Assessment of Vitamin B12 Absorption Based on the Accumulation of Orally Administered Cyanocobalamin on Transcobalamin

Abstract

Background: Vitamin B12, or cobalamin (Cbl), is absorbed in the intestine and transported to the cells bound to transcobalamin (TC). We hypothesize that cyanocobalamin (CNCbl) is absorbed unchanged, thereby allowing measurement of the complex of CNCbl bound to TC (TC–CNCbl) to be used for studying the absorption of the vitamin. Methods: TC was immunoprecipitated from serum samples obtained from healthy donors at baseline and at 24 h after oral administration of three 9-μg CNCbl doses over 1 day. Cbl was released by treatment with subtilisin Carlsberg. The different forms of Cbl were isolated by HPLC and subsequently quantified with an ELISA-based Cbl assay. Results: At baseline, the median TC–CNCbl concentration was 1 pmol/L (range, 0–10 pmol/L); the intraindividual variation (SD) was 1.6 pmol/L (n = 31). After CNCbl administration, the TC–CNCbl concentration increased significantly (P = 0.0003, paired t-test), whereas no major changes were observed in any of the other Cbl forms bound to TC (n = 10). Only a moderate additional increase in TC–CNCbl was observed with prolonged (5 days) CNCbl administration (n = 10). We designed an absorption test based on measuring TC–CNCbl at baseline and 24 h after CNCbl intake and established a reference interval for the increase in TC–CNCbl (n = 78). The median absolute increase was 23 pmol/L (range, 6–64 pmol/L), and the relative increase was >3-fold. Conclusions: Our data demonstrate that CNCbl is absorbed unchanged and accumulates on circulating TC. We suggest that measuring TC–CNCbl will improve the assessment of vitamin B12 absorption.

Forms and Amounts of Vitamin B12 in Infant Formula: A Pilot Study

“…The samples were lyophilized and dissolved in 240 μL PBA-NaOH buffer (120 μl 2M NaOH in 4000 μl 0.1% phosphate buffered albumin) prior to estimating the content of B12. In order to achieve a precise estimate for B12, we used a previously described in-house haptocorrin ELISA [13, 14] with a detection limit of 8 pmol/L B12 rather than the Cobas assay that has a detection limit of 55 pmol/L B12. In brief, 100 μl calibrators or samples were incubated with apo-haptocorrin.…”

The tissue profile of metabolically active coenzyme forms of vitamin B12 differs in vitamin B12-depleted rats treated with hydroxo-B12 or cyano-B12

“…Therefore, we collected fortysix post-column fractions every 20 s (320 µl/fraction) between 9 and 25 min after injection. The samples were lyophilised and dissolved in 240 µl of 0·1 % PBS with 0·1 % bovine serum albumin (PBA) before measurement of B 12 , employing in-house haptocorrin ELISA (15,17) with a detection limit of 8 pmol/l B 12 . In brief, 100 µl (sample or calibrator) was incubated with apohaptocorrin, and excessive apo-haptocorrin was removed with B 12 -coated magnetic beads.…”

Holotranscobalamin, a marker of vitamin B-12 status: analytical aspects and clinical utility

“…On the basis of this observation, we developed a version of the CobaSorb test in which we measure the increase in the amount of cyanocobalamin bound to transcobalamin. We named this version of the test C-Cobasorb (55). Today we recommend the use of CobaSorb to clarify whether a diagnosed vitamin B-12 deficiency is caused by an inability to absorb the vitamin, and recommend the test be judged in the following manner.…”

“…We judge the patient able to absorb vitamin B-12 if holoTC increases >10 pmol/L and >22% after 2 d intake of an oral dose of 3 times 9 μg vitamin B-12 daily. We recommend C-CobaSorb be performed in patients with a baseline holoTC >65 pmol/L and not judged to absorb vitamin B-12 based on the CobaSorb test (55). We stress that currently the CobaSorb tests have been used only to give a “yes” or “no” answer related to the absorption of vitamin B-12.…”