Killing of Giardia lamblia by fresh human milk requires the presence of bile salt, a known activator of bile salt-stimulated lipase, the major lipase in human milk. Purified enzyme did not kill the parasite even in the presence of activator unless milk lipids were also present in the reaction mixture. Free fatty acids had a marked giardiacidal effect, a phenomenon supporting the view that fatty acids, released during hydrolysis of milk triglycerides, are responsible for the killing of G. lamblia by human milk. Bile salt-independent lipolysis took place in milk during storage at 4 C. This lipolysis correlated strongly with activity of lipoprotein lipase, also present in human milk. During such storage, raw human as well as bovine milk developed giardiacidal activity that could be prevented by inactivation or inhibition of the milk lipases by pasteurization or addition of eserine to the milk, respectively, before storage.