Objective To investigate the effect and mechanism of Houpu Sanwu decoction on subcutaneous tumors in C57BL/6J mice. Methods Mouse colorectal cancer MC-38 cells were injected subcutaneously into C57BL/6J mice to establish a subcutaneous tumor model of colorectal cancer. The model mice were then divided randomly into model control, Houpu Sanwu decoction(HPSWD) low dose, HPSWD medium dose, HPSWD high dose, 5-fluorouracil (5-FU), and combination groups (5-FU+HPSWD medium dose) (n=6 mice per group). The mice were treated with normal saline and Houpu Sanwu decoction by gavage, or normal saline and 5-FU by intraperitoneal injection once a day for 24 consecutive days. During the intervention period, the mental state,diet, and other general conditions of the mice, as well as changes in tumor volume, were monitored regularly. At the end of the experiment, the subcutaneous tumors in each group were removed and weighed, and pathological features were examined by hematoxylin and eosin staining. Transmission electron microscopy, Western blot, and nanoparticle tracking analysis were used to identify exosomes derived from colorectal cancer tissues. Tumor mRNA and protein expression levels of Rab27a and the exosome markers CD63, TSG101, and ALIX were detected by reverse transcription-quantitative polymerase chain reaction and Western blot, respectively. Results Houpu Sanwu decoction inhibited tumor growth in mice. Tumor growth was inhibited to varying degrees in all the HPSWD dose groups and in the combination group. The HPSWD medium does group showed the greatest anticancer effect, and the combination group showed a synergistic effect with 5-FU. Houpu Sanwu decoction induced tumor necrosis in a dose-dependent manner, with a similar anticancer effect to 5-FU. Houpu Sanwu decoction also inhibited the protein and mRNA expression levels of Rab27a and the exosome-specific markers CD63, ALIX, and TSG101 in mice. Conclusions Houpu Sanwu decoction inhibits tumor growth and the secretion of exosomes, possibly by regulating Rab27a.