Reactions of halogen-substituted aziridinylbenzoquinones with glutathione: Formation of diglutathionyl conjugates and semiquinones

Cecilia R Giulivi, Angela Forlin, Stefano Bellin, Enrique Cadenas

Research output: Contribution to journalArticle

3 Scopus citations

Abstract

The reaction between glutathione and 2,5-diaziridinyl-1,4-benzoquinones bearing halogen substituents at C3 and C6 was examined in terms of the formation of glutathionyl-quinone conjugates and semiquinones by HPLC with UV detection, mass spectroscopy and EPR. The reactivity of the halogen atoms toward sulfur substitution is the primary reaction leading to the formation of mono- and di-glutathionyl-substituted quinones. The relative formation of these conjugates depended on the GSH/quinone molar ratios. At GSH/quinone molar ratios below unity, the products observed were the reduced form of the parent quinone, a dithiother derivative and GSSG. Disulfide formation accounted for 60-68% of total GSH consumed. EPR analysis of these reaction mixtures showed a 5-line spectrum (1:2:3:2:1 relative intensities) with 2 equivalent N (a(N) = 1.98 G) and assigned to the semiquinone form of dichloro- diaziridinylbenzoquinone. Semiquinone quantification by double integration of the EPR signals and interpolation with an adequate standard revealed that the amount of semiquinone formed per GSH consumed was 0.98. At GSH/quinone molar ratios above unity (4, 10 and 100 molar excess of GSH) a pattern of products emerged consisting of mass spectral analysis. EPR studies revealed that these compounds were minor components of a composite EPR spectrum (a 3-line signal with 1:1:1 relative intensities, 1 equivalent N (a(N) = 1.73 G) and 1 H (a(H) = 1.45 G) or a 3-line signal with 1:2:1 relative intensities and 2 equivalent H (a(H) = 1.4 G). These minor components were assigned to the diglutathionyl conjugates bearing one- or no aziridinyl moiety, respectively. The major component in the EPR signal showed a 3-line spectrum (1:1:1 relative intensity) with 1 equivalent N (a(N) = 1.7 G) and a g shift of -0.96 G. This spectrum was assigned to a triglutathionyl conjugate of a monoaziridinylbenzoquinone. This major component was also observed when GSH/quinone mixtures were incubated with the two-electron transfer flavoprotein NAD(P)H:quinone oxidoreductase. The semiquinone signals were abolished by superoxide dismutase. In the presence of catalase, the contribution of these components to the overall EPR spectrum was equal. These data are discussed in terms of the one-electron transfer steps encompassed by thiol oxidation and semiquinone formation and the two-electron transfers inherent in sulfur substitution and aziridinyl group loss.

Original languageEnglish (US)
Pages (from-to)137-154
Number of pages18
JournalChemico-Biological Interactions
Volume108
Issue number3
DOIs
StatePublished - Jan 30 1998
Externally publishedYes

Keywords

  • Azirinidylbenzoquinones
  • EPR
  • Glutathione
  • Glutathione conjugates
  • NAD(P)H:quinone oxidoreductase
  • Semiquinones

ASJC Scopus subject areas

  • Toxicology

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