Measurement of vitamin B12 in the livers and sera of sheep and cattle and an investigation of factors influencing serum vitamin B12 levels in sheep

Modifications of a radioassay method for the analysis of vitamin B12 using chicken serum as the binder are described. This obviates the need to use individual serum blanks to correct for non-specific binding in vitamin B12 assays of the sera and livers of sheep and cattle. Samples with high vitamin B12 levels can be diluted prior to assay without loss of linearity. Recoveries of added cyanocobalamin or hydroxocobalamin were better than 95% and results correlated significantly with those obtained using a microbiological assay (Poteriochromonas malhamensis). Sera and liver samples stored for four weeks at temperatures ranging from -20 degrees to 22 degrees showed no change in vitamin B12 levels. Withholding food from sheep for 44 hours led to a marked increase in serum vitamin B12. This effect was also evident in sheep eating a limited amount of cut grass. In sheep at pasture there was no evidence of a diurnal variation in serum vitamin B12 levels. Serum vitamin B12 levels in sheep at pasture were shown to be an unreliable indicator of liver vitamin B12.     

Reference curves to diagnose cobalt deficiency in sheep using liver and serum vitamin B12 levels

Reference curves demonstrating the relationship between serum or liver vitamin B12 and weight gain were derived from the examination of 16 published and 48 unpublished N.Z. trials. From these curves probability of obtaining an economic reponse (>10g/day body weight increase) for any serum or liver vitamin B12 can be determined. No significant (P<0.05) weight gain responses occurred to vitamin B12 or cobalt treatment in trials with mean serum vitamin B12 levels above 500 pmol/l or liver vitamin B12 levels greater than 500 nmol/kg. The reference curves were therefore derived from trials with vitamin B12 levels below these levels; 36 trials with serum vitamin B12 and 19 trials with liver vitamin B12 data. The mean vitamin B12 level at the mid point of the weight gain response period was selected from each trial. Examination of serum vitamin B12 reference curves for spring, summer, autumn and winter indicated that curves derived from data closest to the middle of January (summer) adequately reflected response to treatment at any time during the first year of life. Reference curves for liver vitamin B12 also used data closest to middle of January. This was partly because insufficient liver data was available to compare seasonal variations. The fitted response curve approached 0 gram/day at 500 pmol/l for serum vitamin B12 and 375 nmol/kg for liver vitamin B12. The minimum vitamin B12 level at which an economic response to treatment (>10 g/day) is not likely was 336 pmol/l for serum and 282 nmol/kg for liver.     

Serum vitamin B12 and methylmalonic acid determinations in the diagnosis of cobalt deficiency in pregnant ewes

Serum vitamin B12 and methylmalonic acid (MMA) concentrations were used to monitor the development of cobalt (Co) deficiency and repletion from the deficient state in housed pregnant hill sheep. Serum MMA concentrations were less variable and provided a more accurate diagnosis of Co deficiency than serum vitamin B12. This was particularly the case for subclinical disease. However, unlike serum MMA, concentrations of the vitamin in serum could be used prognostically. The most precise diagnosis was provided by serum vitamin B12 and MMA data used in conjunction, but where one technique is to be used, serum MMA determinations are preferred.     

Development of a specific radioimmunoassay for vitamin B12 and its application in the diagnosis of cobalt deficiency in sheep

A radioimmunoassay (RIA) for vitamin B₁₂ is described. Antisera were raised in rabbits using a conjugate between 5-O-succinyl cyanocobalamin and chicken serum albumin. The antibody is largely specific for the (upper) face of the vitamin B₁₂ molecule. The RIA can detect vitamin B₁₂ down to concentrations of 37 pmol/L and correlated well (r=0.980, p<0.001) with a commercial radioassay. The use of this RIA, together with an assay for plasma methylmalonic acid, in the diagnosis of cobalt/vitamin B₁₂ deficiency in sheep is described.     

