Alkoxyl and methyl radical formation during cleavage of tert-butyl hydroperoxide by a mitochondrial membrane-bound, redox active copper pool: An EPR study

Eddy M. Massa, Cecilia R Giulivi

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19 Citations (Scopus)

Abstract

The cleavage of tert-butyl hydroperoxide by submitochondrial particles yielded two distinctive radicals, alkoxyl and methyl radicals, detected by the electron paramagnetic resonance technique in conjunction with the spin trap 5-5′-dimethyl-1-pyrrolyne-N-oxide. Free radical formation was partly sensitive to bathocuproine disulfonate and was augmented upon supplementation of the mitochondrial membranes with copper, thus suggesting that a redox active copper pool in mitochondrial membranes participated actively in the cleavage of the OO bond of tert-butyl hydroperoxide. This view was experimentally substantiated by teh following: first, observation of the maximal EPR signal intensity required the presence of exogenous electron donors, such as succinate or reduced nicotinamide adenine dinucleotide (NADH). Second, copper reduction was accomplished partly by a superoxide radical-dependent mechanism as indicated by the sensitivity of the electron paramagnetic resonance (EPR) signal to superoxide dismutase. Third, the enhancing effec of the respiratory chain inhibitors, antimycin A and rotenone, on free radical yield assessed in the presence of superoxide dismutase pointed to the occurrence of two potential loci in the respiratory chain involved in direct electron transfer to membrane-bound copper: one located between NADH and the rotenone-sensitive site and another, quantitatively less important, between the rotenone- and antimycin-sensitive sites. These results support the notion that a redox pool of cooper tightly bound to the mitochondrial membrane contributes significantly to the reductive cleavage of organic peroxides associated with free radical production.

Original languageEnglish (US)
Pages (from-to)559-565
Number of pages7
JournalFree Radical Biology and Medicine
Volume14
Issue number5
DOIs
StatePublished - 1993
Externally publishedYes

Fingerprint

tert-Butylhydroperoxide
Electron Spin Resonance Spectroscopy
Mitochondrial Membranes
Rotenone
Oxidation-Reduction
Paramagnetic resonance
Copper
NAD
Free Radicals
Membranes
Electron Transport
Superoxide Dismutase
Submitochondrial Particles
Electrons
Antimycin A
Peroxides
Succinic Acid
Superoxides
Oxides
Observation

Keywords

  • Alkoxyl radical
  • Copper
  • Electron transport
  • EPR
  • Free radicals
  • Hydroperoxide
  • Spin trapping
  • Submitochondrial particles

ASJC Scopus subject areas

  • Clinical Biochemistry
  • Medicine(all)
  • Toxicology

Cite this

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title = "Alkoxyl and methyl radical formation during cleavage of tert-butyl hydroperoxide by a mitochondrial membrane-bound, redox active copper pool: An EPR study",
abstract = "The cleavage of tert-butyl hydroperoxide by submitochondrial particles yielded two distinctive radicals, alkoxyl and methyl radicals, detected by the electron paramagnetic resonance technique in conjunction with the spin trap 5-5′-dimethyl-1-pyrrolyne-N-oxide. Free radical formation was partly sensitive to bathocuproine disulfonate and was augmented upon supplementation of the mitochondrial membranes with copper, thus suggesting that a redox active copper pool in mitochondrial membranes participated actively in the cleavage of the OO bond of tert-butyl hydroperoxide. This view was experimentally substantiated by teh following: first, observation of the maximal EPR signal intensity required the presence of exogenous electron donors, such as succinate or reduced nicotinamide adenine dinucleotide (NADH). Second, copper reduction was accomplished partly by a superoxide radical-dependent mechanism as indicated by the sensitivity of the electron paramagnetic resonance (EPR) signal to superoxide dismutase. Third, the enhancing effec of the respiratory chain inhibitors, antimycin A and rotenone, on free radical yield assessed in the presence of superoxide dismutase pointed to the occurrence of two potential loci in the respiratory chain involved in direct electron transfer to membrane-bound copper: one located between NADH and the rotenone-sensitive site and another, quantitatively less important, between the rotenone- and antimycin-sensitive sites. These results support the notion that a redox pool of cooper tightly bound to the mitochondrial membrane contributes significantly to the reductive cleavage of organic peroxides associated with free radical production.",
keywords = "Alkoxyl radical, Copper, Electron transport, EPR, Free radicals, Hydroperoxide, Spin trapping, Submitochondrial particles",
author = "Massa, {Eddy M.} and Giulivi, {Cecilia R}",
year = "1993",
doi = "10.1016/0891-5849(93)90114-A",
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pages = "559--565",
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TY - JOUR

