Cellular adsorption and absorption of antimony (III) by Mycobacterium smegmatis

Adsorption studies of Sb (III) as the antimonyl (125Sb) tartrate complex using acidified, non-proliferative suspensions of Mycobacterium smegmatis show that the uptake of Sb by the biomass increases with the external concentration of antimony ([Sb]) and decreases, at given concentration, with increasing pH. Measurements of Sb uptake in the cells under growth conditions in liquid culture indicate that the cellular concentration factor of antimony is of the order of 10 when the concentration of antimony in the medium is 10 microM, i.e., close to the minimum inhibitory concentration.     

Arsenic transport by the human multidrug resistance protein 1 (MRP1/ABCC1). Evidence that a tri-glutathione conjugate is required

Inorganic arsenic is an established human carcinogen, but its metabolism is incompletely defined. The ATP binding cassette protein, multidrug resistance protein (MRP1/ABCC1), transports conjugated organic anions (e.g. leukotriene C(4)) and also co-transports certain unmodified xenobiotics (e.g. vincristine) with glutathione (GSH). MRP1 also confers resistance to arsenic in association with GSH; however, the mechanism and the species of arsenic transported are unknown. Using membrane vesicles prepared from the MRP1-overexpressing lung cancer cell line, H69AR, we found that MRP1 transports arsenite (As(III)) only in the presence of GSH but does not transport arsenate (As(V)) (with or without GSH). The non-reducing GSH analogs L-gamma-glutamyl-L-alpha-aminobutyryl glycine and S-methyl GSH did not support As(III) transport, indicating that the free thiol group of GSH is required. GSH-dependent transport of As(III) was 2-fold higher at pH 6.5-7 than at a more basic pH, consistent with the formation and transport of the acid-stable arsenic triglutathione (As(GS)(3)). Immunoblot analysis of H69AR vesicles revealed the unexpected membrane association of GSH S-transferase P1-1 (GSTP1-1). Membrane vesicles from an MRP1-transfected HeLa cell line lacking membrane-associated GSTP1-1 did not transport As(III) even in the presence of GSH but did transport synthetic As(GS)(3). The addition of exogenous GSTP1-1 to HeLa-MRP1 vesicles resulted in GSH-dependent As(III) transport. The apparent K(m) of As(GS)(3) for MRP1 was 0.32 microM, suggesting a remarkably high relative affinity. As(GS)(3) transport by MRP1 was osmotically sensitive and was inhibited by several conjugated organic anions (MRP1 substrates) as well as the metalloid antimonite (K(i) 2.8 microM). As(GS)(3) transport experiments using MRP1 mutants with substrate specificities differing from wild-type MRP1 suggested a commonality in the substrate binding pockets of As(GS)(3) and leukotriene C(4). Finally, human MRP2 also transported As(GS)(3). In conclusion, MRP1 transports inorganic arsenic as a tri-GSH conjugate, and GSTP1-1 may have a synergistic role in this process.     

Reduction of pentavalent antimony by trypanothione and formation of a binary and ternary complex of antimony(III) and trypanothione

Several pentavalent antimony compounds have been used for the treatment of leishmaniasis for decades. However, the mechanism of these antimony drugs still remains unclear. One of their targets is thought to be trypanothione, a major low molecular mass thiol inside the parasite. We show that pentavalent antimony (Sb^V) can be rapidly reduced to its trivalent state by trypanothione at mildly acidic conditions and 310 K (k=4.42 M^–1 min^–1 at pH 6.4), and that Sb^III can be bound to trypanothione to form an Sb^III-trypanothione complex. NMR data demonstrate that Sb^III binds to trypanothione at the two thiolates of the cysteine residues, and that the binding is pH dependent and is strongest at biological pH with a stability constant logK=23.6 at 298 K (0.1 M NaNO₃). The addition of low molecular monothiol ligands such as glutathione and cysteine to the Sb^III-trypanothione complex results in the formation of a ternary complex. Thiolates from both trypanothione and monothiol bind to the Sb^III center. The formation of the ternary complex is important, as the antileishmanial properties of the drugs are probably due to a complex between of Sb^III-trypanothione and enzymes. Although thermodynamically stable, the complex is kinetically labile and the free and bound forms of thiolates exchange on the ¹H NMR timescale. Such a facile exchange may be crucial for the transport of Sb^III within parasites.Electronic Supplementary Material Supplementary material is available for this article if you access the article at . A link in the frame on the left on that page takes you directly to the supplementary material.Abbreviations amastigote the parasites culture at pH 5.0 and 310 K to resume the intracellular form - BPR bromopyrogallol - ESI-MS electrospary ionization mass spectrometry - GSH glutathione - pH^* pH meter reading in D₂O without correction for isotope effects - promastigote the parasites culture at pH 7.4 and 298 K to resume the extracellular stage - T(SH)₂ reduced form of trypanothione - T(S-S) oxidized form of trypanothione (disulfide form) - TR trypanothione reductase - tart tartrate     

