Free and Total Vitamin B12 in Cerebrospinal Fluid

Free and total vitamin B₁₂ levels in serum and cerebrospinal fluid (CSF) were bioassayed, since there were no available data on the relationship between free and total vitamin B₁₂ in CSF or between free vitamin in serum and CSF vitamin B₁₂. The subjects were 43 neurological patients. Serum levels were normal in 40 of 43 patients. Values for free and total vitamin B₁₂ in CSF were the same in 42 of 43 patients. Mean CSF vitamin B₁₂ was 21 μμg./ml. In 17 cases CSF vitamin B₁₂ equalled free vitamin B₁₂ level in serum, in 16 cases CSF vitamin B₁₂ was lower than the free level in serum, and in 10 cases CSF vitamin B₁₂ was higher than the free vitamin level in serum. There was no apparent diagnostic correlation. The findings suggest that vitamin B₁₂ is not bound in CSF and that there is some selective control of passage of vitamin B₁₂ across the blood-CSF barrier.     

Vitamin B12 Status in Pregnancy among Immigrants to Britain

Haemoglobin, serum vitamin B₁₂, and serum and red cell folate levels have been measured in 322 pregnant immigrant women in London at their first booking and in a proportion at 34 weeks of gestation and postnatally. The Indian, East-African Indian, and Pakistani and Bangladeshi patients showed significantly lower initial mean serum vitamin B₁₂ levels than the European group, the levels being lower in Hindu and Sikh patients than in Moslems. The patients of West Indian, Indian, and East-African Indian origin showed significantly lower initial mean haemoglobin levels than the immigrants from European countries. Though there was no overall correlation between haemoglobin and serum vitamin B₁₂ level the incidence of hypersegmented polymorphs and macrocytosis in the peripheral blood was highest in the Indian and East-African Indian patients, and both these features were particularly frequent in patients with subnormal serum vitamin B₁₂ levels. Only one patient, however, had overt megaloblastic anaemia due to vitamin B₁₂ deficiency. The Indian patients whose red cell folate levels were less than 200 ng/ml also had a lower mean serum vitamin B₁₂ level than those with red cell folate levels greater than 200 ng/ml. The Indian patients had smaller babies than the Europeans but this was not related to the differences in vitamin B₁₂ status between the two groups. However, out of 39 babies of the Indian group 5 (13%) showed subnormal serum vitamin B₁₂ levels in the first 10 days of life, the lowest level being 120 pg/ml.Though there was an overall statistically significant fall in serum vitamin B₁₂ between first booking and 34 weeks of pregnancy there was no significant fall in serum vitamin B₁₂ in those who initially had subnormal levels. Thus many Indian women are vitamin B₁₂ deficient in pregnancy, and this is associated with morphological blood abnormalities in many cases, but megaloblastic anaemia due to this deficiency is relatively infrequent.     

Serum Folate and Vitamine B12 Levels in Acute and Chronic Renal Disease. Effect of Peritoneal Dialysis

Serum folate and vitamin B₁₂ levels have been measured in 32 patients with renal failure. The initial mean serum folate level was raised above normal in seven patients with acute renal failure whereas the mean level in eight patients severely ill from chronic renal failure was significantly lower than normal. Serum folate levels fell during peritoneal dialysis and rose between dialyses in all these patients and also in one patient who was dialysed for acute pancreatitis.The mean serum B₁₂ level was raised in patients with both acute and chronic renal failure, but there was no consistent change in serum B₁₂ level during dialysis.Hypersegmented polymorphs were present in the peripheral blood film of most of the patients with acute or chronic renal failure. Their presence bore no relation to the clinical state, blood urea, serum folate, or serum B₁₂ level of the patients.     

