This is a continuation of a long topic, addressed by post number rather than by page. Start at post 1 · go to the accepted answer.
Picking up post #60: that is the part I would want checked first.
Deamidation adds approximately one dalton and produces a species that frequently elutes very close to the parent. It is the hardest common impurity to see chromatographically and the easiest to see by mass.
Adding a small correction to the counter-ion content summary above rather than a disagreement with it. The substance holds; one of the figures is out by a factor that matters.
Resolution and mass accuracy are different specifications. An instrument can resolve two species and still assign their masses imprecisely, and the reverse is also possible.
The strength of my opinion here exceeds the strength of my evidence.
Mass error in parts per million is (observed minus theoretical) divided by theoretical, times a million. On a high-resolution instrument a low single-digit figure is unremarkable and expected.
Posting it because the silence on this was starting to look like agreement.
Positional isomers and epimers are mass-identical. Any argument that a mass result rules them out is wrong, and it is the commonest overclaim in this subcategory.
Good question, well framed, and I would like to see it answered properly.
On counter-ion content, I would rather understate and be corrected upward than overstate and be quoted. That is a house style here and it is a good one.
Calibration matters: a high-resolution instrument out of calibration can report mass with ppm error large enough to be uninformative. Check when the instrument was last calibrated before trusting the reported accuracy.
The step people skip is the one I have spelled out.
Post #70 and I disagree about the size of the effect, not about the direction.
Combining a chromatographic result with a mass result is genuinely orthogonal confirmation. Either alone leaves a specific class of problem invisible, and the two classes do not overlap much.
Taking post #70 at face value and following it one step further.
Sample preparation for mass spectrometry can itself introduce modifications, particularly oxidation. A finding at trace level may be telling you about the preparation rather than the material.
Source for the counter-ion content figure, since it was asked for. It is in the discussion rather than the abstract, which is why the version circulating is stronger than the paper is.
Reading the surrounding paragraph is worth the two minutes. The authors are more careful than their summarisers.
Response in electrospray is not proportional to abundance across different species. Using peak intensities from a mass spectrum to estimate proportions is a mistake that looks reasonable.
I am aware this is the third time this month I have made this point.
This follows post #74 rather than contradicting it.
An acylated peptide has a mass that reflects the modification, so comparing against the mass of the unmodified backbone gives a mismatch that is not an error.
Reporting the observation and leaving the explanation open deliberately.
For a compound with no reference standard in circulation, an observed mass is much more useful than an assertion of agreement with a theoretical value nobody can check.
Trifluoroacetate adducts are common in material purified with TFA and are one reason a mass spectrum from a peptide can look busier than expected.
I would treat the number as indicative rather than as a measurement.
A note on how counter-ion content gets discussed rather than on counter-ion content itself: the confident posts get the replies and the careful ones get ignored, and the careful ones have been right more often.
Adding thanks rather than a view. I do not have a view worth the space.
Confirming post #82 from a second method, which matters more than confirming it from a second person.
The honest summary of what a mass result buys you: it narrows the field of what the material could be, considerably. It never closes it, and no certificate should be read as though it had.
The evidence for this is thinner than the way I have phrased it suggests.
I had written a reply contradicting post #80 and deleted it. Here is what survived.
Mass accuracy is expressed in parts per million. It is the difference between observed and theoretical mass divided by theoretical mass, multiplied by a million. A high-resolution instrument in good calibration achieves low single-digit ppm on a peptide of this size.
What mass accuracy establishes: the measured mass is consistent with a specific composition. What it does not establish: purity, sequence order, stereochemistry, or the absence of an isobaric species. Every one of those requires something else.
That is what I would do. It may not be what is correct.
On post #84 — agreed on the reasoning, with one qualification.
Common adducts: sodium adds ≈22, potassium adds ≈38 compared to hydrogen. A [M+Na]+ peak is common and its mass is predictable from the base mass.
Same conclusion as the reply above, reached differently, which is mildly reassuring.
Post #86 is right about the mechanism and I think understates the practical bit.
Tandem mass spectrometry: MS/MS fragments the molecular ion and uses fragment masses to confirm identity and detect modifications. A simple identity confirmation by LC-MS does not address post-translational modifications or impurities with the same or very close mass.
Narrowing post #86, because the general version has more than one answer.
A mass match establishes that the measured mass is consistent with the proposed composition. It does not establish purity, sequence order, or the absence of an isomer, and all three are frequently claimed from it.
The honest answer is that it depends, and here is what it depends on.