We investigated the effect of the pretreatment (sonification or centrifugation) of saliva samples on the concentration of several steroid hormones as measured with highly specific RIA after extraction and chromatography. It appeared that sonification of saliva resulted in significantly higher values for progesterone, cortisone, 17-hydroxyprogesterone, testosterone and oestradiol (10-49% increase), compared with the levels recorded after centrifugation. No differences were demonstrated for the concentrations of cortisol and androstenedione, except that a sex-dependent difference effect was observed in the values for androstenedione: concentrations measured in sonificated male saliva were lower than those measured in supernatant saliva.