Detection of new radiation-induced DNA lesions by liquid chromatography coupled to tandem mass spectrometry

• Published in: Rapid communications in mass spectrometry 2004-10, Vol.18 (19), p.2223-2228
• Main Authors: Regulus, Peggy, Spessotto, Sébastien, Gateau, Mathilde, Cadet, Jean, Favier, Alain, Ravanat, Jean-Luc
• Format: Article
• Language: English
• Subjects: Cell Line; Chemical Sciences; Chromatography, High Pressure Liquid - methods; DNA - analysis; DNA - chemistry; DNA - radiation effects; DNA Damage; DNA Mutational Analysis - methods; Gamma Rays; Humans; Molecular Weight; Monocytes - radiation effects; Organic chemistry; Spectrometry, Mass, Electrospray Ionization - methods
• ISSN: 0951-4198; 1097-0231
• DOI: 10.1002/rcm.1612

High‐performance liquid chromatography coupled to electrospray ionization tandem mass spectrometry (HPLC/ESI‐MS/MS) has been used to search for the formation of as yet unidentified radiation‐induced DNA lesions. For that purpose, the characteristic fragmentation of most of 2′‐deoxyribonucleosides that corresponds to the loss of the 2‐deoxyribose moiety (loss of 116 mass units) has been utilized to specifically detect modified nucleosides. Aerated aqueous solutions of DNA were exposed to ionizing radiation, and subsequently DNA was digested to nucleosides with a cocktail of endo‐ and exonucleases. HPLC/ESI‐MS/MS analysis of the resulting 2′‐deoxyribonucleoside mixture allowed us to detect four novel DNA modifications. In a subsequent step, the sensitivity of the tandem mass spectrometer was used to search for the formation of the newly detected lesions in the DNA of γ‐irradiated cells. Thus, one of the four newly detected lesions was found to be significantly generated in cellular DNA upon exposure to ionizing radiation. In addition, the latter lesion was also shown to be present in untreated cells, indicating that the modified nucleoside could be formed endogenously. Copyright © 2004 John Wiley & Sons, Ltd.