Radical mechanism of aminopyrine oxidation by cumene hydroperoxide catalyzed by purified liver microsomal cytochrome P-450

Brenda Walker Griffin, Charles Marth, Yukio Yasukochi, Bettie Sue Siler Masters

Research output: Contribution to journalArticle

52 Citations (Scopus)

Abstract

Under identical experimental conditions, purified preparations of rabbit liver microsomal cytochrome P-450 and beef heart metmyoglobin were equally effective at stimulating the oxidation of aminopyrine to a free radical species by cumene hydroperoxide. Mannitol had no effect on radical levels produced with either hemeprotein-hydroperoxide system; however, specific ligands of the two hemeproteins, substrates of cytochrome P-450, and phospholipid affected the two systems quite differently. Only the metmyo-globindependent oxidation of aminopyrine was significantly inhibited by fluoride and cyanide. Metyrapone, a specific ligand of cytochrome P-450, and benzphetamine, which was N-demethylated by cumene hydroperoxide only in the presence of cytochrome P-450, inhibited only the cytochrome P-450-stimulated oxidation of aminopyrine. Moreover, only with the solubilized liver hemeprotein was aminopyrine radical generation markedly stimulated by phospholipid. Similar properties of aminopyrine N-demethylation and radical formation by the cytochrome P-450-cumene hydroperoxide system have strongly implicated the radical as a requisite intermediate in product formation. Micromolar concentrations of metyrapone caused parallel inhibition, by at least 50%, of both radical generation and formaldehyde production. These results support a radical pathway of N-demethylation proposed for other hemeprotein-hydroperoxide systems (B. W. Griffin and P. L. Ting, 1978, Biochemistry, 17, 2206-2211), in which the substrate undergoes two successive one-electron abstractions, followed by hydrolysis of the iminium cation intermediate. Thus, for this class of substrates, the experimental data are consistent with the oxygen atom of the product arising from H2O and not directly from the hydroperoxide, which has been previously proposed as a general mechanism for cytochrome P-450 peroxidatic activities.

Original languageEnglish (US)
Pages (from-to)543-553
Number of pages11
JournalArchives of Biochemistry and Biophysics
Volume205
Issue number2
DOIs
StatePublished - 1980
Externally publishedYes

Fingerprint

Aminopyrine
Liver
Cytochrome P-450 Enzyme System
Hemeproteins
Oxidation
Hydrogen Peroxide
Metyrapone
Phospholipids
Substrates
Benzphetamine
Metmyoglobin
Ligands
Beef
Biochemistry
Cyanides
Mannitol
cumene hydroperoxide
Fluorides
Formaldehyde
Free Radicals

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics
  • Molecular Biology

Cite this

Radical mechanism of aminopyrine oxidation by cumene hydroperoxide catalyzed by purified liver microsomal cytochrome P-450. / Griffin, Brenda Walker; Marth, Charles; Yasukochi, Yukio; Masters, Bettie Sue Siler.

In: Archives of Biochemistry and Biophysics, Vol. 205, No. 2, 1980, p. 543-553.

Research output: Contribution to journalArticle

Griffin, Brenda Walker ; Marth, Charles ; Yasukochi, Yukio ; Masters, Bettie Sue Siler. / Radical mechanism of aminopyrine oxidation by cumene hydroperoxide catalyzed by purified liver microsomal cytochrome P-450. In: Archives of Biochemistry and Biophysics. 1980 ; Vol. 205, No. 2. pp. 543-553.
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