Abstracts and comments--Fd Chem. Toxic. Vol. 20. No. 3 345

Ethylene oxide (EO) exposure has been associated with chromosome aberrations in animals and man (Federal Register 1978, 43, 3800; Pesticide & Toxic Chemical News 1980, 8 (26), 18) and three cases of leukaemia have been reported in EO-exposed Swedish workers (Cited in F.C.T. 1979, 17, 686). Sister-chromatid exchange was increased in the lym- phocytes of individuals exposed to a maximum of 36 ppm EO in a hospital sterilization facility (Garry et al. Envir. Mutagen. 1979, 1, 375). That EO concen- trations of only 0'5-1 ppm may also affect the blood cells has now been demonstrated.

The study group consisted of four medical equip- ment sterilizers and one laboratory technician, exposed to 5-10 ppm EO for a total of 1 hr/day, and 12 packers, exposed to 0.5-1 ppm for 8hr/day, 5 days/wk. All were female and had worked in the factory for 0.8-8 yr. Eleven female assembly-line workers from another part of the factory with no ex- posure to EO were used as controls. Unscheduled DNA synthesis (UDS) induced by N-acetoxy-2-acetyl- aminofluorene (AC-AAF) in peripheral lymphocytes was significantly reduced in the packers, and chromo- some aberrations (breaks and/or gaps) were signifi- cantly increased in both exposed groups. In controls from the present study and those from earlier studies UDS was directly correlated with age and/or with the incidence of chromosome aberrations, whereas in exposed individuals UDS was negatively correlated with duration of exposure and with the number of chromosome breaks, indicating an inhibition of DNA-repair capacity by EO. When EO levels in the factory were subsequently reduced to a maximum of 0'5 ppm, after 9 months UDS had returned to normal.

Leucocytes from whole blood exposed in vitro to 0.5-100 mM-EO for 1 hr showed a linear increase in UDS up to 5 mM, after which a reduction occurred. When isolated lymphocytes and leucocytes were exposed to the same concentrations, the maximal level of UDS occurred at only l-2mM. Autoradio- graphic studies on lymphocytes confirmed that UDS was inhibited above 2 raM. whether estimated at 24 or 122 hr. Further studies on lymphocytes indicated that the inhibition of UDS was related to cytotoxicity. Even at a stimulatory dose of 1 raM, although there were only 13% more dead cells than in a control cul- ture after 24hr, after a further day 70% of the cells were dead, compared with only 117/o of control cells. When lymphocytes were exposed to 0"I-I'0mM-EO for 18 hr followed by 10/aM AC-AAF for ! hr, only 5-15% were dead after a further hour. However, after a further 18 hr more than 40% had died at all concen- trations, suggesting that even at 0.1 mM-EO the lym- phocytes were sensitized to subsequent exposure to cytotoxic agents such as AC-AAF. UDS was un- altered in surviving cells at 0'l-0"6mM, but was reduced at 0"8 or 1 mM.

It is calculated that daily exposure to 0.5-1 ppm EO, as in the packers, would lead to a maximum tissue concentration of only 0'1 pM-EO. In contrast, in the in vitro studies effects on UDS were detected only in the mM range, suggesting that repeated exposure must have a pronounced cumulative effect. The corre- lation between years of exposure and UDS inhibition supported this hypothesis. Since the packers also tended to show a negative correlation between leuco-

cyte count and induced UDS, it is suggested that EO first caused a leucopenia in which surviving cells had a reduced UDS. This was followed by a leucocytosis to replace the dead cells, which resulted in a mixing of cells with normal UDS and inhibited UDS capacities. The final result would be an overall inhibited UDS.

Reproductive trouble for 2-methoxyethanol

Nagano, K., Nakayama, E., Oobayashi, H., Yamada, T., Adachi, H., Nishizawa, T., Ozawa, H., Nakaichi, M., Okuda, H., Minami, K. & Yamazaki, K. (1981). Embryotoxic effects of ethylene glycol monomethyl ether in mice. Toxicology 20, 335.

Industrial limits for exposure to the glycolethers may be on the move. The present TLV of 25 ppm set for 2-methoxyethanol (ethylene glycol monomethyl ether) was based on a limited pre-war epidemiological study in which exposed workers were found to der velop encephalopathy and anaemia. There has recently been a renewed interest in the reproductive toxicology of the glycol ethers as a class. Teratogenic effects have been reported in rats, mice and rabbits exposed to ethoxyethanol and testes damage occurred in rats and rabbits exposed to atmospheres containing 2-methoxyethanol (Pesticide & Toxic Chemical News 1981, 9 (17), 8). The mouse study cited above suggests that 2-methoxyethanol may also have teratogenic effects.

