  Sulfur mustard (SM), a chemical warfare agent, is a strong alkylating
compound that readily reacts with numerous biomolecules. The goal of the
present work was to define and validate new biomarkers of exposure to SM that
could be easily accessible in urine or plasma. Because investigations using SM
are prohibited by the Organization for the Prohibition of Chemical Weapons, we
worked with 2-chloroethyl ethyl sulfide (CEES), a monofunctional analog of SM.
We developed an ultra-high-pressure liquid chromatography - tandem mass
spectrometry approach (UHPLC-MS/MS) to the conjugate of CEES to glutathione and
two of its metabolites, the cysteine and the N-acetyl-cysteine conjugates. The
N7-guanine adduct of CEES (N7Gua-CEES) was also targeted. After synthesizing
the specific biomarkers, a solid phase extraction protocol and a UHPLC-MS/MS
method with isotopic dilution were optimized. We were able to quantify
N7Gua-CEES in the DNA of HaCaT keratinocytes and of explants of human skin
exposed to CEES. N7Gua-CEES was also detected in the culture medium of these
two models, together with the glutathione and the cysteine conjugates. In
contrast, the N-acetyl-cysteine conjugate was not detected. The method was then
applied to plasma from mice cutaneously exposed to CEES. All four markers could
be detected. Our present results thus validate both the analytical technique
and the biological relevance of new, easily quantifiable biomarkers of exposure
to CEES. Because CEES behaves very similarly to SM, the results are promising
for application to this toxic of interest.
