Using antibodies against the synthetic opioid peptides BAM-22P and peptide F, immunoreactive (ir-) peptides were measured in bovine brain and adrenal medulla. In addition to the high levels in the adrenal medulla, both ir- peptides were measurable in various areas of the brain with highest concentrations in the anterior hypothalamus. Analysis of the ir- components by gel filtration revealed molecular heterogeneity. Besides peptides with the size of BAM-22P or peptide F, various higher molecular weight species were found. These forms were found in the adrenal medulla in much higher concentrations than in the brain indicating a different processing mechanism for proenkephalin. Synthetic BAM-22P injected intracerebroventricularly into mice produces a substantial analgesia (ED50 6.4 nmole) which is almost as high as that of morphine (ED50 2.8 nmole). This finding and the presence of BAM-22P-like compounds in the brain suggests a role of the enkephalinergic system in pain transmission.