Zinc Trafficking 1. Probing the Roles of Proteome, Metallothionein, and Glutathione

The cellular trafficking pathways that conduct zinc to its sites of binding in functional proteins remain largely unspecified. In this study, the hypothesis was investigated that non-specific proteomic binding sites serve as intermediates in zinc trafficking. Proteome from pig kidney LLC-PK1 cells contains a large concentration of such sites, displaying an average conditional stability constant of 1010-11, that are dependent on sulfhydryl ligands to achieve high affinity binding of zinc. As a result, the proteome competes effectively with induced metallothionein for Zn2+ upon exposure of cells to extracellular Zn2+ or during in vitro direct competition. The reaction of added Zn2+ bound to proteome with apo-carbonic anhydrase was examined as a potential model for intracellular zinc trafficking. The extent of this reaction was inversely dependent upon proteome concentration and under cellular conditions thought to be negligible. The rate of reaction was strictly first order in both Zn2+ and apo-carbonic anhydrase and also considered to be insignificant in cells. Adding the low molecular weight fraction of cell supernatant to the proteome markedly enhanced the speed of this reaction, a phenomenon dependent on the presence of glutathione. In agreement, inclusion of glutathione accelerated the reaction in a concentration-dependent manner. The implications of abundant high affinity binding sites for Zn2+ within the proteome are considered in relation to their interaction with glutathione in the efficient delivery of Zn2+ to functional binding sites and in the operation of fluorescent zinc sensors as a tool to observe zinc trafficking.

Complexation of Cm(III) with blood serum proteins: recombinant human serum albumin (rHSA)

The complexation of Cm(III) with the recombinant human serum albumin (rHSA) (characterized by single deletion of residue Asp-1), is studied in dependence of pH and rHSA concentration using time-resolved laser fluorescence spectroscopy (TRLFS). A Cm(III) rHSA species is formed between pH 6.4 and 10.0 with the conditional stability constant being logK = 6.47 at pH = 7.4. Competition titration experiments with Cu(II) and Zn(II) confirm complexation at the N-terminal binding site (NTS) of rHSA and exclude the involvement of the Multi-Metal Binding Site (MBS). Comparison with a previous study on Cm(III) interaction with native albumin, HSA, points out, that residue Asp-1 is involved in Cm(III) binding to HSA but is not crucial for Cm(III) complexation at the NTS. The results are of major importance for a better understanding of fundamental actinide-protein interaction mechanisms which are highly required for the identification and characterization of relevant distribution pathways of incorporated radionuclides.

BETULIN-3, 28-DIPHOSPHATE SALT COMPLEXES WITH AMINES AND THEIR ANTIOXIDANT ACTIVITY

Objective: Studies of composition, stability and antioxidant properties of the betulin-3, 28-diphosphate complexes with dopamine and trisamine.Methods: The betulin-3, 28-diphosphate (BDP) interaction with amines in a water-alcohol medium was studied by using spectral methods and potentiometric titration. Biochemical indexes such as catalase, superoxide dismutase (SOD), lactate dehydrogenase (LDH) activities and malondialdehyde (MDA) level were estimated in experiments on rats.Results: BDP was synthesized using betulin by POCl3 treatment in the presence of pyridine in dioxane. The complexation of BDP with amines was confirmed by the 31P-NMR and FTIR-spectral data. The stoichiometry of BDP-dopamine complexes was equal to 2:1 and 4:1 and its complexes with trisamine were produced in the ratio 1:1 in a water-alcohol medium. The conditional stability constant К′st of the BDP-trisamine complex is 1130±55 mol∙l-1. BDP-Tris complex improved SOD activity up to 30% and up to 105% in the presence of cytostatic-hydrazine sulfate. The MDA level in erythrocytes decreased up to 57% and in combination with cytostatics (5-fluorouracil and hydrazine sulfate)-up to 11-14%. The catalase activity increased by 44-94% and MDA level in erythrocytes decreased by 22-53% under the action BDP-DA complexes that depends on the dose.Conclusion: The BDP forms stable complexes with trisamine and dopamine that make it possible to use this compound as a component of drug delivery system for high toxicity cytostatics and for readily oxidized catecholamines. It has been shown that both its complexes with amines and the combination with cytostatics enhanced antioxidant activity in an experiment in vitro.

On the nature of dissolved copper ligands in the early buoyant plume of hydrothermal vents

Environmental contextCopper released by deep-sea hydrothermal vents has been recognised to be partly stabilised against precipitation by its complexation with strong Cu binding ligands. Yet, the sources and nature of these compounds in such environments are still not fully understood. This study shows that the Cu ligands detected are hydrothermally sourced and could be mainly inorganic sulfur species. AbstractThe apparent speciation of Cu in the early buoyant plume of two black smokers (Aisics and Y3) from the hydrothermal vent field Lucky Strike (Mid-Atlantic Ridge) was investigated using competitive ligand exchange-adsorptive cathodic stripping voltammetry (CLE–AdCSV). We have assessed the apparent Cu-binding ligand concentration ([L]) and the corresponding conditional stability constant (log K′) for 24 samples. At the smoker Aisics, [L] ranged from 18.2 to 2970 nM. Log K′CuL ranged from 12.4 to 13.4. At Y3, the binding capacity of natural ligands was from 32.5 to 1020 nM, with Log K′CuL ranging from 12.5 to 13.1. Total dissolved Cu ranged from 7.0 to 770 nM and from 12.7 to 409 nM at Aisics and Y3, respectively. Our results show that the amount of ligand L increases with dissolved Mn (dMn) concentrations, suggesting a hydrothermal origin of the Cu-binding ligands detected. In addition, such high concentrations of Cu-binding ligands can only be explained by an additional abiotic source differing from organic processes. Based on the massive in situ concentrations of free sulfides (up to 300 µM) and on the striking similarities between our log K′CuL and the log K′Cu(HS) previously published, we infer that the Cu-binding ligands could be predominantly inorganic sulfur species in the early buoyant plume of the two vent sites studied.

