Mechanistic studies on Pyrobaculum calidifontis porphobilinogen synthase (5-aminolevulinic acid dehydratase).

Porphobilinogen synthase (PBG synthase) gene from Pyrobaculum calidifontis was cloned and expressed in E. coli. The recombinant enzyme was purified as an octamer and was found by mass spectrometry to have a subunit Mr of 37676.59 (calculated, 37676.3). The enzyme showed high thermal stability and retained almost all of its activity after incubation at 70 °C for 16 h in the presence of ß-mercaptoethanol (ß-ME) and zinc chloride. However, in the absence of the latter the enzyme was inactivated after 16 h although it regained full activity in the presence of ß-ME and zinc chloride. The protein contained 4 mol of tightly bound zinc per octamer. Further, 4 mol of low affinity zinc could be incorporated following incubation with exogenous zinc salts. The enzyme was inactivated by incubation with levulinic acid followed by treatment with sodium borohydride. Tryptic digest of the modified enzyme and mass spectrometric analysis showed that Lys257 was the site of modification, which has previously been shown to be the site for the binding of 5-aminolevulinic acid giving rise to the propionate-half of porphobilinogen. P. calidifontis PBG synthase was inactivated by 5-chlorolevulinic acid and the residue modified was shown to be the central cysteine (Cys127) of the zinc-binding cysteine-triad, comprising Cys125, 127, 135. The present results in conjunction with earlier findings on zinc containing PBG synthases, are discussed which advocate that the catalytic role of zinc in the activation of the 5-aminolevulinic acid molecule forming the acetate-half of PBG is possible.

Interaction of metals from group 10 (Ni, Pd, and Pt) and 11 (Cu, Ag, and Au) with human blood δ-ALA-D: in vitro and in silico studies.

Mammalian δ-aminolevulinate dehydratase (δ-ALA-D) is a metalloenzyme, which requires Zn(II) and reduced thiol groups for catalytic activity, and is an important molecular target for the widespread environmental toxic metals. The δ-ALA-D inhibition mechanism by metals of Group 10 (Ni, Pd, and Pt) and 11 (Cu, Ag, and Au) of the periodic table has not yet been determined. The objective of this study was to characterize the molecular mechanism of δ-ALA-D inhibition caused by the elements of groups 10 and 11 using in vitro (δ-ALA-D activity from human erythrocytes) and in silico (docking simulations) methods. Our results showed that Ni(II) and Pd(II) caused a small inhibition (~ 10%) of the δ-ALA-D. Pt(II) and Pt(IV) significantly inhibited the enzyme (75% and 44%, respectively), but this inhibition was attenuated by Zn(II) and dithiothreitol (DTT). In group 11, all metals inhibited δ-ALA-D with great potency (~ 70-90%). In the presence of Zn(II) and DTT, the enzyme activity was restored to the control levels. The in silico molecular docking data suggest that the coordination of the ions Pt(II), Pt(IV), Cu(II), Ag(I), and Au(III) with thiolates groups from C135 and C143 residues from the δ-ALA-D active site are crucial to the enzyme inhibition. The results indicate that a possible mechanism of inhibition of δ-ALA-D by these metals may involve the replacement of the Zn(II) from the active site and/or the cysteinyl residue oxidation.

Modulation of δ-Aminolevulinic Acid Dehydratase Activity by the Sorbitol-Induced Osmotic Stress in Maize Leaf Segments.

Osmotic stress induced with 1 M sorbitol inhibited δ-aminolevulinic acid dehydratase (ALAD) and aminolevulinic acid (ALA) synthesizing activities in etiolated maize leaf segments during greening; the ALAD activity was inhibited to a greater extent than the ALA synthesis. When the leaves were exposed to light, the ALAD activity increased for the first 8 h, followed by a decrease observed at 16 and 24 h in both sorbitol-treated and untreated leaf tissues. The maximum inhibition of the enzyme activity was observed in the leaf segments incubated with sorbitol for 4 to 8 h. Glutamate increased the ALAD activity in the in vitro enzymatic preparations obtained from the sorbitol-treated leaf segments; sorbitol inhibited the ALAD activity in the preparations from both sorbitol-treated and untreated leaves. It was suggested that sorbitol-induced osmotic stress inhibits the enzyme activity by affecting the ALAD induction during greening and regulating the ALAD steady-state level of ALAD in leaf cells. The protective effect of glutamate on ALAD in the preparations from the sorbitol-treated leaves might be due to its stimulatory effect on the enzyme.

