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Functional and morphological aspects of thallium-induced nephrotoxicity in rats.

Until now the effect of thallium (Tl) on renal function has not been investigated systematically. Therefore, the dose (5, 10, 15, 20 mg Tl2SO4/kg body wt., intraperitoneally) and time-dependence of renal damage was investigated in diuresis experiments on conscious rats. Morphology was evaluated after perfusion fixation in situ. Morphologic changes were localized in the thick ascending limb of the loop of Henle, mostly expressed at the 2nd day after Tl administration, which were completely normalized again at the 10th day. Other parameters such as Tl concentration, changes in water content and the activity of Na+/K(+)-ATPase as well as the diuretic effect of furosemide confirmed the Tl effect to be localized in the renal medulla. One single Tl administration is followed by a decrease in glomerular filtration rate (GFR) and urine volume and an increase of proteinuria. Electrolyte excretion was only slightly changed. All changes were reversible within the 10-day investigation period.

Animals↗

The effect of sub-lethal doses of bromadiolone on the breeding performance of house mice (Mus domesticus).

Both laboratory and field strains of Mus were exposed to wheat containing 0.0001% bromadiolone under laboratory and outdoor conditions, respectively. While both strains readily consumed the poisoned wheat, ad libitum sub-lethal doses of this anticoagulant equating to between 20% and 70% of the acute LD50 per feed had little apparent effect on the breeding performance of these mice. The implications of these findings are discussed with respect to the use of anticoagulants as pesticides.

4-Hydroxycoumarins↗

Chemotaxis-mediated biodegradation of cyclic nitramine explosives RDX, HMX, and CL-20 by Clostridium sp. EDB2.

Cyclic nitramine explosives, RDX, HMX, and CL-20 are hydrophobic pollutants with very little aqueous solubility. In sediment and soil environments, they are often attached to solid surfaces and/or trapped in pores and distribute heterogeneously in aqueous environments. For efficient bioremediation of these explosives, the microorganism(s) must access them by chemotaxis ability. In the present study, we isolated an obligate anaerobic bacterium Clostridium sp. strain EDB2 from a marine sediment. Strain EDB2, motile with numerous peritrichous flagella, demonstrated chemotactic response towards RDX, HMX, CL-20, and NO(2)(-). The three explosives were biotransformed by strain EDB2 via N-denitration with concomitant release of NO(2)(-). Biotransformation rates of RDX, HMX, and CL-20 by the resting cells of strain EDB2 were 1.8+/-0.2, 1.1+/-0.1, and 2.6+/-0.2nmol h(-1)mgwet biomass(-1) (mean+/-SD; n=3), respectively. We found that commonly seen RDX metabolites such as TNX, methylenedinitramine, and 4-nitro-2,4-diazabutanal neither produced NO(2)(-) during reaction with strain EDB2 nor they elicited chemotaxis response in strain EDB2. The above data suggested that NO(2)(-) released from explosives during their biotransformation might have elicited chemotaxis response in the bacterium. Biodegradation and chemotactic ability of strain EDB2 renders it useful in accelerating the bioremediation of explosives under in situ conditions.

Aza Compounds↗

Toxicity of hexahydro-1,3,5-trinitro-1,3,5-triazine to larval zebrafish (Danio rerio).

Hexahydro-1,3,5-trinitro-1,3,5-triazine, a cyclonitramine commonly known as RDX, is used in the production of military munitions. Contamination of soil, sediment, and ground and surface waters with RDX has been reported in different places around the world. Acute and subacute toxicities of RDX have been relatively well documented in terrestrial vertebrates, but among aquatic vertebrates the information available is limited. The objective of this study was to characterize the acute toxicity of RDX to larval zebrafish. Mortality (LC50) and incidence of vertebral column deformities (EC50) were two of the end points measured in this study. The 96-h LC50 was estimated at 22.98 and 25.64 mgl(-1) in two different tests. The estimated no-observed-effective-concentration (NOEC) values of RDX on lethality were 13.27+/-0.05 and 15.32+/-0.30 mgl(-1); and the lowest-observed-effective-concentration (LOEC) values were 16.52+/-0.05 and 19.09+/-0.23 mgl(-1) in these two tests, respectively. The 96-h EC50 for vertebral deformities on survivors from one of the acute lethality tests was estimated at 20.84 mgl(-1), with NOEC and LOEC of 9.75+/-0.34 and 12.84+/-0.34 mgl(-1), respectively. Behavioral aberrations were also noted in this acute toxicity study, including the occurrence of whirling movement and lethargic behavior. The acute effects of RDX on survival, incidence of deformities, and behavior of larval zebrafish occurred at the high end of the most frequently reported concentrations of RDX in aquatic environments. The chronic effects of RDX in aquatic vertebrates need to be determined for an adequate assessment of the ecological risk of environmental RDX.

