Named, unnamed and unspecified impurities as regulatory categories — the long version

For anyone arriving from a search: the marked solution above is the direct answer, and the replies underneath it add the caveats that make it safe to use.

Off-target structures: if the sequence synthesis goes wrong, a completely different amino acid can be incorporated. The resulting off-target peptide is a structural isomer with the same mass but a different sequence. No chromatographic purity method detects this without a reference standard.

post #16 is right about the mechanism and I think understates the practical bit.

Off-target structures: if the sequence synthesis goes wrong, a completely different amino acid can be incorporated. The resulting off-target peptide is a structural isomer with the same mass but a different sequence. No chromatographic purity method detects this without a reference standard.

Worth separating two things that post #14 runs together.

Deletion sequences (incomplete coupling during synthesis): lower in mass by one residue. Chromatographically they usually elute earlier or later depending on the residue's hydrophobicity. They are the most common impurity in solid-phase synthesis.

Residual solvents: traces of solvents used in purification. These are usually tested by gas chromatography, not by HPLC. A specification for residual solvents should be stated separately from the purity.

Coming back to post #18, because the follow-up matters more than the original answer.

Acetate content: counter-ion content. Trifluoroacetate or acetate from the salt form of the peptide. Affects mass calculations and should be stated on a complete certificate.

Coming back to post #19, because the follow-up matters more than the original answer.

Two things before anyone answers the substance.

First, the context in the first post is clear and specific. Second, the question is framed so that an answer can actually address it. Both are the norm here and both matter more than they sound.

Disulfide formation: if a peptide contains cysteine, it can form disulfide bonds with itself or with other molecules. Under oxidising conditions multiple species appear. Reducing conditions (like DTT) convert them back.

Dimer and higher-order multimers: two or more peptide molecules bonded together. They appear at double the mass and higher. They may or may not separate from the monomer on HPLC depending on the method.

I read post #23 twice before replying, because I had assumed the opposite.

Truncation products: fragments from incomplete synthesis or from degradation. They elute quite differently from the intact peptide because they are much smaller and have different hydrophobicity. They are usually well separated.

For anyone arriving from a search: the marked solution above is the direct answer, and the replies underneath it add the caveats that make it safe to use.

Oxidation at methionine and tryptophan: adds 16 per oxygen. Usually elutes earlier. Oxidation is common in storage, especially if the solution is exposed to light or if antioxidants are not present.

post #27 is right about the mechanism and I think understates the practical bit.

Deamidation at asparagine and glutamine: adds 1 approximately. Frequently appears as a close-eluting pair. It is a chemical modification that occurs during storage.

Incomplete deprotection: mass higher by the protecting group mass. Usually markedly later eluting. A synthesis artifact from incomplete removal of protecting groups.

Aggregates: multiples of the monomer mass. May not elute at all under a standard reversed-phase method. A species that does not come off the column does not appear in the area percentage.