Chromogenic Detection of the Organophosphorus Nerve Agent Simulant DCP Mediated by Rhodium(II,II) Paddlewheel Complexes.

Organophosphorus nerve agents (OPNAs) pose a great threat to humanity. Possessing extreme toxicity, rapid lethality, and an unassuming appearance, these chemical warfare agents must be quickly and selectively identified so that treatment can be administered to those affected. Chromogenic detection is the most convenient form of OPNA detection, but current methods suffer from false positives. Here, nitrogenous base adducts of dirhodium(II,II) acetate were synthesized and used as chromogenic detectors of diethyl chlorophosphate (DCP), an OPNA simulant. UV-vis spectrophotometry was used to evaluate the sensitivity and selectivity of the complexes in the detection of DCP. Visual limits of detection (LOD) for DCP were as low as 1.5 mM DCP, while UV-vis-based LODs were as low as 0.113 µM. The dirhodium(II,II) complexes were also tested with several potential interferents, none of which produced a visual color change that could be mistaken for OPNA response. Ultimately, the Rh2(OAc)4(1,8-diazabicyclo[5.4.0]undec-7-ene)2 complex showed the best combination of detection capability and interferent resistance. These results, when taken together, show that dirhodium(II,II) paddlewheel complexes with nitrogenous base adducts can produce instant, selective, and sensitive detection of DCP. It is our aim to further explore and apply this new motif to produce even more capable OPNA sensors.

Highly specific and sensitive chromo-fluorogenic detection of sarin, tabun, and mustard gas stimulants: a multianalyte recognition approach.

Nerve agents are the most notorious substances, which can be fatal to an individual because they block the activity of acetylcholinesterase. Fighting against unpredictable terrorist assaults and wars requires the simple and quick detection of chemical warfare agent vapor. In the present contribution, we have introduced a rhodamine-based chemosensor, BDHA, for the detection of nerve gas-mimicking agents diethylchlorophosphate (DCP) and diethylcyanophosphonate (DCNP) and mustard gas-mimicking agent 2-chloroethyl ethyl sulfide (CEES), both in the liquid and vapor phase. Probe BDHA provides the ability for detection by the naked eye in terms of colorimetric and fluorometric changes. It has been revealed that the interaction between nerve agents mimics and probe BDHA facilitates spirolactam ring opening due to the phosphorylation process. Thus, the highly fluorescent and colored species developed while probe BDHA is colorless and non-fluorescent due to the intramolecular spirolactam ring. Moreover, probe BDHA can effectively recognize DCP, DCNP, and CEES in the µM range despite many toxic analytes and could be identified based on the response times and quantum yield values. Inexpensive, easily carried paper strips-based test kits were developed for the quick, on-location solid and vapor phase detection of these mustard gas imitating agents (CEES) and nerve gas mimicking agents (DCP and DCNP) without needing expensive equipment or skilled personnel. More remarkably, the test strips' color and fluorescence can be rapidly restored, exposing them to triethyl amine (TEA) for cyclic use, suggesting a potential application in the real-time identification of chemical warfare agents. To accomplish the on-location application of BDHA, we have experimented with soil samples to find traces of DCP. Therefore, the chromo-fluorogenic probe BDHA is a promising, instantaneous, and on-the-spot monitoring tool for the selective detection of DCP, DCNP, and CEES in the presence of others.

The prediction of hydrolysis and biodegradation of organophosphorus-based chemical warfare agents (G-series and V-series) using toxicology in silico methods.

