Covalent binding of a reactive metabolite derived from propranolol and its active metabolite 4-hydroxypropranolol to hepatic microsomal proteins of the rat.
Repeated administration of propranolol (PL) to rats causes the inhibition of cytochrome P450-2D (P450-2D) enzyme. We recently found that 4-hydroxypropranolol (4-OH-PL) was biotransformed to 1,4-naphthoquinone (1,4-NQ) by superoxide (SO) anions in medium containing rat liver microsomes and NADPH and proposed that the binding of the quinone to P450-2D apoproteins might be one of mechanisms for the enzyme inhibition [Narimatsu et al. (1995) Chem. Res. Toxicol. 8, 721-728]. In this study, we have searched for possible sources of SO for the conversion of 4-OH-PL to 1,4-NQ in rat liver microsomes and determined the radioactivity covalently bound to microsomal proteins after incubation of radioactive PL and 4-OH-PL with rat liver microsomes. Elimination of 4-OH-PL from a mixture containing microsomes and NADPH was suppressed by carbon monoxide. Antibodies raised to P450-2B1 and -3A2 partially, and antibody against NADPH-cytochrome P450 reductase (fp2) markedly suppressed the reaction. 1,4-NQ was formed concomitantly with 4-OH-PL elimination by a reconstituted preparation of fp2. Binding studies using naphthalene ring (NR)- and side chain (SC)-radiolabeled PL and 4-OH-PL showed that radioactivity covalently bound to microsomal proteins was much higher from 4-OH-PL than from PL for the NR-labeled compounds, but higher from PL than from 4-OH-PL for the SC-labeled compounds. These results suggest that the 4-OH-PL formed from PL by P450-2D enzyme is converted to 1,4-NQ with loss of the side chain, and the 1,4-NQ accounts for most of the radioactivity covalently bound to microsomal proteins, including the P450-2D enzymes. The SO for conversion of 4-OH-PL to 1,4-NQ is supplied mainly by fp2 with some contribution by P450 enzymes.
Narimatsu
S
S
Laboratory of Biopharmaceutics, Faculty of Pharmaceutical Sciences, Chiba University, Japan.
Arai
T
T
Watanabe
T
T
Masubuchi
Y
Y
Horie
T
T
Suzuki
T
T
Ishikawa
T
T
Tsutsui
M
M
Kumagai
Y
Y
Cho
A K
AK
D000319:Adrenergic beta-Antagonists / Q000378:metabolism*; D000818:Animals; D001711:Biotransformation; D002248:Carbon Monoxide / Q000494:pharmacology; D003577:Cytochrome P-450 Enzyme System / Q000096:biosynthesis / Q000276:immunology; D004790:Enzyme Induction / Q000187:drug effects; D005260:Female; D066298:In Vitro Techniques; D008297:Male; D008862:Microsomes, Liver / Q000378:metabolism*; D009281:Naphthalenes / Q000737:chemistry; D010634:Phenobarbital / Q000494:pharmacology; D011433:Propranolol / Q000031:analogs & derivatives* / Q000378:metabolism; D011485:Protein Binding; D011817:Rabbits; D051381:Rats; D013481:Superoxides / Q000378:metabolism
D000319:Adrenergic beta-Antagonists; D009281:Naphthalenes; D013481:Superoxides; C001422:4-hydroxypropranolol; D002248:Carbon Monoxide; D003577:Cytochrome P-450 Enzyme System; D011433:Propranolol; D010634:Phenobarbital