In Silico Studies of Mammalian δ-ALAD Interactions with Selenides and Selenoxides.

Previous studies have shown that the mammalian δ-aminolevulinic acid dehydratase (δ-ALAD) is inhibited by selenides and selenoxides, which can involve thiol oxidation. However, the precise molecular interaction of selenides and selenoxides with the active center of the enzyme is unknown. Here, we try to explain the interaction of selenides and the respective selenoxides with human δ-ALAD by in silico molecular docking. The in silico data indicated that Se atoms of selenoxides have higher electrophilic character than their respective selenides. Further, the presence of oxygen increased the interaction of selenoxides with the δ-ALAD active site by O…Zn coordination. The interaction of S atom from Cys124 with the Se atom indicated the importance of the nucleophilic attack of the enzyme thiolate to the organoselenium molecules. These observations help us to understand the interaction of target proteins with organoselenium compounds.

Effects of Zinc and N-Acetylcysteine in Damage Caused by Lead Exposure in Young Rats.

This study investigated the toxicity of rats exposed to lead acetate (AcPb) during the second phase of brain development (8-12 days postnatal) in hematological and cerebral parameters. Moreover, the preventive effect of zinc chloride (ZnCl2) and N-acetylcysteine (NAC) was investigated. Pups were injected subcutaneously with saline (0.9% NaCl solution), ZnCl2 (27 mg/kg/day), NAC (5 mg/kg/day) or ZnCl2 plus NAC for 5 days (3rd-7th postnatal days), and with saline (0.9% NaCl solution) or AcPb (7 mg/kg/day) in the five subsequent days (8th-12th postnatal days). Animals were sacrificed 21 days after the last AcPb exposure. Pups exposed to AcPb presented inhibition of blood porphobilinogen-synthase (PBG-synthase) activity without changes in hemoglobin content. ZnCl2 pre-exposure partially prevented PBG-synthase inhibition. Regarding neurotoxicity biomarkers, animals exposed to AcPb presented a decrease in cerebrum acetylcholinesterase (AChE) activity and an increase in Pb accumulation in blood and cerebrum. These changes were prevented by pre-treatment with ZnCl2, NAC, and ZnCl2 plus NAC. AcPb exposure caused no alteration in behavioral tasks. In short, results show that AcPb inhibited the activity of two important enzymatic biomarkers up to 21 days after the end of the exposure. Moreover, ZnCl2 and NAC prevented the alterations induced by AcPb.

Sulfhydryl-Based Inhibition of δ-ALA-D and Na+ , K+ -ATPase Activities Depends on the Organoselenium Group Bonded to the Isoquinoline.

