The effects of phosphatidic acid (PA) and lysophosphatidic acid (LPA) dispersed in serum-free media on the growth of Chinese hamster ovary (CHO) cells were investigated. After cells were incubated in serum-free media supplemented with PA or LPA for 3 d, the cellular growth was evaluated by the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay. Supplementing PA from egg yolk lecithin to basal synthetic media such as alpha-MEM (minimum essential medium, alpha modification) markedly promoted the growth of CHO cells. PA from egg yolk lecithin also enhanced the cell growth in a serum-free medium containing insulin and transferrin. When PA analogues with different acyl chains of C14-18 were compared, PA with unsaturated acyl chains, dioleoyl-PA (C18 : 1), was most effective far stimulating the growth of CHO cells. Similar results were obtained when LPA was examined. These results suggest that PA and LPA, especially those with unsaturated acyl chains, are promising growth-promoting supplements for use as constituents in a low-protein serum-free medium.