Nerve agents (G- and V-series) are a group of extremely toxic organophosphorus chemical warfare agents that we have had the opportunity to encounter many times on a massive scale (Matsumoto City, Tokyo subway and Gulf War). The threat of using nerve agents in terrorist attacks or military operations is still present, even with establishing the Chemical Weapons Convention as the legal framework. Understanding their environmental sustainability and health risks is critical to social security. Due to the risk of contact with dangerous nerve agents and animal welfare considerations, in silico methods were used to assess hydrolysis and biodegradation safely. The environmental fate of the examined nerve agents was elucidated using QSAR models. The results indicate that the investigated compounds released into the environment hydrolyse at a different rate, from extremely fast (<1 day) to very slow (over a year); V-agents undergo slower hydrolysis compared to G-agents. V-agents turned out to be relatively challenging to biodegrade, the ultimate biodegradation time frame of which was predicted as weeks to months, while for G-agents, the overwhelming majority was classified as weeks. In silico methods for predicting various parameters are critical to preparing for the forthcoming application of nerve agents.

The Search for Antidotes Against Ricin.

The castor plant (Ricinus communis) is primarily known for its seeds, which contain a unique fatty acid called ricinoleic acid with several industrial and commercial applications. Castor seeds also contain ricin, a toxin considered a chemical and biological warfare agent. Despite years of investigation, there is still no effective antidote or vaccine available. However, some progress has been made, and the development of an effective treatment may be on the horizon. To provide an updated overview of this issue, we have conducted a comprehensive review of the literature on the current state of research in the fight against ricin. This review is based on the reported research and aims to address the challenges faced by researchers, as well as highlight the most successful cases achieved thus far. Our goal is to encourage the scientific community to continue their efforts in this critical search.

Ethanolysis of nitrogen mustards: A novel strategy for nitrogen mustard identification in environmental matrices by liquid chromatography-electrospray ionization-tandem mass spectrometry.

RATIONALE: Nitrogen mustards (NMs) are blistering chemical warfare agents. The ability to detect NMs in environmental samples is very important for obtaining forensic evidence. The most common analytical techniques for NM detection are gas chromatography-mass spectrometry, which detects NMs in their intact form but is disadvantaged by high limits of detection (LODs), and liquid chromatography-electrospray ionization-tandem mass spectrometry (LC/ESI-MS/MS) of their hydrolysis products, which do not provide robust evidence to support NM use. METHODS: We developed a novel approach to detect and identify NMs using LC/ESI-MS/MS after chemical derivatization. The method is based on ethoxide-promoted ethanolysis prior to analysis. The effects of reaction time, temperature, ethoxide concentration and chromatography behavior were studied and optimized. In the developed procedure, 0.1% (v/v) sodium ethoxide solution is added to the NMs in ethanol and agitated for 2 h at 50°C, followed by LC/ESI-MS/MS, without any other pretreatment. RESULTS: The ethanolysis reaction efficiencies were evaluated in ethanolic extracts from soil, asphalt, and ethanol contaminated with 0.5% (v/v) diesel fortified with NMs at a five-point calibration curve. The calibration curves showed good linearity in the range of 0.05-1 ng/mL, with an R2 value of 0.99, and were similar to those of LC/MS-grade ethanol, with almost no observable matrix effects. The derivatization products were stable at room temperature, with LODs of 10 pg/mL, in all investigated extracts. CONCLUSIONS: Through this newly developed strategy, the derivatization of active NMs by ethanolysis was achieved for the first time, and these derivatization products can serve as specific indicators for the use and presence of NMs. The methodology can also verify trace levels of NM chemical warfare agents collected in war or terror scenarios in forensic investigations.

Pnictogen bond-driven control of the molecular interaction between organophosphorus and acetylcholinesterase enzyme.

This study addresses a comprehensive assessment of the interaction between chemical warfare agents (CWA) and acetylcholinesterase (AChE) systems, focus on the intriguing pnictogen-bond interaction (PnB). Utilizing the crystallographic data from the Protein Data Bank pertaining to the AChE-CWA complex involving Sarin (GB), Cyclosarin (GF), 2-[fluoro(methyl)phosphoryl]oxy-1,1-dimethylcyclopentane (GP) and venomous agent X (VX) agents, the CWA is systematically displaced by increments of 0.1 Å along the PO bond axis, extending its distance by 4 Å from the original position. The AIM analysis was carried out and consistently revealed the presence of a significant interaction along the PO bond. Investigating the intrinsic nature of the PnB, the NBO and the EDA analysis unearthed the contribution of orbital factors to the overall energy of the system. Strikingly, this observation challenges the conventional σ-hole explanation commonly associated with such interactions. This finding adds a layer of complexity to understanding of PnB, encouraging further exploration into the underlying mechanisms governing these intriguing chemical phenomena.

