Journal Article

NADPH-dependent oxidation of benzidine by rat liver

Vijaya M. Lakshmi, Nathan T. Zenser, F.F. Hsu, Michael B. Mattammal, Terry V. Zenser and Bernard B. Davis

in Carcinogenesis

Volume 17, issue 9, pages 1941-1947
Published in print September 1996 | ISSN: 0143-3334
Published online September 1996 | e-ISSN: 1460-2180 | DOI:
NADPH-dependent oxidation of benzidine by rat liver

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This study used liver microsomes from control and β-naphthoflavone-treated rats to evaluate NADPH-dependent oxidation of benzidine. With microsomes from β-naphtho-flavone-treated rats, the rates of formation of aqueous soluble metabolite (HPLC analysis) and protein and DNA binding were 835 ±81, 14.5 ± 1.8 and 0.71 ± 0.14 pmol/ mg/min respectively. β-Naphthoflavone treatment elicited 12.3-, 1.8- and 14.2-fold increases in benzidine metabolism compared with controls as judged by HPLC and protein and DNA binding respectively. For microsomes from treated animals, Km and Vmax values were 47 ± 6 μM and 1.13 ± 0.16 nmol/mg protein/min respectively. All of the metabolic parameters were inhibited to varying degrees by gluta-thione (1 or 10 mM), N-acetylmethionine (10 mM) and ascorbic acid (10 mM). Following glutathione addition, at least two new metabolite peaks were observed, representing ∼6% of the total radioactivity recovered by HPLC. Neither metabolite was 3-(glutathion-S-yl)benzidine. Cytochrome P450 inhibitors (10 μM) specific for different members of cytochrome gene families 1–3 indicated that benzidine was metabolized by cytochrome P450 1A1/1A2. Ellipticine and α-naphthoflavone, specific 1A1/1A2 inhibitors, elicited 50% inhibition at ∼0.2 and 0.5 μM respectively. Electron impact and negative ion chemical ionization mass spectro-metry identified the aqueous soluble metabolite as 3-hydroxybenzidine. The lability of 3-hydroxybenzidine observed at pH > 7.0 was prevented by ascorbic acid. Thus, cytochrome P450 1A1/1A2 NADPH-dependent metabolism of benzidine to 3-hydroxybenzidine was demonstrated.

Journal Article.  0 words. 

Subjects: Clinical Cytogenetics and Molecular Genetics

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