Oxidative metabolism of the alkaloid rutaecarpine by human cytochrome P450.
Rutaecarpine is the main active alkaloid of the herbal medicine, Evodia rutaecarpa. To identify the major human cytochrome P450 (P450) participating in rutaecarpine oxidative metabolism, human liver microsomes and bacteria-expressed recombinant human P450 were studied. In liver microsomes, rutaecarpine was oxidized to 10-, 11-, 12-, and 3-hydroxyrutaecarpine. Microsomal 10- and 3-hydroxylation activities were strongly inhibited by ketoconazole. The 11- and 12-hydroxylation activities were inhibited by alpha-naphthoflavone, quinidine, and ketoconazole. These results indicated that multiple hepatic P450s including CYP1A2, CYP2D6, and CYP3A4 participate in rutaecarpine hydroxylations. Among recombinant P450s, CYP1A1 had the highest rutaecarpine hydroxylation activity. Decreased metabolite formation at high substrate concentration indicated that there was substrate inhibition of CYP1A1- and CYP1A2-catalyzed hydroxylations. CYP1A1-catalyzed rutaecarpine hydroxylations had V(max) values of 1,388 to approximately 1,893 pmol/min/nmol P450, K(m) values of 4.1 to approximately 9.5 microM, and K(i) values of 45 to approximately 103 microM. These results indicated that more than one molecule of rutaecarpine is accessible to the CYP1A active site. The major metabolite 10-hydroxyrutaecarpine decreased CYP1A1, CYP1A2, and CYP1B1 activities with respective IC(50) values of 2.56 +/- 0.04, 2.57 +/- 0.11, and 0.09 +/- 0.01 microM, suggesting that product inhibition might occur during rutaecarpine hydroxylation. The metabolite profile and kinetic properties of rutaecarpine hydroxylation by human P450s provide important information relevant to the clinical application of rutaecarpine and E. rutaecarpa.
Ueng
Yune-Fang
YF
National Research Institute of Chinese Medicine, 155-1, Li-Nong Street, Sec. 2, Taipei 112, Taiwan, ROC. ueng@nricm.edu.tw
Don
Ming-Jaw
MJ
Jan
Woan-Ching
WC
Wang
Shu-Yun
SY
Ho
Li-Kang
LK
Chen
Chieh-Fu
CF
D000470:Alkaloids / Q000378:metabolism*; D019363:Cytochrome P-450 CYP1A1 / Q000037:antagonists & inhibitors / Q000378:metabolism; D065607:Cytochrome P-450 Enzyme Inhibitors; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004926:Escherichia coli / Q000378:metabolism; D006801:Humans; D006900:Hydroxylation; D066298:In Vitro Techniques; D026121:Indole Alkaloids; D007527:Isoenzymes / Q000037:antagonists & inhibitors / Q000378:metabolism; D007700:Kinetics; D008862:Microsomes, Liver / Q000201:enzymology; D010084:Oxidation-Reduction; D011485:Protein Binding; D011799:Quinazolines; D052999:Quinazolinones
D000470:Alkaloids; D019363:Cytochrome P-450 CYP1A1; D065607:Cytochrome P-450 Enzyme Inhibitors; D003577:Cytochrome P-450 Enzyme System; D004791:Enzyme Inhibitors; D026121:Indole Alkaloids; D007527:Isoenzymes; D011799:Quinazolines; C028632:rutecarpine; D052999:Quinazolinones