Applying a Metallomics Tool to Probe Interactions between Cadmium, Mammalian Blood Plasma Constituents and Chelating Agents
Creators: name: Zeini Jahromi, Elham
Publisher: name: Graduate Studies
Year: 2013
Subjects: subject: Analytical
subject: Cadmium
subject: Plasma proteins
subject: Chelating agent

description: The analysis of biological fluids in a top-down manner can be employed to gain new insight into the bioinorganic chemistry of toxic metals. To this end, a metallomics method has been previously developed to determine all major blood plasma Cu, Fe and Zn metalloproteins based on the direct analysis of plasma by size exclusion chromatography coupled on-line to an inductively coupled plasma atomic emission spectrometer (SEC-ICP-AES). In order to assess the viability of this metallomics method, the effect of various mobile phase buffers on the distribution of Cu, Zn and Fe metalloproteins was studied using rabbit plasma. 100 mM 3-(N-morpholino)propanesulfonic (Mops), 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (Hepes) and tris(hydroxymethyl)-aminomethane (Tris)-buffers perturbed the Fe and/or Zn metalloproteome, whereas phosphate-buffered saline (PBS)-buffer produced results that were most consistent with literature data. This metallomics method was subsequently employed to probe the dose-dependent interaction of Cd2+ (0.05-200 µg/mL) with rabbit and human blood plasma metalloproteins in vitro. At 200 µg Cd/mL, Cd2+ displaced Zn2+ from a 150 kDa protein which likely corresponds to extracellular Cu, Zn superoxide dismutase (Cu, Zn-SOD) based on previous studies by others. In addition, this methodology was used to assess the efficiency of the abstraction of Cd2+ from plasma proteins (2.0 µg Cd/mL plasma) by three different chelating agents, meso-2,3-dimercaptosuccinic acid (DMSA), diethylenetriamine-pentaacetic acid (DTPA) and 2,3-dimercapto-1-propanesulfonic acid (DMPS) in vitro using pharmacologically relevant doses. The unique capability to simultaneously observe the size distribution of Cd2+ and the essential elements Zn2+ and Ca2+ in plasma revealed that Na5DTPA was the most efficient in terms of abstracting Cd2+ from plasma proteins (100% removal, intended), but it also mobilized Zn2+ from plasma proteins and resulted in the dose-dependent formation of a [Ca-DTPA]3- complex (unintended). In order to minimize these unintended effects, the effect of the Zn salt of DTPA (ZnNa3DTPA) on the mobilization of Cd2+ from plasma proteins was investigated. ZnNa3DTPA was as efficient as Na5DTPA in abstracting Cd2+ from plasma proteins, but minimal or no effect on the distribution of Zn and Ca were observed. Thus, in vitro studies combined with the analysis of blood plasma by SEC-ICP-AES revealed an improved selectivity of a chelating agent. This inherent advantage of this metallomics tool can be used to develop more effective and selective chelating agents to mobilize Cd2+ and other toxic metals from afflicted organisms. Finally, the complexes that are formed between Cd2+ and the chelating agents DMSA and DMPS in aqueous solution at pH 7.4 were structurally characterized by using a combination of X-ray absorption spectroscopy (XAS), SEC-ICP-AES, electrospray ionization-mass spectrometry (ESI-MS) and density functional theory (DFT). The results indicate a complex chemistry in which multi-metallic forms are important, but are consistent with both DMPS and DMSA acting as true chelators, using two thiolates for DMPS and one thiolate and one carboxylate for DMSA.