Thiol-induced reduction of antimony(V) into antimony(III): A comparative study with trypanothione, cysteinyl-glycine, cysteine and glutathione

Gluthathione (GSH) has been previously shown to promote the reduction of pentavalent antimony (Sb(V)) into the more toxic trivalent antimony (Sb(III)) in the antimonial drug, meglumine antimonate. However, this reaction occurred at acidic pH (pH 5) but not at the pH of the cytosol (pH 7.2) in which GSH is encountered. The aim of the present study was to further characterize the reaction between thiols and antimonial drugs, addressing the following aspects: (i) the reducing activity of cysteine (Cys) and cysteinyl-glycine (Cys-Gly), expected to be the predominant thiols in the acidic compartiments of mammalian cells; (ii) the reducing activity of trypanothione (T(SH)₂), the main intracelular thiol in Leishmania parasites; (iii) the influence of the state of complexation of Sb(V) on the rate of Sb(V) reduction. We report here that Cys, Cys-Gly and T(SH)₂ did promote the reduction of Sb(V) into Sb(III) at 37 °C. Strikingly, the initial rates of reduction of Sb(V) were much greater in the presence of Cys-Gly, Cys and T(SH)₂ than in the presence of GSH. These reactions occurred at both pH 5 and pH 7 but were favored at acidic pH. Moreover, our data shows that Sb(V) is reduced more slowly in the form of meglumine antimonate than in its non-complexed form, indicating that the complexation of Sb(V) tends to slow down the rate of its reduction. In conclusion, our data supports the hypothesis that Sb(V) is reduced in vivo by T(SH)₂ within Leishmania parasites and by Cys or Cys-Gly within the acidic compartments of mammalian cells.     

As(III) and Sb(III) uptake by GlpF and efflux by ArsB in Escherichia coli

The toxicity of the metalloids arsenic and antimony is related to uptake, whereas detoxification requires efflux. In this report we show that uptake of the trivalent inorganic forms of arsenic and antimony into cells of Escherichia coli is facilitated by the aquaglyceroporin channel GlpF and that transport of Sb(III) is catalyzed by the ArsB carrier protein; everted membrane vesicles accumulated Sb(III) with energy supplied by NADH oxidation, reflecting efflux from intact cells. Dissipation of either the membrane potential or the pH gradient did not prevent Sb(III) uptake, whereas dissipation of both completely uncoupled the carrier protein, suggesting that transport is coupled to either the electrical or the chemical component of the electrochemical proton gradient. Reciprocally, Sb(III) transport via ArsB dissipated both the pH gradient and the membrane potential. These results strongly indicate that ArsB is an antiporter that catalyzes metalloid-proton exchange. Unexpectedly, As(III) inhibited ArsB-mediated Sb(III) uptake, whereas Sb(III) stimulated ArsB-mediated As(III) transport. We propose that the actual substrate of ArsB is a polymer of (AsO)(n), (SbO)(n), or a co-polymer of the two metalloids.     

Formation of methylantimony species by an aerobic prokaryote: <Emphasis Type="Italic">Flavobacterium</Emphasis> sp.

