Analytical interference in serum iron determination reveals iron versus gadolinium transmetallation with linear gadolinium-based contrast agents

Abstract

Objectives: The purposes of this study were to evaluate the risk for analytical interference with gadolinium-based contrast agents (GBCAs) for the colorimetric measurement of serum iron (Fe³⁺) and to investigate the mechanisms involved.

Materials and methods: Rat serum was spiked with several concentrations of all molecular categories of GBCAs, ligands, or "free" soluble gadolinium (Gd³⁺). Serum iron concentration was determined by 2 different colorimetric methods at pH 4.0 (with a Vitros DT60 analyzer or a Cobas Integra 400 analyzer). Secondly, the cause of interference was investigated by (a) adding free soluble Gd³⁺ or Mn²⁺ to serum in the presence of gadobenic acid or gadodiamide and (b) electrospray ionization mass spectrometry.

Results: Spurious decrease in serum Fe³⁺ concentration was observed with all linear GBCAs (only with the Vitros DT60 technique occurring at pH 4.0) but not with macrocyclic GBCAs or with free soluble Gd³⁺. Spurious hyposideremia was also observed with the free ligands present in the pharmaceutical solutions of the linear GBCAs gadopentetic acid and gadodiamide (ie, diethylene triamine pentaacetic acid and calcium-diethylene triamine pentaacetic acid bismethylamide, respectively), suggesting the formation of Fe-ligand chelate.Gadobenic acid-induced interference was blocked in a concentration-dependent fashion by adding a free soluble Gd³⁺ salt. Conversely, Mn²⁺, which has a lower affinity than Gd³⁺ and Fe³⁺ for the ligand of gadobenic acid (ie, benzyloxypropionic diethylenetriamine tetraacetic acid), was less effective (interference was only partially blocked), suggesting an Fe³⁺ versus Gd³⁺ transmetallation phenomenon at pH 4.0. Similar results were observed with gadodiamide. Mass spectrometry detected the formation of Fe-ligand with all linear GBCAs tested in the presence of Fe and the disappearance of Fe-ligand after the addition of free soluble Gd³⁺. No Fe-ligand chelate was found in the case of the macrocyclic GBCA gadoteric acid.

Conclusions: Macrocyclic GBCAs induced no interference with colorimetric methods for iron determination, whereas negative interference was observed with linear GBCAs using a Vitros DT60 analyzer. This interference of linear GBCAs seems to be caused by the excess of ligand and/or an Fe³⁺ versus Gd³⁺ transmetallation phenomenon.

FIGURE 1

Effects of GBCAs and Gd acetate (0.05–5.0 mM in rat serum), Ca-DTPA-BMA (0.0025–0.25 mM), and DTPA (0.0001–0.01 mM) on the iron concentration determined by the colorimetric method with a Vitros DT60 analyzer (percentage recovery) (n = 2, in duplicate).

FIGURE 2

Effects of adding Gd acetate (0.01–5.0 mM in the rat serum) or Mn chloride (1.25 and 5.0 mM) in the presence of gadobenic acid (1.25 mM) on the iron concentration determined by the colorimetric method with a Vitros DT60 analyzer (magnitude of interference) (in duplicate).

FIGURE 3

Effects of adding Gd acetate (2.0 mM) or Mn chloride (2.0, 2.5, 5.0, or 10.0 mM) in the presence of Ca-DTPA-BMA (2.0 mM) on the iron concentration determined by the colorimetric method with a Vitros DT60 analyzer (magnitude of interference) (in duplicate).

FIGURE 4

The ESI/MS spectra for gadodiamide solutions with a zoom for Fe-DTPA-BMA detection (C₁₆H₂₉N₅O₈, the molecular formula for DTPA-BMA ligand).