Reaction of 4-hydroxynonenal with some thiol-containing radioprotective agents or their active metabolites

The rate of reaction of several radioprotective agents or their active metabolites with 4-hydroxynonenal (4HNE) was studied and compared to the rate of reaction with cysteine (Cys) and glutathione (GSH). The agents studied were: mercapto ethylamine (MEA); 2(3-aminopropyl) aminoethanethiol (WR1065); S-2-aminoethylisothiouronium bromide-hydrobromide (AET); 1,4-dithiothreitol (DTT); 1,4-dithioerythritol (DTE); N-2(2-mercaptopropionyl)-glycine (MPG); penicillamine hydrochloride (PA); N-acetylcysteine (NAC); 2–3 dimercapto-1 propane sulfonic acid (DMPS); 2,3-dimercaptopropanol (BAL), and meso 2,3 dimercapto succinic acid (DMS). All of them reacted with 4HNE. MEA and WR1065 were the most reactive thiols, and PA and DMS were the least reactive thiols. All the others reacted at rates comparable to or higher than that of cysteine or GSH. The potential role of this type of interactions in the protective action of these drugs against deleterious effects of radiation or carbon tetrachloride is analyzed.     

Cadmium inhibits delta-aminolevulinate dehydratase from rat lung in vitro: interaction with chelating and antioxidant agents

The effect of cadmium (Cd(2+)) on delta-aminolevulinate dehydratase (delta-ALA-D) activity from rat lung in vitro was investigated. delta-ALA-D activity, a parameter for metal intoxication, has been reported as a target of Cd(2+) in different tissues. The protective effect of monotherapies with dithiol chelating (meso-2,3-dimercaptosuccinic acid (DMSA) and 2,3-dimercaptopropane-1-sulfonic acid (DMPS)) or antioxidant agents (ascorbic acid, diphenyl diselenide (PhSe)(2), and N-acetylcysteine (NAC)) was evaluated. The effect of a combined therapy (dithiol chelatingxantioxidant agent) was also studied. Zinc chloride (ZnCl(2)) and dithiothreitol (DTT) were used to investigate the mechanisms involved in cadmium, chelating and antioxidant effects on delta-ALA-D activity. Cadmium inhibited rat lung delta-ALA-D activity at low concentrations. DTT (3mM), but not ZnCl(2) (100microM), protected the inhibition of enzyme activity caused by Cd(2+). Chelating agents were not effective in restoring the enzyme activity. DMPS and DMSA presented inhibitory effect on enzyme activity. DTT restored the inhibition caused by both chelating agents, but ZnCl(2) restored only the inhibitory effect induced by DMSA. These compounds caused a marked potentiation of delta-ALA-D inhibition induced by Cd(2+). ZnCl(2) did not restore inhibition of enzyme activity caused by Cd(2+) plus chelating agents. Conversely, DTT restored the inhibition induced by Cd(2+)/DMSA, but not by Cd(2+)/DMPS. Antioxidants were not effective in ameliorating delta-ALA-D inhibition induced by Cd(2+), whereas ascorbic acid potentiated the enzyme inhibition induced by this metal. A combined effect of Cd(2+)xDMPSx(PhSe)(2) and Cd(2+)xDMPSxNAC was observed. There was no combined effect of Cd(2+)xchelatorxantioxidants when DMSA was used. This study demonstrated that Cd(2+)inhibited delta-ALA-D activity and chelating and antioxidant agents, alone or combined, did not restore the enzyme activity. In contrast, these compounds potentiated the inhibition induced by Cd(2+) in rat lung.     

Determination and metabolism of dithiol-chelating agents: electrolytic and chemical reduction of oxidized dithiols in urine

The presence of oxidized species of the dithiol-chelating agents, meso-2,3-dimercaptosuccinic acid (DMSA) and 2,3-dimercaptopropane-1-sulfonic acid (DMPS), in human urine was determined by chemical and electrolytic reduction methods. Urine from a human given either DMSA or DMPS was treated with electrolysis, dithiothreitol, or sodium tetrahydridoborate (NaBH4). The SH groups were derivatized with monobromobimane for the determination of unaltered dithiols. Total dithiol (unaltered and oxidized) was determined by reduction followed by derivatization with monobromobimane. The bimane derivatives were identified and quantified by HPLC and fluorescence. Although all three reduction methods gave similar results, electrolytic reduction of oxidized DMSA and chemical reduction with NaBH4 of oxidized DMPS are recommended based upon both day to day reproducibility and recovery of standards. After reduction a 4-fold increase in DMSA and a 20-fold increase in DMPS were found in urine by 12 h after an oral dose of DMSA or DMPS. These new methods for the determination of dithiols and their oxidized forms should lead to a better understanding of the metabolic properties of these increasingly important orally effective chelating agents.     

