Cobalt metabolism in horse. Serum level and biosynthesis of vitamin B12.

The levels of serum vitamin B₁₂ were determined on 16 mature partly warm-blooded, partly Finnish rural-race horses by the radioisotopic competitive inhibition assay method. The mean value from three samplings carried out in dupli- or triplicate was 1.54 ± 0.16 ng/ml. The utilization of serum inorganic cobalt for cyanocobalamin synthesis was studied on two geldings, which received a dose of 200 µCi ⁵⁸CoGl₂ i.v. A Sephadex G-100 gel filtration was carried out with the serum proteins from serial blood samplings at different time intervals 15 min. to 48 hrs. after administration. The gel filtration showed the presence of two labelled proteins in the serum, one of them appearing some time after administration and disappearing almost completely towards the end of the experimental period. The two elution peaks are considered to represent inorganic ⁵⁸Co and ⁵⁸Co labelled vitamin B₁₂. The appearance of labelling in serum vitamin B₁₂ indicates the passing of cobalt into the intestine, and reabsorption into blood in the form of vitamin B₁₂.     

A critical evaluation of serum methylmalonic acid and vitamin B12 for the assessment of cobalt deficiency of growing lambs in New Zealand

AIM: To derive reference ranges for serum methylmalonic acid (MMA) for the diagnosis of cobalt/vitamin B₁₂-responsiveness in lambs and critique existing serum vitamin B₁₂ reference ranges.

METHODS: Individual animal data from earlier supplementation trials, involving 225 ewes, 106 suckling lambs, 301 lambs during the suckling and post-weaning periods and 414 weaned lambs, for which weight gain to supplementation was observed, were used to derive relationships between serum vitamin B₁₂ and MMA, and liveweight gain.

RESULTS: Serum MMA concentrations were rarely elevated above the norm of <2 µmol/L when serum vitamin B₁₂ concentrations were >375 pmol/L, and not elevated into the range where a liveweight response to supplementation occurred (>10 µmol/L) unless serum vitamin B₁₂ concentrations were below 200 pmol/L. Suckling lambs were able to maintain high growth rates despite elevated serum MMA concentrations (>20 µmol/L).

CONCLUSIONS: The current reference ranges used in New Zealand for serum vitamin B₁₂ are set conservatively high. Serum MMA concentrations appear to allow better differentiation of a responsive condition than vitamin B₁₂ concentrations. Serum MMA concentrations <13 µmol/L indicate responsiveness to supplementation whilst concentrations <7 µmol/L indicate unresponsiveness. In the range 7–13 µmol/L, variation in response was observed and predictability of response is less certain, but supplementation is advisable.

CLINICAL RELEVANCE: The current reference ranges for vitamin B₁₂ responsiveness are conservatively high and lead to over-diagnosis of vitamin B₁₂ deficiency in ill-thriftiness of sheep.     

Growth response to increasing doses of microencapsulated vitamin B12 and related changes in tissue vitamin B12 concentrations in cobalt-de cient lambs

AIM: To investigate growth responses of cobalt-deficient lambs to increasing doses of microencapsulated vitamin B₁₂, and to measure associated changes in serum and liver vitamin B₁₂ concentrations over 243 days.

METHODS: From a flock grazing pastures that had low cobalt (Co) levels (about 0.06 mg Co/kg dry matter), 4-6-week-old lambs (n=137) were assigned to four groups and received either no treatment or a subcutaneous injection of 3.0, 4.5 or 6.0 mg of microencapsulated vitamin B₁₂ on Day 1. At approximately monthly intervals, all lambs were weighed and blood samples were collected from a selection (n=10) of monitor animals, up to Day 243. Liver biopsies were also carried out on the monitor lambs (n=8) on Days 1, 124 and 215.

