Microarray technology can be used to study the molecular mechanisms of new chemical entities with the aim to develop effective therapeutics. 7-Hydroxyfrullanolide (7HF) is a sesquiterpene lactone that was found to be efficacious in multiple animal models of inflammation by suppression of pro-inflammatory cytokines; however, its molecular mechanism of action remains unclear. We investigated the effects of 7HF on lipopolysaccharide (LPS)-stimulated human peripheral blood mononuclear cells using microarray-based gene expression studies and explored the molecular targets affected. Gene expression profiles and pathway analysis revealed that 7HF potently suppressed multiple inflammatory pathways induced by LPS. More importantly, 7HF was found to inhibit NF-κB related transcripts. These transcripts were further validated using freshly isolated synovial cells from rheumatoid arthritis patients, thus clinically validating our findings. Cell-based imaging and subsequent Western blot analysis demonstrated that 7HF inhibited the translocation of NF-κB into the nucleus by directly inhibiting the phosphorylation of IKK-β. Since the transcription of adhesion molecules is regulated by NF-κB, further investigation showed that 7HF dose-dependently suppressed ICAM-1, VCAM-1 and E-selectin expression on LPS-stimulated endothelial cells as well as inhibited the adhesion of monocytes to LPS-stimulated endothelial cells. Taken together, our results reveal that 7HF possesses NF-κB inhibitory potential and suggest a likely molecular mechanism of its anti-inflammatory activity. © 2010 Elsevier B.V.