Organoselenium compounds and isoquinoline derivatives have their toxicity linked to induction of pro-oxidant situations. δ-Aminolevulinate dehydratase (δ-ALA-D) and Na+ , K+ -ATPase have sulfhydryl groups susceptible to oxidation. Thus, we investigated toxicological effects of 4-organoseleno-isoquinoline derivatives, cerebral monoamine oxidase B inhibitors, on rat cerebral δ-ALA-D and Na+ , K+ -ATPase activities and the involvement of sulfhydryl groups in vitro. Compounds substituted with fluoro (4-(4-fluorophenylseleno)-3-phenylisoquinoline), chloro (4-(4-chlorophenylseleno)-3-phenylisoquinoline) and trifluoro (4-(3-trifluoromethylphenylseleno)-3-phenylisoquinoline) at the selenium-bonded aromatic ring inhibited δ-ALA-D (IC50 values: 78.42, 92.27, 44.98 µM) and Na+ , K+ -ATPase (IC50 values: 41.36, 89.43, 50.66 µM) activities, possibly due to electronic effects induced by these groups. 3-Phenyl-4-(phenylseleno) isoquinoline (without substitution at the selenium-bonded aromatic ring) and 4-(4-methylphenylseleno)-3-phenylisoquinoline (with a methyl group substituted at the selenium-bonded aromatic ring) did not alter the activity of these enzymes. Dithiothreitol, a reducing agent, restored the enzymatic activities inhibited by 4-(4-fluorophenylseleno)-3-phenylisoquinoline, 4-(4-chlorophenylseleno)-3-phenylisoquinoline and 4-(3-trifluoromethylphenylseleno)-3-phenylisoquinoline, suggesting the involvement of sulfhydryl residues in this effect. However, the release of essential zinc seems not to be related to the δ-ALA-D inhibition by these compounds. According to these data, the effect of oral administration (300 mg/kg, intragastric) of 3-phenyl-4-(phenylseleno) isoquinoline on markers of systemic toxicity in Wistar rats was evaluated. None signs of toxicity was observed during or after treatment. This study suggests that the insertion of electron-withdrawing groups in the aromatic ring bonded to the selenium atom of isoquinolines tested increased its inhibitory effect on sulfhydryl enzymes in vitro. 3-Phenyl-4-(phenylseleno) isoquinoline, which has documented pharmacological properties, had no toxicological effects on the parameters evaluated in this study. J. Cell. Biochem. 118: 1144-1150, 2017. © 2016 Wiley Periodicals, Inc.

Effects of sub-lethal and chronic lead concentrations on blood and liver ALA-D activity and hematological parameters in Nile tilapia.

Liver and blood δ-aminolevulinic acid dehydratase (ALA-D) inhibition by exposure to sub-lethal lead concentrations over time in Nile tilapia (Oreochromis niloticus) were investigated. All three lead concentrations (1mgkg(-1), 10mgkg(-1) and 100mgkg(-1)) significantly inhibited ALA-D activity in blood (319±29.2; 180±14.6 and 172±19µmols(-1)h(-1)L(-1) respectively) and liver (302±5.84; 201±41.4 and 93±22.1µmols(-1)h(-1)L(-1)) 24h after injection relative to controls (blood: 597±37.0µmols(-1)h(-1)L(-1); liver: 376±23.1µmols(-1)h(-1)L(-1)). Blood ALA-D was greatly inhibited in all but the highest lead dose. Fish were then exposed to 1mgkg(-1) lead for 9 days, and presented short-term hyperglycemia, decreased hemoglobin and hematocrit values and time-dependent blood ALA-D activity inhibition, corroborating blood ALA-D activity as being more suitable for investigating lead effects, showing dose and time-dependent ALA-D inhibition after lead exposure. The results of the present study also demonstrated that fish size affects blood ALA-D activity, as fish from the 24-h assay, which were slightly smaller (approximately 200g), showed higher ALA-D inhibition in response to lead exposure when compared to the fish from the 9-day assay (approximately 500g). Thus, fish size should always be taken into account both in the field and in laboratory settings, and efforts should be made to obtain uniform fish size samples for biomarker studies.

Free radical scavenging in vitro and biological activity of diphenyl diselenide-loaded nanocapsules: DPDS-NCS antioxidant and toxicological effects.

Selenium compounds, such as diphenyl diselenide (DPDS), have been shown to exhibit biological activity, including antioxidant effects. However, the use of DPDS in pharmacology is limited due to in vivo pro-oxidative effects. In addition, studies have shown that DPDS-loaded nanocapsules (DPDS-NCS) have greater bioavailability than free DPDS in mice. Accordingly, the aim of this study was to investigate the antioxidant properties of DPDS-NCS in vitro and biological activity in mice. Our in vitro results suggested that DPDS-NCS significantly reduced the production of reactive oxygen species and Fe(II)-induced lipid peroxidation (LPO) in brain. The administration of DPDS-NCS did not result in death or change the levels of endogenous reduced or oxidized glutathione after 72 hours of exposure. Moreover, ex vivo assays demonstrated that DPDS-NCS significantly decreased the LPO and reactive oxygen species levels in the brain. In addition, the highest dose of DPDS-NCS significantly reduced Fe(II)- and sodium nitroprusside-induced LPO in the brain and Fe(II)-induced LPO in the liver. Also, δ-aminolevulinate acid dehydratase within the brain was inhibited only in the highest dose of DPDS-NCS. In conclusion, our data demonstrated that DPDS-NCS exhibited low toxicity in mice and have significant antioxidant characteristics, indicating that nanoencapsulation is a safer method of DPDS administration.

