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This report concerns biosynthetic studies conducted with neurointermediate lobes of the mouse pituitary gland. High performance liquid chromatography was used to resolve newly synthesized peptides after in-vitro incubation of lobes with radioactive amino acids. Among the newly synthesized peptides identified were α-MSH, des-Nα-acetyl-α-MSH, two forms of corticotrophin-like intermediate lobe peptide and β-endorphin. The biosynthesis of a glycosylated γ3-MSH-like peptide was also demonstrated. While no newly synthesized β-MSH could be identified, a peptide designated γ-lipotrophin was found. Pulse-chase analysis revealed that the major biosynthetic pathway leading to the production of α-MSH involved the acetylation of the des-acetyl form of this peptide. Furthermore, it was evident that newly synthesized β-endorphin was largely converted to modified forms of this peptide; most of the terminal product was probably N-acetylated endorphin.