Pure cultures of an aerobically grown Flavobacterium sp. were shown by hydride generation-cold trap-atomic absorption spectrometry to biomethylate inorganic antimony (III) supplied as potassium antimony tartrate. Growth inhibition of the Flavobacterium sp. by antimony (III) over the range 0-30 mg Sb l(-1) was assessed by optimising parameters within an extended logistic growth model. Antimony (III) concentrations over this range influenced both the extent of antimony biomethylation (up to 4.0 microg l(-1)) and the relative proportions of the involatile mono-, di, and trimethylantimony species formed. Provision of inorganic arsenic (III) alongside antimony (III) enhanced formation of the involatile methylantimony species up to eight-fold. The data are consistent with accumulation of involatile intermediates from an antimony or arsenic biomethylation pathway in culture supernatants. Low yields of methylantimony species (<0.03%) suggest that antimony biomethylation by the Flavobacterium sp. was a fortuitous rather than a primary resistance mechanism for this element. These findings demonstrate that anaerobiosis is not an obligate requirement for methylantimony formation in prokaryotes, thus broadening the range of habitats for potential formation of methylantimony species in nature.     

Dual action of antimonial drugs on thiol redox metabolism in the human pathogen Leishmania donovani

Despite extensive use of antimonial compounds in the treatment of leishmaniasis, their mode of action remains uncertain. Here we show that trivalent antimony (Sb(III)) interferes with trypanothione metabolism in drug-sensitive Leishmania parasites by two inherently distinct mechanisms. First, Sb(III) decreases thiol buffering capacity by inducing rapid efflux of intracellular trypanothione and glutathione in approximately equimolar amounts. Second, Sb(III) inhibits trypanothione reductase in intact cells resulting in accumulation of the disulfide forms of trypanothione and glutathione. These two mechanisms combine to profoundly compromise the thiol redox potential in both amastigote and promastigote stages of the life cycle. Furthermore, we demonstrate that sodium stibogluconate, a pentavalent antimonial used clinically for the treatment for leishmaniasis, induces similar effects on thiol redox metabolism in axenically cultured amastigotes. These observations suggest ways in which current antimony therapies could be improved, overcoming the growing problem of antimony resistance.     

Identification of complexes containing glutathione with As(III), Sb(III), Cd(II), Hg(II), Tl(I), Pb(II) or Bi(III) by electrospray ionization mass spectrometry

Electrospray ionization mass spectrometry (ESI-MS) of mixtures containing glutathione (GSH) and nitrates, oxides or chlorides of the heavy metals, arsenic, antimony, cadmium, mercury, thallium, lead or bismuth allows for definitive identification of complexes in the gas phase. In the positive ion mode, spectra show prominent m/z peaks that are assigned to monocations of general formulae [E(GSH)-xH]+ (E = Cd, Hg, Tl, Pb, As, Sb or Bi; x = 0, 1 or 2), [E(GSH)2-xH]+ (E = Hg, As, Sb, or Bi; x = 1 or 2), [E(GSH)3-xH]+ (E = As, Sb or Bi; x = 2), [E2(GSH)-xH]+ (E = Tl or Pb; x = 1 or 3), [E2(GSH)2-xH]+ (E = Bi; x = 5), [E2(GSH)3-xH]+ (E = Bi; x = 5), and/or [E3(GSH)-xH]+ (E = Tl; x = 2). Spectra obtained in the negative ion mode give m/z peaks observed in assigned to monoanionic species that correspond to some of the monocationic species listed above with two protons removed. The results demonstrate the potential application of ESI-MS as a versatile and efficient approach to study toxic heavy metals in biological systems. In addition, the observations provide a foundation database to understand the chemistry of these heavy metals with bio-molecules.     

