This article reports the synthesis and the properties of 5,11,17,23-tetra(t-butyl)-25,27-bis-(ethoxy-1,8-naphthalimide)-26,28-hidroxy-calix[4]arene and the formation of an inclusion compound in organic medium. This functionalized calix[4]arene was conceived as the association of a potential host species with a good fluorophore for optical sensoring purposes. Calix[4]-NI as we will call it, maintains its ‘cone-pinched’ configuration and exhibits typical naphthalimide fluorescence bands in non-polar solvents. Its ability to interact with guest species via hydrogen bonding in its endo-hydrophilic cavity to form inclusion compounds was verified with absorption and fluorescence measurements using N-ethanol-1,8-naphthalimide as guest species, which was projected to fit exactly the host cavity and to interact with its naphthalimide π electrons. For this reason, it was possible to follow the formation of the inclusion compound with electronic spectroscopy.