Powders to Thin Films: Advances in Conjugated Microporous Polymer Chemical Sensors.

Chemical sensing of harmful species released either from natural or anthropogenic activities is critical to ensuring human safety and health. Over the last decade, conjugated microporous polymers (CMPs) have been proven to be potential sensor materials with the possibility of realizing sensing devices for practical applications. CMPs found to be unique among other porous materials such as metal-organic frameworks (MOFs) and covalent organic frameworks (COFs) due to their high chemical/thermal stability, high surface area, microporosity, efficient host-guest interactions with the analyte, efficient exciton migration along the π-conjugated chains, and tailorable structure to target specific analytes. Several CMP-based optical, electrochemical, colorimetric, and ratiometric sensors with excellent selectivity and sensing performance were reported. This review comprehensively discusses the advances in CMP chemical sensors (powders and thin films) in the detection of nitroaromatic explosives, chemical warfare agents, anions, metal ions, biomolecules, iodine, and volatile organic compounds (VOCs), with simultaneous delineation of design strategy principles guiding the selectivity and sensitivity of CMP. Preceding this, various photophysical mechanisms responsible for chemical sensing are discussed in detail for convenience. Finally, future challenges to be addressed in the field of CMP chemical sensors are discussed.

The phosphylated butyrylcholinesterase-derived tetrapeptide GlyGluSerAla proves exposure to organophosphorus agents with enantioselectivity.

We herein present for the first time the phosphylated (*) tetrapeptide (TP)-adduct GlyGluSer198*Ala generated from butyrylcholinesterase (BChE) with proteinase K excellently suited for the verification of exposure to toxic organophosphorus nerve agents (OPNA). Verification requires bioanalytical methods mandatory for toxicological and legal reasons. OPNA react with BChE by phosphonylation of the active site serine residue (Ser198) forming one of the major target protein adducts for verification. After its enzymatic cleavage with pepsin, the nonapeptide (NP) PheGlyGluSer*AlaGlyAlaAlaSer is typically produced as biomarker. Usually OPNA occur as racemic mixtures of phosphonic acid derivatives with the stereocenter at the phosphorus atom, e.g. (±)-VX. Both enantiomers react with BChE, but the adducted NP does not allow their chromatographic distinction. In contrast, the herein introduced TP-adducts appeared as two peaks when using a stationary reversed phase (1.8 µm) in micro-liquid chromatography-electrospray ionisation tandem-mass spectrometry (µLC-ESI MS/MS) analysis. These two peaks represent diastereomers of the (+)- and (-)-OPNA adducted to the peptide that comprises chiral L-amino acids exclusively. Concentration- and time-dependent effects of adduct formation with (±)-VX and its pure enantiomers (+)- and (-)-VX as well as with (±)-cyclosarin (GF) were investigated in detail characterising enantioselective adduct formation, stability, ageing and spontaneous reactivation. The method was also successfully applied to samples from a real case of pesticide poisoning as well as to samples of biomedical proficiency tests provided by the Organisation for the Prohibition of Chemical Weapons.

The prediction of acute toxicity (LD50) for organophosphorus-based chemical warfare agents (V-series) using toxicology in silico methods.

