Many molecules show high affinity to nucleic acids as they interact with DNA by several mechanisms [1]. They can join nucleic acids and inhibit basic functions of the living cells. To analytical chemist, nucleic acids offer a powerful tool in recognition and monitoring of many important compounds, e.g. toxic molecules and anticancer agents [2, 3]. Electrochemical techniques are frequently used for the detection. The goal of this work is to investigate the interactions of methylene blue (MB) with dsDNA using home made screen printed three electrode system and various electrochemical techniques: cyclic voltammetry (CV), differential pulse voltammetry (DPV), and square wave voltammetry (SWV). The explanation of the mechanism of interaction between MB and dsDNA is based on the electrochemical behavior of DNA in the absence and in the presence of MB as well as on the electrochemical behavior of MB in the absence and in the presence of DNA. The DNA presence is detected as signals due to oxidation of guanine and adenine and MB presence is detected as signals arising from oxidation or reduction of MB interacting with the DNA. The dsDNA is immobilized on the surface of the electrode by electrostatic attraction enabling very effective electron transfer between the nucleic acid bases and the electrode. After dsDNA immobilization, the electrode is rinsed with deionized water and then immersed into 10 mmol/L KCl in a pH 4.75 acetic acid buffer and electrochemical signals of oxidation of guanine and adenine are measured (blank). Alternatively, after rinsing, the electrode is immersed into a MB solution of given concentration in 10 mmol/L KCl and acetic acid buffer of pH 4.75, and after rinsing electrochemical signals of oxidation of guanine and adenine in buffered KCl solution are measured (sample). The signals obtained using these two procedures are compared and the difference detected proves the presence of MB interactions with DNA. Electrochemical signals of oxidation and reduction of MB intercalated into DNA are measured and compared with measurements of electrode modified only by MB adsorption. The difference detected proves the presence of MB interactions with DNA. Cyclic voltammetry indicates the existence of the interactions between MB and dsDNA. The similar observations of MB oxidation and reduction current using DPV and SWV indicate some difference when either DNA is present or absent. These suggest that MB can play a role of mediator between DNA and electrode surface. The dependence of SWV peak current and peak potential of the guanine oxidation on MB presence indicates strong interaction between this nucleic acid base and MB and kinetic changes in oxidation reaction of guanine. The dependence of peak current of nucleic bases oxidation in DPV and SWV on MB presence indicates strong interaction between guanine and MB facilitating the oxidation of guanine and weak (DPV, SWV) interactions between adenine and MB, proving the DNA and MB interactions in guanine rich DNA regions.

References:

[1] S. Rauf, J.J. Gooding, K. Akhtar, M.A. Ghauri, M. Rahman, M.A. Anwar, A.M. Khalid, Electrochemical Approach of Anticacer Drugs-DNA Interaction, Journal of Pharmaceutical and Biomedical Analysis, Vol. 37, No. 2, 2005, pp. 205-217.

[2] D. Maciejewska, I. Szpakowska, I. Wolska, M. Niemyjska, M. Mascini, M. Maj-Zurawska, DNA-based Electrochemical Biosensors for Monitoring of bis-indoles as Potential Antitumoral Agents, Chemistry, X-ray Crystolagraphy, Bioelectrochemistry, Vol. 69, No. 1, 2006, pp.1-9.

[3] I. Szpakowska, B. Krassowska-Swiebocka, D. Maciejewska, P. Kazmierczak, W. Jemielita, M. Konrad, J. Trykowska, M. Maj-Zurawska, Electrochemical DNA Biosensor for Testing Pentamidine and Its Analogues as Potential Chemoterapeutics, Electroanalysis, Vol. 18, No. 13-14, 2006, 1422-1430.

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