The structural characteristics of alkali-extracted jujube polysaccharides and their effects of in vitro fermentation on fecal microorganisms were evaluated in this study. Scanning electron microscopy revealed that AJP1 and AJP2 polysaccharides had different morphological characteristics, and their molecular weights were 75465 Da and 268007 Da, respectively. The uronic acid contents of the two polysaccharides were 79.68% and 77.64%. Infrared spectroscopy showed that two alkaline-extracted polysaccharide groups had characteristic absorption peaks of acidic polysaccharides such as C=O and C-H. After 48 h of in vitro fermentation, the total sugar consumption rates of the AJP1 and AJP2 treatments reached 65.40% and 56.24%, respectively. During the fermentation process, the pH values of the polysaccharide-treated groups were significantly lower compared to the control group. In addition, polysaccharides could promote the abundance, diversity, and composition of gut microbiota. These results showed that alkali-extracted jujube polysaccharides could regulate the intestinal microflora composition in feces, and inhibit the growth of pathogenic bacteria Peptoniphilus and Desulfovibrio while promoting the growth of beneficial bacteria such as Megamonas and Allisonella. This study provides a reference for further developing jujube polysaccharide functional food products and the study of the structure-activity relationship between alkali-extracted polysaccharides and intestinal flora.