Potency of irritation by benzylidenemalononitriles in humans correlates with TRPA1 ion channel activation.

We show that the physiological activity of solid aerosolized benzylidenemalononitriles (BMNs) including 'tear gas' (CS) in historic human volunteer trials correlates with activation of the human transient receptor potential ankyrin 1 ion channel (hTRPA1). This suggests that the irritation caused by the most potent of these compounds results from activation of this channel. We prepared 50 BMNs and measured their hTRPA1 agonist potencies. A mechanism of action consistent with their physiological activity, involving their dissolution in water on contaminated body surfaces, cell membrane penetration and reversible thiolation by a cysteine residue of hTRPA1, supported by data from nuclear magnetic resonance experiments with a model thiol, explains the structure-activity relationships. The correlation provides evidence that hTRPA1 is a receptor for irritants on nociceptive neurons involved in pain perception; thus, its activation in the eye, nose, mouth and skin would explain the symptoms of lachrymation, sneezing, coughing and stinging, respectively. The structure-activity results and the use of the BMNs as pharmacological tools in future by other researchers may contribute to a better understanding of the TRPA1 channel in humans (and other animals) and help facilitate the discovery of treatments for human diseases involving this receptor.

Simultaneous measurement of tabun, sarin, soman, cyclosarin, VR, VX, and VM adducts to tyrosine in blood products by isotope dilution UHPLC-MS/MS.

This work describes a new specific, sensitive, and rapid stable isotope dilution method for the simultaneous detection of the organophosphorus nerve agents (OPNAs) tabun (GA), sarin (GB), soman (GD), cyclosarin (GF), VR, VX, and VM adducts to tyrosine (Tyr). Serum, plasma, and lysed whole blood samples (50 µL) were prepared by protein precipitation followed by digestion with Pronase. Specific Tyr adducts were isolated from the digest by a single solid phase extraction (SPE) step, and the analytes were separated by reversed-phase ultra high performance liquid chromatography (UHPLC) gradient elution in less than 2 min. Detection was performed on a triple quadrupole tandem mass spectrometer using time-triggered selected reaction monitoring (SRM) in positive electrospray ionization (ESI) mode. The calibration range was characterized from 0.100-50.0 ng/mL for GB- and VR-Tyr and 0.250-50.0 ng/mL for GA-, GD-, GF-, and VX/VM-Tyr (R(2) ≥ 0.995). Inter- and intra-assay precision had coefficients of variation of ≤17 and ≤10%, respectively, and the measured concentration accuracies of spiked samples were within 15% of the targeted value for multiple spiking levels. The limit of detection was calculated to be 0.097, 0.027, 0.018, 0.074, 0.023, and 0.083 ng/mL for GA-, GB-, GD-, GF-, VR-, and VX/VM-Tyr, respectively. A convenience set of 96 serum samples with no known nerve agent exposure was screened and revealed no baseline values or potential interferences. This method provides a simple and highly specific diagnostic tool that may extend the time postevent that a confirmation of nerve agent exposure can be made with confidence.

The development of an analytical method for urinary metabolites of the riot control agent 2-chlorobenzylidene malononitrile (CS).

The analysis of biomedical samples such as urine and blood can provide evidence of exposure to chemicals for a range of applications including occupational exposure monitoring, detection of drugs of abuse, performance enhancement in sport and investigations of poisoning and incapacitation. This paper reports the development of an analytical method for two suspected urinary metabolites of the riot control agent 2-chlorobenzylidene malononitrile (CS): 2-chlorohippuric acid and 2-chlorobenzyl-N-acetylcysteine. 2-Chlorohippuric acid was identified in all 2h post-exposure samples from a set of urine samples taken from army recruits exposed to low levels of thermally dispersed CS during training. 2-Chlorobenzyl-N-acetylcysteine, a metabolite known to be formed in the rat, was not identified in any of the samples. The lower limit of detection (LLOD) for 2-chlorohippuric acid and 2-chlorobenzyl-N-acetylcysteine was 1ng/ml and 0.5ng/ml in pooled urine from the pre-exposed subjects. 2-Chlorohippuric acid was rapidly excreted but was detectable in the urine of 17 of the 19 subjects tested 20h after exposure.

Analysis of clothing and urine from Moscow theatre siege casualties reveals carfentanil and remifentanil use.

On October 26, 2002, Russian Special Forces deployed a chemical aerosol against Chechen terrorists to rescue hostages in the Dubrovka theatre. Its use confirmed Russian military interest in chemicals with effects on personnel and caused 125 deaths through a combination of the aerosol and inadequate medical care. This study provides evidence from liquid chromatography-tandem mass spectrometry analysis of extracts of clothing from two British survivors, and urine from a third survivor, that the aerosol comprised a mixture of two anaesthetics--carfentanil and remifentanil--whose relative proportions this study was unable to identify. Carfentanil and remifentanil were found on a shirt sample and a metabolite called norcarfentanil was found in a urine sample. This metabolite probably originated from carfentanil.

Two cases of neurofibromatosis presenting with a foot deformity.

Two children presented with an isolated foot and ankle deformity. Examination in each suggested a plexiform neurofibroma although this diagnosis had not been considered before referral. Diagnosis of neurofibromatosis type 1 was confirmed by MRI scanning and on investigation both patients were proved to have widespread disease. One had a plexiform neurofibroma encasing the aorta and oesophagus. Both cases remain under observation and have not undergone surgery for their disease. Neurofibromatosis can present with isolated foot and ankle deformity and when such a diagnosis is suspected thorough investigation is important in a condition in which unsuspected widespread disease may exist.

Biomarkers of organophosphorus nerve agent exposure: comparison of phosphylated butyrylcholinesterase and phosphylated albumin after oxime therapy.

Organophosphorus nerve agents inhibit the activity of cholinesterases by phosphylation of the active site serine. In addition, sarin, cyclosarin, soman and tabun have been shown to phosphylate a tyrosine residue in albumin. Therapies against nerve agent poisoning include the use of oximes to reactivate inhibited cholinesterases by displacement of the phosphyl moiety and hence detectable levels of adducts with cholinesterases may be reduced. Adducts with tyrosine have been shown to be persistent in the guinea pig in the presence of oxime therapy. Plasma samples obtained from an animal study aimed at improving therapy against nerve agent poisoning were used to compare the suitability of tyrosine and butyrylcholinesterase (BuChE) adducts as biomarkers of nerve agent exposure after treatment with therapeutic oximes. Under the terms of the project licence, these samples could be collected only on death of the animal, which occurred within hours of exposure or when culled at 23 or 24 days. Tyrosine adducts were detected in all samples collected following intra-muscular administration of twice the LD50 dose of the respective nerve agent. Aged BuChE adducts were detected in samples collected within a few hours after administration of soman and tabun, but not after 23 or 24 days. No BuChE adducts were detected in animals exposed to sarin and cyclosarin where samples were collected only after 23 or 24 days.