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The oxidation behavior of DNA and RNA nucleotides is studied by an on-line set-up consisting of an electrochemical thin-layer cell (EC) directly coupled to electrospray ionization mass spectrometry (ESI-MS). This set-up allows the generation of nucleotide oxidation products in the electrochemical cell at increasing potentials. Moreover, the products are determined directly, without isolation or derivatization steps, by electrospray ionization time of flight mass spectrometry (ESI-ToF/MS). The dependence of the mass spectra on the applied potential is displayed as mass voltammograms. An advanced set-up, consisting of the electrochemical cell coupled to electrospray ionization tandem mass spectrometry (EC/ESI-MS/MS) allows further structure elucidation based on fragmentation experiments. The electrochemical conversion is performed using a boron doped diamond (BDD) working electrode, which is known to generate hydroxyl radicals at high potentials. The capability of the EC-MS system to generate highly relevant oxidation products which also occur upon oxidative damage in vivo is demonstrated in this study by the formation of well known biomarkers for DNA damage, including 2'-deoxy-8-oxo-guanosine 5'-monophosphate.

It was found that EC/MS is a well-suited approach for the targeted generation of reactive haptens such as PPQD while avoiding detection problems due to the complexity of matrices encountered when conducting conventional in vitro or in vivo experiments.

The pre-hapten PPD was efficiently activated upon EC oxidation and the resulting species were further reacted with different proteins leading to diverse hapten-protein complexes. Thereby, problems related to the complex matrix present in conventional in vitro or in vivo methods could effectively be avoided.

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