An intratumoral hypoxic microenvironment is frequently observed in solid tumors, including breast cancer. Lutein, a plant-derived compound and non-vitamin A carotenoid, has been demonstrated to possess multiple protective properties including anti-inflammation, anti-oxidative stress and antitumor effects. The main objective of the present research was to elucidate the involvement of lutein in the production of reactive oxygen species (ROS) under hypoxia, the activation of hairy and enhancer of split 1 (HES1), and the proliferation, invasion and migration of breast cancer cells. The human breast cancer cell lines MDA‑MB‑157 and MCF‑7 were exposed to hypoxic conditions and various concentrations of lutein. An MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay was performed to examine cell proliferation, and Annexin V-fluorescein isothiocyanate/propidium iodide staining was performed to analyze the apoptosis ratio. The levels of hypoxia inducible factor-1α (HIF‑1α), NOTCH signaling molecules, HES1 and epithelial-mesenchymal transition (EMT)-associated factors were examined by reverse transcription-quantitative polymerase chain reaction and western blot analysis. Wound healing and Transwell invasion assays were used to detect the invasion and migration of breast cancer cells. Intracellular ROS levels were examined using 2,7-dichlorodihydrofluorescein-diacetate and flow cytometry. The results revealed that cell proliferation was inhibited by lutein in a dose-dependent manner, and the apoptosis ratio gradually increased with lutein treatment under hypoxia as evident from flow cytometry-based analysis. Exposure to lutein inhibited hypoxia-mediated activation of HIF‑1α, NOTCH signaling and HES1 expression, and suppressed the hypoxia-induced expression of EMT-associated factors. Lutein markedly inhibited the invasion and migration of breast cancer cells under hypoxia. Hypoxia-induced production of ROS was also decreased by lutein. Furthermore, the ROS scavenger N‑acetylcysteine also suppressed hypoxia inducible factor 1α and HES1 expression in breast cancer cells during hypoxia, but hydrogen peroxide (H2O2) levels were increased. Taken together, the results of the present study suggested that lutein may be a novel candidate for the chemoprevention of breast cancer. Furthermore, HES1 may be crucial in mediating the involvement of lutein in the suppression of hypoxia-driven ROS-induced breast cancer progression.