Amphiphilic diblock copolymers from ethylene oxide and 1,2-butene oxide or 1,2-hexene oxide monomers were synthesized by anionic ring-opening polymerization via the monomer activation methodology. Well defined block copolymers with predictable molecular weights and low polydispersities have been produced by starting the block copolymerization either from the hydrophilic or the hydrophobic monomer. The self-assembly behaviour of poly(ethylene oxide-b-hexene oxide) diblocks, containing segments with the more hydrophobic butyl side groups, was examined in aqueous solutions as a function of the block copolymer composition. Light scattering, atomic force microscopy and fluorescence spectroscopy measurements revealed the presence of self-organized nanostructures. The structure and properties of the nanoassemblies depend on the diblock copolymer composition and the solution preparation protocol.