SEC ICP MS and CZE ICP MS investigation of medium and high molecular weight complexes formed by cadmium ions with phytochelatins

Abstract

Size-exclusion chromatography (SEC) and capillary zone electrophoresis (CZE) coupled with inductively coupled plasma mass spectrometry were applied to characterize low, medium, and high molecular weight cadmium complexes with glutathione and phytochelatins (PCs). The dominant stoichiometry of the complexes formed in vitro was established as 1:1 using electrospray ionization mass spectrometry. Calculated molecular masses of Cd1L1 complexes were used for calibration of the SEC and CZE methods. The results showed a lower (2 kDa) SEC column exclusion limit for cadmium complexes compared with free peptides (10 kDa), and most of the high molecular weight cadmium species were eluted in the void volume of the column. Moreover, the CZE method based on the semiempirical model of Offord to elucidate peptide migration allowed us to show a high propensity of Cd–PC complexes for polymorphism on complexation, which was also observed for extracts of Arabidopsis thaliana treated with cadmium. All the information presented is vital for understanding the mechanism of metal deactivation in plants.FigureEstimation of molecular mass of Cd-thiopeptide complexes by size electrophoretic mobilityElectronic supplementary materialThe online version of this article (doi:10.1007/s00216-013-6868-3) contains supplementary material, which is available to authorized users.

A novel method for the simultaneous analysis of seven biothiols in rice (Oryza sativa L.) using hydrophilic interaction chromatography coupled with electrospray tandem mass spectrometry

“…Such attempts have been previously reported, such as capillary electrophoresis with electrochemical detection (ED) [16], size-exclusion chromatography followed by ICP-MS detection [17] and electrospray ionization (ESI)-MS/MS using reverse-phase columns [18][19][20]. Unfortunately, these methods usually either have low sensitivity, or are not considered suitable for quantification of Cys and other thiols which are not retained on reverse-phase columns, because of severe ion suppression caused by matrix components in ESI-MS/MS [18], thereby greatly limiting their application in practice.…”

Effects of changes in intracellular iron pool on AlkB-dependent and AlkB-independent mechanisms protecting E.coli cells against mutagenic action of alkylating agent

“…This might be explained in two ways, either iron was bound by protein with molecular mass higher than 500 kDa (see supplementary electronic data for calibration of SEC column, Fig. 3S) or stability of iron cytosolic complexes was lower than in the case of other strains leading to creation of high molecular weight agglomerates, as often observed for adducts of peptides and proteins containing cysteine [43,44]. The most considerable difference in the chromatogram was the absence of the peak at 18.5 ml.…”

Metal complexes of amino acids and peptides

“…298 Peptides of Cys are among the most powerful ligands of soft metal ions. [313][314][315] Further studies in this subject provide information on the role of phytochelatins in cadmium detoxification 316 and on the comparison of the metal binding affinities of phytochelatin and its derivatives prepared by protease catalysis. 299 The thiolate residues were separated by alkyl or aryl chains and the effect of increasing distance between the Cys residues was evaluated.…”