Genotoxicity of 1,3-Butadiene and Its Epoxy Intermediates

EPA Grant Number: R832347C144
Subproject: this is subproject number 144 , established and managed by the Center Director under grant R832347
(EPA does not fund or establish subprojects; EPA awards and manages the overall grant for this center).

Center: Health Effects Institute (2005 — 2010)
Center Director: Greenbaum, Daniel S.
Title: Genotoxicity of 1,3-Butadiene and Its Epoxy Intermediates
Investigators: Walker, Vernon E
Institution: University of Vermont , Health Effects Institute
EPA Project Officer: Hunt, Sherri
Project Period: April 1, 2005 through March 31, 2010
RFA: Health Effects Institute (2005) RFA Text |  Recipients Lists
Research Category: Health Effects , Air Quality and Air Toxics , Air

Objective:

1,3-Butadiene (BD) is used extensively in the chemical industry (e.g., for synthetic rubber production) and is also part of motor vehicle exhaust and cigarette smoke. BD is listed by the U.S. Environmental Protection Agency (EPA) as a mobile-source air toxic and was recently reclassified by EPA as a human carcinogen via inhalation exposure. This classification was based on occupational exposures of workers in the butadiene rubber industry who showed higher levels of certain leukemias compared with the general population. In 2002, the EPA cited additional evidence for carcinogenicity from chronic inhalation studies with rodents, but the carcinogenic and mutagenic potency of BD is different in mice and rats, which complicates extrapolating results from rodents to humans for risk assessment purposes. Therefore, toxicologic research in the past decade has focused on how the metabolism of BD differs between mice and rats (and how human metabolism compares) and on assessing the mutagenicity of reactive epoxide metabolites of BD in these two species as well as in human cells in vitro. The major etabolites of interest have been the monoepoxide (BDO), the diepoxide (BDO2), and the epoxydiol (BDO-diol).

Dr. Walker proposed to investigate several unresolved issues surrounding BD metabolites, such as (1) the role of stereoisomerism (differences in the three-dimensional shape of chemical compounds) in the mutagenicity of the metabolites, 2) how much the nonreactive intermediary metabolite BD-diol contributes to the formation of reactive metabolites, and (3) the mutagenic potency of BD in mice and rats exposed to high and low levels of BD. The study would focus on differences in mutagenic potency of BD between rodent species, age groups, and sexes.

Approach:

The study comprised a series of experiments in which male and female F344 rats and B6C3F1 mice, 4 to 5 weeks or 8 to 9 weeks old, were exposed via inhalation to BD or its metabolites BDO2 and BD-diol. Exposures lasted for 6 hours/day, 5 days/week, for 2 to 4 weeks; control groups were exposed to filtered air. Upon completion of the exposures, lymphocytes (T cells) were collected from the spleen to determine mutations in the Hprt gene. (Rare cells with mutations in the Hprt gene can be selected from normal cells by adding 6-thioguanine to the culture medium. It kills normal cells because they incorporate it into their DNA, but does not affect the mutant cells, which are unable to incorporate it.)

First, Hprt mutant frequencies were determined in rodents exposed to 3 or 1250 ppm BD to assess sex and age differences in mutagenic response to BD at low and high concentrations. These data were compared with data obtained from earlier studies by Dr. Walker.

Second, rodents were exposed to 2 or 4 ppm BDOof the mesoform (which does not have optical properties) and killed at different intervals to assess the time course of the formation of Hprt mutant cells and to evaluate differences between sexes and species. These data were compared with data from rats exposed to a mixture of + and − forms of BDO2 (which differ in how they refract light) in earlier studies by Dr. Walker, thereby assessing the role of stereoisomerism of BDO2 in the mutagenicity of BD.

Third, rodents were exposed to BD-diol (6, 18, or 36 ppm) to assess the extent and time course of Hprt mutant frequencies. (Because BD-diol is converted to BDO-diol, exposure to BD-diol allowed that metabolic pathway to be investigated without interference by BDO2, which is also converted to BDO-diol.) Additional data were obtained on levels of BD-diol and other metabolites after exposure to BD (200 ppm) or BD-diol (24 or 36 ppm). The investigators developed a sensitive method to analyze BD-diol levels in plasma specifically for this purpose.

Fourth, cells derived from male rodents exposed to BD (1250 ppm), meso-BDO2 (2 or 4 ppm), or BD-diol (6,18, or 36 ppm) were cloned and propagated for molecular analyses of mutation spectra to assess whether the kinds of mutations (such as point mutations or larger deletions) differ among BD and its metabolites.

Expected Results:

This study has provided important data on the mutagenic potency of BD at low exposure concentrations (3 ppm). In addition, Dr. Walker and colleagues confirmed and extended earlier observations that (1) female rodents are more susceptible than male rodents to BD exposure, (2) mice are more susceptible than rats to inhaled BD, and (3) rodents 4 to 5 weeks old are more sensitive to inhaled BD than animals 8 to 9 weeks old.

The investigators attributed this age-related effect to differences in thymus activity and movement of T cells through the body. Dr. Walker and colleagues showed that in both species the contribution of the metabolites BD-diol and BDO-diol to mutagenicity induced by BD exposure is most prominent at high BD concentrations, whereas the metabolite BDO2 is probably more important at lower concentrations. They found that stereochemistry did not play a role in mutagenicity induced by BDO2; however, the in vivo role of the stereochemistry of the monoepoxide metabolites (BDO and BDO-diol) remains to be established.

Supplemental Keywords:

Health Effects, Air Toxics, VOCs,   indoor air, epidemiology, carcinogens, exposure models, 1,3-Butadiene, inhalation exposure

Relevant Websites:

http://pubs.healtheffects.org/getfile.php?u=509 Exit


Main Center Abstract and Reports:

R832347    Health Effects Institute (2005 — 2010)

Subprojects under this Center: (EPA does not fund or establish subprojects; EPA awards and manages the overall grant for this center).
R832347C135 Mechanisms of Particulate Matter Toxicity in Neonatal and Young Adult Rat Lungs
R832347C136 Uptake and Inflammatory Effects of Nanoparticles in a Human Vascular Endothelial Cell Line
R832347C138 Health Effects of Real-World Exposure to Diesel Exhaust in Persons with Asthma
R832347C140 Extended Follow-Up and Spatial Analysis of the American Cancer Society Study Linking Particulate Air Pollution and Mortality
R832347C141 Air Pollution Effects on Ventricular Repolarization
R832347C143 Measurement and Modeling of Exposure to Selected Air Toxics for Health Effects Studies and Verification by Biomarkers
R832347C144 Genotoxicity of 1,3-Butadiene and Its Epoxy Intermediates
R832347C145 Effects of Concentrated Ambient Particles and Diesel Emissions on Rat Airways
R832347C147 Atmospheric Transformation of Diesel Emissions