In vitro activity of wALADin benzimidazoles against different life cycle stages of Plasmodium parasites.

wALADin1 benzimidazoles are specific inhibitors of δ-aminolevulinic acid dehydratase from Wolbachia endobacteria of filarial nematodes. We report that wALADin1 and two derivatives killed blood stage Plasmodium falciparum in vitro (50% inhibitory concentrations, 39, 7.7, and 12.8 µM, respectively). One of these derivatives inhibited gliding motility of Plasmodium berghei ANKA infectious sporozoites with nanomolar affinity and blocked invasion into hepatocytes but did not affect intrahepatocytic replication. Hence, wALADin1 benzimidazoles are tools to study gliding motility and potential antiplasmodial drug candidates.

Cognitive deficits and ALA-D-inhibition in children exposed to multiple metals.

Children are especially vulnerable to adverse effects of multiple metals exposure. The aim of this study was to assess some metals concentrations such as lead (Pb), arsenic (As), chromium (Cr), manganese (Mn) and iron (Fe) in whole blood, serum, hair and drinking water samples using inductively coupled plasma-mass spectrometry (ICP-MS) in rural and urban children. In addition, evaluate the adverse effects of multiple metals exposure on cognitive function and δ-aminolevulinate dehydratase (ALA-D) activity. The cognitive ability assessment was performed by the Raven's Colored Progressive Matrices (RCPM) test. The ALA-D activity and ALA-D reactivation index (ALA-RE) activity with DTT and ZnCl2 also were determined. Forty-six rural children and 23 urban children were enrolled in this study. Rural children showed percentile IQ scores in the RCPM test significantly decreased in relation to urban children. According to multiple linear regression analysis, the Mn and Fe in hair may account for the cognitive deficits of children. Manganese and Fe in hair also were positively correlated with Mn and Fe in drinking water, respectively. These results suggest that drinking water is possibly a source of metals exposure in children. ALA-D activity was decreased and ALA-RE with DTT and ZnCl2 was increased in rural children in comparison to urban children. Moreover, ALA-D inhibition was correlated with Cr blood levels and ALA-RE/DDT and ALA-RE/ZnCl2 were correlated with levels of Cr and Hg in blood. Thus, our results indicated some adverse effects of children's exposure to multiple metals, such as cognitive deficits and ALA-D inhibition, mainly associated to Mn, Fe, Cr and Hg.

Liver δ-aminolevulinate dehydratase activity is inhibited by neonicotinoids and restored by antioxidant agents.

Neonicotinoids represent the most used class of insecticides worldwide, and their precursor, imidacloprid, is the most widely marketed. The aim of this study was to evaluate the effect of imidacloprid on the activity of hepatic δ-aminolevulinate dehydratase (δ-ALA-D), protective effect of potential antioxidants against this potential effect and presence of chemical elements in the constitution of this pesticide. We observed that δ-ALA-D activity was significantly inhibited by imidacloprid at all concentrations tested in a dose-dependent manner. The IC50 value was obtained and used to evaluate the restoration of the enzymatic activity. δ-ALA-D inhibition was completely restored by addition of dithiotreitol (DTT) and partly by ZnCl2, demonstrating that the inhibition occurs by oxidation of thiol groups and by displacement of the Zn (II), which can be explained by the presence of chemical elements found in the constitution of pesticides. Reduced glutathione (GSH) had the best antioxidant effect against to δ-ALA-D inhibition caused by imidacloprid, followed by curcumin and resveratrol. It is well known that inhibition of the enzyme δ-ALA-D may result in accumulation of its neurotoxic substrate (δ-ALA), in this line, our results suggest that further studies are needed to investigate the possible neurotoxicity induced by neonicotinoids and the involvement of antioxidants in cases of poisoning by neonicotinoids.