Nerve agents are organophosphate chemical warfare agents that exert their toxic effects by irreversibly inhibiting acetylcholinesterase, affecting the breakdown of the neurotransmitter acetylcholine in the synaptic cleft. Due to the risk of exposure to dangerous nerve agents and for animal welfare reasons, in silico methods have been used to assess acute toxicity safely. The next-generation risk assessment (NGRA) is a new approach for predicting toxicological parameters that can meet modern requirements for toxicological research. The present study explains the acute toxicity of the examined V-series nerve agents (n = 9) using QSAR models. Toxicity Estimation Software Tool (ver. 4.2.1 and ver. 5.1.2), QSAR Toolbox (ver. 4.6), and ProTox-II browser application were used to predict the median lethal dose. The Simplified Molecular Input Line Entry Specification (SMILES) was the input data source. The results indicate that the most deadly V-agents were VX and VM, followed by structural VX analogues: RVX and CVX. The least toxic turned out to be V-sub x and Substance 100A. In silico methods for predicting various parameters are crucial for filling data gaps ahead of experimental research and preparing for the upcoming use of nerve agents.

Chemical, Physical, and Toxicological Properties of V-Agents.

V-agents are exceedingly toxic organophosphate nerve agents. The most widely known V-agents are the phosphonylated thiocholines VX and VR. Nonetheless, other V-subclasses have been synthesized. Here, a holistic overview of V-agents is provided, where these compounds have been categorized based on their structures to facilitate their study. A total of seven subclasses of V-agents have been identified, including phospho(n/r)ylated selenocholines and non-sulfur-containing agents, such as VP and EA-1576 (EA: Edgewood Arsenal). Certain V-agents have been designed through the conversion of phosphorylated pesticides to their respective phosphonylated analogs, such as EA-1576 derived from mevinphos. Further, this review provides a description of their production, physical properties, toxicity, and stability during storage. Importantly, V-agents constitute a percutaneous hazard, while their high stability ensures the contamination of the exposed area for weeks. The danger of V-agents was highlighted in the 1968 VX accident in Utah. Until now, VX has been used in limited cases of terrorist attacks and assassinations, but there is an increased concern about potential terrorist production and use. For this reason, studying the chemistry of VX and other less-studied V-agents is important to understand their properties and develop potential countermeasures.

Application of toxicology in silico methods for prediction of acute toxicity (LD50) for Novichoks.

Novichoks represent the fourth generation of chemical warfare agents with paralytic and convulsive effects, produced clandestinely during the Cold War by the Soviet Union. This novel class of organophosphate compounds is characterised by severe toxicity, which, for example, we have already experienced three times (Salisbury, Amesbury, and Navalny's case) as a society. Then the public debate about the true nature of Novichoks began, realising the importance of examining the properties, especially the toxicological aspects of these compounds. The updated Chemical Warfare Agents list registers over 10,000 compounds as candidate structures for Novichoks. Consequently, conducting experimental research for each of them would be a huge challenge. Additionally, due to the enormous risk of contact with hazardous Novichoks, in silico assessments were applied to estimate their toxicity safely. In silico toxicology provides a means of identifying hazards of compounds before synthesis, helping to fill gaps and guide risk minimisation strategies. A new approach to toxicology testing first considers the prediction of toxicological parameters, eliminating unnecessary animal studies. This new generation risk assessment (NGRA) can meet the modern requirements of toxicological research. The present study explains, using QSAR models, the acute toxicity of the Novichoks studied (n = 17). The results indicate that the toxicity of Novichoks varies. The deadliest turned out to be A-232, followed by A-230 and A-234. On the other hand, the "Iranian" Novichok and C01-A038 compounds turned out to be the least toxic. Developing reliable in silico methods to predict various parameters is essential to prepare for the upcoming use of Novichoks.

Decontamination of mustard sulfur and VX by sodium percarbonate complexed with 1-acetylguanidine as a novel activator.

