Probing metal-complexes with metallothioneins by reversed phase microbore chromatography and capillary zone electrophoresis coupled with inductively coupled plasma and electrospray mass spectrometry.

Four different hyphenated techniques: microbore reversed phase (RP) HPLC-ICP MS, CZE-ICP MS, RP HPLC-ES MS and CZE-ES MS were investigated for the characterization of metallothionein-metal complexes under neutral pH conditions. Particular attention was given to the differentiation between metallothionein and artifact signals, identification of mixed-metal complexes, and the validity of the molecular mass as the identification parameter of the different MT iso- and sub-isoforms. Despite the similar morphology of chromatograms and electrophoregrams mass spectrometry revealed different origin of the apparently corresponding peaks. The performance of the four above mentioned techniques was characterized using the example of rabbit liver MT-1 preparation. Reversed-phase HPLC with post-column acidification prior to ES MS was judged to be the most versatile technique for the characterization of metal complexes with metallothioneins but other techniques offer valuable auxiliary information.