Objective To find an ideal animal model of acute respiratory distress syndrome (ARDS) through investigating the characteristics of three "two-hit" animal models of ARDS. Methods Forty-eight SD rats were randomly divided into 4 groups:Control group[2.5 mL/kg normal saline (NS) i.v. given at 0 min and 30 min]; OA+OA group[0.5 mL/kg oleic acid (OA) i.v. given at 0 min and 30 min]; LPS+LPS group[2.5 mg/kg lipopolysaccharide (LPS) i.v. given at 0 min and 30 min]; and OA+LPS group[0.5 mL/kg OA i.v. given at 0 min and 2.5 mg/kg LPS, i.v. given at 30 min]. The samples were collected at 5 h after the second drug injection. White blood cells count (WBC), polymorphonuclear leukocyte ratio (PMN%), total protein concentration, tumor necrosis factor α (TNF-α) level in bronchoalveolar lavage fluid (BALF), arterial blood gas analysis and lung wet-dry weight ratio (W/D) were measured, respectively. Pathological changes in the lung tissues were observed and histological scores were evaluated. Results Compared with those in the control group, PaCO₂, WBC, PMN%, total protein concentration and TNF-α levels in BALF were significantly increased, while PaO₂ was dramatically decreased (P<0.01) in the OA+OA, LPS+LPS and OA+LPS groups. The levels of protein concentration in BALF and lung W/D ratio in the OA+LPS group were significantly higher than these in the LPS+LPS group (P<0.05 for all), but had no statistically significant difference compared with these in the OA+OA group. The levels of WBC, PMN% and TNF-α in BALF in the OA+LPS group were significantly higher than those in the OA+OA group (P<0.05), but not significantly different from those in the LPS+LPS group. The most typical pathological changes and the highest pathological scores were found in the OA+LPS group. Conclusions All the three different Methods including OA+OA, LPS+LPS, and OA+LPS can be used to establish "two-hit" animal models of acute respiratory distress syndrome. The "two-hit" animal model of acute respiratory distress syndrome induced by OA+LPS is more closer to clinical ARDS and is useful for studies on the pathophysiology of ARDS, and is an ideal "two-hit" animal model of ARDS.