title: Glucuronidation of N-hydroxy metabolites of N-acetylbenzidine.
abstract: Glucuronidation of N-hydroxy arylamines is thought to be a necessary step in their initiation of bladder cancer. This was evaluated for the N-hydroxy metabolites of N-acetylbenzidine (ABZ). N'-Hydroxy-N-acetylbenzidine (N'-HA), N-hydroxy-N-acetylbenzidine (N-HA) and N-hydroxy- N,N'-diacetylbenzidine (N-HDA) were synthesized. Except for N'-HA, these compounds were quite stable. Ascorbic acid and/or acidic pH increased the stability of N'-HA. When each N-hydroxy compound was added to reaction mixtures containing [14C]UDP-glucuronic acid, 3 mM ascorbic acid and human liver microsomes a new product was detected by HPLC. Emulgen 911 was a better detergent than Triton X-100 for expressing microsomal activity, with maximal glucuronidation observed with 0.3% Emulgen 911. At 0.125 mM amine the rate of glucuronidation was N-HDA >> N'-HA = benzidine > ABZ > N-HA. In contrast, at 0.5 mM amine the rate of glucuronidation of N-HA was only exceeded by N-HDA. At pH 5.5 and 37 degrees C the t1/2 for the enzymatically prepared glucuronide conjugates of ABZ, N'-HA and N-HA were 7.5 min and 3.5 and 1.8 h respectively. For N-HDA > 90% of this glucuronide remained after 24 h. At pH 7.4 and 37 degrees C the t1/2 for the glucuronide conjugates of ABZ and N-HA were 2.3 and 2 h respectively, with the amounts remaining after 24 h for N'-HA and N-HDA being 75 and 90% respectively. At pH 6.5 the t1/2 for N'-HA was 14 h. Thus only glucuronides of ABZ and N'-HA exhibit pH-dependent changes in t1/2. Compared with ABZ, glucuronides the N-hydroxy metabolites are more stable at acidic pH. Acidic urine would be more likely to hydrolyze the glucuronide conjugate of ABZ than those of its N-hydroxy metabolites. Because these results are different from that hypothesized for arylmonoamines, a new model was developed to explain the role of N-oxidation, N-glucuronidation and N-acetylation in the carcinogenesis of benzidine, an aryldiamine.
authors.affiliation: VA Medical Center, Division of Geriatric Medicine, St Louis, MO 63125, USA.
authors.forename: S R
authors.initials: SR
authors.lastname: Babu
authors.forename: V M
authors.initials: VM
authors.lastname: Lakshmi
authors.forename: F F
authors.initials: FF
authors.lastname: Hsu
authors.forename: T V
authors.initials: TV
authors.lastname: Zenser
authors.forename: B B
authors.initials: BB
authors.lastname: Davis
mesh_terms: D000107:Acetylation; D001560:Benzidines / Q000737:chemistry / Q000378:metabolism*; D002273:Carcinogens / Q000737:chemistry / Q000378:metabolism*; D005965:Glucuronates / Q000378:metabolism*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D006900:Hydroxylation; D008862:Microsomes, Liver / Q000378:metabolism; D001749:Urinary Bladder Neoplasms / Q000139:chemically induced
chemical_list: D001560:Benzidines; D002273:Carcinogens; D005965:Glucuronates; C034938:N-acetylbenzidine