N-ACETYLBENZIDINE-N'-GLUCURONIDATION BY HUMAN, DOG AND RAT-LIVER

While N-glucuronidation is an important pathway for metabolism of arom atic amines, it has not been demonstrated for N-acetylbenzidine. A glu curonide of N-acetylbenzidine was synthesized and identified by mass s pectrometry as N-acetylbenzidine-N'-glucuronide. This N'-glucuronide i s acid labile with a t(1/2) of 4 min at pH 5.3. A similar acid labilit y was also observed with benzidine-N-glucuronide. The formation of N-a cetylbenzidine-N'-glucuronide was assessed with liver slices and micro somes prepared from human, dog and rat. When 0.014 mM [H-3]N-acetylben zidine was incubated with human liver slices a significant amount of N -acetylbenzidine-N'-glucuronide was produced (8-26% of the total radio activity recovered). With higher concentrations of [H-3]N-acetylbenzid ine (1 mM) rat slices also produced N-acetylbenzidine-N'-glucuronide. However, N'-glucuronide formation was not detected with dog liver slic es incubated with either 0.014 or 1 mM [H-3]N-acetylbenzidine. N-acety lbenzidine-N'-glucuronide formation was observed with microsomes prepa red from human, dog and rat. To assess maximum activity four detergent s were used at two concentrations. With or without detergent activatio n the relative amount of glucuronidation was human > > dog > rat. The rate of benzidine N-glucuronide formation was 4.3- and 1.6-fold greate r than N-acetylbenzidine-N'-glucuronide in dog and rat respectively, w hile in human both rates were similar (1.1-fold). With or without dete rgent activation the relative amount of benzidine-N-glucuronide format ion was human > dog > > rat. N-Glucuronidation of [H-3]N,N'-diacetylbe nzidine was not observed. Thus N-acetylbenzidine-N'-glucuronide format ion appears to be an important pathway for metabolism of N-acetylbenzi dine, especially in humans. Due to their acid lability, formation of t he N-glucuronides of N-acetylbenzidine and benzidine provides a mechan ism for hepatic detoxification and accumulation of these carcinogens i n the bladder. A new model is described illustrating the effect of N-g lucuronidation and the influence of N-acetylation on arylmono- and ary ldiamine-induced bladder carcinogenesis.