The peroxide-based decontaminants had attracted great attention for degradation of chemical warfare agents (CWAs) because of their high performance, non-corrosive and environmental-friendly merits. Hydrogen peroxide can be activated by some organic activators to enhance the oxidation ability. In this work, a novel formula based on sodium percarbonate (SPC) complexed with 1-acetylguanidine (ACG) was investigated for decontamination of sulfur mustard (HD) and VX as CWAs. In the experimental results, the active species acetyl peroxide imide acid in the formula aqueous solution was detected in situ by Raman and 13C NMR spectroscopy. The optimized conditions of the decontamination formula (SPC/ACG) were suggested that, the molar ratio of active oxygen and activator ([O]/[ACG]) was 1:1 while the pH value of the formula aqueous solution was about 9. To achieve the decontamination percentage over 99%, the molar ratio of active oxygen to CWA ((O)/(CWA)) needed to be at least 3 for HD and 7 for VX. Meanwhile, the degradation products detected by gas chromatography/mass spectrometry (GC/MS), liquid chromatography/mass spectrometry (LC/MS) and ion chromatography (IC) indicated that the oxidation and elimination reactions should have occurred on HD molecule, while the degradation of VX mainly originate from the nucleophilic substitution and oxidation reactions.

What do we currently know about Novichoks? The state of the art.

Novichok is the name given to the group of nerve agents created stealthily in the later phases of the Cold War by the Soviet Union. Constitute the fourth generation of chemical warfare agents; like other nerve agents, they are organophosphorus compounds designed to be incurable and undetectable. The mechanism of action is based on the non-competitive and irreversible inhibition of acetylcholinesterase. Due to their enormous toxicity, Novichoks have become attractive targets for terrorists. However, little information is known about the identity of nerve agents. Furthermore, these compounds have never been submitted to the Chemical Weapons Convention. Our article aspires to provide a general overview of Novichoks knowledge. As part of this, we reviewed the available literature data to answer the question, what are Novichoks? In addition to the physical and chemical properties of A-agents, synthesis, mechanism of action, and toxicity of nerve agents were also reviewed. We hope that this review will highlight the tremendous threat posed by nerve agents and will inspire further studies on the interdisciplinary aspects of these compounds.

Verification of exposure to chemical warfare agents through analysis of persistent biomarkers in plants.

The continuing threats of military conflicts and terrorism may involve the misuse of chemical weapons. The present study aims to use environmental samples to find evidence of the release of such agents at an incident scene. A novel approach was developed for identifying protein adducts in plants. Basil (Ocimum basilicum), bay laurel leaf (Laurus nobilis) and stinging nettle (Urtica dioica) were exposed to 2.5 to 150 mg m-3 sulfur mustard, 2.5 to 250 mg m-3 sarin, and 0.5 to 25 g m-3 chlorine gas. The vapors of the selected chemicals were generated under controlled conditions in a dedicated set-up. After sample preparation and digestion, the samples were analyzed by liquid chromatography tandem mass spectrometry (LC-MS/MS) and liquid chromatography high resolution tandem mass spectrometry (LC-HRMS/MS), respectively. In the case of chlorine exposure, it was found that 3-chloro- and 3,5-dichlorotyrosine adducts were formed. As a result of sarin exposure, the o-isopropyl methylphosphonic acid adduct to tyrosine could be analyzed, and after sulfur mustard exposure the N1- and N3-HETE-histidine adducts were identified. The lowest vapor exposure levels for which these plant adducts could be detected, were 2.5 mg m-3 for sarin, 50 mg m-3 for chlorine and 12.5 mg m-3 for sulfur mustard. Additionally, protein adducts following a liquid exposure of only 2 nmol Novichock A-234, 0.4 nmol sarin and 0.2 nmol sulfur mustard could still be observed. For both vapor and liquid exposure, the amount of adduct formed increased with the level of exposure. In all cases synthetic reference standards were used for unambiguous identification. The window of opportunity for investigation of agent exposure through the analysis of plant material was found to be remarkably long. Even three months after the actual exposure, the biomarkers could still be detected in the living plants, as well as in dried leaves. An important benefit of the current method is that a relatively simple and generic sample work-up procedure can be applied for all agents studied. In conclusion, the presented work clearly demonstrates the possibility of analyzing chemical warfare agent biomarkers in plants, which is useful for forensic reconstructions, including the investigation into alleged use in conflict areas.

