you need more sample for identity than for purity. plan for that when you post it, not advice obviously
#hplc-massspec 2026-05-14
Thursday41 messages9 participantstimes are UTC
Highlights from this day
- deamidation — relative retention time against a standard is comparable between labs. absolute minutes are not 09:58
- baseline_drift — update from 19 months ago: switched to asking for the chromatogram rather than the number and it changed everything 12:04
- baseline_drift — oxidation is about plus 15.99 Da and is usually methionine or tryptophan 12:06
- VialBot — Channel stats, last 30 days: 108 messages from 50 members. 12:13
- VialBot — Archive lookup: vat_on_import first appears in this channel on 2025-09-01. 14:41
slightly off topic but a shoulder on a peak is information, and usually bad information
i have had a lot where the mass was exactly right and the purity was 96.8. wrong question, right answer
co-elution is the failure mode that a beautiful single peak is most likely to be hiding
tirzepatide is about 4813.5 Da, so 2+ is near 2407.8 and 3+ near 1605.5
retention time is a hint, not a fingerprint. two different peptides can co-elute perfectly happily
can you tell a dimer from the chromatogram alone
relative retention time against a standard is comparable between labs. absolute minutes are not
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plus 0.98 is deamidation
the honest position is that hobby-level interpretation of a chromatogram is worth something but not much, still working it out
the honest position is that hobby-level interpretation of a chromatogram is worth something but not much
214nm sees the amide backbone so it sees everything. 280 only sees aromatics
quick one deamidation is about plus 0.98 Da. it is not a rounding error, it is a degradant
is monoisotopic or average mass the one on most certificates
i was wrong about tailing factor. it is usually the injection or the pH, not the column
integration difference
why does integration change the number so much
does retention time on its own tell you what the compound is
[edited]do you trust an area percent from a shallow gradient
show me the spectrum
if the lab reports area percent without stating the wavelength, the number is less useful than it looks
integration choices move the number by a few tenths easily. that is most of your inter-lab variance
retention time is a hint
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need more sample
update from 19 months ago: switched to asking for the chromatogram rather than the number and it changed everything
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oxidation is about plus 15.99 Da and is usually methionine or tryptophan
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is a tailing factor of anything specific a problem
a dimer usually shows up late and at roughly double the mass. the chromatogram alone will not tell you, anyway
Channel stats, last 30 days: 108 messages from 50 members.
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do the arithmetic
nobody runs enough blanks, including me, i think
[edited]coming back after 20 months, what changed in how you read these
ms/ms or it isnt identity
thats a charge state
shallow gradients hide related substances behind the main peak. a steeper one separates and looks worse, honestly
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i asked Janoshik for the raw trace and they sent it. that is the right answer to that question
within inter-lab range
thats co-elution
Archive lookup: vat_on_import first appears in this channel on 2026-02-09.
Archive lookup: vat_on_import first appears in this channel on 2025-09-01.
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slightly off topic but a warm transit can put a visible degradant peak on a chromatogram. that is what 12 days at ambient does
ms/ms sequencing is the only thing i would call identity. everything else is circumstantial
whats a plus one dalton shift usually