Are delta-aminolevulinate dehydratase inhibition and metal concentrations additional factors for the age-related cognitive decline?

Aging is often accompanied by cognitive impairments and influenced by oxidative status and chemical imbalances. Thus, this study was conducted to examine whether age-related cognitive deficit is associated with oxidative damage, especially with inhibition of the enzyme delta-aminolevulinate dehydratase (ALA-D), as well as to verify the influence of some metals in the enzyme activity and cognitive performance. Blood ALA-D activity, essential (Fe, Zn, Cu, Se) and non-essential metals (Pb, Cd, Hg, As, Cr, Ni, V) were measured in 50 elderly and 20 healthy young subjects. Cognitive function was assessed by tests from Consortium to Establish a Registry for Alzheimer's Disease (CERAD) battery and other. The elderly group presented decreased ALA-D activity compared to the young group. The index of ALA-D reactivation was similar to both study groups, but negatively associated with metals. The mean levels of essential metals were within the reference values, while the most toxic metals were above them in both groups. Cognitive function impairments were observed in elderly group and were associated with decreased ALA-D activity, with lower levels of Se and higher levels of toxic metals (Hg and V). Results suggest that the reduced ALA-D activity in elderly can be an additional factor involved in cognitive decline, since its inhibition throughout life could lead to accumulation of the neurotoxic compound ALA. Toxic metals were found to contribute to cognitive decline and also to influence ALA-D reactivation.

wALADin benzimidazoles differentially modulate the function of porphobilinogen synthase orthologs.

The heme biosynthesis enzyme porphobilinogen synthase (PBGS) is a potential drug target in several human pathogens. wALADin1 benzimidazoles have emerged as species-selective PBGS inhibitors against Wolbachia endobacteria of filarial worms. In the present study, we have systematically tested wALADins against PBGS orthologs from bacteria, protozoa, metazoa, and plants to elucidate the inhibitory spectrum. However, the effect of wALADin1 on different PBGS orthologs was not limited to inhibition: several orthologs were stimulated by wALADin1; others remained unaffected. We demonstrate that wALADins allosterically modulate the PBGS homooligomeric equilibrium with inhibition mediated by favoring low-activity oligomers, while 5-aminolevulinic acid, Mg(2+), or K(+) stabilized high-activity oligomers. Pseudomonas aeruginosa PBGS could be inhibited or stimulated by wALADin1 depending on these factors and pH. We have defined the wALADin chemotypes responsible for either inhibition or stimulation, facilitating the design of tailored PBGS modulators for potential application as antimicrobial agents, herbicides, or drugs for porphyric disorders.

New chemotypes for wALADin1-like inhibitors of delta-aminolevulinic acid dehydratase from Wolbachia endobacteria.

Substituted benzimidazoles of the wALADin1-family have recently been identified as a new class of species-selective inhibitors of delta-aminolevulinic acid dehydratase (ALAD) from Wolbachia endobacteria of parasitic filarial worms. Due to its Wolbachia-dependent antifilarial activity, wALADin1 is a starting point for the development of new drugs against filarial nematodes. We now present several other chemotypes of ALAD inhibitors that have been identified based upon their molecular similarity to wALADin1. A tricyclic quinoline derivative (wALADin2) with a different inhibitory mechanism and improved inhibitory potency and selectivity may represent an improved drug lead candidate.

In vitro glutathione peroxidase mimicry of ebselen is linked to its oxidation of critical thiols on key cerebral suphydryl proteins - A novel component of its GPx-mimic antioxidant mechanism emerging from its thiol-modulated toxicology and pharmacology.