Mouse transcobalamin II metabolism: The effects of antibiotics on the clearance of vitamin B12 from the serum transcobalamin II-vitamin B12 complex and the reappearance of the free serum transcobalamin II in the mouse

The mechanism of the clearance of vitamin B₁₂ from the serum transcobalamin II-vitamin B₁₂ (Tc-II-B₁₂) complex and the reappearance of free Tc-II in mouse have been studied. When a saturating dose of vitamin B₁₂ is given parenterally to normal mice, a portion of the Tc-II-bound vitamin B₁₂ is rapidly cleared and free Tc-II promptly reappears until it reaches a constant level in 6–8 h. The remaining vitamin B₁₂ is cleared slowly from the rest of the Tc-II-B₁₂ complex. In cycloheximide or puromycin-treated mice, free Tc-II fails to reappear and the bound Tc-IIdecreases. Treatment with actinomycin D has no effect on the reappearance of free Tc-II. The probable mechanism of this inhibition is discussed. The results suggest that mouse serum Tc-II has a stable messenger RNA template and a fast turnover. The free Tc-II which reappears in the serum after Tc-II has been saturated with vitamin B₁₂, appears to be newly synthesized.     

Impact of Helicobacter pylori on the development of vitamin B12 deficiency in the absence of gastric atrophy

Background. Cobalamin (vitamin B₁₂) deficiency is associated with Helicobacter pylori infection. This study examined how serum vitamin B₁₂ levels relate to gastric mucosa H. pylori density and histology, and to hematological findings in patients with minimal or no gastric atrophy. A second aim was to confirm that H. pylori eradication therapy increases serum B₁₂. Materials and Methods. Biopsies of the gastric mucosa from a population of dyspeptic patients were graded for level of chronic inflammation, neutrophil activity, atrophy, and H. pylori density. A total of 145 H. pylori‐infected patients with minimal or no atrophy were included in the study. Serum cobalamin level, hemoglobin level, and mean corpuscular volume were measured in the 145 patients before eradication therapy, and in 65 of the subjects after treatment. The hematologic findings before and after eradication therapy and correlations between serum vitamin B₁₂ level and histologic parameters, hematologic findings, and patient age were statistically analyzed. Results. There was no significant correlation between serum cobalamin level and patient age. Before treatment all the histopathological scores were inversely correlated with serum vitamin B₁₂ level (p < .01) on univariate analysis. Only H. pylori density was significantly associated with B₁₂ level on multivariate analysis. Serum hemoglobin and cobalamin levels were significantly increased after treatment, regardless of H. pylori eradication status (p < .001). Conclusion. The findings provide strong evidence that H. pylori infection is associated with cobalamin deficiency, and show that this is true even in patients with nonulcer dyspepsia and minimal or no gastric atrophy.     

Assessment of Deoxyuridine Suppression Test in Diagnosis of Vitamin B12 or Folate Deficiency

Deoxyuridine (dU) suppression tests have been performed on virtually all marrow samples aspirated at this hospital over the past 12 months. Of the 110 samples studied 26 gave abnormal results, and these 26 samples came from patients deficient in either vitamin B₁₂ or folate. The dU suppression test was found to be of particular value in the diagnosis of vitamin B₁₂ or folate deficiency in non-anaemic patients with macrocytosis and equivocal changes in marrow morphology and in patients in whom the serum vitamin B₁₂ or red cell folate levels were within the normal range.     

Smoking in Pregnancy and Vitamin B12 Metabolism

In pregnancy the level of serum vitamin B₁₂ is lower in women who smoke than in non-smokers. This finding occurs independently of social class, parity, or level of haemoglobin. In addition, the mean serum B₁₂ level tends to be less in women who are anaemic and is less in those women who have smaller babies. These findings may be an effect of the cyanide content of tobacco smoke, since cyanide may be detoxified by a mechanism which depletes the stores of vitamin B₁₂ in the body.     

Role of Third Serum Vitamin B12 Binding Protein in Vitamin B12 Transport

A third vitamin B₁₂ binding protein present in normal serum has been shown to participate in transport of labelled vitamin B₁₂ absorbed from the gut. All three vitamin B₁₂ binding proteins in serum were labelled at the same time after oral administration of vitamin B₁₂, implying that “free” vitamin B₁₂ reached the portal blood from the gut mucosa.     

