Opioid-like immunoreactive material was extracted from the pituitary and brain of the Spiny Dogfish Shark Squalus acanthias. The immunoreactive material in the pituitary extracts was purified to apparent homogeneity by reverse phase high performance liquid chromatography and subsequently characterized by amino acid analysis, Edman degradation and fast atom bombardment mass spectrometry. The largest opioid-like peptide isolated contained 30 amino acids and showed 80 percent homology with salmon endorphin-II but less than 50 percent homology with human beta-endorphin. Three structural variants of this molecule were also characterized. These variants were shown to be shorter N-terminal fragments, two of which corresponded to cleavage products at the single basic residues arginine and lysine. Cleavage at a single lysine residue has not been reported for posttranslational processing of beta-endorphin in mammals and could represent a modification seen only in lower vertebrates. The remaining fragment corresponded to a loss of 3 residues from the C-terminus of the parent molecule. No alpha-N-acetylated peptides were detected. These results provide the first unequivocal confirmation of beta-endorphin in an elasmobranch and provide evidence of novel N-terminal variants of beta-endorphin.