RESULTS: The vitamin B₁₂-treated lambs grew significantly faster (p<0.001) than untreated animals. Liveweights after 243 days were 28, 45, 45 and 47 kg for the untreated, 3.0, 4.5 and 6.0 mg vitamin B₁₂-treated lambs, respectively. Of the initial group of untreated lambs, 68% had to be removed before the end of the trial because of substantial weight loss, but none of the treated animals were similarly afflicted. Serum vitamin B₁₂ concentrations increased in all vitamin B₁₂-treated lambs, reaching a peak at Day 25, and those of the 4.5 and 6.0 mg vitamin B₁₂-treated lambs remained significantly higher (except at Day 124) than the untreated lambs to Day 187. However, at Day 124, but not Day 215, the liver vitamin B₁₂ concentrations of treated lambs were two to three times higher than those of controls.

CONCLUSIONS: The growth rates of Co-deficient lambs were markedly improved by injection of 3.0, 4.5 or 6.0 mg of microencapsulated vitamin B₁₂, and liveweights were maintained for at least 243 days. Serum vitamin B₁₂ concentrations were related to this growth response; concentrations of <220 pmol vitamin B₁₂/l were associated with a 95% probability that lambs were Co-deficient and would thus respond to Co/vitamin B₁₂ supplementation. Based on these data, the current New Zealand reference criteria for Co deficiency should be reviewed.

CLINICAL SIGNIFICANCE: An injection of 3 mg microencapsulated vitamin B₁₂2 given to lambs at tailing will treat Co deficiency and will increase and maintain liveweights in a flock for up to 8 months.     

Serum Vitamin B12 Levels in Omani Goats

Blood and faecal samples were studied from 200 Omani goats of both sexes and various age groups, within five different regions in Oman. Survey questionnaires were undertaken on each goat. The serum vitamin B12 values ranged from 105 to >2000 pg/ml, with a mean value of 723.1 pg/ml and a 95% confidence interval of 61-1507 pg/ml. Kids in the age group 1-3 months showed significantly lower levels of serum vitamin B12 than the older animals. Varying levels of serum vitamin B12 were found in different regions of Oman and on farms within the same region. Goats maintained on commercial feed showed higher levels of serum vitamin B12 than those maintained on pasture or fed on household leftover food, as commonly practised in Oman. Coccidial counts were highest in animals with lower levels of serum vitamin B12. The results of this study indicate that there is a wide range of serum vitamin B12 levels in Omani goats and suggests that younger animals, with lower levels of serum vitamin B12, might be more susceptible to vitamin B12 deficiencies than older animals.     

Absence of a weight gain response to Vitamin B12 supplementation in weaned dairy heifers grazing pastures of marginal cobalt content

Aim. To obtain information on serum and liver vitamin B₁₂ and urinary methylmalonic acid concentrations as diagnostic tests to predict a weight gain response to supplementation with vitamin B₁₂ in young dairy cattle when grazing pasture of low cobalt content.

Methodology. Forty dairy cattle (12 Friesian, 14 Friesian × Jersey and 14 Jersey) were allocated to two equal sized groups, treated and untreated, based on liveweight. At monthly intervals for 14 months, all animals were weighed, their serum and urine sampled, their liver biopsied and the pasture sampled from the paddocks they were grazing and going to graze. Serum and liver were assayed for Vitamin B₁₂ concentrations. For the first 5 months of the trial, urine was assayed for methylmalonic acid concentrations. Both washed and unwashed pasture samples were assayed for cobalt concentrations.

Results. No weight gain response occurred to Vitamin B₁₂ supplementation in young growing cattle grazing pasture with a cobalt concentration of 0.04-0.06 mg/kg DM. For 5 months of the trial, liver Vitamin B₁₂ concentrations from untreated calves were in the range 75-220 nmol/kg and serum vitamin B₁₂ concentrations were as low as 72 pmol/1. There was no associated growth response to supplementation.

Conclusion. Further trials involving young cattle grazing pastures with cobalt concentrations less than 0.04 mg/kg DM are required to reliably determine liver and serum Vitamin B₁₂ concentrations at which growth responses to Vitamin B₁₂ or cobalt supplementation are likely under New Zealand pastoral grazing conditions.     

