Acanthamoeba castellanii were incubated in vivo with 1(-14)C linoleic and 1(-14)C-alpha-linolenic acids. The incorporation of the acids into lipid fractions was studied. Labeling was found mainly in triglycerides and phospholipids. Homogenized cells and subcellular fractions separated by centrifugation were incubated with 1-14C-linoleic. 1-14C -alpha-linoleic and 1-14C eicosa-8,11-dienoic acids in the presence of NADH, ATP, and CoA. Different metabolic routes were demonstrated. omega3 and omega6 desaturases of the vegetal type, as well as a delta6 desaturation of alpha-linolenic acid of the animal type were present. The supernatant of 100000 x g contained both types of desaturating enzymes, whereas the corresponding particulated fraction was inactive. The ultrastructure of Acanthamoeba showed the endoplasmic reticulum with a poorly developed membrane component. The metabolic pathways found with Acanthamoeba were compared to Ochromonas danica incubated with linoleic acid in the light and in darkness. Desaturases typical of "vegetal" and "animal" pathways were found in both organisms. In both of them, alpha-linolenic and arachidonic acids could be synthetized. However, alpha-linolenic acid, typical of vegetal synthesis, was only stored in Ochromonas due to the presence of a photosynthetic machinery.