Objective: The inhibition of agar-oligosaccharides on lipopolysaccharide (LPS) -induced inflammatory response in macrophage RAW264.7 and its mechanism were investigated in this study. Methods: The molecular structure of agar-oligosaccharides was analyzed by Fourier transform infrared spectroscopy， nuclear magnetic resonance and high performance liquid chromatography. The inflammatory response model was established using LPS-induced macrophage RAW264.7， and the effects of agar-oligosaccharides on cell viability， intracellular nitric oxide(NO)， tumor necrosis factor-α (TNF-α) and reactive oxygen species (ROS) levels of RAW264.7 were determined. Furthermore， the expression and phosphorylation levels of c-Jun N-terminal kinase (JNK)， extracellular signal-regulated kinase (ERK)， and protein kinase p38 in the mitogen-activated protein kinase (MAPK) signaling pathway were analyzed by Western blot. Results: Molecular structure identification results of Agar-oligosaccharides displayed galactose sugar ring structures with molecular weights ranging of 1065.5-1308.2 u. Agar-oligosaccharides could increase the cell viability of macrophage RAW264.7 with LPS-induced inflammatory response， decreased the intracellular NO， TNF-α and ROS levels in a concentration-dependent manner at 0~250 μg/mL， and inhibited the increase of LPS-induced phosphorylation levels of MAPK family kinases JNK， ERK， and p38. Conclusion: Agar-oligosaccharides can inhibit LPS-induced inflammatory response in macrophage RAW264.7， which may be related to the hindering of MAPK signal transduction.