Objective To investigate the immunological mechanism of Bushen Huoxue prescription in the treatment of endometriosis (EM). Methods The EM mouse model was established by injecting endometrial fragments of donor mice into the abdominal cavity of recipient mice. EM mice were randomly divided into model group, gestrinone group and Bushen Huoxue prescription group, with 8 mice in each group. The mice in gestrinone group were intragastrically administered with gestrinone suspension (0.325 mg/kg) twice a week, while the mice in Bushen Huoxue prescription group were intragastrically administered with Bushen Huoxue prescription (43.68 g/kg) daily for 28 d. After treatment, the mice were sacrificed. The growth of ectopic lesions in mice was observed by gross specimens and hematoxylin-eosin staining. The degree of abdominal adhesion in mice was scored by Blauer adhesion scoring system. The expression of fibrotic markers in mouse lesions was observed by immunohistochemical staining. The levels of inflammatory factors in mouse peritoneal fluid were detected by enzyme-linked immunosorbent assay. The changes of macrophage polarization and phagocytosis were detected by flow cytometry. Results Compared with the model group, the volume of ectopic lesions and abdominal adhesion score of EM mice in the Bushen Huoxue prescription group and gestrinone group were significantly decreased (all P＜0.05), the degree of lesion fibrosis was improved, the levels of tumor necrosis factor-α, transforming growth factor-β1 and interleukin-12 in peritoneal fluid were significantly decreased (all P＜0.05), the M2/M1 ratio of peritoneal macrophages was significantly decreased (both P＜0.01), and the phagocytic function of peritoneal macrophages had no significant change. The above indexes in the Bushen Huoxue prescription group had no significant difference compared with those in the gestrinone group. Conclusion Bushen Huoxue prescription can improve the immune microenvironment by regulating peritoneal macrophages for treating EM in mice.