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Using a combination of As and Se K-edge and Hg L(III)-edge X-ray absorption spectroscopy, 77Se nuclear magnetic resonance spectroscopy, electrospray ionization mass spectrometry and molecular modeling, we have structurally characterized... more
Using a combination of As and Se K-edge and Hg L(III)-edge X-ray absorption spectroscopy, 77Se nuclear magnetic resonance spectroscopy, electrospray ionization mass spectrometry and molecular modeling, we have structurally characterized the novel species methylmercury(II) seleno bis(S-glutathionyl) arsenic(III). This species is formed in aqueous solution from CH3HgOH and the seleno bis(S-glutathionyl) arsinium ion and constitutes an important first step towards characterizing the observed toxicologically relevant interaction between arsenite, selenite and methylmercury which has been previously reported in mammals.
Research Interests: Chemistry, Inorganic Chemistry, Arsenic, Nuclear Magnetic Resonance, X ray absorption spectroscopy, and 13 moreMethylmercury, Magnetic Resonance Spectroscopy, Medicine, Molecular modeling, Selenium, High Pressure Liquid Chromatography, Liquid Chromatography / Electrospray Ionization Mass Spectrometry, X ray Absorption, Aqueous Solution, Biochemistry and cell biology, Nuclear Magnetic Resonance Spectroscopy, Electrospray Ionization, and Electrospray Ionization Mass Spectrometry
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Research Interests: Biophysics, Chemistry, Medicine, Lipids, Humans, and 14 morePhysical sciences, CHEMICAL SCIENCES, Structure activity Relationship, Monolayer, Erythrocytes, Lipid Bilayer, Phosphatidylcholine, A, Phosphatidylserine, Membrane Fluidity, Mercuric Chloride, π, Medical and Health Sciences, and Cadmium Chloride
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The etiology of numerous grievous human diseases, including Alzheimer's and Parkinson's Disease is not well understood. Conversely, the concentration toxic metals and metalloids, such as As,... more
The etiology of numerous grievous human diseases, including Alzheimer's and Parkinson's Disease is not well understood. Conversely, the concentration toxic metals and metalloids, such as As, Cd, Hg and Pb in human blood of the average population is well established, yet we know strikingly little about the role that they might play in the etiology of disease processes. Establishing functional connections between the chronic exposure of humans to these and other inorganic pollutants and the etiology of certain human diseases is therefore viewed by many as one of the greatest challenges in the post-genomic era. Conceptually, this task requires us to uncover hitherto unknown biomolecular mechanisms which must explain how small doses of a toxic metal/metalloid compound (low μg per day) - or mixtures thereof - may eventually result in a particular human disease. The biological complexity that is inherently associated with mammals, however, makes the discovery of these mechanisms a truly monumental task. Recent findings suggest that a better understanding of the bioinorganic chemistry of inorganic pollutants in the mammalian bloodstream represents a fruitful strategy to unravel relevant biomolecular mechanisms. The adverse effect(s) that toxic metals/metalloid compounds exert on the transport of essential ultratrace elements to internal organs appear particularly pertinent. A brief overview of the effect that arsenite and Hg(2+) exert on the mammalian metabolism of selenium is presented.
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The HPLC separation of arsenite, arsenate, methylarsonic acid and dimethylarsinic acid has been studied in the past but not in a systematic manner. The dependence of the retention times of these arsenic compounds on the pH of the mobile... more
The HPLC separation of arsenite, arsenate, methylarsonic acid and dimethylarsinic acid has been studied in the past but not in a systematic manner. The dependence of the retention times of these arsenic compounds on the pH of the mobile phase, on the concentration and the chemical composition of buffer solutions (phosphate, acetate, potassium hydrogen phthalate) and on the presence of sodium sulfate or nickel sulfate in the mobile phase was investigated using a Hamilton PRP‐X100 anion‐exchange column. With a flame atomic absorption detector and arsenic concentrations of at least 10 mg dm−3 all investigated mobile phases will separate the four arsenic compounds at appropriate pH values in the range 4–8. The shortest analysis time (˜3 min) was achieved with a 0.006 mol dm−3 potassium hydrogen phthalate mobile phase at pH 4, the longest (˜10 min) with 0.006 mol dm−3 sodium sulfate at pH 5.9 at a flow rate of 1.5 cm3 min−1. With a graphite furnace atomic absorption detector at the required, much lower, flow rate of ˜0.2 cm3 min−1 acceptable separations were achievable only with the pH 6 phosphate buffer (0.03 mol dm−3) and the nickel sulfate solution (0.005 mol dm−3) as the mobile phase. To become detectable approximately 100 ng arsenic from each arsenic compound (100 μl injection) must be chromatographed with the phosphate buffer, and approximately 10 ng with the nickel sulfate solution.