Chelation therapy in intoxications with mercury,lead and copper

In the present review we provide an update of the appropriate use of chelating agents in the treatment of intoxications with compounds of mercury, lead and copper. The relatively new chelators meso-2,3-dimercaptosuccinic acid (DMSA) and 2,3-dimercapto-propanesulphonate (DMPS) can effectively mobilize deposits of mercury as well as of lead into the urine. These drugs can be administered orally and have relatively low toxicity compared to the classical antidote dimercaptopropanol (BAL). d-Penicillamine has been widely used in copper overload, although 2,3-dimercaptosuccinic acid or tetrathiomolybdate may be more suitable alternatives today. In copper-toxicity, a free radical scavenger might be recommended as adjuvant to the chelator therapy.     

Chelation in metal intoxication—Principles and paradigms

The present review provides an update of the general principles for the investigation and use of chelating agents in the treatment of intoxications by metals. The clinical use of the old chelators EDTA (ethylenediamine tetraacetate) and BAL (2,3-dimercaptopropanol) is now limited due to the inconvenience of parenteral administration, their own toxicity and tendency to increase the neurotoxicity of several metals. The hydrophilic dithiol chelators DMSA (meso-2,3-dimercaptosuccinic acid) and DMPS (2,3-dimercapto-propanesulphonate) are less toxic and more efficient than BAL in the clinical treatment of heavy metal poisoning, and available as capsules for oral use. In copper overload, DMSA appears to be a potent antidote, although d-penicillamine is still widely used. In the chelation of iron, the thiols are inefficient, since iron has higher affinity for ligands with nitrogen and oxygen, but the new oral iron antidotes deferiprone and desferasirox have entered into the clinical arena. Comparisons of these agents and deferoxamine infusions are in progress. General principles for research and development of new chelators are briefly outlined in this review.     

Effect of dithiol chelating agents on [3H]MK-801 and [3H]glutamate binding to synaptic plasma membranes

2,3-Dimercaptopropanol (BAL- British Anti-Lewesite) is a dithiol chelating agent used for the treatment of heavy metal poisoning, however, BAL can produce neurotoxic effects in a variety of situations. Based on the low therapeutic efficiency of BAL other dithiols were developed and DMSA (meso-2,3-dimercaptosuccinic acid) and DMPS (2,3-dimercaptopropane-1-sulfonic acid) are becoming used for treatments of humans exposed to heavy metals. In the present investigation the effect of dithiols in the glutamatergic system was examined. The results showed that BAL inhibited [³H]MK-801 and [³H]glutamate binding in a concentration-dependent manner. At 100 M BAL and DMSA caused a significantly inhibition of [³H]MK-801 binding to brain membranes (p < 0.05 by Duncan's multiple range test). BAL at 100 M caused an inhibition of 40% on [³H]glutamate binding. DMPS and DMSA had no significant effect on [³H]glutamate binding. Dithiotreitol (DTT), abolished the inhibitory effect of BAL on [³H]MK-801 binding. The protection exerted by DTT suggests that BAL inhibit [³H]MK-801 binding by interacting with cysteinyl residues that are important for redox modulation of receptor responses. ZnCl₂ inhibited [³H]glutamate and [³H]MK-801 binding to brain synaptic membrane; nevertheless, the inhibitory effect was slight more accentuated for [³H]MK-801 than [³H]glutamate binding (p < 0.05). The inhibition caused by 10 M ZnCl₂ on [³H]MK-801 binding was attenuated by BAL. The findings present in this study may provide the evidence that BAL affect the glutamatergic system and these effects can contributed to explain, at least in part, why BAL, in contrast to DMPS and DMSA is neurotoxic.     