Animals↗

Ecotoxicological evaluation of sodium fluoroacetate on aquatic organisms and investigation of the effects on two fish cell lines.

Sodium monofluoroacetate (compound 1080) is one of the most potent pesticides. It is also a metabolite of many other fluorinated compounds, including anticancer agents, narcotic analgesics, pesticides or industrial chemicals. Other sources of water contamination are the atmospheric degradation of hydrofluorocarbons and hydrochlorofluorocarbons. However, there is little information available about the adverse effects of sodium fluoroacetate in aquatic organisms. Firstly, the bacterium Vibrio fischeri (decomposer), the alga Chlorella vulgaris (1st producer) and the cladoceran Daphnia magna (1st consumer) were used for the ecotoxicological evaluation of SMFA. The most sensitive models were C. vulgaris and D. magna, with a NOAEL of 0.1 and an EC50 of 0.5 mM at 72 h, respectively. According to the results after the acute exposure and due to its high biodegradation rate and low bioaccumulation potential, sodium fluoroacetate is most unlikely to produce deleterious effects to aquatic organisms. Secondly, two fish cell lines were employed to investigate the effects and mechanisms of toxicity in tissues from 2nd consumers. The hepatoma fish cell line PLHC-1 was more sensitive to SMFA than the fibroblast-like fish cell line RTG-2, being the uptake of neutral red the most sensitive bioindicator. Lysosomal function, succinate dehydrogenase and acetylcholinesterase activities were inhibited, glucose-6-phosphate dehydrogenase activity was particularly stimulated, and metallothionein and ethoxyresorufin-O-deethylase levels were not modified. Intense hydropic degeneration, macrovesicular steatosis and death mainly by necrosis but also by apoptosis were observed. Moreover, sulphydryl groups and oxidative stress could be involved in PLHC-1 cell death induced by SMFA more than changes in calcium homeostasis.

Acetylcholinesterase↗

Phytotoxicity to and uptake of RDX by rice.

Phytoremediation is an emerging strategy to remediate soils contaminated with pollutants like explosives in which plants will uptake, degrade and/or accumulate pollutants. To implement this technology on a site contaminated with RDX, we chose rice, which is able to grow in lagoons, and we tested its ability to grow in soils with high levels of RDX and to decrease RDX concentrations in soil. Rice was grown for 40 days in soil contaminated with increasing [14C]RDX concentrations. Emergence and growth were not affected by RDX. Total chlorophyll content decreased with RDX concentrations of over 500 mg kg(-1). Amounts of chlorophyll were correlated with the appearance of necrosis in leaf extremities. After 40 days, rice translocated 89% of uptaken radioactivity to leaves with 90% in leaf extremities. Analyzes of leaf extracts showed that 95% of radioactivity was RDX in its parent form. Necrosis appears to be a phytotoxic symptom of RDX accumulation.

Biodegradation, Environmental↗

Determination of bromadiolone in whole blood by high-performance liquid chromatography coupled with electrospray ionization tandem mass spectrometry.

A rapid, sensitive and selective high-performance liquid chromatography tandem mass spectrometric method (HPLC/MS-MS) has been developed and validated for the determination of bromadiolone in whole blood using warfarin as an internal standard (IS). Bromadiolone was extracted from the whole blood samples by liquid-liquid extraction with ethyl acetate. Multiple-reaction monitoring (MRM) was used to detect bromadiolone and IS, using precursor --> product ion combinations at m/z 527 --> 465 and 307 --> 161, respectively. The calibration curve was linear (r2=0.998) in the concentration range of 0.5-100.0 ng/mL with a lower limit of quantification of 0.5 ng/mL in whole blood. Intra- and inter-day relative standard deviations (R.S.D.s) were less than 7.5 and 11.9%, respectively. Recoveries of bromadiolone ranged from 82.1 to 85.2%. This method is found to be determined trace bromadiolone in whole blood and can be used in the diagnosis of the poisoned human beings.