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In order to characterize the potentially deleterious effects of toxic Hg(2+) and Cd(2+) on lipid membranes, we have studied their binding to liposomes whose composition mimicked erythrocyte membranes. Fluorescence spectroscopy utilizing... more
In order to characterize the potentially deleterious effects of toxic Hg(2+) and Cd(2+) on lipid membranes, we have studied their binding to liposomes whose composition mimicked erythrocyte membranes. Fluorescence spectroscopy utilizing the concentration dependent quenching of Phen Green SK by Hg(2+) and Cd(2+) was found to be a sensitive tool to probe these interactions at metal concentrations < or =1 microM. We have systematically developed a metal binding affinity assay to screen for the interactions of Hg(2+) or Cd(2+) with certain lipid classes. A biomimetic liposome system was developed that contained four major lipid classes of erythrocyte membranes (zwitterionic lipids: phosphatidylcholine and phosphatidylethanolamine; negatively charged: phosphatidylserine and neutral: cholesterol). In contrast to Hg(2+), which preferentially bound to the negatively charged phosphatidylserine compared to the zwitterionic components, Cd(2+) bound stronger to the two zwitterionic lipids. Thus, the observed distinct differences in the binding affinity of Hg(2+) and Cd(2+) for certain lipid classes together with their known effects on membrane properties represent an important first step toward a better understanding the role of these interactions in the chronic toxicity of these metals.
Research Interests: Biophysics, Chemistry, Fluorescence Spectroscopy, Membranes, Medicine, and 15 moreLipids, Humans, Lipid Membranes, Cholesterol, Liposomes, Membrane Lipids, Liposome, Cadmium, Membrane, Biometals, Erythrocyte Membrane, Chronic Toxicity, Biochemistry and cell biology, Binding affinity, and fluorescent dyes
ABSTRACT When rabbits are simultaneous injected with arsenite and selenite or mercuric chloride and selenite, compounds with As–Se and Hg–Se bonds are formed in the bloodstream. The combined application of liquid... more
ABSTRACT When rabbits are simultaneous injected with arsenite and selenite or mercuric chloride and selenite, compounds with As–Se and Hg–Se bonds are formed in the bloodstream. The combined application of liquid chromatography-inductively coupled plasma atomic emission spectrometry (ICP-AES) and X-ray absorption spectroscopy (XAS) has revealed the molecular structure of these toxicologically important compounds and provided insight into their mechanism of formation. The glutathione-driven formation of these compounds in the bloodstream fundamentally links the metabolism of the environmental pollutants mercuric mercury and arsenite with that of the essential ultratrace element selenium, which establishes a feasible mechanism by which the chronic low-level exposure of various human populations to these toxic metals and metalloid compounds is linked to human diseases, including cancer and neurodegenerative diseases.
