Abstract: The binding characteristics of 3-amino-4-dimethylamino-N-butyl-1,8-naphthalimide (ADBN) with bovine serum albumin (BSA) were investigated by fluorescence spectroscopy, UV-Vis adsorption spectroscopy, IR and circular dichroism (CD). Fluorescence spectroscopy and UV-Vis adsorption spectroscopy revealed that the mechanism of fluorescence quenching of BSA by ADBN was static quenching. The thermodynamic parameters indicated ADBN boned with BSA through hydrogen bonds and van der Waals forces. The binding distance of ADBN with BSA was 4.08 nm. The results of synchronous fluorescence, CD and IR spectra proved that the conformation of BSA changed in the binding process.