Rosemary (Rosmarinus officinalis L.) has been used in folk medicine to treat headaches, epilepsy, poor circulation, and many other ailments. It was found that rosemary could act as a stimulant and mild analgesic and could reduce inflammation. However, the mechanisms underlying the anti-inflammatory effects of rosemary need more study to be established. Therefore, in this study, the effects of rosemary on the activation of nuclear factor kappa beta (NF-kB) and mitogen-activated protein kinases (MAPKs), the expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2), and the production of nitric oxide (NO), prostaglandin E(2) (PGE(2)), and cytokine in lipopolysaccharide (LPS)-stimulated RAW 264.7 cells were investigated. A methanol extract of rosemary and its hexane fraction reduced NO generation with an IC(50) of 2.75 and 2.83 μg/ml, respectively. Also, the methanol extract and the hexane fraction inhibited LPS-induced MAPKs and NF-kB activation associated with the inhibition of iNOS or COX-2 expression. LPS-induced production of PGE(2) and tumour necrosis factor-alpha (TNF-α) were blocked by rosemary. Rosemary extract and its hexane fraction are important for the prevention of phosphorylation of MAPKs, thereby blocking NF-kB activation, which in turn leads to decreased expression of iNOS and COX-2, thus preventing inflammation.