Role of chelating agents and antioxidants in beryllium induced toxicity

The present study was conducted to evaluate the therapeutic effectiveness of chelating agents [glutathione, 2,3 dimercapto propane sulfonic acid (DMPS) and D-penicillamine (DPA)] in combination with antioxidant (sodium selenite) in beryllium induced toxicity in female rats. A bolus dose of 50mg/kg-beryllium nitrate was administered singly followed by chelation therapy with GSH, DMPS + Se and DPA + Se at various durations of 1,3 and 7 days respectively. Results revealed a significant fall in the glycogen content, whereas, a marginal fall in the protein was also observed. The enzymatic activity of alkaline phosphatase and adenosine triphosphatase was depleted; on the contrary, there was a significant rise in the acid phosphatase and glucose-6-phosphatase pattern. A rise in the hepatic lipid peroxidation activity is a direct indication of oxidative damage resulting in free radical generation. The distribution of the metal by atomic absorption spectrophotometry revealed an increased concentration of beryllium in liver and kidney, followed by lung and uterus. The relative ability of three chelating agents to act as antagonists, for acute beryllium poisoning, have been examined in liver, kidney, lungs and uterus. The appreciable change in the beryllium concentration in various organs is duration dependent during the entire period being highly significant at 7 days regimen. Biochemical and distribution studies reveal that DPA + Se was the most effective therapeutic agent followed by DMPS + Se and GSH.     

Factors influencing the efficiency of chelation therapy.

The purpose of the present study was to obtain new data on the effect of age, route, dose and time of metal and chelating agent administration on the efficiency of chelation therapy. The experiments were performed on 1-2 and 6-week-old rats which received radioisotopes of metals--203Pb, 115 mCd, 203Hg and 141Ce intraperitoneally or orally. Chelating agents calcium ethylenediaminetetraacetate (CaEDTA), calcium and zinc diethylenetriaminepentaacetate (CaDTPA, ZnDTPA), 2,3-dimercapto-propane-sulfonate-1 (DMPS), dimercaptosuccinic acid (DMSA) and sodium N-(4-methoxybenzyl)-D-glucamine dithiocarbamate monohydrate (MeOBDCG) were administered twice by intraperitoneal or oral administration as early (immediately and 24 hr after metals) or delayed treatment (24 and 48 or 48 and 72 hr after metals). The animals were killed six days after metal administration and the retention was determined in the whole body, carcass and gut. After intraperitoneal administration of metals and chelating agents chelation therapy had much lower efficacy in younger than older animals. After ingestion of metals oral chelation therapy was more effective in younger than older animals. In suckling rats the treatment effectively reduced metal retention and this was mostly due to decrease in gut retention. This treatment in sucklings was also very effective in condition of late administration. In older rats early oral DMPS treatment after 203Hg ingestion is contraindicated since it increases significantly mercury retention while DMSA and ZnDTPA treatments reduced mercury retention. Delayed oral treatment with ZnDTPA and DMSA caused increased cadmium retention in older rats and decreased retention in sucklings. Opposite to results with CaDTPA, MeOBDCG was effective in reducing cadmium retention also when given as delayed treatment.(ABSTRACT TRUNCATED AT 250 WORDS)     

Response of arsenic-induced oxidative stress,DNA damage,and metal imbalance to combined administration of DMSA and monoisoamyl-DMSA during chronic arsenic poisoning in rats

