This study was conducted to investigate the hypocholesterolemic effect of the hot-water fraction (HW) from cultured mycelia of Cordyceps sinensis in a 5 l fermenter. The composition of HW was mainly carbohydrate (83.9%) and protein (11.8%) on a dry basis, and the carbohydrate of HW consisted of glucose, mannose, galactose, and arabinose in the molecular ratio of 1.0 : 0.8 : 0.5 : 0.1, respectively. In mice fed a cholesterol-free diet and those fed a cholesterol-enriched diet, body and liver weights were not significantly different from those of the controls. The serum total cholesterol (TC) of all mice groups administered HW (150 and 300 mg/kg/d, respectively) with the cholesterol-enriched diet decreased more than in the control group. Among the mice fed the cholesterol-enriched diet, HW also increased the high-density lipoprotein (HDL) cholesterol level, but decreased the very low-density lipoprotein plus low-density lipoprotein (VLDL+LDL) cholesterol level. The changes in HDL- and VLDL+LDL-cholesterol levels consequently decreased the atherogenic value. The results indicate that HW in rats administered a cholesterol-enriched diet decreased the plasma cholesterol level. The 300 mg/kg dose had a significant effect on the serum TC level.