Acid extracts of anterior and intermediate lobes of the pituitary gland from 4 dogs were fractionated by cation-exchange high-performance liquid chromatography and analyzed by radioimmunoassay, using 2 antibodies: one specific for the midregion of beta-endorphin (beta-end) and the other specific for the N-terminal region of N-acetylated beta-END. Identification of peaks of canine beta-END immunoreactivity was based on the retention times relative to those of synthetic human beta-END standards, with predicted variations attributable to differences in the beta-END amino acid sequences between the 2 species. The canine anterior lobe was found to contain almost exclusively beta-END (1-31). By contrast, the intermediate lobe contained substantial amounts of N-acetylated and C-terminally shortened forms of beta-END. The predominant forms of beta-END in canine intermediate lobe were, in decreasing order of abundance: beta-END (1-27), beta-END (1-31), beta-END (1-26), Ac beta-END (1-27), Ac beta-END (1-26) and Ac beta-END (1-31). Individual beta-END immunoreactivity profiles varied, but the general pattern was consistent among the 4 dogs.