The binding studies of imidacloprid to bovine serum albumin (BSA) were investigated by UV-Vis absorption spectrum, fluorescence spectrum and synchronous fluorescence spectrometry. Under the simulative physiological conditions, fluorescence data revealed the presence of a single class of binding site on BSA and the dynamic quenching constants () were 6.851×104 L.mol-1 and 5.813×104 L.mol-1 at 310 and 315 K, respectively, proving the mode of action of imidacloprid with BSA as a static quenching. In addition, according to the Vant Hoff equation, ΔGθ <0 showed="" the="" combination="" of="" imidacloprid="" and="" bsa="" was="" a="" spontaneous="" process="" h="" sup="">θ <0 and="" s="" sup="">θ> 0, indicated an electrostatic interaction process. At the same time, synchronous fluorescence spectrum of BSA could tell us whether the conformation of BSA was changed by imidacloprid.