1 D- and 2D <sup>1</sup>H NMR spectroscopy (500 MHz) has been used to study the self-association of a bifunctional intercalator, ethidium homodimer (EBH), in aqueous solution. A physical model describing the equilibrium ofdifferent associated forms of the homodimer in solution, including the most probable unfolded, folded conformations, dimer and trimer, has been developed. The magnitudes of equilibrium constants and thermodynamical characteristics of the complexation reactions have been obtained from analysis of the concentration and temperature dependences of proton chemical shifts of EBH molecules. Comparative analysis of the self-association parameters of ethidium homodimer and a monointercalator, ethidium bromide, enables to make some conclusions about structural features and energetic characteristics ofaggregates of
aromatic molecules in solution.