Vitamin B12 in the blood and in the liver of normal and of totally gastrectomized pigs

The concentration of vitamin R₁₂ in the blood was studied in five totally gastrectomized pigs followed for 10–18 months, and in four controls followed for 5¼–8 months. The concentration of vitamin B12 in the liver was investigated in three pigs of each group. The content of vitamin B₁₂ in the blood was of the same magnitude in both groups. Thus, removal of the stomach does not compromise the normal presence of this vitamin in the blood. The B₁₂ content in the liver was also unaffected by the operation, whereas it was increased after simultaneous administration of vitamin B₆.     

Reduction of the prevalence of footrot on New Zealand farms by vaccination

AIM: To compare serum analyses of vitamin B₁₂ and methylmalonic acid (MMA) as indices of cobalt/vitamin B₁₂ deficiency in lambs around weaning.

METHODS: Lambs on five properties, considered to be cobalt- deficient, were supplemented with either cobalt bullets, or short- or long-acting vitamin B₁₂ preparations. Blood samples, and in some cases liver biopsies, and liveweights were obtained at monthly intervals. Serum samples were assayed for vitamin B₁₂ and MMA and liver for vitamin B₁₂ concentrations. Pasture cobalt concentrations were measured on three of the properties.

RESULTS: Pasture cobalt concentrations were generally maintained below 0.07 μg/g dry matter (DM) on the properties sampled. Growth responses to supplementation were observed on only 2/5 properties, despite serum vitamin B12 concentrations being within the currently used ’marginal‘ reference range (336–499 pmol/L) for at least 3 months on all properties and in the deficient reference range (0–335 pmol/L) for at least 2 months on all farms except one. Serum MMA concentrations in supplemented lambs were <2 μmol/L, except in those animals sampled 1 month after receiving treatment with a short-acting vitamin B12 injection. Serum MMA concentrations in unsupplemented animals on properties on which no growth response to supplementation occurred generally reached peak levels of between 4 and 7 μmol/L at the nadir of serum vitamin B12 concentration. When a growth response was observed, differences in weight gain between supplemented and unsupplemented lambs occurred as mean serum MMA concentrations increased from 9 to 14 μmol/L. On one property where supplementation commenced before weaning, normal growth rates were maintained despite serum vitamin B12 concentrations of 140 pmol/L and serum MMA concentrations in excess of 40 μmol/L serum.

CONCLUSIONS: The possibility that current serum vitamin B₁₂ references ranges for diagnosis of cobalt deficiency are set too high and lead to over-diagnosis of responsiveness to cobalt/ vitamin B₁₂ supplementation is discussed. The suggestion is made that serum MMA concentrations in excess of 9–14 μmol/L will provide a more reliable diagnostic test for cobalt deficiency. However, there was sufficient variation between properties in the relationships between cobalt concentrations of pasture and serum vitamin B₁₂ or MMA concentrations to require more rigorous testing of the reliability of using serum MMA concentration for this purpose. The possibility that differences in rumen fermentation and therefore propionate and vitamin B₁₂ production could be involved is discussed. The measurement of serum MMA and vitamin B₁₂ appears to be of little value whilst the lamb is still suckling.

CLINICAL SIGNIFICANCE: Serum MMA concentration may offer advantages over serum vitamin B₁₂ concentrations in the diagnosis of a cobalt/vitamin B₁₂ responsiveness in weaned lambs.     

钴和铜对肉用绵羊维生素B_(12)合成、瘤胃发酵及造血机能的影响

为研究不同水平的钴和铜对肉用绵羊维生素B12合成、瘤胃发酵及造血机能的影响,试验选20只体重相近(28.9±1.1)kg、装有永久瘤胃瘘管的杂交一代羯羊,随机均分为5组,定量饲喂对照组日粮,4种不同水平的钴铜(0.3:10、0.3:20、0.6:10和0.6:20)日粮。结果表明:瘤胃维生素B12合成主要取决于钴的摄入量,日粮中同时添加钴、铜及其不同的配比对维生素B12合成未显协同效应和影响。钴与铜合用可增加瘤胃挥发性脂肪酸的浓度,并改变乙酸、丙酸和丁酸比例,用量越多其影响越大。随肉羊日粮中钴与铜合用及其用量增加,可更有效地促进铁的吸收和利用,从而更加明显地改善机体的造血机能。     

