The aim of the present study was to examine the biosynthesis of prostaglandins and to investigate factors conditioning their biosynthesis in pathogenic and nonpathogenic strains of Acanthamoeba spp. We established that the activity of the synthase of prostaglandins was almost identical in pathogenic and non-pathogenic strains and that the synthesis of endoperoxide prostaglandins was similar to that of other organisms up to the point at which prostaglandin H2 was produced. The course of biosynthesis in vitro can be activated by various compounds such as glutathione, albumin, and p-chloromercuribenzoic acid (p-CMB), which are either activators or inhibitors of the enzymes. We suggest that the course of biosynthesis of prostaglandins in vivo is most probably activated by tissues or constitutional liquids surrounding the parasites.