The effects of substituting whey protein isolate (WPI) for butter in the preparation of butter sponge cake were determined by objective and subjective tests. The specific gravity of cake batter, the cooking loss and moisture content of cake were all decreased with increasing amounts of WPI, whereas specific loaf volume was increased. With increasing WPI content, redness of crust and crumb, as well as lightness of crumb were increased, whereas lightness of crust, as well as yellowness of crust and crumb were all decreased. Hardness, chewiness, gumminess, adhesiveness and fracturability were increased significantly with increasing amounts of WPI, however, cohesiveness, springiness, and resilience were decreased. In the sensory evaluation, 20% WPI-substituted cake displayed scores similar to those of control. These results suggest that substitution of 20% WPI could be the best ratio for the preparation of butter sponge cake.