The ability of Sertoli cells harvested from 18-day-old Sprague-Dawley rats to generate reactive oxygen species (ROS) occurs under unstimulated and stimulated conditions. Thus, the generation of ROS and its regulation by stimulating and inhibiting compounds was determined as a lucigenin-dependent chemiluminescence reaction. According to the data, ROS generation was influenced by different cell preparation conditions--stimulating substances such as PMA, FMLP, C5a, A23187, and scavestrogens characterized by antioxidant, radical-scavenging properties. The mechanical homogenization during cell preparation procedures leads to an increase of ROS generation. ROS generation of Sertoli cells was followed by elected substances in the following rank order of efficacy: PMA > FMLP > or = C5a > Ca-ionophore A23187. The registered inhibiting effects of the scavestrogens J811 and J861 were significant. The measured CL counts decreased at 72 and 77%, respectively, of control experiments done without scavestrogens. The generation of reactive oxygen species in Sertoli cells and especially the increase in oxygen free radicals and their effects on cellular structures of spermatids are directly involved in inducing morphological alterations. Sertoli cells play an important role in spermatogenesis. The measurements of ROS may have clinical relevance in the evaluation of infertility.