Twenty-one metabolites were detected in faecal samples collected after infusion of (14C)cortisol into the jugular vein of sheep. Using high-performance liquid chromatography/radiometric analysis plus mass spectrometry. One group of metabolites had molecular weights of between 302 and 308, and another group of 350, which indicates that the substances have a C19O3 or a C21O4 structure. Therefore, an enzyme immunoassay against 5beta-androstane-3alpha-ol-11,17-dione-17-CMO:BSA was established. Faecal samples were collected from 10 cows immediately after transport and then during a course in which non-invasive diagnostic procedures were being taught (course 1). For comparison, faeces were sampled from another 5 cows that were being used for teaching invasive procedures (course 2). Six cows from a university farm served as controls. In the animals used in course 1, the highest concentrations of cortisol metabolites were measured immediately after transport to the university (median value: 2.2 micromol/kg faeces). During the first 5 days at the university, the concentrations decreased to 0.52 micromol/kg (median) and remained at this level during the rest of the course. The median concentration in the samples that were taken during coursc 2 (collected about 2 months after transport) was 0.48 micromol/kg. There was no significant difference in the excretion of cortisol metabolites between these cows and the controls. We conclude from these data that, using the enzyme immunoassay against 5beta-androstane-3alpha-ol-11,17-dione-17-CMO, we were able to detect transport/novel environment stress but not the potential disturbance that cows experience during diagnostic procedures.