One-pot and sustainable liquid-phase peptide extension for synthesis of C-terminal amidation peptides aided by small molecular tags

Abstract

The development of greener and more economical methods for peptide synthesis has been a driving force in promoting the large-scale production of peptide drugs. In this study, we present a series of small-molecule C-terminal amidation tags based on phosphonate or aliphatic moieties. These tags were prepared using a simple method with high yields and employed to facilitate the sustainable preparation of C-terminal amidation peptides via an Fmoc strategy liquid-phase one-pot process. The Fmoc group was deprotected using DEA/ACN and subsequently reacted with mercaptosuccinic acid or cysteine to capture and remove the Fmoc-residue. In the continuous one-pot peptide chain coupling and de-Fmoc reactions, each peptide intermediate can be easily purified through sequential washing with HCl aq. and Na₂CO₃ aq. solutions, allowing subsequent amino acid couplings to proceed in the original reaction solvent. Using the one-pot peptide synthesis strategy, both eptifibatide and the TREM-1 inhibitor LR12 were efficiently synthesized. Compared to LPPS and SPPS, this protocol achieves a more environmentally friendly and cost-effective method for preparing amidated peptides, making it suitable for large-scale production of peptide APIs.