4-Hydroxycoumarins↗

Dissolution kinetics of high explosives particles in a saturated sandy soil.

Solid phase high explosive (HE) residues from munitions detonation may be a persistent source of soil and groundwater contamination at military training ranges. Saturated soil column tests were conducted to observe the dissolution behavior of individual components (RDX, HMX, and TNT) from two HE formulations (Comp B and C4). HE particles dissolved readily, with higher velocities yielding higher dissolution rates, higher mass transfer coefficients, and lower effluent concentrations. Effluent concentrations were below solubility limits for all components at superficial velocities of 10-50 cm day(-1). Under continuous flow at 50 cm day(-1), RDX dissolution rates from Comp B and C4 were 34.6 and 97.6 microg h(-1) cm(-2) (based on initial RDX surface area), respectively, significantly lower than previously reported dissolution rates. Cycling between flow and no-flow conditions had a small effect on the dissolution rates and effluent concentrations; however, TNT dissolution from Comp B was enhanced under intermittent-flow conditions. A model that includes advection, dispersion, and film transfer resistance was developed to estimate the steady-state effluent concentrations.

Algorithms↗

A benchmark dose analysis for sodium monofluoroacetate (1080) using dichotomous toxicity data.

The use of a benchmark dose (BMD) as an alternative to a no-observed-adverse-effect-level (NOAEL) approach was investigated as a means to improve current risk assessment values of sodium monofluoroacetate (1080). The feasibility of implementing the two approaches was investigated for three critical toxicological end points, namely cardiomyopathy, testicular toxicity and teratogenic effects identified from the few available critical studies. The BMD provides better representation of the dose-response relationship, offering an advantage over the current NOAEL approach. The calculated BMDs and lower-bound confidence limits (BMDLs) for the three end points were estimated using the Weibull, probit and quantal linear models for each end point. All models passed the chi2 test statistics (p > or = 0.1) for all three toxicity endpoints tested. A benchmark response (BMR) of 10% (extra risk) was chosen and the Akaike's information criterion (AIC) was used in selecting the appropriate model. The BMDL estimates derived were found to be generally slightly higher but comparable to the NOAEL for those same endpoints. The BMD(10) and BMDL(10) for cardiomyopathy and testicular effects were 0.21 mgkg(-1) bw and 0.10 mgkg(-1) bw, respectively. These values are proposed for use in the eventual determination of the tolerable daily intake (TDI) for 1080.

Abnormalities, Drug-Induced↗

The use of myocardial and testicular end points as a basis for estimating a proposed tolerable daily intake for sodium monofluoroacetate (1080).

This paper presents the development of a tolerable daily intake (TDI) for sodium monofluoroacetate (1080) using the quantal myocardial and testicular toxicity end points derived from the traditional NOAEL and newer benchmark dose (BMD) methods. 1080 is a highly toxic vertebrate pesticide that has been proven to be effective in controlling possums and other pests. By convention, the TDIs are derived using the traditional no-observed-adverse-effect-level (NOAEL) and applying appropriate default uncertainty factors (UF). In addition to the default UF, a statistically derived UF was also employed in deriving the TDI. The TDIs derived from the NOAEL and BMD approach, 0.075 and 0.10 mg/kg bw/day, respectively, were compared. The resulting TDI estimates using the BMDL, a statistical lower confidence bound on the BMD, were generally consistently slightly higher than those derived using the NOAEL approach. Based on the best fit of modelled dose-response data, a TDI of 0.03 micro g/kg bw/day is proposed for human health risk assessment of 1080.

Animals↗

Diaphorase catalyzed biotransformation of RDX via N-denitration mechanism.

