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In-vitro evaluation of the interaction of anticancer drug mitoxantrone with anionic surfactant sodium dodecyl sulfate (SDS), was performed in physiological conditions (phosphate buffer, pH 7.4) by spectral (UV–vis absorption) and electrochemical (cyclic and linear voltammetry) methods.The stoichiometry of interaction, the binding constants and diffusion coefficients of free and bound drug were determined. The partition coefficient of mitoxantrone between aqueous phase and SDS micelles was calculated, and the results indicated that it is strongly dependent on the drug concentration. Both absorption and cyclic voltammetry results have outlined two distinct processes depending on the surfactant concentration: process I in premicellar range, assigned to the interaction of the drug with the surfactant molecules, when electrostatic forces play an important role in the formation of the mitoxantrone-SDS complex with a defined stoichiometry; and process II in micellar range, when the surfactant micelles are formed and the drug is encapsulated in micelles in monomer form. The spectral results have indicated that the drug is most probably located in the micelle surface layer, both electrostatic and polar interactions playing important role in the binding of drug to SDS micelles. Possible application of this system as a micellar drug–carrier was also considered.