Inflammation is one of the basic pathological processes due to the adaption of immune system against infection or irritation. Since non-steroidal anti-inflammatory drugs (NSAIDs) cause many side effects, investigation of anti-inflammatory natural products promising to provide novel therapeutic agents. Here, Launaea sarmentosa (Willd.) Kuntze, a medicinal herb applied to treat inflammation diseases, was investigated for its antiinflammatory components to find new therapies for inflammatory syndromes. This study indicated that ethyl acetate fractional extract reduced the expression of pro-inflammatory cytokines, representing the highest activity for both NO radical-scavenging and NO secretion inhibition during LPS-stimulated RAW264.7 macrophages. Besides, five anti-inflammatory compounds, including succinic acid, quercetin, 2(4-hydroxyphenyl)acetic acid, luteolin-7-O-β-D-glucopyranoside, and quercetin-3-O-rutinoside, were isolated and elucidated the structure according to 1D and 2D- NMR. Among these compounds, succinic acid and 2(4-hydroxyphenyl)acetic acid were first reported in this species. Moreover, this study indicated that the presence of these compounds, typically quercetin and luteolin-7-O-β-D-glucopyranoside, enhanced anti-inflammatory ability via deactivation of NFкB/MAPK pathway to mitigate the expression of IL-6. Hence, this study contributed the initial evidence of antiinflammatory constituents from Launaea sarmentose and highlighted an approach to discovering natural items or phytotherapeutic agents.