The diagnosis and treatment of vitamin B12 deficiency in young lambs

Abstract

Extract

In the course of some earlier experiments, it was observed that vitamin B₁₂ deficiency in 2 hoggets was corrected by injection of hydroxocobalamin (vitamin B_12b) (E. D. Andrews, pers. comm., 1972). Since this form of therapy offered a convenient and effective means of treating young lambs suffering from vitamin B₁₂ deficiency, more detailed investigations were desirable. It has been known for some time that lambs on the cobalt-deficient area at the Ministry of Agriculture and Fisheries Kaitoke Farm lose condition after weaning. Abnormal amounts of methylmalonic acid (MMA) had been found in the urine of some of these lambs, indicating a state of vitamin B12 deficiency Andrews et al, (1970 Andrews, E. D., Hogan, K. G., Stephenson, B. J., White, D. A. and Elliott, D. C. 1970. Cobalt and thiabendazole live-weight responses in grazing sheep, and their relation to the urinary excretion of methylmalonic acid. N.Z. Jl agric. Res., 13: 950–965. [Taylor &; Francis Online], [Web of Science ®] , [Google Scholar]).     

A comparison of vitamin B12 levels in the liver and serum of sheep receiving treatments used to correct cobalt deficiency

The efficacies of four methods, used for the prophylaxis of cobalt deficiency in sheep as measured by the elevation of liver and serum vitamin B₁₂ levels, were compared in marginally deficient sheep over 14 weeks. The methods used were weekly drenches of either cobalt sulphate or cobalt chelate (EDTA); three-weekly injections of hydroxocobalamin, and ruminal cobalt pellets.

On the basis of elevated liver and serum vitamin B₁₂ levels, chelated cobalt was shown to be available to rumen microflora for the synthesis of vitamin B12. However, at no stage were liver and serum vitamin B₁₂ levels of sheep receiving the chelate significantly different from those receiving the same amount of cobalt as the sulphate.

After five, three-weekly injections of hydroxocabalamin liver vitamin B₁₂ levels were significantly higher (p < 0.01) than for the other treatments, with the exception of cobalt sulphate.

Cobalt pellets led to an initial rapid and significant rise in serum vitamin B₁₂ when compared with the other treatments. However, at four weeks there was no significant difference between treatment groups for serum vitamin B₁₂. Fourteen weeks after the administration of cobalt pellets, serum and liver. vitamin B₁₂ levels in this group were not significantly different from those of untreated sheep. At this time, three out of 12 sheep had lost their pellets.     

A comparison of vitamin B12 levels in the liver and serum of sheep receiving treatments used to correct cobalt deficiency

The efficacies of four methods, used for the prophylaxis of cobalt deficiency in sheep as measured by the elevation of liver and serum vitamin B12 levels, were compared in marginally deficient sheep over 14 weeks. The methods used were weekly drenches of either cobalt sulphate or cobalt chelate (EDTA) three-weekly injections of hydroxocobalamin, and ruminal cobalt pellets. On the basis of elevated liver and serum vitamin B12 levels, chelated cobalt was shown to be available to rumen microflora for the synthesis of vitamin B12. However, at no stage were liver and serum vitamin B12 levels of sheep receiving the chelate significantly different from those receiving the same amount of cobalt as the sulphate. After five, three-weekly injections of hydroxocabalamin liver vitamin B12 levels were significantly higher (p<0.01) than for the other treatments, with the exception of cobalt sulphate. Cobalt pellets led to an initial rapid and significant rise in serum vitamin B12 when compared with the other treatments. However, at four weeks there was no significant difference between treatment groups for serum vitamin B12. Fourteen weeks after the administration of cobalt pellets, serum and liver.vitamin B12 levels in this group were not significantly different from those of untreated sheep. At this time, three out of 12 sheep had lost their pellets.     

