Antibiotic trichopolyns (TP)-I and II are 10-residue peptides obtained from the fungus Trichoderma polysporum (TMI 60146) and belong to the class of peptaibols. TPs are characterized by the presence of 2-methyldecanoyl group at the N-terminus and the C-terminal residue protected by trichodiaminol. Three new analogues, TP-III-V, have been isolated together with highly purified TPs-I and II. Their structures have been determined by the combined use of NMR, MS and MS/MS. TPs-III and IV differ from TPs-I and II in the replacement of Aib^9 by Ala, respectively. On the other hand, TP-V has the same amino acid sequence as TP-I, but the N-terminal acyl group is substituted by 3-hydroxy-2-methyldecanoic acid from 2-methyldecanoic acid. Conformational analysis for TPs was carried out by using CD and NMR. Their CD spectra in methanol exhibited two negative Cotton effects at 208 and 222nm, which are typical of a right-handed helical structure. Inter-residual NOE connectivities supported the CD data and, in addition, showed that the helices continue from the N-terminal residue to the C-terminal amino alcohol, trichodiaminol. TP-I exhibited immunosuppressive activity in mouse allogeneic mixed lymphocytic reaction. The activity was stronger than that of cyclosporin A.