Electroanalytical and spectroscopic procedures for examination of interactions between double stranded DNA and intercalating drugs

A method is presented for the electroanalytical characterization of interactions of dsDNA with a drug, under conditions that both agents are dissolved in the phosphate buffer solution and both are electroactive. Normal pulse, square wave, differential pulse, and cyclic voltammetries were employed in the measurements of the drug and dsDNA oxidation signals at carbon electrodes. UV–Vis spectroscopy was used as a non-electrochemical method to support the electroanalytical data. An anticancer drug, C-1311 (5-diethylaminoethyl-amino-8-hydroxyimidazoacridinone), has been selected for the examination. Normal pulse voltammetry was particularly useful in showing that under the conditions employed neither dsDNA nor the drug were adsorbed at the electrode surface. Necessary conditions for the appearance of the well-defined dsDNA voltammetric signal (guanine peak) are: rigorous chemical and biological purity in the cell and appropriate purity of DNA. An analysis of the obtained results confirmed that there were two modes of interaction between C-1311 and dsDNA: by intercalation and electrostatically. In the presence of excess NaCl the electrostatic interactions deteriorate. The binding constants (K ₁ and K ₂, respectively) and the number (n) of nucleic base pairs (bp) and the number (m) of phosphate groups (pg) interacting with one molecule of drug have been determined. For strong interactions (intercalation) the values of the binding constant, K ₁, and the binding-site size, n, equal 3.7 × 10⁴ M⁻¹ and 2.1, respectively. For the weak electrostatic interactions the K ₂ and m parameters equal 0.28 × 10⁴ M⁻¹ and 4.7. The intercalation process is rather slow and its rate (the conditions of pseudo-first-order reaction) was estimated to equal 7 × 10⁻⁴ s⁻¹. The possibility of independent determination of both interacting agents was very useful in the study. Figure Intercalation of C-1311 into a dsDNA fragment