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Metabolism of alpha-naphthoflavone by rat liver microsomes
Abstract:
alpha-Naphthoflavone (ANF) or 7,8-benzoflavone, a synthetic flavonoid, has been widely used in biochemical and biological studies concerning the mechanisms of action of chemical carcinogens. It has been shown previously that ANF inhibits benzo(a)pyrene metabolism by beta-naphthoflavone (BNF)-induced rat liver microsomes but has no inhibitory effects on benzo(a)pyrene metabolism in phenobarbital (PB)-induced rat liver microsomes. This study shows that ANF gives type 1 binding spectra with and is metabolized by both BNF- and PB-induced rat liver microsomes. Specific metabolites identified by ultraviolet and mass spectra and in some cases by cochromatography with authentic standards were: 6-hydroxy-alpha-naphthoflavone, 9-hydroxy-alpha-naphthoflavone, alpha-naphthoflavone-5,6-oxide, and 5,6-dihydro-5,6-dihydroxy-alpha-naphthoflavone. Metabolism at the 5,6 bond of ANF accounted for 73 and 86% of the total organic soluble metabolites produced by PB- and BNF-induced microsomes, respectively. This result is in concert with previous observations on the role of 6 substitution and the loss of inhibitory activity of ANF in BNF-induced rat liver microsomes. Metabolism of ANF is mediated by the cytochrome P-450 mixed-function oxidases, because it is dependent on NADPH and inhibited by carbon monoxide and other cytochrome P-450 inhibitors. BNF-induced microsomes metabolize ANF to 5,6-dihydro-5,6-dihydroxy-alpha-naphthoflavone to a much greater extent than do PB-induced microsomes.
Insights
Alpha-Naphthoflavone (ANF) is metabolized by rat liver microsomes, forming specific hydroxylated and epoxide metabolites. This metabolism, mediated by cytochrome P-450 enzymes, explains ANF's varying inhibitory effects on carcinogen metabolism.
Area of Science:
- Biochemistry
- Pharmacology
- Toxicology
Background:
- Alpha-Naphthoflavone (ANF) is a synthetic flavonoid used in studies of chemical carcinogen mechanisms.
- Previous studies indicated ANF inhibits benzo(a)pyrene metabolism in beta-naphthoflavone (BNF)-induced microsomes but not phenobarbital (PB)-induced microsomes.
Purpose of the Study:
- To investigate the metabolism of ANF by both BNF- and PB-induced rat liver microsomes.
- To identify ANF metabolites and elucidate the role of metabolism in its inhibitory activity.
Main Methods:
- Incubation of ANF with BNF- and PB-induced rat liver microsomes.
- Identification of metabolites using ultraviolet and mass spectrometry.
- Assessment of metabolic pathways and enzyme mediation (cytochrome P-450).
Main Results:
- ANF undergoes metabolism by both BNF- and PB-induced microsomes, exhibiting type 1 binding spectra.
- Key metabolites identified include 6-hydroxy-ANF, 9-hydroxy-ANF, ANF-5,6-oxide, and 5,6-dihydro-5,6-dihydroxy-ANF.
- Metabolism at the 5,6 bond is predominant (73-86%), with BNF-induced microsomes producing more 5,6-dihydro-5,6-dihydroxy-ANF than PB-induced microsomes.
- ANF metabolism is dependent on NADPH and inhibited by carbon monoxide, indicating cytochrome P-450 mediation.
Conclusions:
- ANF metabolism by cytochrome P-450 enzymes is a key factor influencing its differential inhibitory effects.
- The extensive metabolism at the 5,6-position, particularly in BNF-induced microsomes, correlates with reduced inhibitory activity.
- Understanding ANF metabolism provides insights into the mechanisms of carcinogen metabolism and flavonoid-drug interactions.