Vitamin B12 Excretion in Patients with Various Skin Diseases

The excretion in the urine of ⁵⁸Co after an oral dose of ⁵⁸Co vitamin B₁₂ given together with intrinsic factor has been found to be reduced in a number of patients with psoriasis, eczema, and other less common dermatoses. There is a correlation between the abnormality and the extent of the rash. A reduced glomerular filtration rate was found in a few of the patients in whom it was measured, and this must have been responsible, at least in part, for the reduced excretion of vitamin B₁₂ in these patients, but abnormal vitamin B₁₂ excretion also occurred in the absence of impaired renal function. Our evidence is insufficient to show whether malabsorption or increased tissue utilization of vitamin B₁₂ was the explanation in other cases. Certainly a number of patients had steatorrhoea, and in these it is most likely that malabsorption was the major factor. In patients without steatorrhoea a lone malabsorption of vitamin B₁₂ cannot be excluded. A decreased serum concentration of vitamin B₁₂ was found in only one of the patients.     

A comparison between different concentrations and sources of cobalt in goat kid nutrition

There have been extensive studies in sheep and cattle considering cobalt (Co) supplementation and its effects on vitamin B₁₂ concentrations in the body. However, there are limited studies on goats. The aim of this study was to compare two different sources of Co (sulfate v. glucoheptonate) at two different concentrations (0.25 and 0.5 mg/kg dry matter) in goat kid nutrition, and to evaluate the effects of these supplements on performance, serum vitamin B₁₂, blood biochemistry and rumen volatile fatty acids. For this purpose, 30 weaned male goat kids were randomly allotted to five treatments. Serum vitamin B₁₂ increased during the trial in the Co-supplemented groups. Co supplementation increased serum glucose concentrations. On day 35, Co-supplemented groups had greater glucose concentrations compared with control. Propionic+iso-butyric acid concentrations increased only in the 0.5 mg Co glucoheptonate treatment (P<0.05). Our results suggest that, despite the two sources of Co proving mostly similar, the main advantage of Co glucoheptonate compared with Co sulfate was in the ruminal synthesis of vitamin B₁₂. However, although providing Co at National Research Council recommendation levels maintained vitamin B₁₂ above or at normal concentrations, Co supplementation of the Co sufficient basal diet increased vitamin B₁₂ and glucose concentrations.     

Effects of maternal vitamin B12 deficiency from end of gestation to weaning on the growth and haematological and immunological parameters in mouse dams and offspring

Vitamin B₁₂-deficiency may induce specific symptoms as neurological alterations and unspecific symptoms such as anaemia and growth retardation. In this study, maternal vitamin B₁₂ deficiency from end of gestation to weaning was evaluated in mouse dams, which was provoked by feeding a vitamin B₁₂-deficient diet. The animals were divided into two groups (control and deficient). The control group received the vitamin B₁₂-deficient diet supplemented with commercial vitamin B₁₂. Compared to the control, the vitamin B₁₂-deficient dams and their offspring showed a significant decrease of body weight (by 20 and 39%, respectively), serum vitamin B₁₂ concentration (by 61 and 67%, respectively), haematological values as haematocrit (25 and 26%, respectively), and IgA producer cells (by 36 and 54%, respectively). In both, vitamin B₁₂-deficient mouse dams and their offspring, histological alterations of small intestine were observed, whereas growth retardation occurred in the offspring only. This experimental murine model allows assessing the incidence of maternal cobalamin deficiency in offspring and would be useful for evaluating novel adjuncts such as functional foods to prevent vitamin B₁₂ deficiency.     

Mise en évidence et évaluation des “moyennes molécules” de la taille de la vitamine B12 présentes dans les liquides biologiques de sujets normaux et de patients urémiques