The antioxidant mechanism of ebselen in rats brain is largely linked with its glutathione peroxidase (GPx) rather than its peroxiredoxin mimicry ability. However, the precise molecular dynamics between the GPx-mimicry of ebselen and thiol utilization is yet to be fully clarified and thus still open. Herein, we investigated the influence of dithiothreitol (DTT) on the antioxidant action of ebselen against oxidant-induced cerebral lipid peroxidation and deoxyribose degradation. Furthermore, the critical inhibitory concentrations of ebselen on the activities of sulphydryl enzymes such as cerebral sodium pump, δ-aminolevulinic acid dehydratase (δ-ALAD) and lactate dehydrogenase (LDH) were also investigated. We observe that ebselen (at ≥42 µM) markedly inhibited lipid peroxidation in the presence and absence of DTT, whereas it inhibited deoxyribose degradation only in the presence of DTT. Furthermore, under in vitro conditions, ebselen inhibited the thiol containing enzymes; cerebral sodium pump (at ≥40 µM), δ-ALAD (≥10 µM) and LDH (≥1 µM) which were either prevented or reversed by DTT. However, the inhibition of the activities of these sulphydryl proteins in diabetic animals was prevented by ebselen. Summarily, it is apparent that the effective in vitro inhibitory doses of ebselen on the activity of the sulphydryl proteins are far less than its antioxidant doses. In addition, the presence of DTT is evidently a critical requirement for ebselen to effect its antioxidant action against deoxyribose degeradation and not lipid peroxidation. Consequently, we conclude that ebselen possibly utilizes available thiols on sulphydryl proteins to effect its GPx mimicry antioxidant action against lipid peroxidation in rat brain homogenate.

Catechins are not major components responsible for the beneficial effect of Camellia sinensis on the ovarian δ-ALA-D activity inhibited by cadmium.

Cadmium has been associated with a wide spectrum of deleterious effects on the reproductive tissues, including ovary. This investigation evaluated the protective role of Camellia sinensis (green, white and red teas) in the cadmium-induced inhibition of ovarian δ-aminolevulinate dehydratase (δ-ALA-D) activity in vitro and ex vivo. This study demonstrated that green and white teas restored the cow ovary δ-ALA-D activity inhibited by cadmium whereas red tea had no effect in vitro. In addition, green tea was able to restore enzyme activity inhibited after acute cadmium exposure in mice ovary. Teas infusions composition was assessed by HPLC in a quantitative assay for catechins, purine alkaloids and gallic acid as well as total polyphenol content. The greatest effect of green tea observed in vitro as well as the protective role presented in the ex vivo study could be attributed to the major content of phenols, but not catechins. In fact, catechins were not able to restore enzyme activity inhibited by cadmium, demonstrating that these compounds are not major components responsible for the beneficial effect of green tea observed in this study. This study demonstrated the helpful effect of green tea infusion in ameliorating a marker protein of cadmium intoxication in ovarian tissue.

A selective inhibitor of heme biosynthesis in endosymbiotic bacteria elicits antifilarial activity in vitro.

Lymphatic filariasis and onchocerciasis are severe diseases caused by filarial worms and affect more than 150 million people worldwide. Endosymbiotic α-proteobacteria Wolbachia are essential for these parasites throughout their life cycle. Using a high-throughput chemical screen, we identified a benzimidazole compound, wALADin1, that selectively targets the δ-aminolevulinic acid dehydratase (ALAD) of Wolbachia (wALAD) and exhibits macrofilaricidal effects on Wolbachia-containing filarial worms in vitro. wALADin1 is a mixed competitive/noncompetitive inhibitor that interferes with the Mg(2+)-induced activation of wALAD. This mechanism inherently excludes activity against the Zn(2+)-dependent human ortholog and might be translatable to Mg(2+)-responsive orthologs of other bacterial or protozoan pathogens. The specificity profile of wALADin1 derivatives reveals chemical features responsible for inhibitory potency and species selectivity. Our findings validate wALADins as a basis for developing potent leads that meet current requirements for antifilarial drugs.

Plasma succinylacetone is persistently raised after liver transplantation in tyrosinaemia type 1.

