Determination of N7- and O6-methylguanine in Rat Liver DNA after Oral Exposure to Hydrazine by Use of Immunochemical and Electrochemical Detection Methods

Determination of N7- and O6-methylguanine in Rat Liver DNA after Oral Exposure to Hydrazine by Use of Immunochemical and Electrochemical Detection Methods, VAN DELFT, J. H. M., STEENWINKEL, M-J. S. T., DE GROOT, A. T. L., VAN ZEELAND, A. A., EBERLE-ADAMKIEWICZ, G., RAJEWSKY, M. F., THOMALE, J., AND BAAN, R. A. (1997). Fundam. Appl. Toxicol. 35, 131-137. Hydrazine belongs to a group of compounds for which there is evidence that the in vivo genotoxic effects become manifest only upon exposure to toxic dose levels. The present study was per- formed to investigate whether this phenomenon is also reflected in the pattern of DNA methylation. The induction of N7- and O6- methylguanine (MeGua) was studied in liver DNA of rats, 16 hr after treatment with various doses of hydrazine. After DNA isolation, the presence of N7-MeGua in DNA was assessed with an immunochemical method and with a physicochemica l technique (HPLC with electrochemical detection). Application of these two methods resulted in almost identical patterns of dose-dependen t induction of guanine N7-methylation in rats dosed orally with 0.1 to 10 mg hydrazine per kilogram of body weight, increasing from 1.1-1.3 to 39-45 N7-MeGua per 106 nucleotides. At lower dosages a constant adduct level was observed, equivalent to that in un- treated rats (background level). The O6-MeGua level was analyzed by a combination of HPLC separation and competitive radioimmu- noassay. A background level was observed for untreated rats and no increase was visible up to the 0.2 mg/kg dose group. After hydrazine doses from 0.2 to 10 mg/kg, O6-MeGua increased from 0.29 to 134 per 10' nucleotides. These data show that even at dosages below the maximum tolerated dose (0.6 mg/kg/day), for which carcinogenic effects have not been described, DNA adducts are formed. A comparison is made of the data obtained in this study with models that describe the mechanism of hydrazine-in- duced DNA methylation. © lwsocien ofTo«koiog>.