br The reduction of ESTAC
The reduction of 7ESTAC01 showed the first cathodic peak at EIc = −0.38 V, which was lower than the 9-aminoacridine alone. This behaviour hinted a possible synergic activity of 7ESTAC01 due to the significant shift of the first cathodic peak potential. It is worth men-tioning that most of the bioactive compounds are recorded at a less negative potential of − 0.5 V (Bouffier et al., 2012; Dogan-Topal et al., 2014; Nepali et al., 2014; Noh et al., 2015). Thus, 7ESTAC01 represents a promising anti-cancer drug candidate; even though additional bioactivity tests must still be performed.
3.2. Characterization and optimization of SL-DNA/GE and dsDNA/GE
SL-DNA probe contains a thermodynamically stable structure and was used due of its capacity to detect specific site interaction (Nguyen and Wilson, 2009). Both SL-DNA and dsDNA structures were proposed and evaluated, as shown in Scheme 1. The SL-DNA probe, modified with MeB redox marker, was covalently attached to the gold electrode (GE) via a thiol bond (Scheme 1). The cDNA was added to hybridize the SL-DNA probe to form a dsDNA/GE (Scheme 1B). Here, CV was used to evaluate the electrochemical characterization, and subsequently opti-mization of SL-DNA and dsDNA immobilized on the surface of the GE (see Fig. S1 in the Supplementary information). Cyclic voltammograms of the SL-DNA probe in Fig. S1A and Fig. S1B show a high current for MeB due to the close proximity of the redox marker (MeB) to the electrode's surface in the SL-DNA configuration (Scheme 1), since it provides an efficient Dynasore transfer. On the other hand, after hy-bridization with cDNA, the MeB is distant from the surface of the electrode (Scheme 1), consequently decreasing the current of MeB (Fig. S1A and B).
The signal suppression (% SS) as a surface coverage was further evaluated based on equation (Eq. (1)). The % SS of the dsDNA/GE
based on 0.1 µM SL-DNA probe upon hybridization of cDNA, increased in relation to the scan rate, and plateaued around 94% at 100 V s−1 (Fig. S1C). However, when the concentration was at 1.0 µM SL-DNA probe, the %SS reached its maximum at 92% and 600 V s−1. This shows that the proximity of the MeB redox marker supported efficient electron transfer at significantly high scan rates (Fig. S1D). 1.0 µM SL-DNA probe was used to further characterize and optimize the SL-DNA/GE and dsDNA/GE biosensors.
3.3. Electrochemical behaviour of SL-DNA/GE and dsDNA/GE by Differential Pulse Voltammetry
The oxidative DNA damage could be induced in two ways (i) by electro-oxidation and (ii) via oxidizing agents that interact narrowly with DNA (Cadet and Wagner, 2013). Since DNA oxidation is the pro-cess of oxidative injury; in this experiment, the behaviour of SL-DNA/ GE and dsDNA/GE was assessed only under electro-oxidation and without the presence of an oxidizing agent (7ESTAC01). The electrochemical response for SL-DNA/GE and dsDNA/GE were first studied by DPV in acetate buffer at pH 4.2. The modified GE was used for the electrochemical oxidation of adenine (A) and guanine (G). As shown in Fig. 2A, both types of DNA modified GE exhibited peak currents at 1.04 V due to the guanine bases. No adenine peaks were observed for either sensor, which could be explained by the stability of the stem-loop DNA structure through the adenine electro-oxidation (Wei et al., 2011). Otherwise, the electro-oxidation process for modified GE reported oxidation peaks between + 0.85 V to + 0.96 V for guanine (Barman and Jasimuddin, 2014). The DPV peak potentials of the gua-nosine on the ssDNA and dsDNA in acetate buffer reported by Oliveira and Oliveira-Brett (2010) were in agreement with our values. Those results showed that the electrochemical current signal of SL-DNA probe and dsDNA on the GE were mainly attributed to the electrochemical oxidation of guanine bases, which was expected because guanine is the most readily oxidized of the DNA bases (McEwen et al., 2009). Simi-larly, the DPV peak current of guanine on SL-DNA/GE and dsDNA/GE were further evaluated. As indicated in Fig. 2A (inset), SL-DNA/GE, and dsDNA/GE showed a reproducible guanine peak current at 7.94 and 3.86 µA, respectively. The blank response of the GE does not show any peak. All measurements were conducted in triplicates. The S.D of the guanine peak current on the SL-DNA/GE and dsDNA/GE were 0.14 µA and 0.24 µA, respectively (Fig. 2A, inner graph).
The influence of the scan number to the oxidative DNA damage was subsequently examined as an indicator of consecutive DNA lesions under electro-oxidation. DNA oxidation is the process of oxidative in-jury, so it is expected to see higher oxidative damage to the DNA as the number of scans increases. Fig. 2B, prove this hypothesis, wherein the oxidation current of guanine increases with the number of scans, in-dicating that the electron donor was DNA itself.