Five different methods were tested and compared to prepare danofloxacin mesylate liposomes, the ammonium sulfate gradient method with freeze-thawing steps was validated as the best one; the optimal preparation condition confirmed by orthogonal experiment was as follows： EPC-CH ratio was 3 ： 2 and 2.6% SA was added to gain the positive electricity; drug-lipoid was 2 ： 5, the concentration of ammonium sulfate was 250 mmol·L-1, water-oil ratio was 1：5, and they were incubated at 35℃ for 15 min. The prepared liposome products were ivory white semitransparent suspension, the electron microscope appearance was intact and globular or globular-like vesicles with uniformed distribution; the particle size was centralized from 3 to 7 gm, zeta-electric potential valued＋ （15.92＋1.49） mV, pH valued 6.02~0.09; HPLC method was established in quantitative analyses of danofloxacin and reverse dialysis with RP-HPLC method was validated for determination of entrapment efficiency. The entrapment efficiency results were all above 90%. They were stored at 4℃ with satisfied stability. Six months later, the appearance, characters and entrapment efficiency were almost with no change