Abstract: The mechanism of the interaction between bovine serum albumin(BSA) and pyrano[4,3-b] pyran derivative,2-amino-7-methyl-4-(4-nitrophenyl)-5-oxo-4,5-dihydropyrano[4,3-b] pyran-3-carbonitrile,was studied by means of fluorescence spectroscopy under physiological conditions.The results show that this pyrano[4,3-b] pyran derivative observably quenches the fluorescence of BSA in a static quenching mode.The binding constants and the numbers of binding sites at different temperatures were obtained and the effects of some metal ions on the binding constant of the derivative and BSA were also discussed.According to the thermodynamic parameters,the main binding forces between them are hydrogen bonding and Van der Waals interactions.Furthermore,the binding distance and transfer efficiency of the interaction energy between BSA and the pyrano[4,3-b] pyran derivative were obtained according to the theory of Förster non-radiation energy transfer.The effect of this pyrano[4,3-b] pyran derivative on the conformation of BSA was analyzed via the synchronous fluorescence spectra and three-dimensional fluorescence spectra.