Voltammetric behaviour of oligonucleotide lipoplexes adsorbed onto glassy carbon electrodes

Abstract

The voltammetric behaviour of oligonucleotide lipoplexes (ODN-lipoplexes) prepared from short oligodeoxynucleotides (ODN), with different base compositions, and liposomes of the cationic lipid DOTAP, was studied by differential pulse voltammetry with a glassy carbon mini-electrode. It was found that the ODN base composition influences the ODN-lipoplex voltammetric response. Differential pulse voltammograms for ODN-lipoplexes of the ODN adenosine nucleotides present two different features when compared with the differential pulse voltammograms obtained for free ODN: a new peak appeared and the peak attributed to oxidation of adenosine diminished or was absent, depending on whether the ODN sequence had guanosine nucleotides or not. The presence of guanosine nucleotides in the ODN-lipoplex led to a peak due to guanosine oxidation with similar potential and current to the peak obtained for guanosine oxidation in free ODN. No detectable peaks were recorded in the voltammograms obtained with lipoplexes composed of ODN containing only pyrimidine bases. It was possible to show by voltammetry the occurrence of partial denaturation of short double helices of ODN when mixed with DOTAP liposomes to generate lipoplexes. The extent of denaturation was observed to increase with lipoplex (+/)) charge ratio as shown by the increase in the differential pulse voltammetry peak currents. The electrochemical characterisation of lipoplex properties at a charged interface can be important for understanding and development of these gene therapy vectors.

Electrochemical sensing of the behaviour of oligonucleotide lipoplexes at charged interfaces

“…The peak current is proportional to the concentration of the guanosine present in the ODN and it also reflects the accessibility of the guanine nucleotides in the ODN to the surface of the electrode since their adsorption is essential for charge transfer to occur (Oliveira-Brett et al, 1999;Palecek et al, 2002). As expected (Oliveira- Brett and Matysik, 1997;Piedade et al, 2004), the oxidation of the cytosine nucleotides in the ODN was not seen because cytidine can be only detected at high positive potential, for the pH used, and its oxidation peak is usually masked by oxidation of the electrolyte solution.…”

“…It has been shown recently (Piedade et al, 2004) that the ODN base composition and the ODN lipoplex (±) charge ratio influenced the voltammetric response obtained. Therefore, in order to investigate the behaviour of ODN lipoplexes at charged interfaces and to avoid complications due to base sequence effects on the voltammetric response, ODN sequences containing only guanine (G) and cytosine (C) bases (Table 1) were used in the present work.…”

“…The voltammetry of ODN lipoplexes composed of ODN containing all four DNA bases and DOTAP liposomes has been previously studied by differential pulse voltammetry at a glassy carbon electrode surface (Piedade et al, 2004). It was shown that the ODN base composition and the ODN lipoplex (±) charge ratio influenced the voltammetric response confirming the influence of ODN composition on the physicochemical properties of lipoplexes (Meidan et al, 2001).…”

“…After this mechanical treatment the electrode received an electrochemical pre-conditioning that consisted of three steps: first, the electrode was scanned in the electrolyte solution to check the cleanness of the electrode surface; second, a potential of +1.65 V versus Ag/AgCl was applied for 300 s; and finally the electrode was scanned between +0.30 and +1.30 V, until a steady-state base line voltammogram was obtained. This electrochemical pre-conditioning improves the faradaic currents recorded making the assay more sensitive and reproducible (Hong-Ping and Kwok-Keung, 1996;Piedade et al, 2004), and so this pre-conditioning of the electrode surface was employed in all the voltammetric experiments presented.…”

“…1 (full line), corresponding to the oxidation of guanosine present in the ODN (Oliveira- Brett et al, 1999;Piedade et al, 2004), which will be referred to as peak-dG. The peak current is proportional to the concentration of the guanosine present in the ODN and it also reflects the accessibility of the guanine nucleotides in the ODN to the surface of the electrode since their adsorption is essential for charge transfer to occur (Oliveira-Brett et al, 1999;Palecek et al, 2002).…”