Synthesis of a peptide chain on a solid support offers separation of from soluble reagents and simple filtration and washing steps. However, byproducts from incomplete couplings, side reactions, and impurities can build up on the solid support with the desired product. The two most common solid phase peptide synthesis (SPPS) methods currently used are Fmoc/tBu and tBoc/Bzl. The tBoc method is the older method and is based on the original Merrifield approach of graduated acidolysis using TFA to selectively remove temporary and permanent protecting groups. A major disadvantage of the tBoc method is that HF (hydrofluoric acid) is required to cleave the peptide from the solid support. The newer method Fmoc SPPS uses a base-labile N-Fmoc group for a-amino group protection, while acid labile side chain protecting groups and resin-linkage.

Structure of 9-fluorenylmethyloxycarbonyl (Fmoc) protecting group

Removal of the Fmoc protecting group is most often performed with 20-50% piperidine in DMF.

After Fmoc removal, the solution is filtered through a proper filter and washed several times with DMF to ensure complete removal of Fmoc and piperidine from the resin.