This study explores the correlation between human carbonic anhydrase (CA, EC 4.2.1.1) isoforms I and II (hCA I, II) and the inhibitory features of some spirobisnaphthalene derivatives. A group of spirobisnaphthalenes was synthesized and their hCA I and II inhibitory effects was investigated. The Ki values were similar for both CA isoenzymes, the compounds showing good inhibitory activity. Ki values ranged between 1.60 and 460.42 µM for hCA I and between 0.39 and 419.42 µM for hCA II, respectively. The spirobisnaphthalenes derivatives might be useful for designing CA inhibitors belonging to novel chemotypes compared to the highly investigated sulfonamides, sulfamates or coumarins.