Every time you need to purchase quality peptides, one of the greatest concerns you will ever have, is the purity of the peptides. From peptide synthesis, peptides are available with different purity levels, and it is good to have a solid understanding of what the purity is, and how you can apply it when buying or using peptides.
Peptide purity refers to the amount of the target peptide as stipulated by HPLC at 214nm, where the absorption of the peptide bonds take place. Spectrophotometer is what is normally used to determine peptide purity, and it will never detect water and residual salts that might be present within the peptides. The other impurities that might be present in a peptide include truncated sequences, deletion sequences, and incomplete deprotected sequences.
In the determination of peptide purity, salts or water in the samples, are never considered. However, TFA is normally taken into account, and it is usually a result of HPLC amplification. TFA contamination usually arises from the free N-terminus, as well as other side reactions, such as Lys, Arg, and the formation of triflouroacetate. Most peptides delivered as triflouroacetatewill usually contain residual water. Even in lyophilized peptides, the presence of noncovalently bound water will always be detected.
The impurities present in non-purified peptides, may be either peptides or non-peptides, while the impurities present in purified peptides, will mostly consist of peptides, but with modified sequences, with the exception of TFA salts. It is also worth noting that:
- Shorter peptides may lack one or more amino acids of the peptides in the targeted sequence
- Capping steps may be used to avoid the formation of deletion peptides.
- Peptide synthesis or the final cleavage process may also lead to impurities in peptides.
- Impurities may also arise due to reattachment of protecting groups happening at other locations within the peptide sequence.