Separation and identification of platinum adducts with DNA nucleotides by capillary zone electrophoresis and capillary zone electrophoresis coupled to mass spectrometry

Platinum adducts are supposed to be the cytotoxic lesions in DNA after plat inum-containing anticancer therapy. Various adducts are formed upon interac tion of platinum complexes with nucleotides, but contribution of individual adducts to antitumor activity and toxicity of platinum complexes still rem ains to be examined. A capillary zone electrophoresis (CZE) method is descr ibed that is suitable to separate individual platinum adducts. We investiga ted the formation of adducts following the reaction of cis-diamminedichloro platinum (II) (cisplatin) with various DNA nucleotides. Baseline separation of unmodified and modified nucleotides (adducts) was achieved using uncoat ed fused-silica capillaries and basic separation buffers. In order to eluci date the observed peak pattern, a coupled CZE-electrospray ionization-mass spectrometry (ESI)-MS approach was applied. After incubation of mononucleot ides with cisplatin, monochloro, monoaqua and bifunctional adduct species w ere detected. Consequently, the migration order of nucleotides and individu al platinum adducts could be determined. Moreover, the time-dependent conve rsion from monochloro to monoaqua and subsequently to bifunctional adducts was monitored. In conclusion, individual platinum adducts were separated by CZE and identified by CZE-ESI-MS. Formation and conversion of distinct spe cies were confirmed. Potential applications comprise studies of novel plati num complexes, investigations of platinum-adduct formation with DNA, and de termination of platinum-DNA adducts in cells.