Formation of 1,2:3,4-diepoxybutane-specific hemoglobin adducts in 1,3-butadiene exposed workers

Abstract

1,3-Butadiene (BD) is an important industrial chemical that is classified as a human carcinogen. BD carcinogenicity has been attributed to its metabolism to several reactive epoxide metabolites and formation of the highly mutagenic 1,2:3,4-diepoxybutane (DEB) has been hypothesized to drive mutagenesis and carcinogenesis at exposures experienced in humans. We report herein the formation of DEB-specific N,N-(2,3-dihydroxy-1,4-butadiyl)valine (pyr-Val) in BD-exposed workers as a biomarker of DEB formation. pyr-Val was determined in BD monomer and polymer plant workers that had been previously analyzed for several other biomarkers of exposure and effect. pyr-Val was detected in 68 of 81 (84%) samples ranging from 0.08 to 0.86 pmol/g globin. Surprisingly, pyr-Val was observed in 19 of 23 administrative control subjects not known to be exposed to BD, suggesting exposure from environmental sources of BD. The mean ± SD amounts of pyr-Val were 0.11 ± 0.07, 0.16 ± 0.12, and 0.29 ± 0.20 pmol/g globin in the controls, monomer, and polymer workers, respectively, clearly demonstrating formation of DEB in humans. The amounts of pyr-Val found in this study suggest that humans are much less efficient in the formation of DEB than mice or rats at similar exposures. Formation of pyr-Val was more than 50-fold lower than has been associated with increased mutagenesis in rodents. The results further suggest that formation of DEB relative to other epoxides is significantly different in the highest exposed polymer workers compared with controls and BD monomer workers. Whether this is due to saturation of metabolic formation or increased GST-mediated detoxification could not be determined.

FIG. 1.

Overview of BD metabolism and formation of N-terminal valine adducts and urine metabolites.

FIG. 2.

Validation curve for detection of pyr-Val in human globin. Human globin from a volunteer that had no detectable amounts of pyr-Val was spiked with various amounts of synthetic pyr-Val standard peptide (1–11) and analyzed as describe in “Materials and Methods.” Shown are the positive controls containing 5–100 fmol/samples representing 0.1–2 fmol pyr-Val/g globin. The insert shows extended positive controls containing up to 350 fmol pyr-Val/sample.

FIG. 3.

Representative ion-chromatogram of pyr-Val from BD-exposed subject. Globin was isolated and analyzed for presence of pyr-Val by immunoaffinity nano-UPLC-MS/MS as described in “Materials and Methods.” The sample shown is from a monomer worker exposed to 3.7 mg/m³ (1.6 ppm) BD TWA and had 0.23 pmol pyr-Val per g globin.

FIG. 4.

Associations between individual pyr-Val hemoglobin adduct concentrations (ln[pmol/g]) and 4-month average BD exposure (ln[mg/m³]).

FIG. 5.

Associations between individual pyr-Val hemoglobin adduct concentrations (ln[pmol/g]) and HB-Val hemoglobin adducts (ln[pmol/g]).

FIG. 6.

Associations between individual pyr-Val hemoglobin adduct concentrations (ln[pmol/g]) and THB-Val hemoglobin adducts (ln[pmol/g]).

FIG. 7.

Associations between individual pyr-Val hemoglobin adduct concentrations (ln[pmol/g]) and urine M1 concentrations (ln[μg/l]).

FIG. 8.

Associations between individual pyr-Val hemoglobin adduct concentrations (ln[pmol/g]) and urine M2 concentrations (ln[μg/l]).

FIG. 9.

Associations between individual pyr-Val hemoglobin adduct concentrations (ln[pmol/g]) and urine M2/(M1 + M2) concentrations (ln[μg/l]).