Formation of 4-aminophenoxyl free radical from the acetaminophen metabolite N-acetyl-p-benzoquinone imine.
N-Acetyl-p-benzoquinone imine, a hepatic metabolite of acetaminophen, and its analogue, N-acetyl-3,5-dimethyl-p-benzoquinone imine, were metabolized by rat liver microsomes and NADPH to their corresponding 4-aminophenoxyl free radicals. ESR spectra were recorded and unambiguously identified. As indicated by the purple color and confirmed by UV and mass spectroscopy, indophenols were formed as final products. The 4-aminophenoxyl free radical formation could be suppressed by the deacetylase inhibitors, sodium fluoride and paraoxon. Microsomal incubations of N-acetyl-2,6-dimethyl-p-benzoquinone imine and NADPH do not result in a detectable radical concentration; in addition, no indophenol was found. Substitution of NADPH-cytochrome P-450 reductase for rat liver microsomes eliminates the deacetylase activity and results in direct reduction of N-acetyl-3,5-dimethyl-p-benzoquinone imine to the N-acetyl-2,6-dimethyl-4-aminophenoxyl free radical. Neither the incubation of N-acetyl-p-benzoquinone imine nor that of N-acetyl-2,6-dimethyl-p-benzoquinone imine with NADPH-cytochrome P-450 reductase yielded a detectable concentration of the corresponding phenoxyl free radical. When starting material that had been exposed to the atmosphere was used, a previously reported free radical with a splitting constant of approximately 2 G was formed. This spectrum is identical with that of the 2,6-dimethyl-p-benzosemiquinone free radical, implying hydrolysis of the starting material. Neither the N-acetyl-4-aminophenoxyl nor the N-acetyl-2,6-dimethyl-4-aminophenoxyl radical reduces oxygen to form superoxide or react with oxygen in any other detectable way.
D000818:Animals; D016227:Benzoquinones; D003497:Cyclic N-Oxides / Q000378:metabolism; D004578:Electron Spin Resonance Spectroscopy; D005609:Free Radicals; D007097:Imines / Q000378:metabolism*; D066298:In Vitro Techniques; D008862:Microsomes, Liver / Q000378:metabolism; D009251:NADPH-Ferrihemoprotein Reductase / Q000494:pharmacology; D051381:Rats; D013481:Superoxides / Q000378:metabolism
D016227:Benzoquinones; D003497:Cyclic N-Oxides; D005609:Free Radicals; D007097:Imines; D013481:Superoxides; C017245:5,5-dimethyl-1-pyrroline-1-oxide; D009251:NADPH-Ferrihemoprotein Reductase; C028473:N-acetyl-4-benzoquinoneimine