The absorption and fluorescence spectrums of four antibacterial quinolones, namely, ciprofloxacin, norfloxacin, enoxacin, and cinoxacin, were studied in the presence of human serum albumin (HSA). Of the three fluoroquinolones studied, ciprofloxacin and norfloxacin were found to bind efficiently to HSA when irradiated with visible light, whereas the third, enoxacin, bound only moderately. On the other hand, cinoxacin, a nonfluorinated quinolone of the first generation, did not show any interaction with HSA. The findings were inferred by monitoring the evolution of the fluorescence spectrums of the solutions as a function of time. A direct relationship between the capacity of the photo-induced defluorination to produce aryl cation intermediates, and the subsequent binding reaction with HSA, was observed and is discussed.