1,3-Butadiene, a common air pollutant formed in the combustion of organic matter, has been assessed by the U.S. EPA to be a strongly carcinogenic compound. This risk assessment is very uncertain because of the lack of information on the dose of the powerful carcinogenic metabolite diepoxybutane (DEB). This report presents an analytical method for in vivo dose monitoring of a unique marker for DEB. For a large number of alkylating agents in vivo doses are monitored by measurement by gas chromatography/mass spectrometry (GC/MS) of adducts to N-terminal valine in hemoglobin (Hb), using a modified Edman degradation method. This method is applicable to monofunctional epoxides from butadiene. However, in reaction with N-terminal valine, DEB forms an adduct which is ring-closed to a pyrrolidine, N,N-(2,3-dihydroxy-1,4-butadiyl)valine, with a tertiary amino group that prevents detachment of the alkylated valine by the Edman reagent. Therefore a method has been developed based on the analysis by liquid chromatography/electrospray ionization mass spectrometry (LC/ESI-MS) of the N-modified N-terminal peptides enriched after trypsin digestion of globin. In this study Hb samples from mice injected intraperitoneally with (+/-)-DEB were examined qualitatively and quantitatively with regard to the ring-closed adduct. The N-terminal pyrrolidine-heptapeptide was identified in treated mice. The highest adduct levels were obtained in samples from animals given the highest dose of DEB and the adduct levels were below the detection level in control mice.
Copyright 2000 John Wiley & Sons, Ltd.