Topic summary

Racemisation and why it is invisible to a mass measurement

Analytics › Impurities & related substances counter-ion content endotoxin testing HPLC Closed

This is a generated summary. It shows the 9 most-liked posts from a topic of 128, in their original order, with the accepted answer included where one exists. It is a reading aid and it will miss nuance — the full topic is the record.

d.oyelaranTL3Pharmacist8 Jun 2026#4

KLindqvist, post #2: 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. Go to post

Everything in post #2 holds. The case it does not cover is the one I have.

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.

That is all the detail I have. Someone else will have more.

28 likes in reply to #2 2mo

n.moreauTL29 Jun 2026#15

s.leclerc, post #11: 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. Go to post

Post #13 and I disagree about the size of the effect, not about the direction.

Aggregates may be a multiple of the monomer mass and may not elute at all under a standard method. What does not come off the column does not appear in the area percentage.

Reporting the observation and leaving the explanation open deliberately.

29 likes in reply to #11 2mo

ni.kravchenkoTL210 Jun 2026#35

n.rahimi, post #6: Second this, and I would have said it less carefully. Go to post

Truncation products are substantially lower in mass and usually well separated chromatographically, which makes them the easiest class to see and the least likely to be missed.

I am aware this is the third time this month I have made this point.

33 likes in reply to #6 2mo

n.nakamuraTL211 Jun 2026#49

That is the distinction I keep failing to hold on to. Written down now.

30 likes 2mo

BirkelandTL3Regular11 Jun 2026#55

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.

28 likes 2mo

WoodhouseTL2Member12 Jun 2026#61

Post #58 is the version of this I will quote in future. One addition.

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 looked this up rather than remembered it, which is the right order.

28 likes 2mo

TamburelloTL2Member13 Jun 2026#83

That last point is the ceiling on what any purity figure can claim. A method that cannot see a species cannot exclude it, and no certificate says which species its method cannot see.

That holds under the stated conditions and I have stated them.

30 likes 1mo

methods_marginTL3Regular13 Jun 2026#89

s.perrin, post #68: Adding what did not work for me on racemisation, since the failures never get written up and they are half the useful information. Go to post

Freeze-drying itself does not generate impurities in any significant way. Handling before and after it does, which is why transit conditions belong in the discussion.

That holds for the case as described. Change the assumptions and it may not.

32 likes in reply to #68 1mo

q.zhao_qaTL3Quality assurance15 Jun 2026 · edited#127

Truncation products are substantially lower in mass and usually well separated chromatographically, which makes them the easiest class to see and the least likely to be missed.

It reads as pedantry until the day it does not.

29 likes 1mo

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This topic was referenced in

Impurity thresholds: where the common numbers come fromAnalytics › Impurities & related substances · 15 replies
Deamidation and the close-eluting pair it produces — the long versionAnalytics › Impurities & related substances · 3 replies

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