Objective To explore the effect of decoction of Euphorbia fischeriana Steud. and jujuba (DEFSJ) against estrogen receptor (ER) negative (-) and ER positive (+) breast cancer via the PI3k / Akt pathway, and to provide a reference for the targeted treatment of breast cancer. Methods DEFSJ extract was prepared and analyzed using UHPLCTriple Quad. DEFSJ containing serum (CS) was prepared via a serum pharmacology method . Different concentrations of DEFSJ-CS were applied to (ER-) MDA-MB-453 and (ER+) MCF-7 breast cancer cells in vitro for 48 h. The distribution of cells in different stages of the cellular cycle was evaluated using a Flow cytometer. DNA ladder assay was used to assess the degree of apoptosis, and the expression of PI3k / Akt pathway-related proteins was evaluated by Western blot assay. The expression of FoxO3a, FoxO1a, and Bim mRNA was detected by Real-time quantitative PCR. Nuclear transposition of FoxO3a protein was analysed using a confocal laser microscopy. Results Five batches of DEFSJ extract were analyzed using UPLC, and the result showed that the preparation technology was feasible and the quality was controllable, ensuring the accuracy of the pharmacological experiment result. DEFSJ-CS blocked cells in the G2 / M phase (P<0. 05, P<0. 01).Cells treated with DEFSJ-CS displayed the typical apoptotic ladder in the DNA ladder experiment. Compared with the negative control cells, the DEFSJ-CS group cells had decreased protein expression of p-PI3k, p-Akt, p-FoxO3a, and pFoxO1a (P<0. 05, P<0. 01); increased protein expression of Bim (P<0. 05); decreased mRNA expression of FoxO3a and FoxO1a ( P<0. 05,P<0. 01); increased mRNA expression of Bim ( P<0. 05,P<0. 01); and enhanced nuclear translocation of FoxO3a protein. The data showed that DEFSJ-CS had a stronger effect on (ER-) MDA-MB-453 cells than (ER+) MCF-7 cells. Conclusions The regulatory effect of DEFSJ extract on anti-breast cancer involves the PI3k / Akt pathway, and the effect varies with phenotypic differences.