Arsenic and its compounds cause adverse health effects in humans. Current treatment employs administration of thiol chelators, such as meso-2,3-dimercaptosuccinic acid (DMSA) and sodium 2,3-dimercaptopropane 1-sulfonate (DMPS), which facilitate its excretion from the body. However, these chelating agents are compromised by number of limitations due to their lipophobic nature, particularly in case of chronic poisoning. Combination therapy is a new approach to ensure enhanced removal of metal from the body, reduced doses of potentially toxic chelators, and no redistribution of metal from one organ to another, following chronic metal exposure. The present study attempts to investigate dose-related effects of two thiol chelators, DMSA and one of its new analogues, monoisoamyl dimercaptosuccinic acid (MiADMSA), when administered in combination with the aim of achieving normalization of altered biochemical parameters suggestive of oxidative stress and depletion of inorganic arsenic following chronic arsenic exposure. Twenty-five adult male Wistar rats were given 25 ppm arsenic for 10 weeks followed by chelation therapy with the above chelating agents at a dose of 0.3 mmol/kg (orally) when administered individually or 0.15 mmol/kg and 0.3 mmol/kg (once daily for 5 consecutive days), respectively, when administered in combination. Arsenic exposure led to the inhibition of blood δ-aminolevulinic acid dehydratase (ALAD) activity and depletion of glutathione (GSH) level. These changes were accompanied by significant depletion of hemoglobin, RBC and Hct as well as blood superoxide dismutase (SOD) acitivity. There was an increase in hepatic and renal levels of thiobarbituric acid-reactive substances, while GSH:GSSG ratio decreased significantly, accompanied by a significant increase in metallothionein (MT) in hepatocytes. DNA damage based on denaturing polyacrylamide gel electrophoresis revealed significant loss in the integrity of DNA extracted from the liver of arsenic-exposed rats compared to that of normal animals. These changes were accompanied by a significant elevation in blood and soft-tissue arsenic concentration. Co-administration of DMSA and MiADMSA at lower dose (0.15 mmol/kg) was most effective not only in reducing arsenic-induced oxidative stress but also in depleting arsenic from blood and soft tissues compared to other treatments. This combination was also able to repair DNA damage caused following arsenic exposure. We thus recommend combined administration of DMSA and MiADMSA for achieving optimum effects of chelation therapy.     

Comparison of the effectiveness of 2,3-dimercaptopropanol (BAL) andmeso-2,3-dimercaptosuccinic acid (DMSA) as protective agents against mercuric chloride-induced nephrotoxicity in rats

The effectiveness of 2,3-dimercaptopropanol (BAL) andmeso-2,3-dimercaptosuccinic acid (DMSA) on HgCl₂-induced nephrotoxicity was studied in the rat. Seven groups of adult male rats were given a single sc toxic dose of HgCl₂ (0.68 mg/kg) followed by 0.9% saline (positive control group), BAL (15, 30, and 60 mg/kg) or DMSA (50, 100, and 200 mg/kg) administered ip at 0, 24, 48, and 72 h thereafter. Although the renal function of HgCl₂-exposed rats was slightly improved after BAL administration, Hg concentrations in the kidney were only reduced at 60 mg/kg. In addition, the protective effect of BAL was not dose-related. In contrast to BAL, DMSA was effective in increasing the urinary excretion of Hg and in reducing the renal Hg content. These results show that DMSA would be more effective than BAL in preventing or in protecting against inorganic Hg-induced nephrotoxicity.     

Therapeutic potential of meso 2,3-dimercaptosuccinic acid or 2,3-dimercaptopropane 1-sulfonate in chronic arsenic intoxication in rats

The therapeutic efficacy of two thiol chelators, meso 2,3-dimercaptosuccinic acid (DMSA) or 2,3-dimercaptopropane sulfonate (DMPS) in treating chronic arsenic intoxication was investigated in male rats. Both the chelators were effective in promoting urinary arsenic excretion and restoring arsenic induced inhibition of blood -aminolevulinic acid dehydratase activity and hepatic glutathione level. Elevation of urinary -aminolevulinic acid excretion and arsenic concentration in blood, liver and kidneys were reduced significantly by both the chelators. Histopathological lesions induced by arsenic were also effectively reduced by the above chelators. DMSA being more effective than DMPS. The results suggest DMSA and DMPS to be effective antidotes for treating chronic arsenic toxicity in experimental animals.     

