Human semen contains large amounts of authentic beta-endorphin and other beta-endorphin immunoreactive peptides. To eliminate the non-specific effects of semen in the beta-endorphin RIA, the beta-endorphin immunoreactive peptides were extracted with octadecylsilane-C18 reverse phase beads. beta-Endorphin immunoreactivity was parallel to the RIA standard curve. Sephadex G-50 gel chromatography showed that 93% of the immunoreactive material coeluted with beta-endorphin. Reversed phase high pressure liquid chromatography of semen extract demonstrated several peaks of beta-endorphin immunoreactivity; one of these was coincident with synthetic h-beta-endorphin61-91.