Norfloxacin (NFX) is a fluorescent antibiotic of broad bacteriological spectrum, member of the fluoroquinolone (FQ) family. The interaction of FQs with metal ions can increase the bactericidal action and act against antibiotics bacterial resistance. Several examples of mixed-ligand norfloxacin Cu (II) complexes can be found in the literature. Understanding the molecular mechanisms of interaction of these antibiotics with different cell components and with gold nanoparticles as drug transporters is extremely important. To clarify these interactions, we used different spectroscopic techniques, such as steady-state and time-resolved fluorescence spectroscopy, UV-Visible light absorption, and electron paramagnetic resonance (EPR). The interaction of NFX and the ternary complex Cu(II):L:NFX, where the L is the ligand 1,10-phenanthroline (Phen) or 2,2-bipyridine (Bipy), with micelles and small unilamellar liposomes of egg phosphatidylcholine (PC) was studied using steady-state and time-resolved fluorescence spectroscopy. The stability of the ternary complexes formed in micelles was studied, and stability constants were obtained inside SDS micelles, which showed values much larger than in buffer. The EPR spectra gave further details on the structure of the complexes, and confirmed the formation of the ternary complex inside the micelles. The interactions of FQs and their copper complexes with PC liposomes prepared with different surface densities of negative electrical charge were studied. In the study of the interaction of FQs with gold nanoparticles synthesized by laser ablation (nanocomposites, AuNCs), NFX showed greater changes than FQs ciprofloxacin (CFX) and levofloxacin (LFX) in both absorbance and fluorescence. The results suggest changes in the surface refractive index of the AuNCs and/or cluster formation, as result of the interaction with the drug. A slow but significant quenching of NFX fluorescence was also observed, with no change in peak position, indicating that NFX maintains its initial state of protonation by binding to the surface of the AuNCs. Release of FQs attached to the surface of AuNCs by thiols has also been observed, which causes partial recovery of FQ fluorescence. Finally, as the anionic surfactant SDS showed to be promising in the interaction with NFX, compared to cationic and neutral surfactants, and because the AuNCs are stable in SDS, we studied the interaction of NFX with AuNCs synthesized in the presence of SDS and, for comparison, with AuNCs placed in SDS solution after synthesis.