Characterization of Glutathione Uptake in Broad Bean Leaf Protoplasts

Transport of reduced glutathione (GSH) and oxidized glutathione (GSSG) was studied with broad bean (Vicia faba L.) leaf tissues and protoplasts. Protoplasts and leaf discs took up GSSG at a rate about twice the uptake rate of GSH. Detailed studies with protoplasts indicated that GSH and GSSG uptake exhibited the same sensitivity to the external pH and to various chemical reagents. GSH uptake was inhibited by GSSG and glutathione conjugates. GSSG uptake was inhibited by GSH and GS conjugates, and the uptake of metolachlor-GS was inhibited by GSSG. Various amino acids (L-glutamic acid, L-glutamine, L-cysteine, L-glycine, L-methionine) and peptides (glycine-glycine, glycine-glycine-glycine) affected neither the transport of GSH nor GSSG. Uptake kinetics indicate that GSH is taken up by a single saturable transporter, with an apparent Km of 0.4 mM, whereas GSSG uptake exhibits two saturable phases, with an apparent Km of 7 [mu]M and 3.7 mM. It is concluded that the plasma membrane of leaf cells contains a specific transport system for glutathione, which takes up GSSG and GS conjugates preferentially over GSH. Proton flux measurements and electrophysiological measurements indicate that GSH and GSSG are taken up with proton symport. However, a detailed analysis of these measurements suggests that the ion movements induced by GSSG differ from those induced by GSH.     

Reactions of [PtCl(dien)]Cl with glutathione,oxidized glutathione and S-methyl glutathione. Formation of an S-bridged dinuclear unit

The reaction of the monofunctional platinum compound [PtCl(dien)]Cl with the tripeptide glutathione (GSH), oxidized glutathione (GSSG) and S-methyl glutathione (GS-Me) has been investigated by ¹H, ¹³C and ¹⁹⁵Pt magnetic resonance spectroscopy and by potentiometric titrations. It appears that platinum binds with a high degree of specificity to the GSH sulfhydryl group. The reaction of platinum with GSH proceeds in two steps. In the first step only one platinum binds to the sulfur atom and, in the second step, another [Pt(dien)]²⁺ unit binds to [Pt(dien)GS]⁺ forming an S-bridged dinuclear unit [{Pt(dien)}₂GS]³⁺. The rate of the first binding step is pH-dependent, whereas the rate of the second step is not. At pH < 7 the rate of the first binding step is slow compared to the rate of the second binding step. At pH > 10, on the other hand, the rate of the first binding step is faster than the rate of the second binding step. Consequently, at pH < 7 one can only isolate the [{Pt(dien)}₂GS]³⁺ complex. In the presence of free GSH, at pH > 7, one [Pt(dien)]²⁺ unit of [{Pt(dien)}₂GS]³⁺ dissociates forming [Pt(dien)GS]⁺. The mechanism of the pH-dependent rate of the first platinum binding step and the ligand-exchange reaction are discussed. GSSG reacts with [Pt(dien)]²⁺, also forming the S-bridged dinuclear unit [{Pt(dien)}₂GS]³⁺, probably through a redox disproportionation reaction with a catalytic function of [PtCl(dien)]Cl. GS-Me reacts with [Pt(dien)]²⁺ forming the S-coordinated [Pt(dien)GS-Me]²⁺. [Pt(dien)GS-Me]²⁺ exists as a pair of diastereomers due to different configurations about sulfur. The rate of the inversion of configuration at the coordinated sulfur atom is slow on the NMR time-scale.     

Influence of arsenic on antimony methylation by the aerobic yeast<Emphasis Type="Italic"> Cryptococcus humicolus</Emphasis>

The anamorphic basidomycetous yeast Cryptococcus humicolus was shown by hydride generation-gas chromatography-atomic absorption spectrometry to methylate inorganic antimony compounds to mono-, di-, and trimethylantimony species under oxic growth conditions. Methylantimony levels were positively correlated with initial substrate concentrations up to 300 mg Sb l^–1 as potassium antimony tartrate (K-Sb-tartrate). Increasing concentrations of K-Sb-tartrate increased the ratio of di- to trimethylantimony species, indicating that methylation of dimethylantimony was rate limiting. Antimony methylation capability in C. humicolus was developed after the exponential growth phase and was dependent upon protein synthesis in the early stationary phase. Inclusion of inorganic arsenic (III) or (V) species alongside antimony in culture incubations enhanced antimony methylation. Pre-incubation of cells with inorganic arsenic (III) further induced antimony methylation capability, whereas pre-incubation with inorganic antimony (III) did not. Exposure of cells to inorganic arsenic—either through pre-incubation or provision during cultivation—influenced the antimony speciation; involatile trimethylantimony species was the sole methylated antimony species detected, i.e. mono- and dimethylantimony species were not detected. Competitive inhibition of antimony methylation was observed at high arsenic loadings. These data indicate that antimony methylation is a fortuitous process, catalysed at least in part by enzymes responsible for arsenic methylation.     

