Elucidation and prevention of an unexpected methylamine byproduct formed during trifluoroacetic acid-mediated side chain deprotection of azido-containing peptides

TL;DR

This paper identifies and prevents a methylamine byproduct formed during a chemical process used in peptide synthesis.

Contribution

The study introduces a new method using a Teoc-protected compound to prevent methylamine byproduct formation.

Findings

The byproduct is formed via a Schmidt rearrangement involving azide and t-butyl cations.

Using a Teoc-protected compound significantly reduces methylamine byproduct formation.

Abstract

Purpose
We found that during TFA-mediated cleavage of azide-containing peptides during solid-phase peptide synthesis a byproduct characterized by a difference of 12 mass units is formed. This study identifies this byproduct, provides a rationale for its formation and a solution for its inhibition.
Method
NMR and HRMS analyses as well as chemical synthesis is employed to identify the byproduct and probe the mechanism of its formation.
Results
Data is consistent with the conversion of azide to methylamine which occurs during peptide cleavage likely via a Schmidt rearrangement involving nucleophilic azide attack of t-butyl cations generated during side chain deprotection of Boc- and t-butyl ether groups.
Conclusion
Installing azide-containing residues using the newly reported 2-(trimethylsilyl)ethoxycarbonyl (Teoc)-protected compound
3a
proved effective at significantly thwarting the formation of methyl amine byproduct.