Arsenic induced blood and brain oxidative stress and its response to some thiol chelators in rats

Chronic arsenic toxicity is a widespread problem, not only in India and Bangladesh but also in various other regions of the world. Exposure to arsenic may occur from natural or industrial sources. The treatment that is in use at present employs administration of thiol chelators, such as meso 2,3-dimercaptosuccinic acid (DMSA) and sodium 2,3-dimercaptopropane 1-sulfonate (DMPS), which facilitate its excretion from the body. However, these chelating agents are compromised with number of limitations due to their lipophobic nature, particularly for their use in cases of chronic poisoning. During chronic exposure, arsenic gains access into the cell and it becomes mandatory for a drug to cross cell membrane to chelate intracellular arsenic. To address this problem, analogs of DMSA having lipophilic character, were examined against chronic arsenic poisoning in experimental animals. In the present study, therapeutic efficacy of meso 2,3-dimercaptosuccinic acid (DMSA), sodium 2,3-dimercaptopropane 1-sulfonate (DMPS), monoisoamyl DMSA (MiADMSA) were compared in terms of reducing arsenic burden, as well as recovery in the altered biochemical variables particularly suggestive of oxidative stress. Adult male Wistar rats were given 100-ppm arsenic for 10 weeks followed by chelation therapy with the above chelating agents at a dose of 50 mg/Kg (orally) once daily for 5 consecutive days. Arsenic exposure resulted in marked elevation in reactive oxygen species (ROS) in blood, inhibition of ALAD activity and depletion of GSH. These changes were accompanied by significant decline in blood hemoglobin level. MiADMSA was the most effective chelator in reducing ROS in red blood cells, and in restoring blood ALAD compared to two other chelators. Brain superoxide dismutase (SOD) and glutathione peroxidase (GPx) decreased, while ROS and TBARS increased significantly following arsenic exposure. There was a significant increase in the activity of glutathione-S-transferase (GST) with a corresponding decline in its substrate i.e. glutathione. Among all the three chelators, MiADMSA showed maximum reduction in the level of ROS in brain. Additionally, administration of MiADMSA was most effective in counteracting arsenic induced inhibition in brain ALAD, SOD and GPx activity. Based on these results and in particular higher metal decorporation from blood and brain, we suggest MiADMSA to be a potential drug of choice for the treatment of chronic arsenic poisoning. However, further studies are required for the choice of appropriate dose, duration of treatment and possible effects on other major organs.     

Safety and efficacy of oral DMSA therapy for children with autism spectrum disorders: Part B - Behavioral results

Background

This study investigated the effects of oral dimercapto succinic acid (DMSA) therapy on the behavioural symptoms of children with autism spectrum disorders (ASD) ages 3-8 years.

Methods

Phase 1 involved 65 children with ASD who received one round of DMSA (3 days). Participants who had high urinary excretion of toxic metals were selected to continue on to phase 2. In phase 2, 49 participants were randomly assigned in a double-blind design to receive an additional 6 rounds of either DMSA or placebo.

Results

The groups receiving one round and seven rounds of DMSA had significant improvements on all the assessment measures. For the seven round group, the degree of improvement on the assessment measures could be partially explained by a regression analysis based on excretion of toxic metals and changes in glutathione (adjusted R² of 0.28-0.75, p < 0.02 in all cases). One round of DMSA had nearly the same benefit as seven rounds. The assessment measures correlated reasonably with one another at the beginning of the study (r = 0.60-0.87) and even better at the end of the study (r = 0.63-0.94).

Conclusion

Overall, both one and seven rounds of DMSA therapy seems to be reasonably safe in children with ASD who have high urinary excretion of toxic metals, and possibly helpful in reducing some of the symptoms of autism in those children.     

In vitro assessment of chelating agents with regard to their abstraction efficiency of Cd(2+) bound to plasma proteins

