Analysis for Plasma Protein Biomarkers Following an Accidental Human Exposure to Sulfur Mustard

Abstract

Following an accidental human exposure to a vesicating agent, plasma samples were analyzed for specific biomarkers of sulfur mustard. One individual suffered chemical burns over 6.5% of the body surface area and required hospitalization; the second individual developed a single, small blister. Plasma specimens from both individuals were examined using two different assays. The first assay targeted sulfur mustard adducts to cysteine-34 of albumin using affinity chromatography, enzyme digestion, and analysis of the alkylated peptide fragment using liquid chromatography-tandem mass spectrometry. The second assay targeted alkylation sites of glutamic and aspartic acids of plasma proteins. Following precipitation of plasma proteins, the sulfur mustard adducts were cleaved from the protein using base, derivatized, and analyzed using gas chromatography-mass spectrometry. Samples obtained over a 42-day period from the individual requiring hospitalization produced positive results for sulfur mustard adducts using both assays. Observed levels of the sulfur mustard biomarker decreased by approximately 75% between days 2 and 42 for both assays. Samples obtained over a six-day period from the individual with a single, small blister produced positive results for the albumin adduct assay. Observed levels were much lower than levels from the hospitalized patient. Blood samples from suspected human exposures to sulfur mustard have only rarely been made available for analysis by sensitive and specific laboratory assays. The data presented here add significantly to the small database of information that currently exists on human biomarkers of sulfur mustard exposure, linking a well-documented exposure event with levels of plasma protein adducts.

Mass spectrometry of peptides and proteins from human blood

“…Oxidation levels of blood proteins may reveal physiological conditions (Aldini et al, 2006(Aldini et al, , 2008Morgan et al, 2007;Bruschi et al, 2008) and might be detected by biotin hydrazide to selectively derivatize carbonyl groups in oxidized proteins followed by avidin affinity chromatography (Mirzaei et al, 2008). Protein adducts may form in response to physiological processes (Szapacs et al, 2006(Szapacs et al, , 2008 or may be indicative of the presence of specific mutagens, toxins, drug binding, or chemical weapons (Yike et al, 2006;Carol-Visser et al, 2008;Lohmann, Hayen, & Karst, 2008;Myers & Ali, 2008;Scholl & Groopman, 2008;Smith et al, 2008;Noort et al, 2008a,b;Will, Wolters, & Sheldrick, 2008;Zhang et al, 2008a;Moreno-Gordaliza et al, 2009;Preston et al, 2009;Winther et al, 2009). Protein serotonylation may have a physiological function since transglutaminase activity was observed as a cystamine-inhibitable incorporation of the free amine pentylamine-biotin into arterial proteins integral to contractility and the cytoskeleton (Schaub et al, 2009).…”

Sulfur Mustard: Persisting Threat

“…Jedním z nejvíce studovaných analytů jsou adukty s N-terminálním valinem (hydroxyethylthioethyl-valin) nebo histidinem (hydroxyethylthioethylhistidin). Tyto analyty lze stanovit metodou GC-MS až 120 dní po expozici, tedy po dobu životnosti erytrocytu (31,32). V krevní plazmě lze též stanovit alkylační adukt s albuminem.…”

“…Místem alkylace je aminokyselina cystein na 34. pozici. Vzniklý adukt S-2-hydroxyethylthioethyl-albumin je pro účely analýzy následně štěpen na tripeptid S-2-hydroxyethylthioethyl-Cys-Pro-Phe, který lze následně stanovit metodou HPLC-MS. Adukty s albuminem jsou stanovitelné v plazmě až 50 dní po expozici sirnému yperitu (30,32).…”

An improved method for retrospective quantification of sulfur mustard exposure by detection of its albumin adduct using ultra-high pressure liquid chromatography-tandem mass spectrometry

“…[S-HETE]-ALB adduct has become another important biomarker for long-term retrospective detection of exposure in spite of the shorter half-life of 20-25 d [9,13]. Many plasma samples collected from HD exposure accidents or conflicts had been analyzed for the [S-HETE]-ALB adduct to verify HD exposure [13][14][15].…”

“…Many plasma samples collected from HD exposure accidents or conflicts had been analyzed for the [S-HETE]-ALB adduct to verify HD exposure [13][14][15]. The established assay methods for the stable [S-HETE]-ALB adduct have been based on the fact that enzymatic digestion of the adduct leads to the formation of [S-HETE]-Cys-Pro-Phe ([S-HETE]-CPF) tripeptide adduct, which can be analyzed by liquid chromatographytandem mass spectrometry (LC-MS/MS) [14].…”