Speciation analysis of trace elements in the brains of individuals with Alzheimer's disease with special emphasis on metallothioneins

Abstract

A speciation analysis of protein-bound elements in the cytosol of human brain was achieved by size exclusion chromatographical separation of the biomolecules and on-line detection of the metal profiles in the eluate by hyphenated inductively coupled plasma-mass spectrometry. Post-mortem samples from Alzheimer's disease brains and from brains of a control group were investigated to elucidate changes in the trace element distribution during the pathological process. Special attention was paid to the metallothioneins (MT) - cysteine-rich, metal-binding proteins of low molecular weight, existing in several isoforms. The isoform MT-3 is found especially in the brain and has a growth inhibition function on neurons. The MT peaks were identified in the element profiles. For this purpose, the metal binding capability and the heat stability of MT were taken into consideration. For verification, a comparison with pure MT-3 was carried out and further biochemical and analytical methods were applied to the fractions of the chromatographical run. A comparison between Alzheimer's disease and control brains showed a significant difference concerning the MT-1/-2 and MT-3 metal levels, leading to the assumption that there were oxidative processes having taken place in the Alzheimer's brain samples.

Identification and quantification of metallothionein isoforms and superoxide dismutase in spiked liver extracts using HPLC-ESI-MS offline coupling and HPLC-ICP-MS online coupling

“…It demonstrates the parallel use of ICP-MS and ESI-MS detection for both well-known commercial preparations and spiked liver extracts with clear assignment of the data from both MS modes. The quantitative data evaluation was especially considered (metal stoichiometry, calibration, recovery, and metal balance) because this aspect is of course vital for the validation of the method; in spite of this, it is neglected in most MT publications with hyphenated techniques [3,21].…”

“…These species are known to be involved in essential or pathological processes like the detoxification of radical oxygen species or heavy metals in cells, but the knowledge in this area is quite limited for many trace elements. Recent advances in speciation analysis have made a variety of promising techniques available including HPLC-ICP-MS, CE-ICP-MS, HPLC-ESI-MS, and CE-ESI-MS. A series of applications for biomolecule characterization with these coupling techniques has been published [1,2,3,4], but few use the powerful combination of molecule-specific and element-specific detection [5,6,7]. Besides this, some methods are developed only with purified preparations of the species and are not applied to real samples.…”

Trace element speciation by ICP-MS in large biomolecules and its potential for proteomics

“…Speciation analysis of MTs and MLPs by HPLC coupled to ICP-MS seems to be adequate for detecting metal-biomolecule complexes in cytosols from animal tissues [48,49,50,51,52,53]. The majority of such studies concern the speciation of Cd, Zn and Cu in kidney [54,55,56], liver [46,49,57,58,59,60,61,62,63,64] and brain [65] of mammals (rat, rabbit, bovine, humans) or tissues of bream [66], carps [67], eels [68] and mussels [69,70,71]. MT isoforms have been separated by size SEC, anionic exchange-HPLC and reverse phase (RP)-HPLC chromatography.…”

Hyphenated techniques for the characterization and quantification of metallothionein isoforms

“…Although SE-HPLC cannot distinguish between MT-1 and MT-2, an MT-3 fraction can be separated from the MT-1/MT-2 fraction. Richarz and Brätter [51] studied the MT-3 isoform in human brain cytosol by using the SE-HPLC-ICP-MS coupling with the aim of comparing the MT-3 contents in the brains of human patients suffering from Alzheimer's disease with those from a control group.…”