Use of cloud-point preconcentration for spectrophotometric determination of trace amounts of antimony in biological and environmental samples

This work presents a cloud-point extraction process using the micelle-mediated extraction method for simultaneous preconcentration and determination of Sb(III) and Sb(V) species in biological and environmental samples as a prior preconcentration step to their spectrophotometric determination. The analytical system is based on the selective reaction between Sb(III) and 3-dichloro-6-(3-carboxy-2-hydroxy-1-naphthylazo)quinoxaline (DCHNAQ) in the presence of cetyltrimethylammonium bromide (CTAB) and potassium iodide at pH 4.5. Total Sb concentration was determined after reduction of Sb(V) to Sb(III) in the presence of potassium iodide and ascorbic acid. The optimal reaction conditions and extraction were studied, and the analytical characteristics of the method (e.g., limits of detection and quantification, linear range, preconcentration, improvement factors) were obtained. Linearity for Sb(III) was obeyed in the range of 0.2–20 ng ml⁻¹. The detection and quantification limits for the determination of Sb(III) were 0.055 and 0.185 ng ml⁻¹, respectively. The method has a lower detection limit and wider linear range, inexpensive instrument, and low cost, and is more sensitive compared with most other methods. The interference effect of some anions and cations was also studied. The method was applied to the determination of Sb(III) in the presence of Sb(V) and total antimony in blood plasma, urine, biological, and water samples.     

Speciation of antimony in natural waters : the determination of Sb(III) and Sb(V) by continuous flow hydride generation-atomic absorption spectrometry

Abstract

A new procedure for the speciation of dissolved antimony is described. This makes use of complexation with citrate to prevent, preferentially, the formation of hydride from Sb(V) and allow the selective determination of Sb(III) to be made by continuous flow hydride generation - atomic absorption spectrometry. When the citric acid (12% m/V) is replaced by potassium iodide (3% m/V), total antimony is determined and the concentration of Sb(V) can be obtained by difference. The determination of the antimony species is dominated in this new procedure by the complexation of Sb(V) with citrate and the effect of pH is limited to a minor, re-inforcing role. This permits acidification to be made with hydrochloric acid. The principal interfering species in the determination of total antimony and Sb(III) is Fe³⁺, with Fe²⁺, Cu²⁺ and Ni²⁺ showing lesser effects on Sb(III). The technique is applied successfully to synthetic mixtures and to natural waters from the environment of a disused antimony mine.

The characteristic concentration obtained for antimony was 0.7 ng mL^–1 and the detection limit 1 ng mL^–1.     

Effect of pH and Salinity on Sorption of Antimony (III and V) on Mangrove Sediment,Sundarban, India

The extent of toxic metalloid retention and bioavailability and mobility in the sediment is of interest for understanding their biogeochemical cycling and for accurate risk assessment in an ecosystem. Intensification of monsoon and rainfall, believed to be related to global warming, could drive future changes of temperature, salinity, and pH distribution pattern affecting antimony cycling in the Sundarbans. This study investigated sorption kinetics of antimony (Sb) (III and V) as a function of temperature, salinity, and pH following the Langmuir model, and demonstrated that clayey silt type mangrove sediment was an effective adsorbent with higher efficiency for Sb (V) than Sb (III). Background level of Sb in the sediment was 0.35–0.78% of the maximum adsorption capacity (Γ_m). Out of the two distinct type of sorption sites governing mobility and bioavailability of Sb in the sediment, site 1 (Humic acid) showed higher affinity for Sb than the site II (oxyhydroxide). Sb adsorption was strongly influenced by temperature, salinity, and pH, which may be altered by long-term changes in climate and rainfall pattern.     