BACKGROUND: Tyrosinaemia type 1 (HT1) is a rare disorder leading to accumulation of toxic metabolites such as succinylacetone (SA) and a high risk of hepatocellular carcinoma. Children with HT1 traditionally required liver transplantation (OLT) and while the need for this has been reduced by the introduction of nitisinone some still require OLT. SA inhibits the enzyme porphobilinogen (PBG) synthase and its activity can be used as a marker of active SA. Elevated urinary SA post OLT has been reported previously. This study describes a novel finding of elevated plasma SA following OLT for HT1. METHODS: A retrospective analysis was performed of patients treated for HT1 at our institution from 1989-2010. RESULTS: Thirteen patients had an OLT for HT1. In patients who received nitisinone prior to OLT, mean urinary and plasma SA were elevated prior to treatment but normalised by the time of OLT (p ≤ 0.01). Mean PBG synthase activity increased from 0.032 to 0.99 nkat/gHb (ref range 0.58-1.25) at the time of OLT (p < 0.01). Mean urinary SA in patients not treated with nitisinone was also elevated prior to OLT; plasma levels and PBG synthase activity were not available prior to OLT for this group. Following OLT, mean urinary and plasma SA were elevated in all for the duration of follow-up and associated with low-normal PBG synthase activity. CONCLUSION: Urinary and plasma SA levels are elevated following OLT for HT1. Low-normal PBG synthase activity suggests the plasma SA may be active. The clinical significance of this is unclear.

Blood lead levels, δ-ALAD inhibition, and hemoglobin content in blood of giant toad (Rhinella marina) to assess lead exposure in three areas surrounding an industrial complex in Coatzacoalcos, Veracruz, Mexico.

The Coatzacoalcos Region in Veracruz, Mexico houses one of the most important industrial complexes in Mexico and Latin America. Lead is an ubiquitous environmental pollutant which represents a great risk to human health and ecosystems. Amphibian populations have been recognized as biomonitors of changes in environmental conditions. The purpose of this research is to measure exposure to lead and evaluate hematological and biochemical effects in specimens of giant toads (Rhinella marina) taken from three areas surrounding an industrial complex in the Coatzacoalcos River downstream. Lead levels in toads' blood are between 10.8 and 70.6 µg/dL and are significantly higher in industrial sites. We have found a significant decrease in the delta-aminolevulinic acid dehydratase (δ-ALAD) activity in blood from 35.3 to 78 % for the urban-industrial and industrial sites, respectively. In addition, we have identified a strong inverse relationship between the δ-ALAD activity and the blood lead levels (r = -0.84, p < 0.001). Hemoglobin and mean corpuscular hemoglobin levels, as well as the condition factor, are found to be lower at industrial sites compared with the reference sites. Our results suggest that the R. marina can be considered a good biomonitor of the δ-ALAD activity inhibition and hematological alterations at low lead concentrations.

Cadmium inhibits the ovary δ-aminolevulinate dehydratase activity in vitro and ex vivo: protective role of seleno-furanoside.

Cadmium (Cd) toxicity is a concern to the tobacco-smoking sub-population which includes millions of people worldwide. Although this metal may cause severe damage to embryos and the reproductive organs, the precise mechanisms underlying its toxicity remain unclear. In the present study, the Cd effect on ovary δ-aminolevulinate dehydratase (δ-ALA-D) activity was investigated in vitro and ex vivo. We observed that low concentrations of Cd inhibited cow ovary δ-ALA-D activity in vitro and the IC50 value obtained was 19.17 µM. Furthermore, the protective effect of a novel organic selenium compound (seleno-furanoside) in restoring enzyme activity was evaluated. Seleno-furanoside (10, 50, 100, 200, 400 and 1000 µM) did not reverse the Cd toxicity in bovine ovarian tissue in vitro. According to the in vitro reults, acute Cd exposure (2.5 and 5 mg kg(-1)) caused a significant inhibition in ovary δ-ALA-D activity in mice (around 27% and 34%, respectively). Therapy with seleno-furanoside (100 µmol kg(-1)) was able to restore enzyme activity. Thus, we demonstrated for the first time that δ-ALA-D activity from ovary is inhibited by Cd both in vitro and ex vivo. Additionally, seleno-furanoside therapy was effective in restoring ovarian enzyme activity inhibited by Cd exposure in mice, but it did not reverse the in vitro metal effect. This study detected a new toxicity marker of Cd toxicity on ovarian tissue as well as the beneficial effect of a new compound to manage the metal effect after acute exposure.