Electrochemical Detection of Guanine‐methylation Using Glassy Carbon Electrode

Abstract

This work presents a study of the electrochemical oxidation of 7‐methylguanine (7‐mGua) in aqueous solution at glassy carbon electrode by cyclic voltammetry, differential pulse voltammetry, square wave voltammetry and electrochemical impedance spectrometry. The anodic behaviour of 7‐mGua was compared with the electro‐oxidation of guanine and 7‐methylguanosine. The results demonstrated that the methyl and ribose groups are not electroactive but strongly influence the oxidation mechanism of these species. The oxidation of 7‐mGua occurred in a single pH‐dependent step, with the withdrawal of two electrons and two protons of C8, to form 8‐oxo‐7‐methylguanine, while the electro‐oxidation of 7‐methylguanosine also occurred in a single pH‐dependent step, however, with the withdrawal of one electron and one proton of C8 to form a hydroxylated product, since its oxidation to 8‐oxo‐7‐methylguanosine is hindered by the presence of the pendant groups. In addition, the oxidation of 7‐mGua was investigated in the presence of DNA and DNA‐bases, leading to the conclusion that the formation of 7‐mGua, from an interaction of DNA with an alkylating agent, would cause an increase on the deoxyguanosine peak current of the DNA‐biosensor, with no interference of any free DNA bases, which demonstrated that DNA‐electrochemical biosensors find application on detecting DNA methylation, opening a new avenue for applications of DNA biosensors.

Voltammetric Studies of 3‐Nitro‐tyrosine Electro‐oxidation at Solid Electrodes and its Interaction with DNA

“…DP voltammograms were recorded in a 1 μmol L −1 guanine (Gua) solution and in a Gua+dsDNA mixture (1 μmol L −1 /200 μg mL −1 ), prepared in phosphate buffer, pH=7.0, and detected only a single anodic peak, at E pGua =+0.69 V, with currents close to 14 nA [7, 42, 43], Figure 9, thus demonstrating, as expected, that the dsDNA molecule does not interfere in the Gua electro‐oxidation process. These experiments are important to demonstrate and confirm that the disappearance of the anodic peak of 3‐NO 2 ‐Tyr in the presence of dsDNA occurs in fact and exclusively due to the interaction of these molecules, hindering the access of 3‐NO 2 ‐Tyr to the GCE surface and its electro‐oxidation.…”

“…All DP voltammograms of 3‐NO 2 ‐Tyr detected only the anodic peak 1a, at E p1a =+0.81 V, due to the oxidation of the phenolic group [1], with currents close to 16 nA, Figure 6A. The DP voltammograms of dsDNA detected, as expected, two consecutive oxidation peaks, Figure 6B, the first anodic peak corresponds to the oxidation of deoxyguanosine residues (dGuo), at E pdGuo =+0.91 V and the second is associated with the oxidation of deoxyadenosine (dAdo) residues at E pdAdo =+1.17 V [7, 42, 43].…”

“…The low affinity of the dsDNA molecule with the GCE surface is well known in the literature, with reports on the complexity of its structure and its high hydrophilicity [7, 42, 43]. However, the electro‐oxidation of guanine in the absence/presence of the dsDNA was investigated in the present study as a control/model experiment, to demonstrate that the dsDNA molecule does not cause the blocking of the GCE surface, which could hinder new charge transfer processes.…”

“…In this way, the effects of the 3‐NO 2 ‐Tyr‐dsDNA interaction were monitored by DPV, as described in Section 2.1 , comparing the variations of the 3‐NO 2 ‐Tyr oxidation peak 1a in the absence/presence of the dsDNA, the variations of the DNA anodic peaks in the absence/presence of the 3‐NO 2 ‐Tyr and monitoring the detection of the 8‐oxoguanine oxidation peak, as an indicator of dsDNA oxidative damage [7, 42, 43].…”