Nitric oxide decreases the stability of DMPO spin adducts

Krzysztof J. Reszka, Michael L. McCormick, Garry R. Buettner, C. Michael Hart, Bradley E Britigan

Research output: Contribution to journalArticle

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Abstract

The effect nitric oxide (NO{radical dot}) on the stability of 5,5-dimethyl-1-pyrroline-N-oxide (DMPO) adducts has been investigated using EPR spectroscopy. We report that the DMPO/HO{radical dot} adduct, generated by porcine pulmonary artery endothelial cells in the presence of H2O2 and DMPO, or by a Fenton system (Fe(II) + H2O2) is degraded in the presence of the NO{radical dot}-donor, 2-(N,N-diethylamino)-diazenolate-2-oxide (DEANO) or by bolus addition of an aqueous solution of NO{radical dot}. A similar effect of DEANO was observed on other DMPO adducts, such as DMPO/{radical dot}CH3 and DMPO/{radical dot}CH(CH3)OH, generated in cell-free systems. Measurements of the loss of DMPO/HO{radical dot} in the presence of DEANO in aerated and oxygen-free buffers showed that in both of these settings the process obeys first-order kinetics and proceeds with similar efficacy. This indicates that direct interaction of the nitroxide with NO{radical dot}, rather than with NO2{radical dot} (formed from NO{radical dot} and O2 in aerated media), is responsible for destruction of the spin adduct. These results suggest that the presence of NO{radical dot} may substantially affect the quantitative determination of DMPO adducts. We also show that NO2{radical dot} radicals, generated by a myeloperoxidase/H2O2/nitrite system, also degrade DMPO/HO{radical dot}. Because DMPO is frequently used to study generation of superoxide and hydroxyl radicals in biological systems, these observations indicate that extra caution is required when studying generation of these species in the presence of NO{radical dot} or NO2{radical dot} radicals.

Original languageEnglish (US)
Pages (from-to)133-141
Number of pages9
JournalNitric Oxide - Biology and Chemistry
Volume15
Issue number2
DOIs
StatePublished - Sep 1 2006

Fingerprint

Nitric Oxide
Oxides
5,5-dimethyl-1-pyrroline-1-oxide
Cell-Free System
Endothelial cells
Biological systems
Nitrites
Superoxides
Hydroxyl Radical
Pulmonary Artery
Peroxidase
Paramagnetic resonance
Spectrum Analysis
Buffers
Swine
Endothelial Cells
Spectroscopy
Oxygen
Kinetics

Keywords

  • DMPO
  • EPR
  • Hydroxyl radical
  • Myeloperoxidase
  • Nitric oxide
  • Nitrogen dioxide
  • Spin trapping

ASJC Scopus subject areas

  • Biochemistry
  • Physiology
  • Clinical Biochemistry
  • Cancer Research

Cite this

Nitric oxide decreases the stability of DMPO spin adducts. / Reszka, Krzysztof J.; McCormick, Michael L.; Buettner, Garry R.; Michael Hart, C.; Britigan, Bradley E.

In: Nitric Oxide - Biology and Chemistry, Vol. 15, No. 2, 01.09.2006, p. 133-141.

Research output: Contribution to journalArticle

Reszka, Krzysztof J. ; McCormick, Michael L. ; Buettner, Garry R. ; Michael Hart, C. ; Britigan, Bradley E. / Nitric oxide decreases the stability of DMPO spin adducts. In: Nitric Oxide - Biology and Chemistry. 2006 ; Vol. 15, No. 2. pp. 133-141.
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AU - Britigan, Bradley E

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N2 - The effect nitric oxide (NO{radical dot}) on the stability of 5,5-dimethyl-1-pyrroline-N-oxide (DMPO) adducts has been investigated using EPR spectroscopy. We report that the DMPO/HO{radical dot} adduct, generated by porcine pulmonary artery endothelial cells in the presence of H2O2 and DMPO, or by a Fenton system (Fe(II) + H2O2) is degraded in the presence of the NO{radical dot}-donor, 2-(N,N-diethylamino)-diazenolate-2-oxide (DEANO) or by bolus addition of an aqueous solution of NO{radical dot}. A similar effect of DEANO was observed on other DMPO adducts, such as DMPO/{radical dot}CH3 and DMPO/{radical dot}CH(CH3)OH, generated in cell-free systems. Measurements of the loss of DMPO/HO{radical dot} in the presence of DEANO in aerated and oxygen-free buffers showed that in both of these settings the process obeys first-order kinetics and proceeds with similar efficacy. This indicates that direct interaction of the nitroxide with NO{radical dot}, rather than with NO2{radical dot} (formed from NO{radical dot} and O2 in aerated media), is responsible for destruction of the spin adduct. These results suggest that the presence of NO{radical dot} may substantially affect the quantitative determination of DMPO adducts. We also show that NO2{radical dot} radicals, generated by a myeloperoxidase/H2O2/nitrite system, also degrade DMPO/HO{radical dot}. Because DMPO is frequently used to study generation of superoxide and hydroxyl radicals in biological systems, these observations indicate that extra caution is required when studying generation of these species in the presence of NO{radical dot} or NO2{radical dot} radicals.

AB - The effect nitric oxide (NO{radical dot}) on the stability of 5,5-dimethyl-1-pyrroline-N-oxide (DMPO) adducts has been investigated using EPR spectroscopy. We report that the DMPO/HO{radical dot} adduct, generated by porcine pulmonary artery endothelial cells in the presence of H2O2 and DMPO, or by a Fenton system (Fe(II) + H2O2) is degraded in the presence of the NO{radical dot}-donor, 2-(N,N-diethylamino)-diazenolate-2-oxide (DEANO) or by bolus addition of an aqueous solution of NO{radical dot}. A similar effect of DEANO was observed on other DMPO adducts, such as DMPO/{radical dot}CH3 and DMPO/{radical dot}CH(CH3)OH, generated in cell-free systems. Measurements of the loss of DMPO/HO{radical dot} in the presence of DEANO in aerated and oxygen-free buffers showed that in both of these settings the process obeys first-order kinetics and proceeds with similar efficacy. This indicates that direct interaction of the nitroxide with NO{radical dot}, rather than with NO2{radical dot} (formed from NO{radical dot} and O2 in aerated media), is responsible for destruction of the spin adduct. These results suggest that the presence of NO{radical dot} may substantially affect the quantitative determination of DMPO adducts. We also show that NO2{radical dot} radicals, generated by a myeloperoxidase/H2O2/nitrite system, also degrade DMPO/HO{radical dot}. Because DMPO is frequently used to study generation of superoxide and hydroxyl radicals in biological systems, these observations indicate that extra caution is required when studying generation of these species in the presence of NO{radical dot} or NO2{radical dot} radicals.

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