Benzidine glucuronidation in dog liver.
Dog is an animal model for assessing aromatic amine-induced bladder cancer, and hepatic N-glucuronidation is proposed as an important pathway leading to initiation of carcinogenesis. Therefore, benzidine N-glucuronidation was evaluated with dog liver microsomes and slices. Microsomal benzidine UDP-glucuronosyltransferase activity was increased with a variety of detergents. For kinetic analysis, native microsomal preparations were separated into treated (detergent treated, not centrifuged) or soluble (detergent treated, centrifuged) fractions. The detergents Triton X-100, Lubrol PX, Emulgen 911 and CHAPS increased the specific activity of treated fractions relative to the native microsomes 3- to 6-fold. The specific activities of the soluble fractions were highest with Emulgen 911 and CHAPS at a detergent-to-protein ratio of 1. Subsequent studies used Emulgen 911 or CHAPS. Similar results were observed with either preparation. For treated preparations, the Km and Vmax values were 0.142 +/- 0.006 mM and 0.65 +/- 0.1 nmol/mg protein/min respectively. A variety of chemicals were tested for their effect on benzidine N-glucuronide formation. At 0.1 mM, the only effective inhibitors (< 50% of control) were 2-aminofluorene, estriol, 17-epiestriol, 2-OH-estrone, and 4-OH-estrone. With Emulgen-treated microsomes, the Ki values for 2-aminofluorene, 4-aminobiphenyl and estriol were 0.114 +/- 0.014, 0.347 +/- 0.032 and 0.047 +/- 0.003 mM respectively. 2-Aminofluorene and estriol were non-competitive inhibitors, while 4-aminobiphenyl was a competitive inhibitor. Slices incubated with these chemicals exhibited an inhibition profile similar to that observed with microsomes. Thus, N-glucuronidation of benzidine may be an important metabolic pathway in dog. Inhibition of benzidine N-glucuronidation by estriol and catechol estrones may be important in vivo events in aromatic amine-induced carcinogenesis.
VA Medical Center, St. Louis, MO.
S R
SR
Babu
V J
VJ
Wongsurawat
T V
TV
Zenser
B B
BB
Davis
D000611:Aminobiphenyl Compounds / Q000494:pharmacology; D000818:Animals; D001560:Benzidines / Q000378:metabolism*; D003902:Detergents / Q000494:pharmacology; D004285:Dogs; D004964:Estriol / Q000494:pharmacology; D005449:Fluorenes / Q000494:pharmacology; D005965:Glucuronates / Q000378:metabolism*; D066298:In Vitro Techniques; D007700:Kinetics; D008099:Liver / Q000378:metabolism*; D008862:Microsomes, Liver / Q000201:enzymology*; D017137:Nonoxynol / Q000494:pharmacology; D013739:Testosterone / Q000494:pharmacology
D000611:Aminobiphenyl Compounds; D001560:Benzidines; D003902:Detergents; D005449:Fluorenes; D005965:Glucuronates; C006757:4-biphenylamine; D017137:Nonoxynol; C029876:benzidine; C012177:2-aminofluorene; D013739:Testosterone; C075855:benzidine N-glucuronide; D004964:Estriol