Previously, we hypothesized that hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) can be biotransformed by anaerobic sludge via three different routes: (1) direct ring cleavage via alpha-hydroxylation of a-CH(2) group, (2) reduction of one of the -NO(2) groups to -NO, (3) N-denitration prior to ring cleavage. The present study describes biotransformation of RDX via route 3 by a diaphorase (EC 1.8.1.4) from Clostridium kluyveri using NADH as electron donor. The removal of RDX was accompanied by the formation and accumulation of nitrite ion (NO(2)(-)), formaldehyde (HCHO), ammonium (NH(4)(+)), and nitrous oxide (N(2)O). None of the RDX-nitroso products were detected. The ring cleavage product methylenedinitramine was detected as the transient intermediate. Product stoichiometry showed that each reacted RDX molecule produced one nitrite ion and the product distribution gave a carbon (C) and nitrogen (N) mass balance of 91 and 92%, respectively, supporting the occurrence of a mono-denitration step prior to the ring cleavage and decomposition. Severe oxygen mediated inhibition (92% inhibition) of RDX biotransformation and superoxide dismutase-sensitive cytochrome c reduction indicated the potential involvement of an anion radical RDX(.-) prior to denitration. A comparative study between native- and apo-enzymes showed the possible involvement of flavin mononucleotide (FMN) in catalyzing the transfer of a redox equivalent (e/H(+)) from NADH to RDX to produce RDX(.-) responsible for secondary decomposition.

Catalysis↗

Echinococcus multilocularis: secondary poisoning of fox population during a vole outbreak reduces environmental contamination in a high endemicity area.

This paper describes the role of fox population level on Echinococcus multilocularis infection in foxes in a highly endemic area in eastern France. Fox population level was monitored by spotlight survey at Le Souillot from 1989 to 2000, and from 1992 to 2000 at Chemin, a control site located in a low endemic area. The infection level of the fox population was estimated at Le Souillot from winter 1995 to winter 1999 using a coproantigen ELISA performed on faeces collected in the field. Population biomass of intermediate hosts (Microtus arvalis and Arvicola terrestris) was monitored using index methods from 1995 to 1999. At Le Souillot, a significant decline in the fox population level was recorded after spring 1997 (P<0.001), and the population level remained low until 2000. The decline occurred when 31% of the grassland area was treated with bromadiolone, an anticoagulant used at a large scale for the control of A. terrestris population outbreaks. No decline of population was recorded at Chemin, where bromadiolone was not used for rodent control. Significant differences among ELISA OD distributions in fox faeces were recorded for the five winters under study at Le Souillot (P=0.0004). The median of ELISA OD distribution was 0.209 and 0.207 before the population decline (winter 1995 and 1996, respectively), significantly increased to 0.306 just after the decline (winter 1997), and then significantly decreased to 0.099 and 0.104 afterwards (winter 1998 and 1999, respectively). Therefore, the decrease in infection level occurred during winter 1998, 1 year after the population decline, when the intermediate host biomass in the field was at its highest. These results suggest a complex dependence between the fox population level and E. multilocularis infection in a high endemicity area. Alternative ways to control fox population as a way to reduce E. multilocularis transmission in a given area are discussed.

4-Hydroxycoumarins↗

Screening for coumatetralyl in soft drinks by solid-matrix extraction and high-performance liquid chromatography with diode-array detection.

A method was developed for the rapid determination coumatetralyl in cola- and orange-type soft drinks, which includes extraction using solid-matrix column, clean-up by silica cartridge chromatography and reversed-phase high-performance liquid chromatography with diode-array detection. The recovery of coumatetralyl from 50 ml of soft drinks was better than 80% at spiking levels down to 50 micrograms/kg (ppb).

4-Hydroxycoumarins↗

Reversed-phase ion-pair liquid chromatographic determination of chlorophacinone residues in animal tissues.

A sensitive and specific high-performance liquid chromatographic method for the identification and quantitation of chlorophacinone in animal tissues has been developed. Residues were extracted with chloroform-acetone (1:1, v/v). Clean-up of extracts was accomplished with a combined gel permeation-adsorption chromatographic procedure using Bio-Beads SX-3 and incorporating an on-line Sep-Pak silica cartridge. Residues were determined by ion-pair liquid chromatography, with the tetrabutylammonium ion as counter-ion, using a fixed-wavelength UV detector at 280 nm or a diode array detector at 285 nm. Recoveries from spiked liver tissue were around 90% at levels from 0.05 to 1 mg kg-1. A detection limit of 0.001 mg kg-1 could be achieved in animal tissues. The diode array detector confirmed identification by matching spectra for residues down to 0.1 mg kg-1 and below this level by multi-wavelength monitoring.

Animals↗