Adduct of the blistering warfare agent sesquimustard with human serum albumin and its mass spectrometric identification
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Adduct of the blistering warfare agent sesquimustard with human serum albumin and its mass spectrometric identification for biomedical verification of exposure Marc-Michael Blum 1 & Annika Richter 2 & Markus Siegert 2,3 & Horst Thiermann 3 & Harald John 3 Received: 10 July 2020 / Revised: 7 August 2020 / Accepted: 25 August 2020 # The Author(s) 2020
Abstract Apart from the well-known sulfur mustard (SM), additional sulfur-containing blistering chemical warfare agents exist. Sesquimustard (Q) is one of them and five times more blistering than SM. It is a common impurity in mustard mixtures and regularly found in old munitions but can also be used in pure form. Compared to the extensive literature on SM, very little experimental data is available on Q and no protein biomarkers of exposure have been reported. We herein report for the first time the adduct of Q with the nucleophilic Cys34 residue of human serum albumin (HSA) formed in vitro and introduce two novel bioanalytical procedures for detection. After proteolysis of this HSA adduct catalyzed either by pronase or by proteinase K, two biomarkers were identified by high-resolution tandem mass spectrometry (MS/HR MS), namely a dipeptide and a tripeptide, both alkylated at their Cys residue, which we refer to as HETETE-CP and HETETE-CPF. HETETE represents the Q-derived thioalkyl moiety bearing a terminal hydroxyl group: “hydroxyethylthioethylthioethyl.” Targeting both peptide markers from plasma, a micro liquid chromatography–electrospray ionization tandem mass spectrometry method working in the selected reaction monitoring mode (μLC-ESI MS/MS SRM) was developed and validated as well suited for the verification of exposure to Q. Fulfilling the quality criteria defined by the Organisation for the Prohibition of Chemical Weapons, the novel methods enable the detection of exposure to Q alone or in mixtures with SM. We further report on the relative reactivity of Q compared to SM. Based on experiments making use of partially deuterated Q as the alkylating agent, we rule out a major role for six-membered ring sulfonium ions as relevant reactive species in the alkylation of Cys34. Furthermore, the results of molecular dynamics simulations are indicative that the protein environment around Cys34 allows adduct formation with elongated but not bulky molecules such as Q, and identify important hydrogen bonding interactions and hydrophobic contacts.
Keywords Hydroxyethylthioethyl . Protein adduct . Sulfur mustard . Verification . Vesicant Abbreviations Atr-d3 Atropine, triple deuterated BioPT OPCW biomedical proficiency test Electronic supplementary material The online version of this article (https://doi.org/10.1007/s00216-020-02917-w ) contains supplementary material, which is available to authorized users. * Harald John [email protected] 1
Blum – Scientific Services, Björnsonweg 70d, 22587 Hamburg, Germany
2
Department of Chemistry, Humboldt-Universität zu Berlin, Brook-Taylor-Straße 2, 12489 Berlin, Germany
3
Bundeswehr Institute of Pharm
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