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1.
J Anal Toxicol ; 28(5): 364-71, 2004.
Artículo en Inglés | MEDLINE | ID: mdl-15239857

RESUMEN

One of the most appropriate biomarkers for the verification of organophosphorus nerve agent exposure is the conjugate of the nerve agent to butyrylcholinesterase (BuChE). The phosphyl moiety of the nerve agent can be released from the BuChE enzyme by incubation with fluoride ions, after which the resulting organophosphonofluoridate can be analyzed with gas chromatography-mass spectrometry (GC-MS). This paper describes recent improvements of the fluoride-induced reactivation in human plasma or serum samples by enhancing the sample preparation with new solid-phase extraction cartridges and the MS analysis with large volume injections. Analysis is performed with thermal desorption GC with either mass selective detection with ammonia chemical ionization or high-resolution MS with electron impact ionization. The organophosphorus chemical warfare agents analyzed in this study are O-ethyl S-2-diisopropylaminoethyl methylphosphonothiolate, ethyl methylphosphonofluoridate, isopropyl methylphosphonofluoridate (sarin, GB), O-ethyl N,N-dimethylphosphoramidocyanidate, ethyl N,N-dimethylphosphoramidofluoridate, and cyclohexyl methylphosphonfluoridate. Detection limits of approximately 10 pg/mL plasma were achieved for all analytes, which corresponds to 0.09% inhibition with GB on a sample with normal BuChE levels.


Asunto(s)
Sustancias para la Guerra Química , Reactivadores de la Colinesterasa , Monitoreo del Ambiente/métodos , Fluoruros/farmacología , Cromatografía de Gases y Espectrometría de Masas/métodos , Biomarcadores/sangre , Butirilcolinesterasa/metabolismo , Sustancias para la Guerra Química/análisis , Sustancias para la Guerra Química/farmacocinética , Sustancias para la Guerra Química/envenenamiento , Exposición a Riesgos Ambientales/análisis , Eritrocitos/efectos de los fármacos , Eritrocitos/enzimología , Fluoruros/metabolismo , Cromatografía de Gases y Espectrometría de Masas/instrumentación , Humanos , Reproducibilidad de los Resultados , Sensibilidad y Especificidad
2.
Arch Toxicol ; 78(9): 508-24, 2004 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-15170525

RESUMEN

The utility was explored of a new approach to detect retrospectively exposure to nerve agents by means of conversion of the inhibitor moiety bound to the active site of the enzyme BuChE in plasma with fluoride ions into a phosphofluoridate which is subsequently analyzed by means of gas chromatography (GC). This quantifies >or=0.01% inhibition of BuChE and identifies the structure of the inhibitor except for the original leaving group. A three-tiered approach was followed involving the five classical nerve agents GA, GB, GF, GD, and VX, as well as the active metabolite of parathion, i.e., paraoxon: in vivo experiments in rhesus monkeys after iv administration of a sign-free dose of agent and concomitant in vitro experiments in plasma of rhesus monkeys and humans should allow an assessment of in vivo retrospectivity in humans. A systematic investigation was performed in order to find a single set of reaction conditions which yields a maximum amount of phosphofluoridate for all nerve agents. Fluoride-induced reactivation at 25 degrees C at a final concentration of 250 mM KF during 15 min in a pH-range between 4 and 6 appears to be effective. The in vitro decrease with time in reactivatibility of inhibited BuChE in plasma from humans and rhesus monkeys was largely due to aging of the phosphyl moiety, except for VX where spontaneous reactivation was a major cause. The decrease followed first-order except for a biphasic course in the case of GF in human and rhesus monkey plasma as well as of GD in rhesus plasma. In vitro half-lifes in human plasma ranged between ca. 14 h for GB and ca. 63 h for GA. A comparison of the in vivo data from rhesus monkeys and the in vitro data is complicated by the observation that the in vivo decrease with time of fluoride-reactivated phosphofluoridate is biphasic for all nerve agents. The terminal in vivo phase pertains to a small fraction of the amount of initially regenerated phosphofluoridate but is responsible for a considerable degree of retrospectivity, ranging between 14 and 56 days for GF and GB, respectively. The new procedure can be used in a variety of practical applications, e.g., (i) biomonitoring in health surveillance at exposure levels that are several orders of magnitude lower than presently possible; (ii) diagnosis in case of alleged exposure to nerve agents in time of war or after terrorist attacks; (iii) in forensic cases against suspected terrorists that have handled organophosphate anticholinesterases; and (iv) in research applications such as investigations on lowest observable effect levels of exposure to nerve agents.


Asunto(s)
Butirilcolinesterasa/sangre , Sustancias para la Guerra Química/análisis , Inhibidores de la Colinesterasa/análisis , Reactivadores de la Colinesterasa/química , Fluoruros/química , Compuestos Organofosforados/análisis , Animales , Butirilcolinesterasa/metabolismo , Sustancias para la Guerra Química/farmacocinética , Inhibidores de la Colinesterasa/farmacocinética , Cromatografía de Gases , Femenino , Semivida , Humanos , Inyecciones Intravenosas , Macaca mulatta , Masculino , Compuestos Organofosforados/farmacocinética , Paraoxon/análisis , Paraoxon/farmacocinética , Estándares de Referencia , Factores de Tiempo
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