Studies on the macromolecular binding of benzidine.

3H Benzidine, injected i.p. into male Wistar rats, was shown to bind to liver macromolecules including DNA. Binding to DNA was persistent, significant amounts of radioactivity being detected four weeks after injection. Enzymic digestion of the DNA resulted in the separation of five peaks of radioactivity following Sephadex LH20 chromatography. The final and major peak appears from preliminary data to result from reaction of benzidine, via a nitrenium ion, with deoxyguanosine in DNA. An identical chromatographic profile was obtained when DNA, which had been reacted in vitro with N-benzoyloxybenzidine, was treated in a similar manner. Acid hydrolysis of DNA, obtained from either in vivo or in vitro reaction with benzidine and N-benzoyloxybenzidine, respectively, resulted in the separation of only one major peak on both LH20 chromotography and reversed phase HPLC. In each case this peak eluted very close to the major nucleoside adduct peak following enzymic digestion and was therefore presumed to be the corresponding base adduct.