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ABSTRACT The pH-dependent retention behavior of arsenobetaine, arsenocholine, trimethylarsine oxide, tetramethylarsonium iodide (cationic arsenic compounds), arsenite, arsenate, methylarsonic acid, and dimethylarsinic acid (anionic... more
ABSTRACT The pH-dependent retention behavior of arsenobetaine, arsenocholine, trimethylarsine oxide, tetramethylarsonium iodide (cationic arsenic compounds), arsenite, arsenate, methylarsonic acid, and dimethylarsinic acid (anionic arsenic compounds) was studied on a Hamilton PRP-1 reversed-phase column (250×4.1 mm I.D.) with 10 mM aqueous solutions of benzensulfonic acids (X-C6H4SO3−; X=H, 4-HO, 3-CO2H; 4-HO-3-HO2C-C6H3SO3−) as ion-pairing reagents in the pH range 2–5 using flame atomic absorption spectrometry as the arsenic-specific detector. The dependencies of the k′-values of the ‘cationic’ arsenic compounds was rationalized on the basis of the protonation/deprotonation behavior of the arsenic compounds and of the four benzenesulfonates. The results provided evidence for the formation of a cationic species from trimethylarsine oxide below pH 3. Benzenesulfonate is the most hydrophobic ion-pairing reagent causing strong retention of the cationic arsenic compounds and consequently impeding their rapid separation. With the less hydrophobic, substituted benzenesulfonates the cationic arsenic compounds had retention times not exceeding 6 min. At a flow-rate of 1.5 cm3 min−1 10 mM aqueous 3-carboxy-4-hydroxybenzenesulfonate solution adjusted to pH 3.5 allowed the separation of arsenate, methylarsonic acid, arsenobetaine, trimethylarsine oxide, the tetramethylarsonium ion, and arsenocholine within 3 min. Dimethylarsinic acid coelutes with arsenobetaine at pH 3.5, but can be separated from arsenobetaine with the same mobile phase at pH 2.5. At pH 2.5 the signals for trimethylarsine oxide, the tetramethylarsonium ion, and arsenocholine are too broad to be useful for quantification. Arsenite and methylarsonic acid cannot be separated under these conditions.
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Research Interests: Chemistry, Mass Spectrometry, Metabolomics, Proteomics, Medicine, and 10 moreHumans, Animals, Gel Permeation Chromatography, Metalloproteins, Inductively Coupled Plasma, Atomic Absorption Spectrophotometry, Inductively Coupled Plasma Atomic Emission Spectroscopy, Proteome, Blood Proteins, and Biochemistry and cell biology
Since mercuric mercury (Hg(2+)) and methylmercury (CH(3)Hg(+)) display different toxicological properties in mammals, methods for their quantification in dietary items must be available. Employing Hg-specific detection, we have developed... more
Since mercuric mercury (Hg(2+)) and methylmercury (CH(3)Hg(+)) display different toxicological properties in mammals, methods for their quantification in dietary items must be available. Employing Hg-specific detection, we have developed a rapid, isocratic, and affordable RP-HPLC separation of these mercurials using thiol-containing mobile phases. Optimal separation was achieved with a 50mM phosphate-buffer containing 10mM L-cysteine at pH 7.5. The separation is driven by the on-column formation of complexes between each mercurial and L-cysteine, which are then separated according to their different hydrophobicities. The developed method is compatible with inductively coupled plasma atomic emission spectrometry and was applied to analyze spiked human urine.
Research Interests: Engineering, Chemistry, Technology, Chromatography, Methylmercury, and 11 moreMedicine, Humans, Mercury, Inductively Coupled Plasma Mass Spectrometry, High Performance Liquid Chromatography, High Pressure Liquid Chromatography, CHEMICAL SCIENCES, Cysteine, Thiol, Hydrogen-Ion Concentration, and Chromatographic separation
Research Interests: Chemistry, Analytical Chemistry, Physical Chemistry, Chromatography, Arsenic, and 15 moreMedicine, Humans, Liver, Glutathione, Temperature, Gel Permeation Chromatography, Buffers, Atomic Absorption Spectrophotometry, Atomic Absorption, Erythrocytes, Arsenite, Chelating Agents, Hydrogen-Ion Concentration, Size Exclusion Chromatography, and Phosphate Buffer Saline
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The iron chelation therapy drugs desferrioxamine B (DFO) and deferiprone (DFP) are used to treat iron overload patients, but not much is known about their adverse effects on other essential metals in vivo. After the addition of a... more
The iron chelation therapy drugs desferrioxamine B (DFO) and deferiprone (DFP) are used to treat iron overload patients, but not much is known about their adverse effects on other essential metals in vivo. After the addition of a clinically relevant dose of DFP or an equimolar dose of DFO to human plasma in vitro, the mixtures were analyzed by size exclusion chromatography (SEC) coupled to an inductively coupled plasma atomic emission spectrometer (ICP-AES). Simultaneous detection of the emission lines of copper, iron and zinc allowed the visualization of changes that these drugs exerted at the metalloprotein level. After the addition of DFP, a <10 kDa novel Fe-peak was detected and identified as (DFP)(3)Fe, whereas DFO resulted in the elution of a much smaller amount of Fe in this elution range. In fact, DFP was approximately 8-times more efficient than DFO regarding the removal of Fe from plasma proteins. The addition of both iron chelators also resulted in the elution of a <10 kDa novel Zn-peak. DFP abstracted twice as much Zn from plasma proteins compared to DFO. The identification of one of these peaks as (DFP)(2)Zn establishes a feasible biomolecular basis for the etiology of Zn-deficiency in patients that undergo long-term treatment with these drugs. Our results demonstrate that the analysis of plasma by SEC-ICP-AES can simultaneously provide insight into the efficacy of chelation therapy drugs and their adverse health effects at the metalloprotein level. Thus, SEC-ICP-AES emerges as a useful analytical tool to visualize health-relevant bioinorganic chemistry-related reactions of medicinal drugs in blood plasma in vitro.