Determination of “middle molecules” presenting vitamin B₁₂ molecular size in normal and uremic body fluidsUremic solutes with the molecular size of vitamin B₁₂ are assumed to be toxic. An analytical method is proposed to detect and separate these solutes in body fluids using two combined techniques: gel filtration on Sephadex G-15 and ion-exchange chromatography on DEAE-Sephadex A-25. The vitamin B₁₂ molecular size has been localized by ultrafiltration through membranes with a defined cut-off. Normal and uremic body fluids (urine, plasma, hemodialysis fluid) have been separated into 9 ultraviolet-absorbing peaks (a to i) by high-speed gel filtration. Peaks b and e present the molecular size of vitamin B₁₂, 10–15 Å molecular diameter in pH 7 aqueous solution. Peak b, which correlates with uremic neuropathy, is separated into 6 sub-peaks (b₁ to b₆) by ion-exchange chromatography, sub-peak b_4.2 is the only one to correlate with uremic neuropathy. The coefficient of variation in the integrated area of a single peak is 16%. This method gives the chromatographic profile of the vitamin B₁₂ molecular size content from 500 μl of uremic plasma or 100 μl of normal urine within one hour.     

Vitamin B12, homocysteine and carotid plaque in the era of folic acid fortification of enriched cereal grain products

Background

Carotid plaque area is a strong predictor of cardiovascular events. High homocysteine levels, which are associated with plaque formation, can result from inadequate intake of folate and vitamin B₁₂. Now that folic acid fortification is widespread in North America, vitamin B₁₂ has become an important determinant of homocysteine levels. We sought to determine the prevalence of low serum levels of vitamin B₁₂, and their relation to homocysteine levels and carotid plaque area among patients referred for treatment of vascular disease since folic acid fortification of enriched grain products.

Methods

We evaluated 421 consecutive new patients with complete data whom we saw in our vascular disease prevention clinics between January 1998 and January 2002. We measured total carotid plaque area by ultrasound and determined homocysteine and serum vitamin B₁₂ levels in all patients.

Results

The patients, 215 men and 206 women, ranged in age from 37 to 90 years (mean 66 years). Most were taking medications for hypertension (67%) and dyslipidemia (62%). Seventy-three patients (17%) had vitamin B₁₂ deficiency (vitamin B₁₂ level < 258 pmol/L with homocysteine level > 14 μmol/L or methylmalonic acid level > 271 nmol/L). The mean area of carotid plaque was significantly larger among the group of patients whose vitamin B₁₂ level was below the median of 253 pmol/L than among those whose vitamin B₁₂ level was above the median: 1.36 (standard deviation [SD] 1.27) cm² v. 1.09 (SD 1.0) cm²; p = 0.016.

Conclusions

Vitamin B₁₂ deficiency is surprisingly common among patients with vascular disease, and, in the setting of folic acid fortification, low serum vitamin B₁₂ levels are a major determinant of elevated homocysteine levels and increased carotid plaque area.Elevated plasma total homocysteine levels are a strong, graded independent risk factor for stroke and myocardial infarction.^1,2 Mechanisms by which homocysteine may cause vascular disease include a propensity for thrombosis and impaired thrombolysis, increased production of hydrogen peroxide, endothelial dysfunction and increased oxidation of low-density lipoproteins and Lp(a) lipoproteins.³ Folic acid fortification of enriched cereal grain products began in North America in March 1996 and was made mandatory in 1998. Fortification has reduced the number of neural tube defects by half,⁴ which is clearly a beneficial outcome, but so far it has had little impact on cardiovascular mortality.⁵Carotid plaque area is a strong predictor of cardiovascular events.⁶ High homocysteine levels, which are associated with plaque formation, can result from inadequate intake of folate and vitamin B₁₂. Now that folic acid fortification is widespread, vitamin B₁₂ has become an important determinant of homocysteine levels.⁷ We sought to determine the prevalence of low serum levels of vitamin B₁₂, and their relation to homocysteine levels and carotid plaque area among patients referred for treatment of vascular disease since folic acid fortification of enriched grain products.     

