Comprehensive gas chromatography with Time of Flight MS and large volume introduction for the detection of fluoride-induced regenerated nerve agent in biological samples

Recently, several methods have been developed to verify exposure to nerve agents. Most of these methods, such as the fluoride reactivation technique and the analysis of inhibited phosphonylated butyrylcholinesterase (BuChE), are based on mass spectrometry. The high specificity of the mass spectrometer might also imply a disadvantage, because the acquisition mass, i.e. the identity of the analyte must be known beforehand in order to direct the MS analysis in the most sensitive mode. In real cases, the identity of the nerve agent is not always known beforehand and the mass spectrometer should be operated in a scanning mode, with the consequence that sensitivity of the method will be lower. Comprehensive GC, or GC × GC, is a technique which offers enhanced separation. The implied larger selectivity of the GC separation allows mass spectrometry to be conducted in a less specific, scanning, mode. By the use of this configuration, the identity of the nerve agent does not have to be known beforehand but can be traced. In order to be able to detect lower concentrations and assess lower exposure levels, a large volume injection technique was developed allowing sample sizes up to 100 μL. The technique was tested with plasma samples that had been inhibited with various nerve agents. Subsequently, the cholinesterase-bound nerve agent was regenerated by the fluoride reactivation technique. Using the newly developed comprehensive GC–MS method it was possible to detect nerve agent at an exposure level of 1% BuChE inhibition, which is approximately 70 pg nerve agent/mL. These low exposure levels cannot be verified with a cholinesterase (ChE) activity assay. Moreover, the identity of the regenerated nerve agent was verified by the mass spectrum that was generated by the TOF mass spectrometer. This paper presents a technique able to deliver full-scan data on the analysis of nerve agents in biomedical samples at relevant exposure levels (1% BuChE inhibition). This full-scan data meets for a large part the forensic requirements that are in place for the analysis of biomedical samples in the context of alleged use of Chemical Warfare Agents.