Research Interests: Chemistry, Inorganic Chemistry, Medicine, Humans, EDTA Chelation, and 14 moreCopper, Zinc, Iron, Gel Permeation Chromatography, Iron Overload, Metalloproteins, Inductively Coupled Plasma, Atomic Absorption Spectrophotometry, Inductively Coupled Plasma Atomic Emission Spectroscopy, Serum albumin, Deferoxamine, binding sites, Elution, and Deferiprone
Since no chelating agent has been clinically approved for the treatment of Cd(2+) intoxicated humans, we have previously investigated diethylenetriaminepentaacetic acid (Na5DTPA) to abstract this toxic metal from plasma proteins in vitro.... more
Since no chelating agent has been clinically approved for the treatment of Cd(2+) intoxicated humans, we have previously investigated diethylenetriaminepentaacetic acid (Na5DTPA) to abstract this toxic metal from plasma proteins in vitro. In addition to the complete mobilization of Cd(2+), an inadvertent abstraction of Zn(2+) from metalloproteins was also observed. Therefore, we have evaluated the effect of ZnNa3DTPA on the plasma distribution of Ca, Cd, Cu, Fe and Zn after its addition to Cd(2+)-spiked rabbit plasma. ZnNa3DTPA was as efficient as Na5DTPA in abstracting Cd(2+) (100% removal), but it did not abstract Zn(2+). Thus, complexes between essential metals (e.g. Zn(2+)) and known chelating agents emerge as feasible candidates to decrease potential adverse side effects in patients.
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Meso-2,3-dimercaptosuccinic acid (DMSA) and 2,3-dimercaptopropane-1-sulfonic acid (DMPS) are chelating agents which have been used clinically to treat patients suffering from Pb(2+) or Hg(2+) exposure. Cd(2+) is a related environmental... more
Meso-2,3-dimercaptosuccinic acid (DMSA) and 2,3-dimercaptopropane-1-sulfonic acid (DMPS) are chelating agents which have been used clinically to treat patients suffering from Pb(2+) or Hg(2+) exposure. Cd(2+) is a related environmental pollutant that is of increasing public health concern due to a demonstrated dose-response between urinary Cd level and an increased risk of diabetes. However, therapeutically effective chelating agents which enhance the excretion of Cd(2+) from humans have yet to be identified. Here we present a structural characterization of complexes of DMSA and DMPS with Cd(2+) at physiological pH using a combination of X-ray absorption spectroscopy, size exclusion chromatography and density functional theory. 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.
