The non-covalent interaction of a series of new water-soluble benzo- and tetrahydrobenzofurocoumarins with salmon testes DNA has been studied using flow linear dichroism, circular dichroism, contact fluorescence energy transfer and ethidium bromide displacement assay. The new derivatives are characterised by having an alkyl amino side chain protonated at physiological pH; this fact strongly enhances the solubility in aqueous media and the affinity for the macromolecule. The results show significant difference in the affinity and the mode of binding among the examined compounds depending on the nature of the fourth condensed ring and the position of the alkylamino side chain. Benzofurocoumarins derivatives bind DNA by undergoing intercalation inside the duplex macromolecule, whereas tetrahydrobenzofurocoumarins derivatives show a substantial tilt relative to the base planes. Molecular modeling studies have been performed to characterise in detail the intercalation mechanism of these benzofurocoumarins to DNA.