Objective: Misoprostol is a synthetic prostaglandin that is related structurally to naturally occurring prostaglandin (PG), and it has been acknowledged as an effective inhibitor of gastric acid secretion when administered intravenously. Methods: In the present study, the novel application of 1,8-Diazabicyclo[5.4.0]undec-7-ene, a cyclic unsaturated amine [DBU] for the conversion of A-type Misoprostol (A-MP) to B-type Misoprostol (B-MP) via intramolecular isomerization. Results: The intramolecular isomerization process was successfully applied for the separation of enantio-pure isomers with no impurities using DBU. Conclusion: The chemical structures of A-MP and B-MP were confirmed using spectral analyses of 1H-NMR, 13C-NMR and Mass spectroscopy.