The contents of total lipids, total sterols, total and individual phospholipids: lysophosphatidyl choline, lysophosphatidyl ethanolamine, phosphatidyl choline, phosphatidyl ethanolamine and alkali-stable phospholipids I and II, were significantly lower in mature cysts of Acanthamoeba culbertsoni strain A-1 than in trophozoites. The major constituent sterols of trophozoites, viz. ergosterol, 7-dehydrostigmasterol and 7,22,25-tridehydrostigmasterol which resolved as a single spot on thin-layer chromatography, disappeared in the cysts. When trophozoites grown in the presence of [14C]acetate were transferred to encystation medium, there was a steady decrease in the radioactivity in the lipids and acid-soluble fraction (glycogen) of the encysting cells while the radioactivity increased in the akali-insoluble fraction (cellulose). The activity of isocitrate lyase in the encysting cells steadily increased up to 24 h of encystation, followed by a slight decrease during 24-32 h, but the activity of isocitrate dehydrogenase steadily decreased up to 32 h of encystation, beyond which time no enzyme activity was detected. Inhibitors of isocitrate lyase, glycolate and maleate at 8.5 to 34 mM concentration inhibited encystation by 18-67%.