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Title
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Alkylation of DNA by melphalan : investigation of capillary liquid chromatography electrospray tandem mass spectrometry in the study of adducts at the nucleoside level
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Author
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Abstract
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| Nitrogen mustards are among the oldest cancer chemotherapeutic agents and remain the drugs of choice for treatment of many human cancers. A serious complication of treatment with nitrogen mustards is the increased risk of a secondary leukaemia in long-term survivors because not all alkylating agent interactions with DNA result in cell death. In an earlier study 2'-deoxy-5'-mononucleotide/melphalan adducts have been analysed by us by LC-ES MSMS. In this work we want to present the first results of the analysis of the corresponding 2'-deoxynucleoside/melphalan adducts from DNA hydrolysates by column switching/capillary LC-ES tandem mass spectrometry. Nucleosides, compared to nucleotides, give better chromatographic results and show a good sensitivity under electrospray (+) [ES(+)] ionisation. Several adducts were identified under ES(+) conditions. Mono-alkylated nucleoside adducts alkylated at the base moiety were identified for dGuo, dCyd and dAdo. Structures were identified by recording the low-energy CAD product ion scans. Also a mono-alkylated nucleotide pdA with alkylation position at the phosphate moiety could be detected. This proves that in the case of phosphate alkylation the enzymatic dephosphorylation reaction was inhibited. A Jurkat cell suspension was treated with melphalan (I mM) and incubated at 37 degreesC (5% CO2). After 6 and 48 h, the DNA was isolated and enzymatically hydrolysed. The corresponding nucleoside pool was evaluated with the developed LC-MS method. In the 48-h experiment, one adduct could be identified as a N-7 alkylated dGuo. In the 6-h experiment, no adducts could be found. Additional experiments were done wherein Jurkat-DNA, isolated from a non-treated cell culture, was treated with melphalan. These results were analogous with the data found in melphalan-treated calf thymus DNA. Additionally, we tried to determine the exact alkylation position by interpreting high-resolution fragmentation spectra. (C) 2002 Elsevier Science B.V. All rights reserved. |
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Language
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English
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Source (journal)
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Journal of chromatography : B: analytical technologies in the biomedical and life sciences. - Amsterdam, 2002, currens
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Publication
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Amsterdam
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Elsevier
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2003
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ISSN
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1570-0232
[print]
1873-376X
[online]
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DOI
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10.1016/S1570-0232(02)00851-6
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Volume/pages
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75
:1
(2003)
, p. 21-37
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ISI
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000180592700002
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Full text (Publisher's DOI)
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Full text (publisher's version - intranet only)
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