Effect of ingestion of soil on the iodine, copper, cobalt (vitamin B12) and selenium status of grazing sheep

AIM: To determine the impact of ingestion of soil on the iodine (I), selenium (Se), copper (Cu) and cobalt (Co; vitamin B12) status of young sheep. METHODS: Twenty young sheep were divided into two groups; one group was fed lucerne pellets, while the other group was fed lucerne pellets plus 100 g soil, for 63 days. At the end of the study the animals were blood-sampled, slaughtered, and the liver removed, and concentrations of I, Cu, vitamin B12 and Se were determined. RESULTS: The ingestion of soil significantly increased concentrations of I and vitamin B12 in serum, but had no effect on concentrations of Cu and Se in serum/blood and liver, and vitamin B12 in liver. CONCLUSION: Ingested soil can be a significant source of I and Co (vitamin B12) for grazing sheep.     

Ceruloplasmin as an indicator of copper status in cattle and sheep.

The relationship between ceruloplasmin, a metalloenzyme with oxidase activity, and copper was investigated in cattle and sheep. The oxidase activity of ceruloplasmin correlated closely with the serum or plasma copper concentrations in cattle. The respective correlation coefficients were 0.83 and 0.60. In sheep serum, a correlation coefficient of 0.92 was obtained. In each instance, the relationship remained linear from the deficient to the high normal ranges of copper. Comparison of the linear regression relationships indicated the ceruloplasmin activity in bovine serum was statistically lower than the activity in bovine plasma (P less than 0.0001), through the intercepts from the regression lines of the two relationships were similar (P = 0.412). Comparisons of ovine and bovine serum-ceruloplasmin relationships indicated that a significant species difference was present. Ovine ceruloplasmin activity increased more rapidly as compared to the corresponding bovine activity over the range of copper concentrations investigated (P less than 0.0001). The intercept from the ovine regression relationship was also lower (P less than 0.0001). A correlation coefficient of 0.35 was observed between the serum ceruloplasmin activity and hepatic copper concentrations in cattle indicating that the mathematical relationship was not as well defined. Ceruloplasmin activity appears to correlate more closely with serum or plasma copper concentrations as compared to corresponding liver copper concentrations.     

The distribution of copper, vitamin B12 and zinc in the livers of sheep

Six livers from one-year-old sheep were each sampled at 27 different sites and the concentrations of copper, vitamin B12 and zinc determined for each sample. The distribution of these components was uniform throughout the liver, indicating that analysis of a single liver biopsy sample would give an accurate assessment of their hepatic concentration.     

The distribution of copper,vitamin B12 and zinc in the livers of sheep

Six livers from one-year-old sheep were each sampled at 27 different sites and the concentrations of copper, vitamin B,, and zinc determined for each sample. The distribution of these components was uniform throughout the liver, indicating that analysis of a single liver biopsy sample would give an accurate assessment of their hepatic concentration.     

Rickets associated with vitamin D deficiency in young sheep

An outbreak of rickets in sheep under a year old (hoggs) appeared clinically as stiffness and rotation of the carpal joints. Histological studies confirmed the diagnosis and biochemical analyses of blood demonstrated a primary vitamin D deficiency.     

维生素B12的研究概况

维生素B12是人和动物体内非常重要的水溶性维生素之一,参与体内一碳基团的代谢,主要起转移甲基的作用。体内缺乏时会导致食欲下降、贫血、表皮炎、毛发粗糙、废绝以致死亡。本文就其近些年的研究情况做一概述,以引起营养研究对这种维生素的关注。维生素B12为粉红色结晶,在弱酸性水溶液中相当稳定。在pH值3以下和9以上则及易分解,日光、氧化剂、或还原剂均易破坏维生素B12的活性。维生素B12的活性是由一组化合物构成的,其中包括一个由四个吡咯核连接而成的卟啉环,其氮原子与分子中心的钴原子相连(McDowell,1989h)。吡咯核连接而成的这种结…