Echinacea preparations are widely used herbal medicines for the prevention and treatment of colds and minor infections. There is little evidence for the individual components in Echinacea that contribute to immune regulatory activity. Activity of an ethanolic Echinacea extract and several constituents, including cichoric acid, have been examined using three in vitro measures of macrophage immune function - NF-kappaB, TNF- alpha and nitric oxide (NO). In cultured macrophages, all components except the monoene alkylamide (AA1) decreased lipopolysaccharide (LPS) stimulated NF-kappaB levels. 0.2 microg/ml cichoric acid and 2.0 microg/mL Echinacea Premium Liquid (EPL) and EPL alkylamide fraction (EPL AA) were found to significantly decrease TNF-alpha production under LPS stimulated conditions in macrophages. In macrophages, only the alkylamide mixture isolated from the ethanolic Echinacea extract decreased LPS stimulated NO production. In this study, the mixture of alkylamides in the Echinacea ethanolic liquid extract did not respond in the same manner in the assays as the individual alkylamides investigated. While cichoric acid has been shown to affect NF-kappaB, TNF-alpha and NO levels, it is unlikely to be relevant in the Echinacea alterations of the immune response in vivo due to its non- bioavailability - i.e. no demonstrated absorption across the intestinal barrier and no detectable levels in plasma. These results demonstrate that Echinacea is an effective modulator of macrophage immune responses in vitro.