Influence of heavy metal ions on the spectra and charge characteristics of DOM of municipal sewage secondary effluent

The spectral and charged characteristics of dissolved organic matter (DOM) in municipal sewage secondary effluent influenced by heavy metal ions were studied by three-dimensional fluorescence spectroscopy, ultraviolet spectrometry, etc. As the Cu(II) concentration increased, the fluorescence intensity of the DOM's fluorescence peak reduced. When the Cu(II) concentration was 100 μmol/L, the fluorescence intensity of the humic-like acid in the DOM reduced by 19.5%–27.6%. However, there was no obvious change in the fluorescence intensity when Cd(II) was added. According to the modified Stern–Volmer equation, the conditional stability constant lgK and the proportion of the fluorescent functional groups of the DOM binding with Cu(II) were 2.19–2.69 and 16%–48%, respectively. The UV253/UV203 value changed little with the additional dosage of metal ions below 10 μmol/L, which increased as the concentration of Cu(II) or Pb(II) increased from 10–100 μmol/L. As the metal concentrations increase, the absolute values of the Zeta potential and the particle sizes of the DOM basically increased. The results showed that Cu(II) had the strongest binding ability to the groups that emitted fluorescence in the secondary effluent DOM compared to Pb(II) and Cd(II).

Investigation of a new “N2S2O2” chelating agent with high Po(iv) affinity

A water-soluble complexing agent for 210-polonium decorporation was designed and synthesised with a high conditional stability constant.

Pd(II) complexes of acetylcholinesterase reactivator obidoxime

ABSTRACT The ability of the acetylcholinesterase reactivator obidoxime (H2L2+) to bind palladium(II) cations was evaluated spectrophotometrically at different reaction conditions (pH, reaction time, metal-to-ligand molar ratio). The results showed that immediately after mixing the reagents, pH 7.4, complex species of composition [PdHL]3+ existed predominantly with a value of conditional stability constant lgβ'=6.52. The reaction was completed within 24 hours affording the formation of species [Pd2L]4+ with significantly increased stability (lgβ'=9.34). The spectral data suggest that obidoxime coordinates metal(II) ions through the oximate functional groups. The in vitro reactivation assay of paraoxon-inhibited rat brain acetylcholinesterase revealed that the new complex species were much less active than the non-coordinated obidoxime. The lack of reactivation ability could be explained by the considerable stability of complexes in solution as well as by the deprotonation of oxime groups essential for recovery of the enzymatic activity.

Simple and Selective Spectrophotometric Determination of Ofloxacin in Pharmaceutical Formulations Using Two Sulphonphthalein Acid Dyes

Two new simple and sensitive extraction-free spectrophotometric methods have been established for the determination of ofloxacin (OFX). The methods are based on ion-pair complex formation reaction between OFX and acidic sulphonphthalein dyes, bromocresol purple (method A), and bromocresol green (method B) in dichloromethane. The experimental variables such as reaction medium, reaction time, and reagent concentration have been carefully optimized to achieve the highest sensitivity. Both dyes react spontaneously with OFX to give yellow-colored complexes. Beer’s law is obeyed over the concentration ranges of 1.0–16 µg ml−1 OFX with correlation coefficient of 0.999 in both methods. The molar absorptivity values are calculated to be and l mol−1 cm−1, for method A and method B, respectively, with corresponding Sandell's sensitivity values of 0.015 and 0.019 µg cm−2. The limits of detection (LOD) and quantification (LOQ) are also reported. A Job’s plot of the absorbance versus the molar ratio of OFX to each of dyes under consideration indicated (1 : 1) ratio and the conditional stability constant () of the complexes have been calculated. The proposed methods were applied successfully to the determination of OFX in tablets with good accuracy and precision and without interference from common additives. The results obtained by the proposed methods were compared favorably with those of the reference method.

The solubility of Ca(OH)2 in extremely concentrated NaOH solutions at 25°C

The solubility of Ca(OH)2 in aqueous NaOH solutions up to 12.50 M at 25°C has been determined. The solubility data obtained for NaOH concentrations lower than 3 M was compared with those published in the literature. The solubility of Ca(OH)2 steadily decreases with the increasing NaOH concentration. The solubility data obtained at a constant ionic strength (I = 1 M Na(Cl,OH)) enabled the determination of the conditional solubility product of Ca(OH)2(s) (lgLCa(OH)2 = − 4.10 ± 0.02). Formation of the hydroxo complex CaOH+(aq) was invoked to describe the variation of [Ca2+]T with [OH−]T. Its conditional stability constant was found to be lgKCaOH+ = 0.97 ± 0.02. The experimental protocol employed was proven to be suitable for accurate solubility determinations in rapidly equilibrating systems comprising of highly concentrated, alkaline solutions and containing analytes in the ppm range.