br A showing an acceptable reproducibility Nevertheless base
2.51 µA, showing an acceptable reproducibility. Nevertheless, based on biosensor performance, the best S.D was obtained for 2 h of interaction. All further experimentation was carried out at the optimized two hours of interaction time.
3.5. Interaction of SL-DNA/GE and dsDNA/GE with 7ESTAC01 and detection of DNA damage
All the following experiments were carried out for different con-centrations of 7ESTAC01 with 2 h of interaction time. As depicted in Fig. 4, after the addition of 7ESTAC01, the strong binding interaction between 7ESTAC01 with guanine and adenine base of SL-DNA/GE and dsDNA/GE occurred. Therefore, as the concentration of 7ESTAC01 in-creases, the oxidation peak current of SL-DNA probe and dsDNA in-creased (Fig. 4A and B). These results are similar to those obtained by Lucarelli et al. (2002). Lucarelli and co-workers utilized screen-printed electrodes for the detection of apolipoprotein E, where an increase of the electrochemical signal of the guanine base resulted from the non-specific interaction of the apolipoprotein E and the DNA immobilized on the electrode. The increasing oxidation peak current we report here is consistent with other electrochemical DNA/GE biosensors testing a redox-active intercalator, such as in the case of anthraquinone mono sulfonic 33069-62-4 (AQMS) (Wong and Gooding, 2006). Electron transfer from the DNA to AQMS intercalated into DNA duplexes reported the growth of the peak current signal with time. These last results support the electrochemical behaviour of our biosensors in the presence of 7ESTAC01, which is intercalated into the double-stranded DNA.
To evaluate the level of DNA damage with the dsDNA/GE and SL-DNA/GE, we compared guanine and adenine peak current in the same conditions. The adenine oxidation was recorded for SL-DNA/GE and dsDNA/GE, each yielding different sensitivity levels. The SL-DNA/GE-7ESTAC01 showed adenine oxidation for 100 µM (Fig. 3B) and 400 µM 7ESTAC01 (Fig. 4A). Notoriously, an adenine peak for 100 and 400 µM 7ESTAC01 (Fig. 4B, red histograms) with the dsDNA/GE-7ESTAC01, reached higher adenine peak current at 12.36 µA and 18.63 µA, re-spectively. On the other hand, as shown in Fig. 4 (blue histograms), the guanine oxidation was seen for dsDNA/GE and SL-DNA/GE with the presence of the minimum concentration of 7ESTAC01. The guanine peak current for 40 µM 7ESTAC01 with SL-DNA/GE and dsDNA/GE, reached 9.412 µA and 11.35 µA, respectively (Fig. 4A and B, inner graph). These results not only validated the strong interaction between 7ESTAC01 and purine bases but also emphasized the higher sensitivity
Fig. 4. Detection of the DNA damage product of the interaction between SL-DNA/GE and dsDNA/GE with 7ESTAC01 expressed by the DPV peak currents. DPV peak currents responses under electro-oxidation on SL-DNA/GE (A) and dsDNA/GE (B) in presence of 7ESTAC01 at concentrations of (a) 40, (b) 100 and (c) 400 µM for 2 h of interaction. Histograms represent the guanine and adenine peak current for SL-DNA/GE, and dsDNA/GE extrapolated from the DPV signal of each biosensor (A and B, inner graph). Error bars represent standard deviations (S.D). The S.D of the guanine peak current on the SL-DNA/GE for 40 and 400 µM 7ESTAC01 were 0.31 µA and 0.91 µA, respectively. The S.D of the guanine peak current on the dsDNA/GE for 40, 100 and 400 µM 7ESTAC01 were 0.44 µA, 0.52 µA and 1.32 µA, respectively. DPV signal in acetate buffer for SL-DNA/GE (*) and dsDNA/GE (+) without the presence of 7ESTAC01.
and more damage induced by employing the dsDNA/GE versus the SL-DNA/GE. The dsDNA and SL-DNA sequences exhibit 11 and 5 guanines, respectively. It is possible that 7ESTAC01-DNA intercalation can lead to breaking hydrogen bonds and exposing guanine and adenine bases to the surface of the GE. Therefore, it is likely that the higher oxidation peak current exhibited by the dsDNA/GE-7ESTAC01 system is due to a higher quantity of available guanine and adenine bases in comparison to the SL-DNA/GE-7ESTAC01 system (Fig. 4A). In fact, two critical parameters must be followed to improve the electron transfer effi-ciency, (i) type of DNA structure, and (ii) distance between guanine base and electrode surface (Brett et al., 2003; McEwen et al., 2009; Ibañez et al., 2015). Similar results were observed in a single stranded DNA modified GE biosensor, wherein electron transfer efficiency was the highest when guanine bases were in close proximity and exposed to the surface of the GE (Huang et al., 2016). In addition, the high charge migration along the DNA duplex in the presence of a DNA intercalator like 7ESTAC01, can also account for the higher oxidation peak current seen in the dsDNA platform (Liu and Barton, 2005; Elias et al., 2008).