Abstract: To investigate the relationship between the diversities of plasma protein binding rate of Difloxacin (DIF) and its pharmacokinetic in Carassais auratus gibebio, ultrafilteration and high performance liquid chromatographic (HPLC) methods were applied to determine the plasma protein binding rate and pharmacokinetics of DIF in both infected by Aerominas hydrophila bacteria (infected group) and healthy Carassais auratus gibebio (healthy group), and the relationships between the diversities of plasma protein binding rate of DIF and its pharmacokinetic were analyzed in this paper. The results showed that the plasma protein binding rates of the drug in infected group was significantly higher than that of the healthy group at the corresponding time points. The plasma protein binding rates had logarithmic relationships with the total drug concentration and their equations were y=-9.01ln x+74.34(Infected group)and y=-4.81ln x+65.15(Healthy group), while the equations between the plasma protein binding rates and free concentration of the drug were y=-6.36ln x+64.91(Infected group) and y=-4.36ln x+ 60.63(Healthy group) in these two groups. Meanwhile the concentration time curve of the two groups were all in accordance with the two-compartment open model. The absorptions and eliminations of DIF in infection group were slower while its apparent distribution volumes and the area under the concentration-time curves were greater than that of the healthy group. The results indicate that the values of the plasma protein binding rates of DIF could be increased by Aeromonas hydrophila in crucian carp, and this may be one of the reasons that leads to limit distribution of drugs, elimination of slowing down and a long stay in the blood in crucian carp which infected by Aeromonas hydrophila.