T1 - Alkoxyl and methyl radical formation during cleavage of tert-butyl hydroperoxide by a mitochondrial membrane-bound, redox active copper pool

T2 - An EPR study

AU - Massa, Eddy M.

AU - Giulivi, Cecilia R

PY - 1993

Y1 - 1993

N2 - The cleavage of tert-butyl hydroperoxide by submitochondrial particles yielded two distinctive radicals, alkoxyl and methyl radicals, detected by the electron paramagnetic resonance technique in conjunction with the spin trap 5-5′-dimethyl-1-pyrrolyne-N-oxide. Free radical formation was partly sensitive to bathocuproine disulfonate and was augmented upon supplementation of the mitochondrial membranes with copper, thus suggesting that a redox active copper pool in mitochondrial membranes participated actively in the cleavage of the OO bond of tert-butyl hydroperoxide. This view was experimentally substantiated by teh following: first, observation of the maximal EPR signal intensity required the presence of exogenous electron donors, such as succinate or reduced nicotinamide adenine dinucleotide (NADH). Second, copper reduction was accomplished partly by a superoxide radical-dependent mechanism as indicated by the sensitivity of the electron paramagnetic resonance (EPR) signal to superoxide dismutase. Third, the enhancing effec of the respiratory chain inhibitors, antimycin A and rotenone, on free radical yield assessed in the presence of superoxide dismutase pointed to the occurrence of two potential loci in the respiratory chain involved in direct electron transfer to membrane-bound copper: one located between NADH and the rotenone-sensitive site and another, quantitatively less important, between the rotenone- and antimycin-sensitive sites. These results support the notion that a redox pool of cooper tightly bound to the mitochondrial membrane contributes significantly to the reductive cleavage of organic peroxides associated with free radical production.

AB - The cleavage of tert-butyl hydroperoxide by submitochondrial particles yielded two distinctive radicals, alkoxyl and methyl radicals, detected by the electron paramagnetic resonance technique in conjunction with the spin trap 5-5′-dimethyl-1-pyrrolyne-N-oxide. Free radical formation was partly sensitive to bathocuproine disulfonate and was augmented upon supplementation of the mitochondrial membranes with copper, thus suggesting that a redox active copper pool in mitochondrial membranes participated actively in the cleavage of the OO bond of tert-butyl hydroperoxide. This view was experimentally substantiated by teh following: first, observation of the maximal EPR signal intensity required the presence of exogenous electron donors, such as succinate or reduced nicotinamide adenine dinucleotide (NADH). Second, copper reduction was accomplished partly by a superoxide radical-dependent mechanism as indicated by the sensitivity of the electron paramagnetic resonance (EPR) signal to superoxide dismutase. Third, the enhancing effec of the respiratory chain inhibitors, antimycin A and rotenone, on free radical yield assessed in the presence of superoxide dismutase pointed to the occurrence of two potential loci in the respiratory chain involved in direct electron transfer to membrane-bound copper: one located between NADH and the rotenone-sensitive site and another, quantitatively less important, between the rotenone- and antimycin-sensitive sites. These results support the notion that a redox pool of cooper tightly bound to the mitochondrial membrane contributes significantly to the reductive cleavage of organic peroxides associated with free radical production.

KW - Alkoxyl radical

KW - Copper

KW - Electron transport

KW - EPR

KW - Free radicals

KW - Hydroperoxide

KW - Spin trapping

KW - Submitochondrial particles

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