The degradation of organophosphorus compounds CVX [O-butyl S-(2-diethylaminoethyl)methylphosphonothioate] and RVX [O-isobutyl S-(2-diethylaminoethyl)methylphosphonothioate] in soils is described. The work included the determination of the extraction efficiency of different types of soil matrices with respect to time for these organophosphorus substances. Extraction of the contaminant from the soil achieved the highest yields using dichloromethane. The most significant decreases of both RVX and CVX were recorded within the first 24 h, while the total decrease of the analytes in the studied soils varied significantly according to their composition. Gas chromatography–mass spectrometry was used to determine the extraction efficiency of organophosphorus substances from soil matrices. The proposed method presented excellent linearity for RVX and CVX calibration curves with correlation coefficients exceeding 0.99. To increase the sensitivity, selected ion monitoring with a limited mass-to-charge ratio range was employed to characterize the extraction efficiency.
Using an immobilized acetylcholinesterase–tabun enzyme–inhibitor complex, the reactivation efficacy of a homologous series of bispyridinium reactivators with increasing length of the alkylene chain between the pyridinium rings has been studied. The number of the alkylene groups in the chain ranged from one to six. N,N′-Monomethylenebis(4-pyridiniumaldoxime) dibromide (MMB-4) and N,N′-trimethylenebis(4-pyridiniumaldoxime) dibromide (TMB-4) are the most efficient reactivators of the series.
Reactivation efficacy of three homologous and three isomeric series of pralidoxime-type reactivators with aldoxime group in position 2, 3 and 4 of the heterocycle was tested in reactivation of tabun-inhibited AChE. The experiments were performed with immobilized and stabilized porcine brain AChE. The enzyme acticity was measured by Ellman method. Reactivation efficacy was determined by measurement of indicator fabric coloration intensity as a measure of AChE activity. Of the studied group of nine reactivators, isomers with the functional group in position 2 were the most effective. The highest value (30 %) for reactivation of inhibited AChE was found for 2PAE after treatment for 15 min at concentration 0.5 mg/cm3. The efficacy of the isomers decreased in the order ortho > para > meta. No marked effect on the efficacy of the reactivators was observed on prolongation of the reactivation time. The reactivators efficacy decreased with decreasing concentration of their solutions.
A simple colorimetric biosensor, which uses the modified Ellman's reaction, enables selective analysis of nerve agents based on a different ability of bispyridinium oximes to reactivate enzyme-inhibitor complexes in the phase before dealkylation.The analysis was made based on spectral data of reflectance of the surface of a cotton cloth reaction zone of biosensor with immobilized and stabilized enzyme after inhibition and subsequent reactivation.The evaluation of measured data was made based on a method of artificial neuronal networks.The individual inhibitors from the groups of three nerve agents were identified: sarin, soman, tabun, cyclosarin, agent VX and its Russian analogue, R-33.
Organophosphorus compounds are a wide-spread group of agents which can be used among others as pesticides, especially insecticides, or chemical warfare agents. These extremely toxic compounds irreversibly inhibit the enzymes of hydrolases class, which have the catalytic capability of hydrolyzing their specific neurotransmitters in synaptic clefts of the nervous system. The organophosphorus pesticides and carbamates are agents used in agriculture very often, because of their relatively low stability. Many of them, however, have very high acute toxicity for warm-blooded animals. The group of organophosphorus compounds also includes nerve agents with their dominant position among chemical warfare agents. The discovery of their effects was made more or less accidentally in the 30-ies of the last century during the research of fluoroorganic compounds in IG Farben, a German company, by Dr. Gerhard Schrader. The first synthesized agent was ethyl-(dimethylamido)phosphorocyanidate, designated as trilon 83 or tabun. However, it was not the first known agent with a cholinergic effect. Back in 1854 a French chemist, Phillip de Clermont, synthesized the first organophosphate – tetraethyl pyrophosphate (de Clermont, 1855, as cited in Holmstedt, 2000). Gradually more agents with N-P, P-CN or C-F bonds were synthesized in order to produce insecticides and later also nerve agents (Holmstedt, 2000). After verifying the effects of tabun for warm-blooded animals, a synthesis of other, even more toxic, agents followed (Pitschmann, 1999). In 1939 isopropyl-methylphosphonofluoridate was discovered, also called trilon 46 or sarin, and in 1944 (3,3-dimethylbutane-2-yl)-methylphosphonofluoridate, the so-called soman, followed. These agents belong to the so-called G-series of nerve agents. The origin of a new V-series dates back to the end of 50-ies. This group of agents has increased toxicity when penetrating the skin. The principal representative is S-[2-(diisopropylamino)ethyl]-O-ethylmethylphosphonothioate, also known as VX, and S-[2-(diethylamino)ethyl]-O-isobutylmethylphosphonothioate with the code designation R-33. In the 70-ies up to 90-ies another group of nerve agents was discovered in the former USSR within the so-called Foliant program. These are compounds based on phosphorylated and phosphonylated oximes and amidases. According to unauthorized sources they reach at least the toxicity of VX, some are even 5 to 8 times more toxic. In a case like this there could be a problem with their detection.
