Total Synthesis of Nocardiotide A by Using a Combination of Solid- and Solution-Phase Methods

Nocardiotide A has been successfully synthesized using a combination of solid- and solution-phase synthesis. Peptides 1 and 2 of nocardiotide A, with different C and N termini, were synthesized on 2-chlorotrityl chloride resin. A combination of N,N,N ',N '-tetramethyl-O-(1H-benzotriazol-1-yl)uronium hexafluorophosphate and hydroxybenzotriazole (HBTU/HOBt) was employed as coupling reagent and the release of the linear peptides from the resin was undertaken using AcOH and trifluoroethanol (TFE) reagents resulting in linear hexapeptides with protected side chains in good yields (83.4 % and 76 %, respectively). Peptide 1, which has Ala as C terminus and Trp as N terminus, was the only peptide that was successfully cyclized without epimerization under a dilute condition (1.25x10(-3) M) using HBTU (3 equiv.) and 1 % N,N-diisopropylethylamine (DIPEA) in dichloromethane. A combination of a small residue at the C terminus and a larger residue at the N terminus was found to be effective for the cyclization. The protecting group of the crude cyclic product was then deprotected by 95 % TFA in water and the crude was then purified to give white solid product with 16 % overall yield. The peptide was then characterized by HR-ToFMS, H-1- and C-13-NMR spectroscopy. In order to ensure the absence of epimerization during cyclization, the D-analogue of nocardiotide A was synthesized using similar synthetic protocol. The NMR data of D-analogue was found to have a larger difference compared to the synthetic and natural product, showing there is no epimerization at C-terminus during cyclization.

Automated summary

Nocardiotide A was successfully synthesized using a combination of solid- and solution-phase synthesis, with detailed experimental procedures described for the synthesis and cyclization process. The absence of epimerization during cyclization was confirmed through comparison of NMR data between the synthetic and natural product, as well as the synthesis of a D-analogue of Nocardiotide A.