The fatty acid composition of the total and polar lipid fractions of Choanephora cucurbitarum grown under different cultural conditions were analyzed by thin-layer and gas-liquid chromatography. It was observed that temperature, age, pH, and light influenced the degree of unsaturation, this being due mainly to changes in the gamma-linolenic acid concentration. The conditions used in this study did not alter the qualitative profile of fatty acids normally present in the organism. Neither did these conditions stimulate the production of further long-chain fatty acids (C20-C26) beyond gamma-linolenic acid (C18:3) as reported earlier using growth media containing glutamic acid. The fatty acid pattern of lipid fractions though the same qualitatively, differed quantitatively. The polar lipid fractions, phosphatidyl choline, phosphatidyl ethanolamine, and diphosphatidyl glycerol showed an appreciable variation in gamma-linolenic acid content under different cultural conditions. The degree of unsaturation of the various lipid fractions decreased with increases in temperature, light intensity, and pH, but within each treatment the same pattern of decreasing degree of unsaturation with increasing age was observed. The significance of these observations is discussed.