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We report a new synthesis of the seleno-bis (S-glutathionyl) arsinium anion, [(GS)2AsSe]−. An aqueous solution of bis-glutathionylarsenous acid, (GS)2AsOH, prepared from stoichiometric glutathione and arsenite, was reacted in situ with a... more
We report a new synthesis of the seleno-bis (S-glutathionyl) arsinium anion, [(GS)2AsSe]−. An aqueous solution of bis-glutathionylarsenous acid, (GS)2AsOH, prepared from stoichiometric glutathione and arsenite, was reacted in situ with a solution of sodium hydrogen selenide, prepared from elemental selenium and sodium borohydride. Analysis of the arsenic and selenium K-edge X-ray absorption spectra indicated virtually quantitative formation of [(GS)2AsSe]−, with
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Hg(2+) and CH(3)Hg(+) are frequently encountered in the environment either as free ions or complexed with organic matter, such as humic acids. The majority of the reported HPLC-based separations of environmental mercury species, however,... more
Hg(2+) and CH(3)Hg(+) are frequently encountered in the environment either as free ions or complexed with organic matter, such as humic acids. The majority of the reported HPLC-based separations of environmental mercury species, however, separate Hg(2+) from CH(3)Hg(+) in which the former species elutes close to the void volume. To detect mercury-species in environmental waters that may have so far escaped detection, a separation method is needed that sufficiently retains both Hg(2+) and CH(3)Hg(+). One way to develop such a method is to increase the retention of Hg(2+) and CH(3)Hg(+) using existing HPLC separations. We here report on the improvement of a previously reported RP-HPLC-based separation of Hg(2+) and CH(3)Hg(+) that employed a 100 % aqueous mobile phase [10 mM L-cysteine (Cys) in 50 mM phosphate buffer (pH 7.5)]. To increase the retention of Hg(2+), Cys was replaced by the comparatively more hydrophobic N-acetylcysteine (N-Cys). To achieve a compromise between an increased retention of Hg(2+) and its baseline separation from CH(3)Hg(+) in the shortest possible analysis time, the retention behavior of both mercurials was investigated on two RP-HPLC columns with mobile phases that contained mixtures of Cys and N-Cys in which the overall thiol concentration was maintained at 10 mM. An optimal separation of both mercurials could be achieved in ∼540 s using a Gemini C(18) HPLC column (150 × 4.6 mm I.D.) and a mobile phase comprised of 7.5 mM N-Cys and 2.5 Cys in 50 mM phosphate buffer (pH 7.4). Coupling the developed HPLC separation with an inductively coupled plasma mass spectrometer should allow one to detect mercury species other than Hg(2+) and CH(3)Hg(+) in environmental waters. The detection of such species is critical to better understand the mobilization of mercury species from natural and anthropogenic pollution sources.
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It has been shown that the seleno-bis (S-glutathionyl) arsinium ion, [(GS)(2)AsSe](-), is the major arsenic and selenium excretory product in bile of rabbits treated with arsenite and selenite [Gailer, J., Madden, S., Buttigieg, G. A.,... more
It has been shown that the seleno-bis (S-glutathionyl) arsinium ion, [(GS)(2)AsSe](-), is the major arsenic and selenium excretory product in bile of rabbits treated with arsenite and selenite [Gailer, J., Madden, S., Buttigieg, G. A., Denton, M. B., and Younis, H. S. (2002) Appl. Organomet. Chem. 16, 72-75]. To investigate the in vivo interaction between the other environmentally common oxy-anions of arsenic and selenium in mammals, we have intravenously injected rabbits with different combinations of the arsenic and selenium oxo-anions (arsenite + selenate, arsenate + selenite, and arsenate + selenate) and analyzed the collected bile and whole blood samples by X-ray absorption spectroscopy. Only the injection of arsenite and selenate led to the biliary excretion of [(GS)(2)AsSe](-) within 25 min. Whole blood collected from these animals (25 min postinjection) contained predominantly unchanged selenate, which suggests the presence of a mammalian selenate reductase in the liver. The lack of any significant biliary excretion of [(GS)(2)AsSe](-) in the other treatment groups implies that arsenate was not reduced in the liver on the time scale of our experiments. The relevance of these results for the human toxicology of arsenic and selenium is discussed.
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Understanding the fate of metallodrugs in the bloodstream is critical to assess if the parent drug has a reasonable probability to reach the intended target tissue and to predict toxic side-effects. To gain insight into these processes,... more
Understanding the fate of metallodrugs in the bloodstream is critical to assess if the parent drug has a reasonable probability to reach the intended target tissue and to predict toxic side-effects. To gain insight into these processes, we have added pharmacologically relevant doses of metallodrugs to blood plasma and applied an LC-method to directly analyze the latter for metallodrug metabolites. Using human or rabbit plasma, this LC-method was employed to gain insight into the metabolism of clinically used as well as emerging anticancer metallodrugs and to unravel the mechanisms by which small molecular weight compounds that - when co-administered with a metallodrug - decrease the toxic side-effects of the metallodrug by modulating its metabolism. The results suggest that the developed LC-method is useful to probe the fate of biologically active novel metal-complexes in plasma to help select those which may be advanced to animal/clinical studies to ultimately develop safer metallodrugs.