Nutritional Anemia and Megaloblastosis in Pregnancy

Macrogranulocytic and/or erythroid megaloblastic bone marrow changes which could not be accurately predicted from the hematologic findings in the blood were present in 25% of 305 mildly to moderately anemic pregnant women attending a public antepartum clinic in Montreal. Iron deficiency was the primary cause of anemia in most instances. Serum folate activity of less than 4.1 ng./ml. and/or serum vitamin B₁₂ levels of less than 100 pg./ml. were present in 90% of the 77 patients having these bone marrow changes, whereas approximately one-third of 228 patients with normoblastic marrow had these low values. Red cell folate did not correlate as well as serum folate activity with bone marrow changes. After treatment with oral folic acid in the range of 0.2 mg. to 0.8 mg., daily, for seven to 14 days, the megaloblastic and macrogranulocytic changes in patients with low serum folate activity and normal serum vitamin B₁₂ values disappeared in 15 of 21 patients. Of five women having both low folate and vitamin B₁₂ values, three failed to respond and two showed only partial improvement after 0.4 mg. of folic acid daily, per os, for 10 days. The average diet of these anemic women was suboptimal in folate and in iron.     

Development and application of an α-face-specific radioimmunoassay for vitamin B12

The first development of an α-face-specific radioimmunoassay for vitamin B₁₂ is described. Sheep, fed a cobalt-deficient diet, and immunized with a conjugate between Co-β carboxypropyl cobalamin and keyhole limpet hemocyanin, were used to produce antisera. The antisera crossreacted with Co-β derivatives of vitamin B₁₂, but did not crossreact with the α-face vitamin B₁₂ analog cobinamide. The antisera were used to develop a sensitive and reproducible radioimmunoassay that was free from contamination with the nonspecific vitamin B₁₂ binding protein, R-protein. Both the radioimmunoassay and measurements of plasma concentrations of methylmalonic acid were applied to the diagnosis of cobalt/vitamin B₁₂ deficiency in sheep. The assay correlated well with a commercially available radioassay and did not falsely detect normal vitamin B₁₂ concentration in plasma samples containing elevated concentrations of methylmalonic acid.     

Vitamin B12 Content of Circulating Leukocytes as an Aid in the Differentiation of the Acute Leukemias

From 25 patients with acute leukemia 116 specimens of leukocytes were assayed microbiologically for total vitamin B₁₂ to determine if variation in vitamin B₁₂ content would help in differentiating the acute leukemias. The mean cell vitamin B₁₂ levels (μμg./10⁸ cells) in the different types of leukemia were: lymphoblastic 464, myeloblastic 1058 and monocytic 200. Cell vitamin B₁₂ levels above the normal range (100-800 μμg./10⁸ cells) are suggestive of myeloblastic leukemia. The only elevated cell vitamin B₁₂ levels comparable to those found in myeloblastic leukemia were in reticulum cell leukemia, and this type of leukemia was not difficult to diagnose morphologically. Blast cells contained more vitamin B₁₂ than mature cells of the same series; there was a significant positive correlation between the percentage of blast cells and cell levels of total vitamin B₁₂ in both lymphoblastic and myeloblastic leukemia.     

Transient Fluctuation of Serum Melatonin Rhythm is Suppressed Centrally by Vitamin B

Vitamin B₁₂ has been reported to improve sleep-wake rhythm disorders. Although the mechanism is still unclear, a change in the sensitivity of the circadian clock system to photic input is thought to be a possible mechanism of the effect. In this study, the effect of the vitamin B₁₂ on the circadian aspect of the electroretinogram (ERG) and serum melatonin level was analyzed in rats. Vitamin B₁₂, α-(5,6-dimethylbenzimidazolyl)-co-methyl-cobamide was daily administrated subcutaneously for 8 weeks to adult male Wister rats in the experimental group, and saline was given to the control group. The ERGs were recorded under dark adaptation during the night and day, and under light adaptation (0.1 lux) during the night. Blood was drawn before and after ERG recording. The amplitudes of the a-wave, fc-wave, and trough-to-peak of both waves and latencies of ERG were analyzed following various exposures to stimuli of light intensity. These parameters in the group treated with vitamin B₁₂ showed similar characteristics to the control group, and no significant difference was observed between the two groups. The melatonin levels of both groups before the measurement of ERG were similar under each measurement condition. The elevated serum melatonin concentration in the control group under dark adaptation at night was suppressed after the series of 10-msec light stimuli used for measurement of ERG. However, this suppressing effect of light pulses on melatonin level was significantly inhibited in the group treated with vitamin B₁₂. Under light adaptation during the night and under dark adaptation during the day, melatonin levels after the measurement of ERG were not different between the groups. From these results, it is suggested that vitamin B₁₂ is effective in suppressing melatonin rhythm disturbances introduced by transient light stimulation, and it affects the site more central than the retinal level. (Chronobiology International, 14(6), 549–560, 1997)     