牛羊4种寄生虫病联合诊断技术的研究与应用

疫区牛羊同时寄生有捻转血矛线虫、日本血吸虫、伊氏锥虫和肝片吸虫等多种寄生虫。目前血清学诊断方法对这 4种寄生虫病的检测需 2～ 4次操作。将捻转血矛线虫、日本血吸虫、伊氏锥虫和肝片吸虫抗原分别以 0 1 5 ,0 0 4 ,0 0 6和 0 1 3μg/ μL的最佳浓度依次定点在同一硝酸纤维素膜条上 ,制成 4种寄生虫病联合诊断膜。用黄牛IgG ,水牛IgG和山羊IgG联合免疫兔 ,获得了高效价兔抗黄牛、水牛和山羊 (简称兔抗牛羊 )IgG抗血清。应用兔抗牛羊IgG抗血清连接酶标SPA ,建立了 1份血纸能同时用于检测牛、羊捻转血矛线虫、日本血吸虫、伊氏锥虫和肝片吸虫病抗体技术。经浙江、湖北、四川、安徽等省应用 ,该项技术具有省工、省时、经济、实用等优点。适宜于牛、羊捻转血矛线虫、日本血吸虫、伊氏锥虫和肝片吸虫病流行区的普查或监测     

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.     

The effect of low levels of dietary cobalt on the chemiluminescence response of polymorphonuclear leukocytes of goats

Twenty ten-week-old newly weaned male Batinah goats were randomly assigned to a control (n = 10) and a treated (n = 10) group and were fed a diet containing 0.1 mg/kg DM cobalt (Co). Goats in the treated group received bi-monthly subcutaneous injections of 2000 μg of hydroxycobalamin. The phagocytic function of the polymorphonuclear leukocytes (PMN) were tested using a luminol-dependent chemiluminescence assay with opsonized zymosan as the phagocytic target. One month after the onset of the experiment PMN from the control group exhibited a significantly (p < 0.05) lower CL response, which continued for the second month. The results of the present study demonstrated that low levels of dietary cobalt leads to an early impairment of phagocytic function. This may at least in part, be an explanation as to why at the field level in Oman young goats fed diets containing low levels of Co appear to be more susceptible to infections.     

Investigation on the growth of coprophagy-prevented rats with supplemented vitamin B12

The growth of coprophagy-prevented rats was compared under administration of normal levels of vitamin B12 and supplemented amounts. Two experiments in which supplemented amounts of vitamin B12 were administered were conducted under different conditions. Six rats per group were fed under coprophagy-allowed (conventional feeding) and coprophagy-prevented conditions respectively. In the first experiment, coprophagy-prevented rats were fed only feed containing recommended vitamin B12 level and forced fed hydrous faeces, vitamin B12 and folic acid respectively. In the second experiment, coprophagy-prevented rats were fed AIN-93G at the recommended vitamin B12 level (25 microg/kg diet), at 100 times the level and at 1000 times the level respectively. Body weight, feed consumption and amounts of each faeces type were determined in both experiments. In a comparison of body weight gain, we learned that coprophagy prevention reduced the values, but that there was no significant difference in the forced feeding group in the first experiment. Similar results were recognized in the second experiment. Vitamin B12 supplementation was not able to raise feed intake significantly and hence it obviously was not a severely limiting factor under the respective experimental condition which depressed feed intake.     

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.     

重离子注入VB12生产菌诱变高产菌株

维生素B12生产菌2009-106株经重离子50 Gy诱变处理得106-8株,其摇瓶单位较出发菌株2009-106提高了31%,对其发酵培养基优化后,应用于120吨发酵罐生产,明显提高产量。     

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]).