Development and Validation of Butadiene Trihydroxybutyl-Valine Globin Peptide Standards for Quantitation of Butadiene Hemoglobin Adducts

Exposure to Butadiene (BD) has been associated with multi-site carcinogenesis in rodents and with an increased incidence of leukemia and lymphoma in humans included in a blinded study of occupationally exposed workers. Metabolites of BD have been shown to induce alkylated adducts of the N-terminal Valine (Val) of hemoglobin (Hb). These adducts can be measured and used as biomarkers of the internal dose resulting from exposure to BD. We have developed methods for the synthesis and quantification of N-terminal peptide standards containing the BDspecific adduct N-(2,3,4-trihydroxybutyl)-valine (THB-Val). The synthesis entails direct alkylation of standard peptides, and the resulting adducts are purified by HPLC, characterized by LC/MS/MS peptide fragmentation, and quantified by UPLC/MS/MS based amino acid quantitation. Analyte and internal standards of THB-Val peptides were prepared for use in the quantitative assessment of these BD-specific adducts from globin samples subjected to immunoaffinity enrichment (IA).