Intermediates produced in the reaction of chromium(VI) with dehydroascorbate cause single-strand breaks in plasmid DNA

Diane M Stearns, Karen E. Wetterhahn

Research output: Contribution to journalArticle

26 Citations (Scopus)

Abstract

Ascorbate (vitamin C) is a biological reductant of the human carcinogen chromium(VI). The product of this reaction is presumed to be dehydroascorbate. However, we have found that chromium(VI) can also react with dehydroascorbate. This reaction was monitored by UV/visible and electron paramagnetic resonance (EPR) spectroscopies. In sodium acetate buffer at pH 3.8, the reaction of chromium(VI) and excess dehydroascorbate produced chromium(V) and chromium(IV) intermediates. At high reaction concentration, the chromium(V) intermediate formed an EPR silent dimer, which dissociated upon dilution to lower concentration. UV/visible experiments at pH 3.8 demonstrated that manganese(II) catalyzed the disproportionation of chromium(IV) to chromium(V) and chromium(III). The ability of the reaction intermediates to induce strand breaks in pBR322 DNA was determined at pH 3.8 and pH 5.8. At pH 3.8, chromium(IV) appeared to be the major species responsible for induction of strand breaks because the time course for formation of strand breaks did not parallel that of chromium(V), and strand breaks were decreased in the presence of the chromium(IV) scavenger manganese(II). At pH 5.8, fewer strand breaks were observed; however, the time course for their formation followed that of chromium(V). There has been much effort devoted to identification of the intermediate responsible for the induction of strand breaks during reactions of chromium(VI) with biological reductants. The current results suggest that it is not a single type of species that universally produces the DNA strand breaks observed in different chromium(VI) systems and that the reactivity of intermediates will depend on the chosen experimental conditions. Understanding this variability in chromium(VI) reactions may help to resolve the conflicting results from in vitro studies that are aimed at deciphering mechanisms of chromium(VI)induced cancers.

Original languageEnglish (US)
Pages (from-to)271-278
Number of pages8
JournalChemical Research in Toxicology
Volume10
Issue number3
DOIs
StatePublished - Mar 1997
Externally publishedYes

Fingerprint

Chromium
Plasmids
DNA
Reducing Agents
Electron Spin Resonance Spectroscopy
Manganese
Paramagnetic resonance
chromium hexavalent ion
Sodium Acetate
Reaction intermediates
DNA Breaks
Carcinogens
Dimers
Dilution
Ascorbic Acid
Spectrum Analysis
Buffers
Spectroscopy

ASJC Scopus subject areas

  • Drug Discovery
  • Organic Chemistry
  • Chemistry(all)
  • Toxicology
  • Health, Toxicology and Mutagenesis

Cite this

Intermediates produced in the reaction of chromium(VI) with dehydroascorbate cause single-strand breaks in plasmid DNA. / Stearns, Diane M; Wetterhahn, Karen E.

In: Chemical Research in Toxicology, Vol. 10, No. 3, 03.1997, p. 271-278.

Research output: Contribution to journalArticle

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abstract = "Ascorbate (vitamin C) is a biological reductant of the human carcinogen chromium(VI). The product of this reaction is presumed to be dehydroascorbate. However, we have found that chromium(VI) can also react with dehydroascorbate. This reaction was monitored by UV/visible and electron paramagnetic resonance (EPR) spectroscopies. In sodium acetate buffer at pH 3.8, the reaction of chromium(VI) and excess dehydroascorbate produced chromium(V) and chromium(IV) intermediates. At high reaction concentration, the chromium(V) intermediate formed an EPR silent dimer, which dissociated upon dilution to lower concentration. UV/visible experiments at pH 3.8 demonstrated that manganese(II) catalyzed the disproportionation of chromium(IV) to chromium(V) and chromium(III). The ability of the reaction intermediates to induce strand breaks in pBR322 DNA was determined at pH 3.8 and pH 5.8. At pH 3.8, chromium(IV) appeared to be the major species responsible for induction of strand breaks because the time course for formation of strand breaks did not parallel that of chromium(V), and strand breaks were decreased in the presence of the chromium(IV) scavenger manganese(II). At pH 5.8, fewer strand breaks were observed; however, the time course for their formation followed that of chromium(V). There has been much effort devoted to identification of the intermediate responsible for the induction of strand breaks during reactions of chromium(VI) with biological reductants. The current results suggest that it is not a single type of species that universally produces the DNA strand breaks observed in different chromium(VI) systems and that the reactivity of intermediates will depend on the chosen experimental conditions. Understanding this variability in chromium(VI) reactions may help to resolve the conflicting results from in vitro studies that are aimed at deciphering mechanisms of chromium(VI)induced cancers.",
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N2 - Ascorbate (vitamin C) is a biological reductant of the human carcinogen chromium(VI). The product of this reaction is presumed to be dehydroascorbate. However, we have found that chromium(VI) can also react with dehydroascorbate. This reaction was monitored by UV/visible and electron paramagnetic resonance (EPR) spectroscopies. In sodium acetate buffer at pH 3.8, the reaction of chromium(VI) and excess dehydroascorbate produced chromium(V) and chromium(IV) intermediates. At high reaction concentration, the chromium(V) intermediate formed an EPR silent dimer, which dissociated upon dilution to lower concentration. UV/visible experiments at pH 3.8 demonstrated that manganese(II) catalyzed the disproportionation of chromium(IV) to chromium(V) and chromium(III). The ability of the reaction intermediates to induce strand breaks in pBR322 DNA was determined at pH 3.8 and pH 5.8. At pH 3.8, chromium(IV) appeared to be the major species responsible for induction of strand breaks because the time course for formation of strand breaks did not parallel that of chromium(V), and strand breaks were decreased in the presence of the chromium(IV) scavenger manganese(II). At pH 5.8, fewer strand breaks were observed; however, the time course for their formation followed that of chromium(V). There has been much effort devoted to identification of the intermediate responsible for the induction of strand breaks during reactions of chromium(VI) with biological reductants. The current results suggest that it is not a single type of species that universally produces the DNA strand breaks observed in different chromium(VI) systems and that the reactivity of intermediates will depend on the chosen experimental conditions. Understanding this variability in chromium(VI) reactions may help to resolve the conflicting results from in vitro studies that are aimed at deciphering mechanisms of chromium(VI)induced cancers.

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