The exposure of various human populations to Cd(2+) is of increasing health concern. After its gastrointestinal absorption into the bloodstream, Cd(2+) binds to α(2)-macroglobulin and serum albumin. Although animal studies have demonstrated that meso-2,3-dimercaptosuccinic acid (DMSA) and diethylenetriamine pentaacetic acid (DTPA) can effectively mobilize Cd(2+) to urine and decrease the Cd concentrations of the kidneys, the liver and the brain, not much is known about the abstraction of Cd(2+) from blood plasma proteins. We prepared a stock of Cd(2+) spiked rabbit plasma (2.0 μg of Cd(2+)/mL) and analyzed aliquots by size exclusion chromatography coupled on-line to an inductively coupled plasma atomic emission spectrometer (SEC-ICP-AES) while simultaneously monitoring the emission lines of Ca, Cd, Cu, Fe, and Zn. After the addition of 0.33 mM, 0.66 mM or 0.99 mM of DMSA, DTPA, 2,3-dimercapto-1-propanesulfonic acid (DMPS) or N-acetyl-l-cysteine (NAC) to plasma aliquots, the obtained mixtures were analyzed by SEC-ICP-AES after 5 min and 30 min. None of the investigated compounds adversely affected the plasma distribution of Fe at all investigated doses. At 0.33 mM, DTPA was most effective at mobilizing plasma protein bound Cd(2+) to a ～5 kDa Cd-species (100% removal), followed by DMPS (94%), DMSA (83%) and NAC (3%). All investigated compounds also mobilized Zn(2+) from plasma proteins to ～5 kDa Zn-species (DTPA: 80% removal; DMPS: 63%; DMSA: 29% and NAC: 3%). The addition of DTPA resulted in the dose-dependent elution of a [Ca-DTPA](3-) complex. Based on these results, 0.33 mM DMSA represents the best compromise that can be achieved between maximizing the abstraction of Cd(2+) from plasma proteins (83%), while minimizing the mobilization of Zn(2+) from plasma proteins (29%), and avoiding the complexation of Ca(2+).     

Gas chromatographic analysis of urinary dimercaptosuccinic acid

The therapeutic use of disulfhydryl compounds such as 2,3-dimercaptosuccinic acid (DMSA) for the treatment of heavy metal poisoning has generated a requirement for specific and sensitive methods to determine those compounds in biological media. We have developed a gas chromatographic assay for DMSA in urine. The use of capillary column technology eliminates the requirement for a preliminary clean-up step. Samples are first reduced electrochemically to liberate DMSA present as disulfides. The reduced product is then extracted into ethyl acetate and the organic phase removed by evaporation. The residue is derivatized with N,O-bis(trimethylsilyl)acetamide for gas chromatography. The silylated DMSA derivative is then detected with a flame ionization detector. The detection limit for DMSA is 1.9 nmol per 1-μl aliquot of derivatized extract injected on column (detector sensitivity at 1·10⁻¹¹ A/mV). The utility of the method was demonstrated by analyzing the urine of rats orally dosed with DMSA.     

Evaluation of the developmental effects on mice after prenatal, or pre- and postnatal exposure to 2,3-dimercaptopropane-1-sulfonic acid (DMPS)

The sodium salt of 2,3-dimercaptopropane-1-sulfonic acid (DMPS), a water soluble metal complexing agent, was administered to four groups of pregnant Swiss mice at 0, 70, 210, and 630 mg/kg/day by two dosing schedules: gestation day 14 until birth (prenatal exposure), and gestation day 14 until postnatal day 21 (pre- and postnatal section). Dams were allowed to deliver and the number of live and dead pups recorded. Each pup was sexed and weighed on days 0, 4, 14, and 21. Also, pinna detachment, incisor eruption and eye opening were monitored. No adverse effects on offspring survival or development were evident in either exposures at doses employed in this study. The "no observable effect level" (NOEL) for health hazard to the developing fetus or pup was 630 mg DMPS/kg/day. This dose is much higher than the amounts of DMPS usually administered in human heavy metal poisoning.     

Interactions of the chelating agent 2,3-dimercapto-propane-1-sulfonate with red blood cells in vitro. I. Evidence for carrier mediated transport

Uptake of the water soluble 1,2-dimercaptopropanol (BAL) derivative 2,3-dimercapto-1-sulfonate (DMPS) into human red blood cells was found in vitro and the mode of penetration studied in detail. The compound entered erythrocytes in a concentration dependent manner. In contrast to sealed ghosts where inside and outside concentrations reached the same value, DMPS accumulated in intact erythrocytes. Since no binding of DMPS could be detected, the reason for accumulation was assumed to be a conversion of DMPS into chelates or metabolites which penetrated the membrane in a slower rate. A facilitated transport of DMPS mediated by the anion carrier protein was concluded on the basis of the following similarities with the anion transport: inhibition of [¹⁴C]DMPS-uptake by N-ethylmaleimide (NEM), tetrathionate (90%), sulfate (50%), 5,5′-dithio bis(2-nitrobenzoic acid) (DTNB) (25%); inhibition of uptake and efflux by 4,4′-diisothiocyano-2,2′-stilbene disulfonate (DIDS) (80%), dipyridamole (55%); temperature dependency (activation energy 24 K_cal/mol); pH-dependency (pH optimum about 6.9); counter-transport; activation of uptake by preincubation with DMPS (transmembrane effect).     

