What is peptide purity? ---Peptide FAQ from Pepmic Co., Ltd

Peptide purity is the amount of the target peptide as determined by HPLC at 214 nm, where the peptide bond absorbs. Water and residual salts are not detected by UV spectrophotometer. Other impurities that can be found in the content include deletion sequences (shorter peptides lacking one or more amino acids of the target sequence), truncated sequences (generated by capping steps to avoid the formation of deletion peptides), and incompletely deprotected sequences (generated during the synthesis or the final cleavage process).
Peptide purity does not include any water or salts in the sample. TFA results from HPLC purification. The free N terminus and other side chains such as Arg, Lys, and His form trifluoroacetates and this allows small amounts of TFA to contaminate the peptides. Peptides are usually delivered as trifluoroacetates containing residual water. Even in lyophilized peptides, varying amounts of noncovalently bound water still exist.
What are other substances (impurities) in the peptides?

Impurities	Non-Purified Peptides	Purified Peptides (HPLC)
Deletion sequences1	✓	✓
Truncation sequences2	✓	✓
Incompletely deprotected sequences3	✓	✓
Sequences modified during cleavage4	✓	✓
DTT (dithiothreitol)	✓	☓
TFA (trifluoroacetic acid)	✓	✓
Acetic acid	✓	☓
Peptides that have undergone side reactions such as proline isomerization or isoaspartimide formation, etc.	✓	✓

The impurities in non-purified peptides are both peptides and non-peptides, the impurities in purified peptides are mostly peptides with modified sequences, except for TFA salt.

• Shorter peptides lacking one or more amino acids of the target sequence
• Generated by capping steps to avoid the formation of deletion peptides
• Generated during the synthesis or the final cleavage process
• Reattachment of protecting groups at other locations on the peptide

Eva He
eva@pepmic.com

Pepmic Co., Ltd.

www.pepmic.com