A 1H-NMR study on the blue copper protein amicyanin from Thiobacillus versutus. Resonance identifications, structural rearrangements and determination of the electron self-exchange rate constant

A number of resonances in the 1H-NMR spectra of reduced and oxidised amicyanin from Thiobacillus versutus have been identified by one- and two-dimensional NMR techniques. The second-order electron self-exchange rate constant (8.5 x 10(4) M-1.s-1; pH = 7.4; T = 308.5 K) was determined by measuring the line broadening of six singlets in slightly oxidised solutions of the protein. A large increase in electron exchange rate is observed in the presence of ferrocyanide. The copper atom in the reactive centre of the protein appears to be coordinated by nitrogens from two histidines and sulfurs from a methionine and a cysteine. One of the ligand histidines becomes protonated at low pH [pK*a = 6.74 (+/- 0.02)], the asterisk indicating value uncorrected for the deuterium isotope effect] in reduced amicyanin. This is the first example of a non-photosynthetic blue copper protein in which a ligand histidine becomes protonated at low pH. A small pH-independent conformational rearrangement occurs upon oxidation.     

Effects of trivalent antimony on human erythrocyte glutathione-S-transferases

Trivalent antimony (SB3+) in the form of potassium antimony tartrate was found to be an inhibitor of glutathione-S-transferases (GST) from human erythrocytes with a 50% inhibition concentration (IC50) of 0.05 mM. The inhibition was, however, incomplete with 15-20% of the GST activity remaining unaffected. In comparison, ethacrynic acid, a known inhibitor of GST, was tenfold more potent and affected close to 100% inhibition. Pentavalent antimony (SB5+) in the form of sodium stibogluconate had no effect on GST. Group V metalloids such as arsenite was slightly inhibitory, and arsenate was noninhibitory. When compared with five heavy metals, the inhibitory potency followed the order of SB3+ > Hg2+, Cu2+ > Cd 2+ > Cr3+ > Fe2+ x SB3+ inhibition of GST was competitive against the substrate 1-chloro-2,4-dinitrobenzene (CDNB) with an apparent Ki of 0.018 mM. Increasing the glutathione (GSH) concentration, however, produced a biphasic response: at concentrations below 1 mM, GSH was noncompetitive against SB3+, but at 1 mM and higher it was apparently competitive. A concurrent study of interactions between GSH, CDNB, and SB3+ showed that there was a significant nonenzymatic conjugation of CDNB at high GSH concentrations, which was suppressed by SB3+. The presence of albumin (500 mg/dL), or up to 5 mM N-acetylcysteine, cysteine, or ethylenediamine tetraacetic acid (EDTA) did not protect GST from the inhibitory effect of SB3+. The ability of erythrocyte GST to conjugate CDNB, which was measured directly by the formation of dinitrophenyl-glutathione (DNP-glutathione), was reduced by approximately 20 and 33%, respectively, in the presence of 2 and 10 mM SB3+, and nearly abolished with the addition of 0.2 mM ethacrynic acid. Based on these inhibition characteristics and the preferential accumulation of SB3+ in mammalian erythrocytes, it may be deduced that in the case of high antimonial intake, for example, during therapeutic treatment of Leishmaniasis, SB3+ levels in erythrocytes may be high enough to depress GST activity, which might compromise the ability of erythrocytes to detoxify electrophilic xenotbiotics.     

Non-enzymatic glutathione conjugation of 2-nitroso-6-methyldipyrido [1,2-a: 3',2'-d] imidazole (NO-Glu-P-1) in vitro: N-hydroxy-sulfonamide, a new binding form of arylnitroso compounds and thiols

In order to study the possible detoxification mechanisms of the carcinogenic arylamine, 2-amino-6-methyldipyrido[1,2-a: 3',2'-d]imidazole (Glu-P-1), the in vitro non-enzymatic reaction of 2-nitroso-6-methyldipyrido[1,2-a: 3',2'-d]imidazole (NO-Glu-P-1) with reduced glutathione (GSH) was examined at pH 7.4 under both aerobic and anaerobic conditions. Two GSH-arylamine adducts were isolated and found to contain the Glu-P-1 and GSH moieties in a 1:1 molar ratio via an N-S linkage. Their structures were assigned as sulfinamide (-NH-SO-) and N-hydroxy-sulfonamide (-N(OH)-SO2-) by their behaviour under acidic and basic conditions and by UV-VIS, 1H-NMR, infrared and mass spectrometries. Also, a N-hydroxy-sulfonamide adduct was produced when NO-Glu-P-1 and cysteine were reacted at pH 7.4. The N-hydroxy-sulfonamide structure is a new binding form between arylnitroso compounds and thiols. The formation of these adducts may also take place in vivo as a detoxification of toxic arylamines since GSH is abundant in organs such as liver or kidney.     