Groups of 21-24 pregnant ICR mice were given daily doses of 0, 31'25, 62-5, 125, 250, 500 or 1000 mg 2-methoxyethanol/kg body weight by gavage on days 7 to 14 of gestation and then killed on day 18. In- creased foetal deaths occurred at maternal doses of 250 mg/kg and above. All the litters were dead in the 1000 mg/kg group and there was only one live foetus among the 500 mg/kg group. The first indication of a reduction in foetal body weight was observed at 125 mg/kg. A high incidence of gross abnormalities (exencephaly, abnormal fingers and umbilical hernia) was found at 250mg/kg. There was a dose-related increase in the incidence of malformations at doses of 62"5 mg/kg and above. Skeletal variations such as cer- vical ribs, asymmetry of the sternebrae, and bificur- ated or split cervical vertebrae were also increased at all doses in a dose-related manner. There was evi- dence of a retardation in bone ossification in all treat- ment groups. Examination of the maternal animals-- limited to clinical examination, measurement of weight gain and leucocyte sampling--indicated that doses of 250 mg/kg and above were toxic, reducing maternal weight gains. Decreased leucocyte count was limited to the dams of the highest dose group.

Does DNA binding reflect carcinogenic potency?

Pereira, M. A., Lin, L.-H. & Chang, L. W. (1981). Dose-dependency of 2-acetylaminofluorene binding to liver DNA and hemoglobin in mice and rats. Toxic. appl. Pharmac. 60, 472.

Many chemical carcinogens or their active metab- olites have been shown to bind to DNA. To deter- mine whether the extent of such binding is correlated

346 Abstracts and comments--Fd Chem. Toxic. Vol. 20, No. 3

with carcinogenic potency, a study of its dose-depen- dency has been undertaken with 2-acetylaminofluor- erie (2-AAF) for which the relationship between dose and liver turnout induction has been established in mice (Littlefield et al. J. envir. Path. Toxicol. 1979, 3 (3), 17) and rats (Albert et al. Cancer Res. 1972, 32, 2172). The dose-dependency of haemoglobin binding was also studied, because this has been proposed as a systemic dose monitor of DNA binding in target organs.

Rats and mice given [9-14C]2-AAF by garage at doses of 1, 10 or 100 pmol/kg body weight were killed 24 hr later, and DNA-2-AAF adducts in the liver were determined chromatographically after enzymatic degradation of DNA to deoxyribonucleosides. The elution profile from rat liver contained four major peaks, of which one was identified as the DNA adduct of N-(deoxyguanosine-8-yl)-N-AAF and another (the major peak) appeared to be the adduct of deacetylated 2-AAF. The relative contribution of these four peaks to the total acylation of DNA was independent of dose, and together they contained more than 90~ of the radioactivity. Binding in both mice and rats was a linear function of dose, and in rats was about seven- fold greater than in mice. The liver DNA of female mice bound about four times as much '4C as that of male mice, whereas in rats no sex difference was apparent. In the haemoglobin of treated rats, more than 95% of the radioactivity represented 2-AAF

metabolites covalently bound to amino acids. In both species binding was again directly dependent on dose (0"1, 1, l0 or 100/lmol 2-AAF/kg body weight), and in rats was 2'5 times greater than in mice, but there were no apparent sex differences. It was calculated that rats and mice bound 0.07% and 0"03~o, respectively, of an oral dose of 2-AAF to haemoglobin.

The findings indicated that over the dose range studied the formation and DNA binding in the liver of the active metabolites of 2-AAF was linearly related to dose, and that haemoglobin binding could be used to predict DNA in the liver. However, in the long-term rat and mouse studies (Albert et al. Ioc. cir. Littlefield et al. Ioc. cir.) the potency of 2-AAF was 246-fold greater in rat than in mouse liver, a far greater difference than the sevenfold discrepancy in DNA binding. It is suggested that the extent of DNA binding may correlate better with initiation than with carcinogenic potency, and that the latter depends on the subsequent influence of other factors. Moreover, although female mice and male rats are more suscep- tible to 2-AAF-induced hepatocarcinogenesis than male mice and female rats, these differences were re- flected only in the higher binding to. mouse-liver DNA in females than in males. In rats the difference in susceptibility has been associated with testosterone (Toh, Adv. Cancel" Res. 1973, 18, 155) and in this case would appear to depend not on relative DNA binding but on promotion by sex steroids.