Tokyo subway samples

Recently, Noort et al developed a procedure that is based on straightforward isolation of adducted BuChE from plasma by means of affinity chromatography with a procainamide column, followed by pepsin digestion and LC/electrospray tandem MS analysis of a specific nonapeptide containing the phosphonylated active site serine-198 residue (5). This method surpasses the limitations of the fluoride-reactivation method, since it can also deal with dealkylated ( aged ) phosphonylated BuChE. The method allowed the positive analysis of several serum samples of Japanese victims of the terrorist attack in the Tokyo subway in 1995. Furthermore, the method could be applied for detection of ChE modifications induced by, e.g., diethyl paraoxon and pyridostigmine bromide, illustrating the broad scope of this approach. This new approach... 

The only reported incidents of nerve agent poisoning, where biomedical samples have been obtained, are those resulting from terrorist dissemination of sarin in Matsumoto (1994) and the Tokyo subway (1995), plus an assassination using VX, also in Japan (60). In contrast to the CW incidents involving sulfur mustard, many of the biomedical samples associated with these terrorist attacks were collected within hours of the event. 

Application of the fluoride-reactivation method to serum samples of victims from the Tokyo subway attack, and of the Matsumoto incident, yielded, sarin concentrations in the range of 0.2-4.1ng/ml serum (44). Evidently, these casualties had been exposed to an organophosphate with the formula i-PrO(CH3)P(0)X, presumably with X = F (sarin). 

Finally, mass spectrometric determination of the phosphonylated peptic nonapeptide from butyrylcholinesterase allowed the positive identification of sarin-inhibited enzyme in serum samples from several Japanese victims of the Tokyo subway attack (see Figure 10). 

Tissue sample collection is obviously the most invasive and usually limited to a deceased casualty. Consequently, tissue collection is not normally used for diagnosis to medically respond but for forensic analysis, such as the formalin-fixed brain tissues from the Tokyo subway attack victims, to verify GB as the agent employed (Matsuda et al., 1998). 

Although the police investigation collected enough evidence to prove that sarin was used by AUM Shinrikyo terrorists in Tokyo, the final scientific proof came this summer, when two laboratories independently determined sarin metabolites in blood and urine samples drawn from Tokyo subway attack victims. The Holland group of investigators liberated them from plasma butyrylcholineste-rases [14], while the Japanese group used urine instead [30],... 

A new method for identifying exposure to nerve agents was introduced by Polhuijs et al. (1997). The metirod is based on tire finding that incubation of sarin-inhibited BChE witir 2M potassium fluoride at pH 4 results in the release of sarin. Sarin is then extracted and analyzed by CC-MS. Polhuijs et al. (1997) applied their new metirod to positively identify sarin in serum samples from Japanese victims of the Tokyo subway attack. This method has tire advantage that it positively identifies nerve agents and other OPs (van der Schans et al., 2004). The method has been validated by Adams et al. (2004), who found that potassium fluoride released sarin and soman from human BChE, as well as from covalent attachment to human albunrin. 

Today, everyone is aware of terrorist activity. A well-known terrorist attack involving chemicals occurred in 1995 with the release of the nerve gas sarin in the subway systems of Tokyo, resulting in the death of 12 people and injury of many more. Should chemicals again be used for terrorist activity, forensic pharmacologists and toxicologists may be called upon to analyze bodily samples in order to identify the chemical, determine its mechanism of action, and propose antidotes and preventive measures. 

Application of fluoride reactivation to serum samples of casualties of the Matsumoto and Tokyo incidents yielded sarin concentrations in the range 0.2 1.1 ng/ml serum (Polhuijs et al, 1997). Hydrolytic displacement of the phospho-nyl residue identified /PrMPA at levels sufficient for full-scan mass spectra to be obtained from samples collected from casualties who died (Nagao et al, 1997) MPA was also identified. MPA was detected in formalin-fixed brain tissues some two years later using a similar procedure (Matsuda et al, 1998). The phosphonylated peptic nonapeptide from BuChE was identified in serum samples from several casualties of the subway attack (Fidder et al, 2002). 

Chemical warfare nerve agents pose a potential threat to the general public as well as the military, as evidenced by several incidents. Between 1980 and 1988, sarin (GB) was used by Iraq in the war with Iran, with the most notable incident occurring in 1988 when a Kurdish city in northern Iraq was bombarded with chemicals, possibly including GB, tabun (GA) and 0-ethyl 5-[2-(diisopropylamino)ethyl] methylphosphonothioate (VX). In 1994 and 1995, the Aum Shinrikyo sect attacked subways in Matsumoto and Tokyo with GB, and also attacked individuals with VX in Osaka and Tokyo. One of these individual attacks resulted in the death of the intended victim. The victim had VX deposited on his neck and exhibited symptoms typical of organo-phosphate poisoning, but confirmation of the nerve agent used could only be achieved after his death with the testimony from one of the suspected attackers and detection of VX metabolites [ethyl methylphosphonic acid (EMPA) and 2-(diisopropylamino-ethyl)methyl sulfide (DAEMS)] in a blood sample taken approximately 1 h after the attack. 

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