Soy protein preparations were reported to have hypocholesterolemic actions in experimental animals and humans, while the active components and the mechanism by which this occurs are not clarified yet. The objective of this study is to address these issues by using exogenously hypercholesterolemic rats which are susceptible to dietary cholesterol. Two groups of five rats (male, 12-wk-old) were fed on AIN 93G-based diet with soy protein isolate (SPI) or ethanol-extracted SPI (EE-SPI) for 2 wk. EE-SPI was prepared by ethanol extraction to remove isoflavones and other components. Concentrations of serum and liver total cholesterol were lower in rats fed SPI than in those fed EE-SPI. The abundances of mRNA for 7alpha-hydroxylase and low density lipoprotein receptor in the liver were lower in EE-SPI group than those in SPI group. These results suggest that the ethanol extract from SPI has a factor(s) to alleviate hypercholesterolemia by increasing the removal of cholesterol from serum through the receptor pathway and then from liver through enhancement of bile acid synthesis.