Cephalonoplos segetum has been used as an herbal remedy, and is considered to have anti-inflammatory potential. However, its biological mechanism in this treatment process remains unknown. Therefore, the anti-inflammatory activity of the ethyl acetate fraction of C. segetum extracts (CSE-EA), more active than C. segetum extracts (CSE) in murine macrophages, was investigated. Production levels of nitric oxide (NO), prostaglandin $E_2$ ($PGE_2$), tumor necrosis factor-${alpha}$ (TNF-${alpha}$), and interleukin-$1{eta}$ (IL-$1{eta}$) by lipopolysaccharide (LPS)-induced RAW 264.7 macrophages were measured by ELISA. In addition, protein expression levels of inducible NO synthase (iNOS) and cyclooxygenase (COX)-2, and the phosphorylation of mitogen-activated protein kinases (MAPKs) in the LPS-induced macrophages were investigated by Western blotting. The CSE-EA (50, 100 or 200 ${mu}g/mL$) significantly inhibited NO, $PGE_2$, TNF-${alpha}$, and IL-$1{eta}$ production in LPS-induced macrophages in a dose-dependent manner with 50% inhibitory concentration values of 80.4, 104.7, 91.3, and 46.7 ${mu}g/mL$, respectively. Similarly, CSE-EA reduced protein expression of iNOS and COX-2 and led to the attenuated activation of kinases ERK1/2 and JNK in the macrophages. Results of the present study suggest that the anti-inflammatory effects of CSE-EA are likely due to the down-regulation of NO, $PGE_2$ TNF-${alpha}$, and IL-$1{eta}$ and the reduced expression of iNOS and COX-2 via suppression of MAPK signaling pathways in LPS-induced murine macrophages.