Ginkgo biloba L. is among the oldest species of trees on earth. Various parts of the tree have a history of use in prevention of cardiovascular and neurodegenerative diseases. However, most of the current scientific research is focused on the Ginkgo leaf extract. Thus, we tested the effect of Ginkgo nuts on lipid metabolism as a therapeutic for cardiovascular diseases. Four preparations of Ginkgo nuts (whole nuts, their methanol extract and its lipid and water soluble fractions) were tested using Human Hepatoma cells (Hep G2) and in vivo mouse feeding study. Ginkgo nut preparation may modulate serum cholesterol levels by modulating Apolipoprotein B secretion as well as low-density lipoprotein receptor in the liver. From mouse feeding trial, Ginkgo nut supplementation, as whole nuts, reduced the levels of hepatic cholesterol while increasing serum cholesterol compared to control. The lipid soluble fraction was responsible for decreased hepatic cholesterol, while the water soluble fraction may contribute to increased serum cholesterol. The lipid soluble fraction of Ginkgo nut may have potential to be used for the purpose of prevention of cardiovascular diseases. This is the first report investigating the effects of Ginkgo nuts on cholesterol metabolism.