Serum albumin is the most abundant protein in plasma. It can bind with many intrinsic and extrinsic materials. The study of the interaction between serum albumin and drugs is a very important task in life science and chemistry. Quinolone drug is a kind of antibacterial drugs which have been used widely in clinical medicine, but its pharmacology and toxicology still have to be studied further. Sinafloxacin is a new quinolone antibiotics. The interaction between sinafloxacin and bovine serum albumin (BSA) at different temperatures and pH was investigated by fluorescence and UV absorption spectroscopy. The experimental results demonstrate that the fluorescence quenching of BSA by sinafloxacin is a result of the formation of sinafloxacin-BSA complex and the quenching mechanism is mainly static quenching. The interaction association constants of BSA and sinafloxacin were determined from the double reciprocal Lineweaver-Burk plot. The binding distance(r = 3.64 Rnm) and energy transfer efficiency (E = 0.163) between donor (BSA) and acceptor (sinafloxacin) were obtained based on Förster's non-radiative energy transfer theory. There is a strong interaction between sinafloxacin and BSA. From thermodynamic coordination it can be judged that the binding force between sinafloxacin and BSA is mainly electro-static force. The effect of sinafloxacin on the conformation of BSA was analyzed by synchronous fluorescence spectrometry and three-dimensional fluorescence spectrometry. The emission maximum of tyrosine residues does not show a significant shift, while the small blue shift of tryptophan residues indicates that the hydrophobicity of microenvironment was increased. In addition, in the plasma, there are some metal ions, which can participate in many important vital actions and affect the reactions of the drugs with the serum albumins. So the effect of Cu(II), Fe(III), Zn( II) and Mg(II) on the binding constant of sinafloxacin to BSA was also discussed and it was shown that the interaction between BSA and sinafloxacin was decreased. In short, the results offer a reference for the studies on the biological effects and action mechanism of sinafloxacin with albumins in vivo.