Reactivation with bis quaternary aldoxime HI-6, chemical formula 1-(2-hydroxyamino-methylpyridinium)-3-(4-carbamoylpyridinium)-2-oxapropane dichloride of immobilized enzyme acetylcholinesterase inhibited by nerve agent type "G" was studied. This aldoxime is effective in reactivation of sarin-inhibited acetylcholinesterase. Substantially lower reactivation potency was observed with cyclosarin-inhibited enzyme and almost no effect was found for that acetylcholinesterase is the enzyme complex. HI 6 is completely ineffective towards the soman-inhibited enzyme: After a 2-minute inhibition of the enzyme with soman no ability to define reactivator the inhibited enzymes and complexes.
Abstract Construction of Detehit (nerve-agent detector), a colorimetric biosensor of nerve agents, enables its utilization for selective analysis of nerve agents based on the different ability of nucleophilic reagents to reactivate enzyme-inhibitor complexes in the phase before dealkylation. For this purpose mono- and bispyridinium aldoximes, common in treatment of nerve agent poisoning, are used. For a positive identification of a pair of organophosphates, a neural network analysis was used. The analysis was processed based on spectral data of the intensity of color changes of the surface of a cotton cloth reaction zone with immobilized and stabilized enzyme acetylcholinesterase. Color changes are based on Ellman's reaction. Intensity of these changes depends on activity of the enzyme after inhibition and reactivation of enzyme-inhibitor complex. The following nerve agents were identified: sarin, soman, tabun, cyclosarin, VX, and R-33. Keywords: Acetylcholinesterase reactivatorColorimetric biosensorNerve agentNeural analysisAbbreViations: GB, sarinGD, somanGF, cyclosarinGA, tabunVX, agent VXR-33, agent R-33VVPS, multilayer perceptron networkRBF, Radial Basis Function-type network Acknowledgments VOP (Military Repair Plant) Šternberk is a state-owned company. Notes Measurement conditions: HI-6 reactivator at a concentration of 0.05 mg · cm−3 (1.393 × 10−4 M) and 10-minute treatment of acetylcholinesterase-tabun and acetylcholinesterase-VX complexes. Linear: linear network with characterization of the layers; VVPS: multilayer perceptron network with characterization of the layers; RBF: Radial Basis Function-type network with characterization of the layers; GF: cyclosarin; VX: compound VX. GA: tabun; GB: sarin; GD: soman; GF: cyclosarin; R-33: agent R-33; VX: agent VX.
OBJECTIVES: Quantification of efficacy of monopyridinium isomers and homologs derived from clinically used Pralidoxime within reactivation of acetylcholinesterase inhibited with organophosphorus nerve agents. METHODS: This work uses the colorimetric biosensor called Detehit - cotton cloth with immobilized enzyme acetylcholinesterase. Biosensor is based on the modificated Ellman's method. RESULTS: The highest reactivation was observed with sarin-inhibited acetylcholinesterase. Substantially lower reactivation was found with the cyclosarin-inhibited enzyme whereas AChE, inhibited by soman could not be effectively reactivated under the given conditions (enzyme inhibition for 2 minutes and subsequent treatment with the reactivator for 15 minutes). CONCLUSION: Our work gives comparison of efficacy of reactivators in dependence on the lenght of alkylene chain and position of aldoxime functional group. Evaluation of effectivity of aldoxime reactivators is provided by simple means. The method allows rapid in vitro evaluation of the reactivators without being disturbed by excess of the organophosphate or reactivator.
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