The other common method for introducing the Fmoc group is through 9-fluorenylmethylsuccinimidyl carbonate (Fmoc-OSu), which may itself be obtained by the reaction of Fmoc-Cl with the dicyclohexylammonium salt of N-hydroxysuccinimide.[4]
Reacting with 9-fluorenylmethyloxycarbonyl azide (itself made by reacting Fmoc-Cl with sodium azide) in sodium bicarbonate and aqueous dioxane is also a method to install Fmoc group.[1]
Because the fluorenyl group is highly fluorescent, certain UV-inactive compounds may be reacted to give the Fmoc derivatives, suitable for analysis by reversed phase HPLC. Analytical uses of Fmoc-Cl that do not use chromatography may be limited by the requirement that excess Fmoc-Cl be removed before an analysis of fluorescence.
Cleavage & Deprotection
The Fmoc group is rapidly removed by base. Piperidine is usually preferred for Fmoc group removal as it forms a stable adduct with the dibenzofulvene byproduct, preventing it from reacting with the substrate.[5][6]
Role in Peptide Synthesis
The use of Fmoc as a temporary protecting group for amine at the N-terminus in solid phase synthesis is very widespread for Fmoc/tBu approach, because its removal with piperidine does not disturb the acid-labile linker between the peptide and the resin.[7] A typical SPPS Fmoc deprotection is performed with a solution of 20% piperidine in N,N-dimethylformamide (DMF).[8]
Common deprotection cocktails for Fmoc during SPPS
20% piperidine in DMF (Fmoc group has an approximate half life of 6 seconds in this solution)[8]
5% piperazine, 1% DBU and 1% formic acid in DMF. This method avoids the use of strictly controlled piperidine.[9] No side product was observed for a peptide with 9 residues synthesized with this method.[10]
