We extracted and purified three polysaccharides fromEchinacea purpureausing pectinase-assisted extraction to obtain crude preparations and optimized the method using an orthogonal analysis. We obtained three polysaccharide fractions (EPPS-1, -2 and -3) using DEAE ion exchange and gel filtration chromatography. The homogeneity of the fractions was confirmed using high performance gel permeation chromatography. EPPS-3 administered to mice in a LPS-induced septicemia model effectively counteracted the effects of LPS resulting in significantly less lung damage. This trend was also seen in the serum and lung cytokine levels where EPPS-3 significantly decreased the levels of TNF-alpha and IL-6 and increased IL-10. Particularly, we fully characterized the structure of the EPPS-3 polysaccharide using a series of technologies. This polysaccharide structure was mainly composed of -> 4)-alpha-Glcp-(1 ->, -> 4)-alpha-Galp-(1 ->, T-alpha-Araf-(1 ->, -> 3,4)-beta-GalpA-(1 -> glycosidic linkages at a certain proportion. In sum, EPPS-3, with a clear structure, has potent anti-inflammatory activities and is a candidate for further development as an anti-inflammatory agent for clinical development.