Investigation of the covalent binding of styrene-7,8-oxide to DNA in rat and mouse

Abstract

Styrene-7,8-oxide (SO), the main intermediate metabolite of styrene, induces hyperkeratosis and tumors in the forestomach of rats and mice upon chronic administration by gavage. The aim of this study was to investigate whether DNA binding could be responsible for the carcinogenic effect observed. [7?H]SO was administered by oral gavage in corn oil to male CD rats at two dose levels (1.65 or 240 mg/kg). After 4 or 24 h, forestomach, glandular stomach and liver were excised, DNA was isolated and its radioactivity determined. At the 4 h time point, the DNA radioactivity was below the limit of detection in the forestomach and the liver. Expressed in the units of the covalent binding index, CBI = (/unol adduct/mol DNA nucleotide)/(mmol chemical administered/kg body wt), the DNA-binding potency was below 2.6 and 2.0 respectively. In the glandular stomach at 4 h, and in most 24 h samples, DNA was slightly radiolabeled. Enzymatic degradation of the DNA and separation by HPLC of the normal nucleotides showed that the DNA radioactivity represented biosynthetic incorporation of radiolabel into newly synthesized DNA. The limit of detection of DNA adducts in the glandular stomach was 1.0. In a second experiment, [T-TIISO was administered by i.p. injection to male B6C3F1 mice. Liver DNA was analyzed after 2 h. No radioactivity was detectable at a limit of detection of CBI < 0.6. In agreement with the relatively long half-life of SO in animals, the chemical reactivity of SO appears to be too low to result in a detectable production of DNA adducts in an in vivo situation. Upon comparison with the DNA-binding of other carcinogens, a purely genotoxic mechanism of tumorigenic action of SO is unlikely. The observed tumorigenic potency in the forestomach could be the result of strong tumor promotion by high-dose cytotoxkity followed by regenerative hyperplasia.

Covalent binding of styrene to DNA in rat and mouse

“…A similar discrepancy was noted in the experiments with SO: a CBI value of 2.2 was deduced from Byfalt Nordqvist et al (8), while our values were below a limit of detection of CBI <0.6 under identical experimental conditions (12). Byfalt Nordqvist et al apparently did not purify the DNA to constant specific activity.…”

“…DNA binding by SO was not detectable in vivo but the limit of detection was not as good as in the present experiments with S: a CBI <0.6 was found for mouse liver after i.p. injection (12). The CBI = 0.1 measured here for S is therefore not in contradiction to the negative findings with SO.…”

“…HPLC analysis of the nucleotides revealed, in all eight samples, a small but significant amount of radioactivity in fractions eluting after the natural nucleotides ( Figure IB). The elution time was similar to the early-eluting S-nucleotide adduct(s) generated from DNA plus SO in vitro (12). Expressed in the units of the CBI, the DNAbinding potency was at the extremely low level of 0.05-0.09 and 0.07-0.18 for males and females respectively.…”

“…HPLC profiles with rat liver DNA showed that all radioactivity eluted with normal nucleotides (Figure 1A). At elution times known for S-nucleotide adducts (12), no radioactivity was measured at a detection limit of 1.1 c.p.m. Conversion of this detection limit to the units of the CBI showed a maximum possible DN A-binding potency of CBI < 0.1 for both males (Table I) and females (Table II).…”

Adduct formation on DNA and haemoglobin in mice intraperitoneally administered with styrene

“…Moreover, O 6 -and A^-styrene specific guanine DNA adducts have been detected in humans occupationally exposed to styrene (4)(5)(6)(7). Adduct formation with haemoglobin, albumin, and DNA has been demonstrated in animals treated with styrene and styrene oxide (9)(10)(11)(12)(13)(14). Styrene oxide covalently bound to haemoglobin is usually detected by analysing the adducts on A'-terminal valine and cysteine (15,16).…”