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Vitamin B12

Vitamin B12, which is also called cyanocobalamin, is a coenzyme synthesized by microorganisms that is required for nucleic acid synthesis. It is released after protein digestion and bound to intrinsic factor in the stomach. The intrinsic factor‑vitamin B12 complex is then rapidly absorbed in the ileum. Once absorbed, Vitamin B12 is transported in the blood by transcobalamin 1, which is released by granulocytes. The daily requirement for Vitamin B12 is approximately 2 to 5 ug and the total body stores are approximately 2 to 5 mg.

Vitamin B12 measurement is useful in the evaluation of patients with macrocytic anemia (MCV >100), motor and/or sensory neuropathies, and unexplained neuropsychiatric symptoms. Some patients with B12 may not present with macrocytic anemia because it is masked by a coexisting disorder such as iron deficiency or thalassemia.

The most common cause of vitamin B12 deficiency is pernicious anemia, which is characterized by a defect in intrinsic factor secretion from the gastric mucosa. Other causes of vitamin B12 deficiency include gastrectomy, bariatric surgery, ileal resection, inflammatory bowel disease, pancreatic insufficiency and atrophic gastritis. Vitamin B12 deficiency secondary to decreased dietary intake is extremely rare, because vitamin B12 stores last between 3 and 6 years. However, it can occur in individuals adhering to a strict vegan diet. A low vitamin B12 level may be associated with folic acid deficiency. Therefore, in evaluation of macrocytic anemia, vitamin B12 levels should be done in conjunction with serum folate levels. Combined deficiencies are rare.

Metformin is an oral hypoglycemic medication that improves insulin action and is used to treat diabetes. Metformin reduces intestinal absorption of vitamin B12 in up to 30 percent of patients and lowers serum vitamin B12 concentrations in 5 to 10 percent, but only rarely causes megaloblastic anemia. The dose and duration of use of metformin correlates with the risk of vitamin B12 deficiency (Arch Int Med 2006; 166:1075).

Vitamin B12 and folate are required for metabolism of homocysteine to methionine, whereas vitamin B12 but not folate is required for metabolism of methylmalonyl–coenzyme A (methylmalonic acid linked to coenzyme A) to succinyl–coenzyme A. Thus, isolated folate deficiency can produce an elevation in the homocysteine level but not in the methylmalonic acid level. In contrast, isolated vitamin B12 deficiency can produce an elevation in both the methylmalonic acid level and the homocysteine level

Laboratory evaluation of vitamin B12 deficiency includes measurement of serum vitamin B12 (cobalamin) and serum folate concentrations, measurement of methylmalonic acid and homocysteine in patients with low normal vitamin B12 levels, and antibodies to intrinsic factor for the diagnosis of pernicious anemia. In patients with anemia, the peripheral blood smear should be examined for macrocytic red blood cells (MCV >100 fL) and hypersegmented neutrophils containing more than 5 lobes per nucleus. The probability of folate and/or Vitamin B12 deficiency is <25 percent when the MCV is normal (ie, 80 to 100 fL). Bone marrow examination for megaloblastic erythropoiesis is usually unnecessary.

Low vitamin B12 concentrations are not necessarily indicative of deficiency. Low B12 levels may be due to granulocytopenia, because transcobalamin 1 is released from granulocytes. Serum vitamin B12 concentrations commonly fall during pregnancy, but these patients do not exhibit hematologic evidence of deficiency.

Serum vitamin B12 levels can fluctuate significantly over time in the same individual. Median intraindividual variation is 23 percent (range: 0 to 119 percent). Absolute differences of more than 100 pmol/L have been seen in as many as 20% of patients during repeat testing.

High levels of intrinsic factor blocking antibodies may cause spuriously high values for vitamin B12 with some immunoassays. Testing for the presence of IF-blocking antibodies is recommended if vitamin B12 results do not support the clinical impression.

Laboratories use different methods to measure vitamin B12. As a result, there are many different normal ranges. Although vitamin B12 assays have not been standardized, results can generally be interpreted using the following guidelines.

  • Greater than 300 pg/mL or >221 pmol/L is a normal result and probability of Vitamin B12 deficiency is less than 5%
  • 200 to 300 pg/mL or 148 to 221 pmol/L is a borderline result and vitamin B12 deficiency is possible.
  • Less than 200 pg/mL or less than148 pmol/L) is low and consistent with vitamin B12 deficiency

Serum B12 levels less than 400 pg/mL are considered suboptimal because 5% to 10% of patients with neurologic symptoms related to B12insufficiency present with levels from 200 to 400 pg/mL. Neurologic symptoms, most commonly parasthesias, may be observed at serum levels as high as 280 mg/mL and often precede hematological abnormalities, such as macrocytic anemia or hyper- segmented neutrophils

Patients with vitamin B12 values at the lower end of the normal range may be truly deficient. Serum concentrations of homocysteine and methylmalonic acid are elevated in functional vitamin B12 deficiency due to a decreased rate of metabolism. Measurement of homocysteine and methylmalonic acid is helpful in diagnosing which patients with borderline vitamin B12 results are likely to respond to therapy.

Serum or plasma collected in EDTA is acceptable.  Heparin plasma should not be used because heparin binds to the vitamin. Also, the lithium salt of heparin may stimulate the release of vitamin B12 binding proteins from granulocytes, which interfere with the assay. Use of serum separator tubes may be associated with spuriously elevated serum vitamin B12 concentrations in some assays. The effect of sunlight on vitamin B12 concentrations in plasma is controversial. Contradictory studies have been published regarding the stability of vitamin B12 in samples exposed to sunlight.

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