Molecular Dosimetry of DNA and Hemoglobin Adducts in Mice and Rats Exposed to Ethylene Oxide

Abstract

Experiments involving ethylene oxide (ETO) have been used to support the concept of using adducts in hemoglobin as a surrogate for DNA adducts in target tissues. The relationship between repeated exposures to ETO and the formation of N-(2-hydroxyethyl)valine (HEtVal) in hemoglobin and 7-(2-hydroxyethyl)guanine (7-HEG) in DNA was investigated in male rats and mice exposed by inhalation toO, 3,10,33, or 100 ppm ETO for 6 hr/day for 4 weeks, or exposed to 100 ppm (mice) or 300 ppm (rats) for 1, 3, 5,10, or 20 days (5 days/week). HEtVal was detennined by Ednan degradation, and 7-HEG was quantitated by HPLC separation and fluorescence detection. HEtVal formation was linear between 3 and 33 ppm ET1O and increased in slope above 33 ppm. The dose-response curves for 7-HEG in rat tissues were linear between 10 and 100 ppm ETO and increased in slope above 100 ppm. In contrast, only exposures to 100 ppm ETO resulted in significant accumulation of 7-HEG in mice. Hemoglobin adducts were lost at a greater rate than predicted by normal erythrocyte life span. The loss of 7-HEG from DNA was both species and tissue dependent, with the adduct half-lives ranging from 2.9 to 5.8 days in rat tissues (brain, kidney, liver, lung, spleen, testis) and 1.0 to 2.3 days in all mouse tissues except kidney (tlm = 6.9 days). The concentrations of HEtVal were similar in concurrently exposed rats and mice, whereas DNA from rats had at least 2-fold greater concentrations of 7-HEG than DNA from mice. Due to differences in formation, persistence, repair, and chemical depurination, the relationships between HEtVal and 7-HEG concentrations varied with length of exposure, interval since exposure, species, and tissue. Thus, it appears unlikely that HEtVal adducts in hemoglobin will provide accurate predictions ofDNA adducts in specific tissues of humans under conditions where actual exposure scenarios are unknown.

The formation and biological significance of N7-guanine adducts

“…The metabolism of ET to EO in mice saturates at ∼1000 ppm ET [130, 131], producing in liver ∼3.5 N 7-HE-Gua/ 10 7 nnt (Table 2, Figure 4) [33, 132, 133]. In contrast, a linear dose-response for N 7-HE-Gua has been found in experiments with mice or rats exposed to EO over a wide range of exposures (Figure 4)[133, 134]. The molecular dose of N 7-HE-Gua can be orders of magnitude greater for exposures to EO than can ever be achieved by ET.…”

Rapid and sensitive on‐line liquid chromatographic/tandem mass spectrometric determination of an ethylene oxide‐DNA adduct, N7‐(2‐hydroxyethyl)guanine, in urine of nonsmokers

“…If not detoxified by enzyme glutathione transferase or epoxide hydrolase, EtO as a direct alkylating agent can attack nucleophilic sites of biological molecules in vivo , such as proteins and DNA bases, to form protein and DNA adducts 6–9. The formation of adducted DNA bases is considered as one of early events in chemical carcinogenesis and DNA adducts are considered as risk‐associated biomarkers 10, 11.…”

“…The formation of adducted DNA bases is considered as one of early events in chemical carcinogenesis and DNA adducts are considered as risk‐associated biomarkers 10, 11. The abundance of EtO‐DNA adducts in order is N7‐(2‐hydroxyethyl)guanine (N7‐HEG), N3‐(2‐hydroxyethyl)adenine, and N 6 ‐(2‐hydroxyethyl)deoxyadenosine,7, 12 and, with the limitations of analytical methods, the most studied adduct is N7‐HEG. Several methods have been developed to analyze N7‐HEG such as a 32 P‐postlabelling method,13 gas chromatography/mass spectrometry (GC/MS) based methods,9, 14–16 liquid chromatography (LC)/fluorescence,17 and LC/MS‐based methods 18–20.…”

Understanding the importance of low‐molecular weight (ethylene oxide‐ and propylene oxide‐induced) DNA adducts and mutations in risk assessment: Insights from 15 years of research and collaborative discussions

“…Inhalation studies with E or P in mice and rats have established adduct dose–response curves supportive of the saturation of CYP450‐mediated metabolic activation to their respective reactive oxides, which then form the DNA adducts (Bolt et al, , Bolt and Filser, , Wu et al, , Pottenger et al, , Rusyn et al, ). For example, the metabolism of E to EO in mice saturates at ~1000 ppm E (Bolt et al, , Bolt and Filser, , Wu et al, , Rusyn et al, ), producing in liver ~3.5 N 7‐HEG/ 10 7 normal nucleotides (Filser et al, , Walker et al, ). In contrast, direct exposure to the oxide metabolites produces a linear dose–response for N 7‐HEG/ N 7‐HPG (Filser and Bolt, , Filser et al, , Rios‐Blanco et al, , Walker et al, , ).…”