A., AI-Ajmi, D.S., Ritchie, A., Annamalai, K., AIAlhali, A.S., 2003. Effect of low levels of dietary cobalt on apparent nutrient digestibility in Omani goats. Anim. Feed Sci. Technol. 109, 209–216.
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