AbstractExperiments were conducted to determine the effect ofβ‐carotene on human colon cancer cells in vitro.β‐Carotene solubilized in tetrahydrofuran (THF) was determined to be cytotoxic for three different cell lines: LS 180, SW620, and HCT‐15. The number of LS 180 and SW 620 cells surviving treatment with 2.9μ?β‐carotene was significantly reduced relative to THF‐treated cells, and a similar reduction was achieved in HCT‐15 cells with use of 5.8μ?β‐carotene. These concentrations are in the range achieved in serum of individuals supplemented withβ‐carotene at 30 mg/day. There was noβ‐carotene cytotoxicity in the concentration range that characterizes serum of unsupplemented individuals. Vitamin ? at>200μ? was not cytotoxic and at higher concentrations slightly stimulated proliferation of all three cell lines. Exposure of cells to vitamin ? did not diminish the cytotoxicity ofβ‐carotene, suggesting that the toxic effect ofβ‐carotene is not due to prooxidant activity. Percent cytotoxicity was increased by extending the duration of exposure of cells toβ‐carotene. Interestingly,β‐carotene cytotoxicity decreased with increasing cell density. This density‐dependent toxicity was attributable to a higherβ‐carotene concentration per cell for cells plated at lower densities. Thus toxicity ofβ‐carotene for colon cancer cells is dose, time, and cell density dependent and occurs in vitro at concentrations that can be achieved safely in humans.