Online Peptides choose standard stepwise solid phase peptide synthesis (SPPS) procedures, including both Boc and Fmoc chemical synthesis technique, the current and most popular method of peptide synthesis.
About Solid Phase Peptide Synthesis (SPPS)
Solid phase peptide synthesis (SPPS), developed by R. B. Merrifield, was a major breakthrough allowing for the chemical synthesis of peptides and small proteins. The first stage of the technique consists of peptide chain assembly with protected amino acid derivatives on a polymeric support. The second stage of the technique is the cleavage of the peptide from the resin support with the concurrent cleavage of all side chain protecting groups to give the crude free peptide.
The general principle of SPPS is one of repeated cycles of coupling-deprotection. The free N-terminal amine of a solid-phase attached peptide is coupled to a single N-protected amino acid unit. This unit is then deprotected, revealing a new N-terminal amine to which a further amino acid may be attached.
There are two major used forms of solid phase peptide synthesis – Fmoc (base labile alpha-amino protecting group) and t-Boc (acid labile protecting group). Each method involves different resins and amino acid side-chain protection and consequent cleavage/deprotection steps. Fmoc chemistry is known for generating peptides of higher quality and in greater yield than t-Boc chemistry. Impurities in t-Boc-synthesized peptides are mostly attributed to cleavage problems, dehydration and t-butylation. After cleavage from the resin, peptides are usually purified by reverse phase HPLC using columns such as C-18, C-8, and C-4. The primary advantage of SPPS is its high yield. As peptides consists of many amino acids, if the yield for each amino acid addition is much less than 100%, overall peptide yields are negligible. For example, if each amino acid addition has a 90% yield then the overall yield of a 50 amino acid peptide is only 0.5%. Modern SPPS instrumentation pushes coupling and deprotection yields to greater than 99.99%, giving an overall yield of greater than 99% for a 50 amino acid peptide.