THE EFFECT OF VITAMIN B12 ON THE ANEMIA AND COMBINED SYSTEM DISEASE OF ADDISONIAN PERNICIOUS ANEMIA

The effect of the parenteral administration of vitamin B₁₂ has been observed in eight patients with Addisonian pernicious anemia.Vitamin B₁₂ in initial doses of 50 micrograms or 25 micrograms induced satisfactory reticulocyte response and a return of erythrocyte count to within normal range in 60 days.In only two of the patients were secondary reticulocyte responses induced on a second injection of vitamin B₁₂.Concurrently with the hemopoietic response, the bone marrow changed from megaloblastic hyperplasia to normoblastic distribution.The paresthesias associated with combined system disease as well as disturbances in position sense and locomotor function may be entirely relieved or greatly diminished following injections of vitamin B₁₂.Maintenance injections of vitamin B₁₂ may be from 30 to 50 micrograms at intervals of one month, the amount depending upon the individual case.Vitamin B₁₂ may be used without untoward symptoms in patients previously sensitive to liver extract.     

Selective vitamin B12 malabsorption in two siblings

Two siblings with megaloblastic anemia responsive to parenteral vitamin B₁₂ were studied to elucidate the cause of the B₁₂ deficiency. Gastric juice from both contained acid and functional intrinsic factor. Serum contained transcobalamin II and lacked antibodies to intrinsic factor. Schilling tests showed vitamin B₁₂ malabsorption uncorrected by hog intrinsic factor or pancreatic extract. Other parameters of small intestinal function were normal. Proteinuria was initially present in both but cleared in one following treatment with B₁₂. These patients with “familial selective vitamin B₁₂ malabsorption” are the first reported from Canada. Only 37 cases have been reported in the world literature to date.     

Higher Prevalence of Metformin-Induced Vitamin B12 Deficiency in Sulfonylurea Combination Compared with Insulin Combination in Patients with Type 2 Diabetes: A Cross-Sectional Study

Long-term and high-dose treatment with metformin is known to be associated with vitamin B₁₂ deficiency in patients with type 2 diabetes. We investigated whether the prevalence of B₁₂ deficiency was different in patients treated with different combination of hypoglycemic agents with metformin during the same time period. A total of 394 patients with type 2 diabetes treated with metformin and sulfonylurea (S+M group, n = 299) or metformin and insulin (I+M group, n = 95) were consecutively recruited. The vitamin B₁₂ and folate levels were quantified using the chemiluminescent enzyme immunoassay. Vitamin B₁₂ deficiency was defined as vitamin B₁₂≤300 pg/mL without folate deficiency (folate>4 ng/mL). The mean age of and duration of diabetes in the subjects were 59.4±10.5 years and 12.2±6.7 years, respectively. The mean vitamin B₁₂ level of the total population was 638.0±279.6 pg/mL. The mean serum B₁₂ levels were significantly lower in the S+M group compared with the I+M group (600.0±266.5 vs. 757.7±287.6 pg/mL, P<0.001). The prevalence of vitamin B₁₂ deficiency in the metformin-treated patients was significantly higher in the S+M group compared with the I+M group (17.4% vs. 4.2%, P = 0.001). After adjustment for various factors, such as age, sex, diabetic duration, duration or daily dose of metformin, diabetic complications, and presence of anemia, sulfonylurea use was a significant independent risk factor for B₁₂ deficiency (OR = 4.74, 95% CI 1.41–15.99, P = 0.012). In conclusion, our study demonstrated that patients with type 2 diabetes who were treated with metformin combined with sulfonylurea require clinical attention for vitamin B₁₂ deficiency and regular monitoring of their vitamin B₁₂ levels.