Follow-up: Attributing a new peak: degradation product or excipient?

Attributing a new peak: degradation product or excipient? I have a specific reason for asking rather than idle curiosity, and the context is below.

A follow-up question about Attributing a new peak that I did not know to ask the first time.

The earlier thread answered what I asked. What I should have asked is below, and I think it is the one that matters.

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.

On the opening post — agreed on the reasoning, with one qualification.

Where I have landed on Attributing a new peak, having got it wrong once in public: the direction is clear, the magnitude is not, and anyone quoting a precise magnitude has borrowed it from somewhere that did not measure it.

The confident answers on Attributing a new peak and the well-sourced answers are not the same answers, which is the most useful thing I have learned reading this category.

Incomplete deprotection leaves a protecting group attached, raising the mass substantially and usually pushing retention much later. A late-eluting peak on a peptide chromatogram is worth asking about.

Two things can be true about Attributing a new peak at once: the mechanism is plausible and the evidence for the size of the effect is thin. Most of the argument here is people defending the first against attacks on the second.

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.

I checked the source rather than the summary, and they differ.

Everything in post #5 holds. The case it does not cover is the one I have.

Answering the Attributing a new peak question as asked, then the question I think is meant. As asked: yes, with the qualification below. As meant: it depends on how the first measurement was taken.

Narrowing post #9, because the general version has more than one answer.

On Attributing a new peak: the maintained page in the documentation commons covers the general case with citations and a review date, which is more reliable than any reply here including this one.

The arithmetic in post #10 is right; the assumption feeding it is the part to check.

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.

The interesting part of this is the exception, and I do not understand the exception.

Answering the question post #8 raises rather than the one it answers.

Deletion sequences arise from incomplete coupling and are lower by one residue mass. Where they elute depends on the hydrophobicity of the residue that is missing, so they can appear on either side of the main peak.

Written from notes rather than memory, which is why the numbers are specific.

Relative response factors mean impurities are not detected in proportion to how much of them is present. A one per cent peak is not one per cent by mass unless the response factors happen to match.

Reading back through the Attributing a new peak threads from last year, the same three questions come up every time and only one of them has ever been answered properly. That seems like a documentation gap rather than a knowledge gap.