Pulsatilla decoction is a famous traditional Chinese herbal formula for clearing heat and treating dysentery of animals or human. To elucidate its mechanism, many active components have been studied, however, the roles of its polysaccharides still remain unclear. This study aimed to explore effects of polysaccharides from Pulsatilla decoction(PPD) on the microvascular endothelial glycocalyx(e GC). The polysaccharides were extracted from PPD by water extraction and alcohol precipitation method. Mice were administered with PPD for 4 wk, and were then anesthetized with ether inhalation and were fixed by cardiac perfusion with gradient concentration alcian blue solution. The jejunum was sampled and jejunal mucosa was prepared for ultrathin sections by routine method and was analyzed by transmission electron microscope. The results indicated that the e GC was observed as a strong electron-dense smooth linear margin or nonuniform conglomerates coating cell membranes, and PPD significantly increased its thickness from(21.85±1.87) to(28.71±3.61) nm and improved its integrity. This study suggested that PPD may express their biological activities and protect against pathogenic factor damages by influencing the e GC. Pulsatilla decoction is a famous traditional Chinese herbal formula for clearing heat and treating dysentery of animals or human. To elucidate its mechanism, many active components have been studied, however, the roles of its polysaccharides still remain unclear. This study aimed to explore effects of Polysaccharides from Pulsatilla decoction (PPD) on the microvascular endothelial glycocalyx (e GC). The polysaccharides were extracted from PPD by water extraction and alcohol precipitation method. Mice were administered with PPD for 4 wk, and were then anesthetized with ether inhalation and were fixed by cardiac perfusion with gradient concentration alcian blue solution. The jejunum was sampled and jejunal mucosa was prepared for ultrathin sections by routine method and was analyzed by transmission electron microscope. The results indicated that the e GC was observed as a strong electron-dense smooth linear margin or nonuniform conglomerates coating cell membranes, and PPD significantly in This study suggests that PPD may express their biological activities and protect against pathogenic factor damages by influencing the e GC.