Identification of cadmium-bioinduced ligands in rat liver using parallel HPLC-ICP-MS and HPLC-electrospray MS

An analytical strategy has been developed for the characterization of Cd complexes in rat liver tissue. The sample preparation included the isolation of the Cd-macromolecular fraction by semi-preparative size-exclusion chromatography, followed by a preconcentration step by lyophilization and desalting. The analysis was carried out by microbore reversed-phase HPLC with parallel ICP-MS and ESMS detection. The HPLC-ICP-MS interface was based on a Micromist nebulizer and a low dead volume cyclonic spray chamber. The HPLC-ESMS interface allowed the measurement of the molecular mass of the ligand protein owing to the dissociation of the complex by the on-line post-column acidification of the eluate. The approach allowed the detection of two major metallothionein (MT) isoforms and their identification on the basis of the molecular mass as rat liver MT-1 and MT-2. The actual number of peaks in the chromatograms was larger because of the formation of mixed Cd-Cu complexes of the same MT isoform that showed different hydrophobicities.