Isolation of human TRPA1 channel from transfected HEK293 cells and identification of alkylation sites after sulfur mustard exposure.

Transient receptor potential (TRP) channels are important in the sensing of pain and other stimuli. They may be triggered by electrophilic agonists after covalent modification of certain cysteine residues. Sulfur mustard (SM) is a banned chemical warfare agent and its reactivity is also based on an electrophilic intermediate. The activation of human TRP ankyrin 1 (hTRPA1) channels by SM has already been documented, however, the mechanism of action is not known in detail. The aim of this work was to purify hTRPA1 channel from overexpressing HEK293 cells for identification of SM-induced alkylation sites. To confirm hTRPA1 isolation, Western blot analysis was performed showing a characteristic double band at 125 kDa. Immunomagnetic separation was carried out using either an anti-His-tag or an anti-hTRPA1 antibody to isolate hTRPA1 from lysates of transfected HEK293 cells. The identity of the channel was confirmed by micro liquid chromatography-electrospray ionization high-resolution tandem-mass spectrometry. Following SM exposure, hTRPA1 channel modifications were found at Cys462 and Cys665, as well as at Asp339 and Glu341 described herein for the first time. Since Cys665 is a well-known target of hTRPA1 agonists and is involved in hTRPA1 activation, SM-induced modifications of cysteine, as well as aspartic acid and glutamic acid residues may play a role in hTRPA1 activation. Considering hTRPA1 as a target of other SM-related chemical warfare agents, analogous adducts may be predicted and identified applying the analytical approach described herein.

Cooperative Catalysis between Dual Copper Centers in a Metal-Organic Framework for Efficient Detoxification of Chemical Warfare Agent Simulants.

Chemical warfare agents (CWAs) are among the most lethal chemicals known to humans. Thus, developing multifunctional catalysts for highly efficient detoxification of various CWAs is of great importance. In this work, we developed a robust copper tetrazolate metal-organic framework (MOF) catalyst containing a dicopper unit similar to the coordination geometry of the active sites of natural phosphatase and tyrosinase enzymes. This catalyst aided in phosphate ester bond hydrolysis and hydrogen peroxide decomposition, ultimately achieving high detoxification efficiency against both a nerve agent simulant (diethoxy-phosphoryl cyanide (DECP)) with a half-life of 3.5 min and a sulfur mustard simulant (2-chloroethyl ethyl sulfide (CEES)) with a half-life of 4.5 min, making it competitive with other reported materials. The dicopper sites in ZZU-282 provide versatile binding modes with the substrates, thereby promoting the activation of substrates and enhancing the catalytic efficiency. A combination of postmodified metal exchange control experiments, density functional theory calculations, and catalytic evaluations confirmed that dual Cu sites are the active centers promoting the catalytic reaction. This study offers a new design perspective to achieve advanced catalysts for CWA detoxification.

Selective Photodetoxification of a Sulfur Mustard Simulant Using Plasmonic Aluminum Nanoparticles.

Plasmonic nanostructures have attracted increasing interest in the fields of photochemistry and photocatalysis for their ability to enhance reactivity and tune reaction selectivity, a benefit of their strong interactions with light and their multiple energy decay mechanisms. Here we introduce the use of earth-abundant plasmonic aluminum nanoparticles as a promising renewable detoxifier of the sulfur mustard simulant 2-chloroethylethylsulfide through gas phase photodecomposition. Analysis of the decomposition products indicates that C-S bond breaking is facilitated under illumination, while C-Cl breaking and HCl elimination are favored under thermocatalytic (dark) conditions. This difference in reaction pathways illuminates the potential of plasmonic nanoparticles to tailor reaction selectivity toward less hazardous products in the detoxification of chemical warfare agents. Moreover, the photocatalytic activity of the Al nanoparticles can be regenerated almost completely after the reaction concludes through a simple surface treatment.

