DETECTION OF SULFUR MUSTARD-INDUCED DNA MODIFICATIONS

Sulfur mustard is acutely toxic to the skin, eyes, and respiratory tra ct, and is considered carcinogenic to humans by the IARC. Since all of these toxicities are thought to be initiated by DNA alkylation, the l evel of DNA damage should serve as a biomarker for exposure. To develo p methods of detecting this damage, DNA was modified by [C-14]-labeled sulfur mustard and DNA adducts were released by mild acid hydrolysis. Radioactivity co-eluted on HPLC analysis with marker 7-(2-hydroxyethy lthioethyl) guanine and 3-(2-hydroxyethylthioethyl) adenine synthesize d from 2-chloroethyl 2-hydroxy-ethyl sulfide. Unambiguous identificati on of the major adduct, 7-(2-hydroxy-ethylthioethyl) guanine, was prov ided by gas chromatography combined with mass spectrometric detection. The most abundant adduct, 7-(2-hydroxyethyl-thioethyl) guanine, accou nted for 61% of the total alkylation and could be detected as a fluore scent HPLC peak with a detection limit of IO pmol. To demonstrate the applicability of this method to biological samples, DNA was extracted from the white blood cells of human blood exposed to 131 mu M sulfur m ustard in vitro and shown to contain 470 pmol of 7-(2-hydroxyethylthio -ethyl) guanine per mg of DNA.