Abstract: Eucommia ulmoides Oliv. seed oil liposomes were prepared by a combined technology of ethanol injection and ultrasound, and the stability of liposomes was also evaluated. The optimized conditions and the encapsulation efficiency of E. ulmoides seed oil liposomes were investigated by Plackett-Burman design, the steepest ascent design and response surface methodology.The results showed that the optimal conditions selected were as follows:the mass ratio of phosphatidylcholine to sitosterol (X₁)3.1:1, the mass ratio of phosphatidylcholine to E. ulmoides seed oil (X₂) 4:1, dosage of tween-80(X₃) 20%, phosphate buffer solution pH value (X₄) 6.9, ultrasonic time (X₅) 13 min, ultrasonic power (X₆) 180 W. Under this condition, the actual encapsulation efficiency, average particle size, average potential and poly dispersity index (PDI) of E. ulmoides seed oil liposomes were 75.26%, 137.4 nm, -19.3 mV and 0.219, respectively. The particles showed irregular oval shape and smooth and complete morphology via the transmission electron microscopy observation. The stability experiments showed that E. ulmoides seed oil liposomes were significantly affected by the temperature, and the stability of all indexes at 4℃ was better than that at 25℃ and 40℃.When the E. ulmoides seed oil liposomes were stored at 4℃ for 15 days, the encapsulation efficiency decreased from 75.26% to 61.45%, the pH value decreased slightly, the average particle size increased from 137.4 nm to 160.9 nm, and malondialdehyde (MDA) increased from 0.023 2 mg/g to 0.033 1 mg/g.