Gonadotropin-releasing hormone (GnRH)-like material was measured by radioimmunoassay in acid-ethanol-extracted human seminal plasma using radiolabeled D-[Leu6] GnRH ethylamide as labeled ligand, authentic GnRH as standard, and antibody raised against D-[Lys6] GnRH analog. The mean amount of GnRH-like material measured in the seminal plasma of semen samples with sperm counts greater than 20 X 10(6)/ml was 229.0 +/- 66 pg/ml, with sperm counts less than 20 X 10(6)/ml was 213 +/- 42 pg/ml, and from vasectomized samples was 252 +/- 36 pg/ml. There was no significant difference among the three groups. Scatchard analysis of radioreceptor binding data demonstrated significant displacement of GnRH agonist ligand from castrated male rat pituitary membrane preparations. Ultrafiltration and gel column chromatography of pooled extracted seminal plasma identified two compounds with apparent molecular weights of 2600 and 5000 that differ chemically and immunologically from native GnRH. Further characterization using affinity column chromatography suggests that at least one of these GnRH-like factors is a glycosylated protein.