In this study, factors affecting anti-inflammatory effect of chitooligosaccharides (COSs) in lipopolysaccharides (LPS)-induced RAW264.7 macrophage cells were investigated. The inhibition of NO secretion by COSs revealed that 90-COSs (90% N-deacetylation) significantly inhibited NO secretion than those of 50-COSs (50% N-deacetylation), and 90-HMWCOS (5000-10,000Da) in the 90-COSs showed the highest inhibition activity. Furthermore, 90-HMWCOSs also found to inhibit LPS-stimulated production of PGE(2), TNF-alpha and IL-6, as well as the expression of iNOS, COX-2, TNF-alpha, and IL-6. These results suggested that 90-HMWCOS may have anti-inflammatory effect via down-regulation of transcriptional and translational expression levels of TNF-alpha, IL-6 and iNOS and COX-2.