During the course of screening fermentation broths for new inhibitors of dipeptidyl peptidase II (EC 3.4.14.2), we discovered epostatin (C23H33N3O5) from Streptomyces sp. MJ995-OF5. The taxonomy of producing strain, fermentation, isolation, physico-chemical properties and biological properties have been described in the preceding paper1). Herein we report on the structure determination of epostatin on the basis of spectroscopic studies. The molecular weight of epostatin was measured by FAB-MS (m/z 432 [M+H+], m/z 430 [M-H-]), and the molecular formula of C23H33N3O5 was determined by interpretation of the HRFAB-MS, NMR spectral analyses and elemental analysis1). The structure of epostatin was elucidated on the basis of 1D and 2D NMR experiments in DMSO-d6. The geometry of the double bond at C-9 was determined to be 9E by the coupling constant, J9 ,10 = 15.0 Hz. Other stereochemistry of epostatin remains to be determined. Thus, epostatin was determined to be Nα-[4-(2 hexyl-1a, 2, 3, 4, 4a, 7-hexahydro-cyclopent[b]oxireno[c] pyrid-7-ylidine)-2-butenoyl]-L-glutamine.