Several studies have demonstrated that certain essential fatty acids present a specific cytotoxicity for tumor cells. However, no investigation of this type has been performed on human colon cancer cells to date. This study investigated the effect of gamma-linolenic acid (GLA), eicosapentaenoic acid (EPA) and prostaglandin (PG) E1 on the proliferation and metabolism of three human colon cancer cell lines: HT 29, HRT 18, and CACO 2. GLA, EPA and PGE1 all inhibited the proliferation of the three cell lines, but with a decreasing gradient of sensitivity: HRT 18 > HT 29 > CACO 2, and with different IC50 values. PGE1 was markedly less effective than the other two. GLA and EPA increased lipid peroxidation and membrane fluidity in a dose-dependent manner. The presence of indomethacin did not modify the effects of GLA and EPA. In addition, PGE1 had little effect on membrane fluidity and lipid peroxidation. The antitumoral effect thus does not appear to be mediated by PGE1. Addition of vitamin E decreased the effects of GLA and EPA, which supports the hypothesis of direct action by these fatty acids. In conclusion, while EPA and GLA have an antitumoral effect in vitro, their effect on primary cultures of normal human colon cells must be investigated to determine whether this effect is specific to tumoral cells, as has been observed for other cell types.