Determination of the binding of trenbolone and zeranol to rat-liver DNA in vivo as compared to 17 beta-oestradiol and testosterone.

The in vivo interactions of trenbolone and zeranol with rat-liver DNA were assayed by determining their covalent-binding indices (CBI), which correlate well with the hepatocarcinogenic potencies of chemicals. As controls, 17 beta-oestradiol and testosterone were used. Sixteen hours after intraperitoneal injection of the [3H]-labelled compounds (specific radioactivity approximately 50 Ci/mmol) the animals were sacrificed and the DNA was isolated from the livers and deproteinized by several Sevag extractions and proteinase digestion. Contaminating RNA was eliminated by pancreatic RNAase digestion. Finally, the DNA was further purified by chromatography on hydroxylapatite and by centrifugation in a CsCl gradient. Decreased CBI values after each step of purification showed that extensive purification was essential. The results obtained for the CBI were 11.4 for 17 beta-oestradiol, 4.8 for testosterone, 5.6 for trenbolone and 1.65 for zeranol, when, respectively, 15, 19, 17 and 60 micrograms/kg of anabolics were administered 16 h before the animals were killed. The CBI values for the anabolics decreased when the amount administered increased. When determined as a function of time, the CBI increased up to 24 h and decreased subsequently. After 96 h the CBI for zeranol was only 0.37 and that for trenbolone, 1.11.