Objective: The aim of the research was to examine the potential cytotoxic effect of Lavandula x intermedia essential oil on two breast cancer cell lines: MDA-MB-231 and MCF-7. The assumption was that after exposure of cells to essential oil, the number of metabolically active cells will decrease compared to the control group. Materials and methods: Cytotoxic activity was tested using the MTT test, which serves to assess cell viability. Both cell lines were incubated with lavender essential oil in concentrations of 1, 5, 20, 50, 100 and 200 µg/mL and the effects were observed after 4, 24, 48 and 72 hours. By comparing the absorbance of the resulting formazan product in cells exposed to essential oil and the control group, the percentage of living, metabolically active cells was determined. Results: The results of the investigation of the cytotoxic effect of lavender essential oil on breast cancer cells are shown graphically as a dependence of cell viability and the concentration of essential oil. The best effect on the MDA-MB-231 cell line was observed after 48 hours of incubation with essential oil at a concentration of 200 µg/mL, where cell viability was 33.54%. The greatest effect on the MCF-7 cell line was also observed at a concentration of 200 µg/mL, but after 72 hours of incubation when the cell viability decreased to 47.73%. Conclusion: The essential oil of Lavandula x intermedia shows a cytotoxic effect on breast cancer cell lines MDA-MB-231 and MCF-7. There is a correlation between cytotoxic activity and both incubation time of the cells and the concentration of the essential oil. Greater cytotoxic activity of the essential oil is observed on the cell line MDA-MB-231. IC50 values were achieved on both cell lines: after 48 and 72 hours on the MDA-MB-231 cell line and after 72 hours on the MCF-7 cell line.