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Forensic evidence of sulfur mustard exposure in real cases of human poisoning by detection of diverse albumin-derived protein adducts

  • Toxicokinetics and Metabolism
  • Published:
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Abstract

We present the forensic analyses of plasma samples of human victims exposed to sulfur mustard (SM) in a crisis region in the Middle East in 2015. A few hours after exposure, poisoned persons showed typical signs and symptoms of percutaneous SM exposure including erythema and later on blisters and hardly healing skin wounds. Blood samples were collected 15 days after poisoning to be analyzed for the presence of long-lived protein-adduct biomarkers to verify SM poisoning. We applied a novel bioanalytical toolbox targeting four human serum albumin-derived biomarkers that were made accessible after plasma proteolysis. These adducts contained the SM-specific hydroxyethylthioethyl moiety either bound to the thiol group of a cysteine residue (C34*) or to the side-chain carboxylic group of a glutamic acid residue (E230*). Peptide biomarkers were produced from plasma of the victims using proteinase K (C34*PF), pronase (C34*P) and pepsin (AE230*VSKL and LQQC34*PFEDHVKL) for enzymatic protein cleavage. Separation and detection were carried out by selective micro-liquid chromatography–electrospray ionization high-resolution tandem mass spectrometry (µLC–ESI MS/HR MS). In addition to this site-specific adduct detection, a general approach after alkaline hydrolysis of the plasma protein fraction was applied. Liberated thiodiglycol (TDG) was derivatized with heptafluorobutyric anhydride and detected by gas chromatography–electron ionization mass spectrometry (GC–EI MS). The different bioanalytical methods yielded congruent results confirming SM poisoning for all patients who showed clinical signs and symptoms. This is the first time that real cases of SM poisoning were confirmed and presented by such a broad compilation of protein-derived biomarkers.

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Abbreviations

ACN:

Acetonitrile

CWA:

Chemical warfare agents

CWC:

Chemical weapons convention

d3-Atr:

Triple deuterated atropine

FA:

Formic acid

GC–EI MS:

Gas chromatography–electron ionization mass spectrometry

HSA:

Human serum albumin

HETE:

Hydroxyethylthioethyl

HFBA:

Heptafluorobutyric anhydride

iPrOH:

Iso-propanol

TDP:

Thiodipropanol

µLC–ESI MS/HRMS:

Micro-liquid chromatography–electrospray ionization high-resolution tandem mass spectrometry

SIM:

Selected ion monitoring

SM:

Sulfur mustard

SPE:

Solid-phase extraction

TDG:

Thiodiglycol

TOF:

Time-of-flight

UF:

Ultrafiltration

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Acknowledgements

We thank Erwin Wagner (InstPharmToxBw) for his excellent technical assistance.

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Authors and Affiliations

  1. Bundeswehr Institute of Pharmacology and Toxicology (InstPharmToxBw), Neuherbergstr. 11, 80937, Munich, Germany

    Harald John, Marianne Koller, Franz Worek, Horst Thiermann & Markus Siegert

  2. Department of Chemistry, Humboldt-Universität zu Berlin, Brook-Taylor-Strasse 2, 12489, Berlin, Germany

    Markus Siegert

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  1. Harald John
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  2. Marianne Koller
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  3. Franz Worek
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  4. Horst Thiermann
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  5. Markus Siegert
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Correspondence to Harald John.

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John, H., Koller, M., Worek, F. et al. Forensic evidence of sulfur mustard exposure in real cases of human poisoning by detection of diverse albumin-derived protein adducts. Arch Toxicol 93, 1881–1891 (2019). https://doi.org/10.1007/s00204-019-02461-2

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