Abstract: The adenosine triphosphate (ATP) liposome, prepared with the methods of film dispersion and ion-pairing was evaluated for its therapeutic effect on hypoxic brain damage. The appropriate formulation is adenosine disodium triphosphate, hexadecyl trimethyl ammonium bromide, soybean phospholipid, cholesterol with mass ratio of 1∶1.98∶8∶3. The encapsulation efficiency of ATP liposome was (81.50 ±0.82) % and the loading efficiency was (6.79 ±0.07) %. In vitro release test and rheology test were conducted to investigate the physicochemical properties of ATP liposomes and empty gels respectively. The blank methylcellulose gel, followed with ATP liposome and ATP aqueous solution added to the methycellulose gel, were used for nasal administration in mice respectively. All experiments were approved by the Ethics Committee for Experimental Research in Academy of Military Medical Sciences. After 9 days of continuous administration, ATP liposome hydrogel increased the values of red blood cells and hemoglobin (P<0.01) compared to ATP hydrogel and blank gel. And the ATP liposome hydrogel significantly increased the standard hypoxia tolerance time in mice compared to ATP hydrogel and blank gel after 13 days of nasal administration (P<0.05). The immunohistochemical staining of mice hippocampus for the proapoptotic gene p53 showed that ATP liposome hydrogel was capable of protecting brain tissue in hypoxia. It is indicated that the prophylactic administration of ATP liposome nasal gel can significantly improve the hypoxia tolerance of mice, and the ATP liposome nasal gel was proved to be a promising anti-hypoxia preparation.