ИСТИНА

The development, production, stockpiling and use of chemical weapons are prohibited under the Chemical Weapons Convention [1]. In cases of alleged use of chemical warfare (CW) agents, environmental samples may be collected and analyzed for agents and their degradation products presented as a supporting evidence of a CW attack. Biomedical samples, e.g. blood and urine, may be analyzed for biological markers of poisoning as evidence that individuals have been exposed to a CW agent [2]. Biomedical sample analysis also has applications in exposure monitoring, e.g. in individuals engaged in demilitarization activities, and for the diagnosis of poisoning prior to the administration of medical countermeasures. In general, nerve agents are rather volatile and easy to degrade. Once exposed to the environment nerve agents readily degrade by rapid hydrolysis to the corresponding alkyl methylphosphonic acids (AMPAs) which never exist in nature. Sarin to isopropyl methylphosphonic acid (IMPA), Soman to pinacolyl methylphosphonic acid (PMPA), VX to ethyl methylphosphonic acid (EMPA), Russian VX to isobutyl methylphosphonic acid (i-BuMPA). As the degradation productions are more stable than their parent compounds, they have been used as surrogates to indicate the presence of the original nerve agents. Several analytical methods have measured AMPAs to determine nerve agents exposure, etc. the authors [3] determined alkyl phosphonic and phosphonothioic acids by using GC-MS for the pentafluorobenzyl derivatives. Several methods, based on the detection of nerve agents degradation products, have been reported for the identification of nerve agents in biological [4,5] samples. These approaches have advantages but may require derivatization that add to sample preparation time. Liquid chromatography–mass spectrometry (LC–MS) thus seems to be an attractive alternative to GC–MS for the analysis of chemical warfare agent degradation products, which are frequently nonvolatile, highly polar and very water soluble, since it allows the direct analysis of aqueous samples or aqueous extracts with little or no sample preparation. In recent years, with the development of new interfaces and ionization methods, more and more papers have been published on the analysis of chemical warfare agent degradation products using LC–MS. The main purpose of our work was to develop a rapid, sensitive technique for EMPA, i-BuMPA, IMPA and PMPA determination in urine samples. We present a method for the simultaneous detection of EMPA, i-BuMPA, IMPA and PMPA in human urine spiked with EMPA, i-BuMPA, IMPA and PMPA that is based on the materials used by liquid chromatography interfaced to negative ion-electrospray ionization-tandem mass spectrometry. Columns under gradient conditions were used. By automating and optimizing several sample cleanup steps, the method provides the LC–MS/MS instrument with sufficiently clean concentrated samples, at a rate comparable to that at which it quantifies them. The use of fast, automated sample preparation steps, employing separate solid-phase extraction analyte isolation, provides samples that are sufficiently clean and the 0.8 ng/ml for EMPA, 0.6 ng/ml for IMPA, 0.5 ng/ml for i-BuMPA and 0.1 ng/ml for PMPA limit of detections to be achieved in urine samples. Thus, the method is well suited for the purpose of the biomonitoring of nerve agents exposure in the event of a mass-casualty terrorist incident or in case of an accident at CW storage units characterized by high sample loads and the low concentrations to be detected. References 1. Convention on the Prohibition of the Development. Production. Stockpiling and Use of Chemical Weapons and on their Destruction. Technical Secretariat for the Organization for the Prohibition of Chemical Weapons. The Hague. 1997. 2. T.P. Logan, J.R. Smith, E.M. Jakubowski, R.E. Nielson, Toxicol. Methods 9 (1999) 275. 3. M. Palit, A.K. Gupta, Rajeev Jain, S.K. Raza, J. Chromatogr. A 1043 (2004) 275. 4. M. Koller, C. Becker, H. Thiermann, F. Worek, J. Chromatogr. B 878 (2010) 1226. 5. L. N. McDaniel, N. A. Romero, J. Boyd, G. Coimbatore, G. P. Cobb, Talanta 81 (2010) 1568.