Reactions of nitrosobenzene with reduced glutathione.

Nitrosobenzene (NOB) formed acid labile conjugates with reduced glutathione (GSH) and hemoglobin within red cells. In vitro, NOB rapidly reacted with GSH with formation of phenylhydroxylamine (PH), oxidized glutathione (GSSG), and a water-soluble compound identified as glutathionesulfinanilide (GSO-AN). Free aniline (AN), aminophenols and azoxybenzene were not detected. The proportion of PH formed increased with increasing GSH concentration and at higher pH values. Spectroscopic analysis revealed the formation of a labile adduct following a second order reaction (K = 5 x 10(3) M-1 . sec-1 at pH 7.4 and 37 degrees). This reaction was reversible because nearly all NOB could be extracted with ether from the labile intermediate. On the other hand, the labile intermediate was transformed into GSO-AN (with increasing rate at lower pH values) or it was cleaved by GSH with formation of GSSG and PH. Intermediate formation of NOB and thiol radicals was ruled out by analysis of the equilibrium data. A tentative scheme is presented for the proposed reaction mechanism.     

Syntheses, crystal structures and antimicrobial activities of 6-coordinate antimony(III) complexes with tridentate 2-acetylpyridine thiosemicarbazone, bis(thiosemicarbazone) and semicarbazone ligands

Five novel antimony(III) complexes with the mono- and bis(thiosemicarbazone) ligands of 2N1S or 4N2S donor atoms, N'-[1-(2-pyridyl)ethylidene]morpholine-4-carbothiohydrazide (Hmtsc, L1) and bis[N'-[1-(2-pyridyl)ethylidene]]-1,4-piperazinedicarbothiohydrazide (H(2)ptsc, L7), and the tridentate semicarbazone ligand of 2N1O donor atoms, 2-acetylpyridine semicarbazone (Hasc, L2b), were prepared by reactions of SbCl(3) or SbBr(3), and characterized by elemental analysis, TG/DTA, FT-IR and (1)H NMR spectroscopy. The crystal and molecular structures of five antimony(III) complexes were determined by single-crystal X-ray structure analysis. The neutral, 6-coordinate antimony(III) complexes ([Sb(mtsc)Cl(2)] 1, [Sb(mtsc)Br(2)] 2, [Sb(asc)Cl(2)] 3 and [Sb(asc)Br(2)] 4) are depicted with one electron pair (5s(2)) of the antimony(III) atom, deprotonated forms of multidentate thiosemicarbazone or semicarbazone ligands, and two monodentate halogen ligands, respectively. In the dimer complex 5 ([Sb(2)(ptsc)Cl(4)]) with the ligand in which two tridentate thiosemicarbazone moieties are connected by the piperazine moiety, each antimony(III) was also described as a neutral 6-coordinate structure. These antimony(III) complexes were thermally stable around 200 degrees C. Water-soluble antimony(III) complexes 1 and 2 showed moderate antimicrobial activities against Gram-positive (Bacillus subtilis and Staphylococcus aureus) and -negative bacteria (Escherichia coli and Pseudomonas aeruginosa), yeasts (Candida albicans and Saccharomyces cerevisiae) and molds (Aspergillus niger and Penicillium citrinum). Complex 5 showed moderate antimicrobial activities against four bacteria, and two molds, while the ligand itself showed only modest antimicrobial activities against selected bacteria (B. subtilis, E. coli and S. aureus). The molecular structures and antimicrobial activities of antimony(III) complexes were compared with those of bismuth(III) complexes in the same 15 group in the periodic table.