Controllable synthesis of conjugated microporous polymer films for ultrasensitive detection of chemical warfare agents.

Nerve agents, one of the most toxic chemical warfare agents, seriously threaten human life and public security. The high toxicity of nerve agents makes the development of fluorescence sensors with suitable limit of detection challenging. Here, we propose a sensor design based on a conjugated microporous polymer film for the detection of diethyl chlorophosphate, a substitute of Sarin, with low detection limit of 2.5 ppt. This is due to the synergy of the susceptible on-off effect of hybridization and de-hybridization of hybrid local and charge transfer (HLCT) materials and the microporous structure of CMP films facilitating the inward diffusion of DCP vapors, and the extended π-conjugated structure. This strategy provides a new idea for the future development of gas sensors. In addition, a portable sensor is successfully integrated based on TCzP-CMP films that enables wireless, remote, ultrasensitive, and real-time detection of DCP vapors.

Software-Assisted Automated Detection and Identification of "Unknown" Analogues: Implementation on V-Type Nerve Agents.

V-type nerve agents are among the most toxic organophosphorus chemical warfare agents, and they are under strict regulation and supervision by the OPCW (Organization for the Prohibition of Chemical Weapons). The V-type class of materials refers to a potentially large number of analogues and isomers. In order to expose instances of unfulfillment of the OPCW treaty, it is essential to have the ability to detect and identify "unknown" analogues of this family, even in the absence of an analytical standard. This work demonstrates a new automated tool for the detection and identification of V-type analogues, using high-resolution-accurate-mass LC-MS analysis, followed by "Compound Discoverer" software data processing. This software, originally developed for metabolism and metabolomics screening, is used here to automatically detect various V-type analogues by picking peaks and comparing them to "in-silico" calculated modifications made on a predefined basic V-backbone structure (according to the OPCW definitions for V-type agents). Subsequently, a complete structural elucidation for the proposed molecular formula is obtained by MS/MS data analysis of the suspected component, for both the V-type analogue (using ESI(+) analysis) as well as its hydrolysis product (using ESI(-) analysis) for a better elucidation of the phosphonate "head" structure. This method was found to be useful for the detection and identification of several "unknown" analogues, at low ng/mL levels in soil extracts.

Highly stable peptide adducts from hard keratins as biomarkers to verify local sulfur mustard exposure of hair by high-resolution mass spectrometry.

In the recent past, the blister agent sulfur mustard (SM) deployed by the terroristic group Islamic State has caused a huge number of civilian and military casualties in armed conflicts in the Middle East. The vaporized or aerolized agent might be inhaled and have direct contact to skin and hair. Reaction products of SM with plasma proteins (adducts) represent well-established systemic targets for the bioanalytical verification of exposure. The SM-derived hydroxyethylthioethyl (HETE)-moiety is attached to nucleophilic amino acid side chains and allows unambiguous adduct detection. For shipping of common blood and plasma samples, extensive packaging rules are to be followed as these matrices are considered as potentially infectious material. In contrast, hair is considered as non-infectious thus making its handling and transportation much less complicated. Therefore, we addressed this matrix to develop a procedure for bioanalytical verification. Following optimized lysis of SM-treated human scalp hair and pepsin-catalyzed proteolysis of adducts of keratin type I and II, microbore liquid chromatography-electrospray ionization high-resolution tandem-mass spectrometry (µLC-ESI MS/HR MS) was used to detect three alkylated keratin-derived biomarker peptides: AE(-HETE)IRSDL, FKTIE(-HETE)EL, and LE(-HETE)TKLQF simultaneously. All bear the HETE-moiety bound to a glutamic acid residue. Protein adducts were stable for at least 14 weeks at ambient temperature and contact to air, and were not affected by washing the hair with shampoo. The biomarker peptides were also obtained from beard, armpit, abdominal, and pubic hair. This is the first report introducing stable local peptide adduct biomarkers from hair, that is easily accessible by a non-invasive sampling process.