The purpose of this study was to investigate the effect on the achievement of knowledge by animation-based learning program(ALP) in environmental education. The topic covered by ALP was "meaning and kinds of alien animals and plants". Four classes were selected and divided into two groups. The one was control group, and the other was treatment. 121 female the eighth graders(60 control and 61 ALP students) were piloted. Whereas control students learned using Powerpoint presentation-based materials, ALP students learned using web-based flash animations and animated database with images and movie clips. The score of knowledge achievement of ALP students was compared to control group students using ANCOVA, and then the score of ALP students was significantly higher than that of control group students(p<0.05). The ALP using animations and animated database was effective on the understanding of the alien animals and plants.