Original ArticlesStrain Differences in CYP3A-Mediated C-8 Hydroxylation (1,3,7-Trimethyluric Acid Formation) of Caffeine in Wistar and Dark Agouti Rats: Rapid Metabolism of Caffeine in Debrisoquine Poor Metabolizer Model Rats
Section snippets
Chemicals
Chemicals were obtained from the following sources: caffeine from Wako; theobromine, theophylline, and 8-chlorotheophylline from Tokyo Kasei; paraxanthine and 1,3,7-trimethyluric acid from Funakoshi; TAO, 3-methylcholanthrene, phenobarbital, dexamethasone, and pregnenolone 16α-carbonitrile from the Sigma Chemical Co.; anti-rat CYP3A2 serum for inhibition studies from the Daiichi Pure Chemical Co.; and glucose-6-phosphate, glucose-6-phosphate dehydrogenase, and NADPH from the Oriental Yeast Co.
Results and Discussion
Caffeine N-demethylase and C-8 hydroxylase activities were assayed in liver microsomes from Wistar and DA rats of both sexes (7 weeks old). We determined the amounts of theobromine (N-1 demethylation), paraxanthine (N-3 demethylation), theophylline (N-7 demethylation), and 1,3,7-trimethyluric acid (C-8 hydroxylation) by HPLC after incubating caffeine with rat liver microsomes.
The main metabolite of caffeine in rats was 1,3,7-trimethyluric acid (C-8 hydroxylation formation), which amounted to
Acknowledgements
We thank Dr. H. Iwata, Y. Yamamoto, T. Tasaki, M. Ishizuka, H. Hoshi, Y. Shimamoto, M. Akamatsu, I. Chiba, and K. Hirose for many helpful discussions, Professor W. Jones for useful advice on the writing of this manuscript, and A. Takahashi and S. Yoshino for their secretarial assistance in the preparation of this manuscript. This study was supported, in part, by a Grant-in-Aid for scientific research from the Ministry of Education, Sciences, Sports and Culture of Japan.
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Present address: Drug Metabolism and Pharmacokinetics Lab., Chugai Pharmaceutical Co., Ltd., 3-41-8, Takada, Toshima-ku, Tokyo 171, Japan.