Changes in composition of fatty acids under various culture conditions were studied in branches or sporelings of four species of Gracilaria. The two dominant fatty acids in all four species were the saturated palmitic acid (16 : 0) and the polyunsaturated arachidonic acid (20 : 4omega6). In G. tikvahiae sporelings and apices cultured at 10, 15, and 18-degrees-C there were higher levels of 18 : 1omega9 and 20 : 4omega6 than in those grown at 20 and 26-degrees-C, responses similar to that reported in hardening of flowering plants. In contrast, G. verrucosa, G. domingensis, and G. conferta did not show an increase in unsaturation of fatty acids when cultured in lower temperatures. Thus, the broader geographical distribution of G. tikvahiae and G. verrucosa compared to the narrower distribution of the other two species also could not be used to explain the differences in fatty acid changes with regard to temperature. There were no differences in fatty acid composition for any of the four species grown under increasing levels of photon fluence. Low salinity (15 parts per thousand) resulted in an increased level of 20 : 4omega6 fatty acid only in G. tikvahiae sporelings. Branches of G. tikvahiae cultured in seawater enriched with sodium nitrate had higher levels of 20 : 4omega6 than those in sterile seawater. A positive relationship between weekly growth rates and unsaturated fatty acids occurred in G. verrucosa and G. domingensis sporelings but not in G. conferta tips. There was no common pattern in fatty acid alterations with relation to temperature, salinity or nitrogen enrichment.