Carcinogenesis, Vol 18, 2385-2391, Copyright © 1997 by Oxford University Press
T Douki, R Martini, JL Ravanat, RJ Turesky and J Cadet
Hydroxyl radical-mediated modifications of nucleobases are among the main
deleterious processes induced in DNA by ionizing radiation. Determination
of the underlying mechanisms in both isolated and cellular DNA requires the
development of accurate assays for the detection of modified bases.
High-performance liquid chromatography associated with electrochemical
detection (HPLC-EC) and gas chromatography coupled to mass spectrometry
(GC-MS) are the two main methods used for this purpose. In the present
work, HPLC-EC was applied to the measurement of
8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodGuo) following enzymatic
digestion of DNA. Evidence was provided for the quantitative aspect of the
latter step. Moreover, a GC-MS assay was designed for the detection of
2,6-diamino-4-hydroxy-5- formamidopyrimidine (FapyGua). It was first shown
that the widely used hot formic acid hydrolysis of DNA induced a complete
decomposition of the lesion. A similar observation was also made for
4,6-diamino-5- formamidopyrimidine (FapyAde). A milder DNA hydrolyzing
agent, HF in pyridine, was substituted to HCOOH for the quantitative
release of FapyGua from DNA. The radiation-induced formation of 8-oxodGuo
and FapyGua in aqueous solution of DNA was compared. Insights in the
mechanistic aspects were inferred from the results of gamma-irradiation
studies carried out under different gas saturating conditions. It was
clearly shown that the formation of both lesions is mediated by HO.. In
addition, the presence of oxygen favors the formation of 8-oxodGuo at the
expense of FapyGua.
ARTICLES
Measurement of 2,6-diamino-4-hydroxy-5-formamidopyrimidine and 8-oxo- 7,8-dihydroguanine in isolated DNA exposed to gamma radiation in aqueous solution
Departement de Recherche Fondamentale sur la Matiere Condensee, SCIB/Laboratoire des Lesions des Acides Nucleiques, CEA/Grenoble, France.
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