The aim of this work was to evaluate the preparation of matrine ethosome and the percutaneous permeation in vitro and the anti-inflammatory activity in vivo in the rat skin. The matrine ethosomes were prepared by the ethanol injection-sonication method. The particle size of the ethosomes was measured by a laser particle-size analyzer, and the entrapment efficiency was detected by ultracentrifugation. The anti-inflammatory activity in vivo of the matrine formulations was determined by a reflection spectrophotometer. In this study, we found that the average particle size of matrine ethosomes was in the range of 50-200 nm with a narrow distribution, and the entrapment efficiency was in the range of 40-90%. Compared with other formulations, matrine ethosomes had the largest 24-hour accumulative permeation quantity (60.5%) and with no permeation lag time. Matrine ethosomes were able to make the induced erythema disappear more rapidly than the nonethosomes formulations of matrine. This study reveals that the average particle size of matrine ethosomes decreases with the increase of ethanol concentration and increases with the increase of phospholipid concentration, while the entrapment efficiency increases with the increase of the concentration of both ethanol and phospholipid. Matrine ethosomes can increase the percutaneous permeation of matrine in the experiment in vitro and improve the anti-inflammatory activity of matrine in vivo in rat skin.