Identification and quantification of 8,5 '-cycle-2 '-deoxyadenosine in DNAby liquid chromatography/mass spectrometry

Recent studies suggested that 8,5'-cyclo-2' -deoxyadenosine may play a role in diseases with defective nucleotide-excision repair. This compound is on e of the major lesions, which is formed in DNA by hydroxyl radical attack o n the sugar moiety of 2'-deoxyadenosine. It is likely to be repaired by nuc leotide-excision repair rather than by base-excision repair because of a co valent bond between the sugar and base moieties. We studied the measurement of 8,5'-cyclo-2'-deoxyadenosine in DNA by liquid chromatography/isotope-di lution mass spectrometry. A methodology was developed for the analysis of 8 ,5'-cyclo-2'-deoxyadenosine by liquid chromatography in DNA hydrolyzed to n ucleosides by a combination of four enzymes, i.e., DNase I, phosphodiestera ses I and II, and alkaline phosphatase. Detection by mass spectrometry was performed using atmospheric pressure ionization-electrospray process in the positive ionization mode. Results showed that liquid chromatography/isotop e-dilution mass spectrometry is well suited for identification and quantifi cation of 8,5'-cyclo-2'-deoxyadenosine in DNA. Both (5'R)- and (5'S)-diaste reomers of 8,5'-cyclo-2'-deoxyadenosine were detected. The level of sensiti vity of liquid chromatography/mass spectrometry with selected-ion monitorin g amounted to 2 fmol of this compound on the column. The yield of 8,5'-cycl o-2'-deoxyadenosine was measured in DNA in aqueous solution exposed to ioni zing radiation at doses from 2.5 to 80 Gray. Gas chromatography/mass spectr ometry was also used to measure this compound in DNA. Both techniques yield ed similar results. The yield of 8,5'-cyclo-2'-deoxyadenosine was comparabl e to the yields of some of the other major modified bases in DNA, which wer e measured using gas chromatography/mass spectrometry. The measurement of 8 ,5'-cyclo-2'-deoxyadenosine by Liquid chromatography/mass spectrometry may contribute to the understanding of its biological properties and its role i n diseases with defective nucleotide-excision repair. (C) 2001 Elsevier Sci ence Inc.