Evaluation of methyl mercury chelating agents using red blood cells and isolated hepatocytes

The relative efficacy of thiol-containing mercurial scavengers was assayed by using cellular suspensions of erythrocytes or isolated hepatocytes. The blood cells incubated in a buffer (pH 7.4) containing 1 mM glucose (10% hematocrit) were exposed to 5 μM methyl mercuric chloride. In the absence of extracellular thiols the red blood cells took up more than 90% of methyl mercury from the surrounding medium during 5–10 min. This uptake was almost completely inhibited by dimercaptosuccinic acid (DMSA) (1 mM) and the same chelant could rapidly remove 80% of the mercury from ‘pre-loaded’ erythrocytes. Hepatocytes prepared according to the method of Seglen [11] in a suspension of 10⁶ cells/ml in a buffer containing 5 mM glucose and 5 mg/ml of bovine serum albumin were also exposed to methyl mercuric chloride (4 μM). Almost 50% of the mercurial was taken up by the cells slowly during the incubation period of 240 min. DMSA (1 mM) almost completely blocked the methyl mercury binding by the hepatocytes. 2-Mercaptopropionylglycin (Thiola) or mercaptosuccinic acid (MSA) was almost as effective mercurial scavengers as DMSA in hepatocytes and in red blood cells. Diethyldithiocarbamate (DDC) and dimercaptopropanol (BAL) were considerably less effective than DMSA to inhibit the mercurial binding to hepatocytes. Experiments in vivo have shown that DMSA is a better mercurial chelator than Thiola or MSA, whereas DDC and BAL may both be considered to be inapplicable in methyl mercury poisonings. Our cellular assay provides preliminary information of the efficiency of chelating thiols and may serve as a useful first approximation when planning further experiments.     

Interaction Between Metals and Chelating Agents Affects Glutamate Binding on Brain Synaptic Membranes

The present study investigates the possible effects of Hg²⁺, Pb²⁺, and Cd²⁺ on [³H]-glutamate binding. To better understand the role of the thiol-disulfide status on the toxicity of such metals toward glutamatergic neurotransmission, we used three thiol chelating agents, 2,3-dimercaptopropanol (BAL), 2,3-dimercaptopropane 1-sulfonate (DMPS), and meso-2,3-dimercaptosuccinic acid (DMSA). Dithiotreitol (DTT) was tested for its ability to prevent metals-induced inhibition on [³H]-glutamate binding. Hg²⁺, Pb²⁺, and Cd²⁺ showed a concentration-dependent inhibition on [³H]-glutamate binding, and mercury was the most effective inhibitor. BAL did not prevent [³H]-glutamate binding inhibition by Hg²⁺, Cd²⁺, and Pb²⁺. However, DMPS and DMSA prevented the inhibition caused by Cd²⁺ and Pb²⁺, but not by Hg²⁺. DTT did not prevent the inhibition on [³H]-glutamate binding caused by 10 M Hg²⁺. In contrast, it was able to partially prevent [³H]-glutamate binding inhibition caused by 40 M Pb²⁺ and Cd²⁺. These results demonstrated that the heavy metals present an inhibitory effect on [³H]-glutamate binding. In addition, BAL was less effective to protect [³H]-glutamate binding inhibition caused by these metals than other chelating agents studied.     

On the interaction of vanadium species with meso-2,3-dimercaptosuccinic acid

The interaction of the VO²⁺ cation with meso-2,3-dimercaptosuccinic acid (DMSA) was investigated by electron absorption spectroscopy in aqueous solution at different pH values. The spectral behavior, complemented with a spectrophotometric titration, shows the generation of a [VO(DMSA)₂]²⁻ complex in which the oxocation interacts with two pairs of deprotonated-SH groups of the acid. It was also found that DMSA rapidly reduces VO₃ ⁻ to VO²⁺, which might be chelated by an excess of the acid. DMSA can also produce the partial reduction of a V₂O₅ suspension at pH=5.2. The results of this study suggest that DMSA might be a potentially useful detoxification agent for vanadium.