This study delved into the effects of Chuanxiong Rhizoma polysaccharides (CRP) on immune regulation and intestinal flora in mice. CRP was first experimentally prepared and its chemical composition was analyzed. Male Kunming mice were chosen to establish an immunosuppression model through intraperitoneal injection of cyclophosphamide. After 12 days of intervention with CRP, serum hemolysin and other immune function related indexes were determined, and 16S rRNA gene high-throughput sequencing technology was used to determine the species composition, abundance and diversity of intestinal flora. In addition, combined with bioinformatics analysis methods, we further explored the metabolic pathways and correlation analysis of the intestinal flora community structure. The molecular weight of CRP was found to be in the range of 45.0 to 220.0 kDa, suggesting potential immune activity. Further experiments showed that mice treated with CRP improved immune organ index and morphology, serum hemolysin and cytokine levels, as well as macrophage phagocytic function, which pointed to a favorable trend in immune balance regulation. Moreover, the abundance and diversity of intestinal flora partially recovered, as beneficial bacteria, including Lachnospiraceae and Akkermansia, increased in relative abundance. And the richness of intestinal microbiota was effectively improved from the perspective of diversity index. The correlation heatmap results also showd that these changes in intestinal microbiota were closely related to the improvement of immune function in mice. These findings suggest the immunomodulatory effect of CRP may be related to  the regulation of intestinal microbiota structure and the improvement of the intestinal microecological environment.