Second pass at: What a 2 ppm match establishes, and what it does not — setting out what I have, and where I think it stops being reliable.
Two things I would like separated before anyone answers on 2 ppm match, because they get bundled and then argued about as one thing.
The first is descriptive: what has actually been observed, by whom, and how. The second is causal: why. I am asking about the first only.
Building on the opening post rather than restating it.
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.
I have written this out at length because the short version keeps being misread.
Answering the question the opening post raises rather than the one it answers.
2 ppm match was covered in the wiki last year and the page has a review date on it, which is a better starting point than my memory of a thread.
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.
Post #3 and I disagree about the size of the effect, not about the direction.
Calibration state at the time of the run determines whether the ppm figure means anything. A report that states when the instrument was last calibrated is unusual and is worth more than one that does not.
Two people can read the same figure differently here and both be reasonable.
Resolution: "high resolution" commonly means <5 ppm across the mass range. Unit-resolution instruments achieve ±1 Da at best and cannot distinguish two species differing by less than 1 Da in total mass.
Post #9 answers the question as asked. The question underneath it is different.
2 ppm match looks different depending on whether you are reading the primary literature or the summaries of it, and the difference is not in our favour.
I read post #11 twice before replying, because I had assumed the opposite.
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.
The practical version of 2 ppm match is three sentences long. The rigorous version is three pages and reaches the same conclusion with the conditions attached.
Purity and identity are different questions: LC-MS establishes that the species at a retention time has the expected mass. It does not establish how much of the sample is that species (that is what LC-UV purity answers).
If the premise is wrong, everything after it is decoration.
Bookmarking this. I will come back when I have something worth adding.
Coming back to post #13, because the follow-up matters more than the original 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 general case is well covered; this is the awkward specific one.
This follows post #16 rather than contradicting it.
Where I would push back on the 2 ppm match consensus is the confidence, not the direction. The direction looks right. The confidence is borrowed.
Having read the whole 2 ppm match thread before replying: the question in the first post has not actually been answered yet, and three of us have answered a nearby one instead.
Quantitation by MS: most quantitation is done by LC-UV detection at 214 nm, not by MS, because extinction coefficients are better known. MS can quantify if an internal standard is used but that requires preparation.
I would not lead a decision with this, but I would not ignore it either.
Distinguishing three things in the 2 ppm match discussion that keep getting used interchangeably: the observation, the proposed mechanism, and the recommendation that gets attached to both.
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 evidence for this is thinner than the way I have phrased it suggests.
I had written a reply contradicting post #20 and deleted it. Here is what survived.
Summarising the 2 ppm match thread so far, since it is long and the answer is buried: the first reply has the method, the fourth has the correction to it, and the rest is people agreeing at length.
One more thing on 2 ppm match that took me far too long to see: the two figures people quote are not measuring the same quantity. Once you notice that, the apparent contradiction disappears.
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.
Thank you for the correction. I would rather find out here than later.
Where I part company with post #23, and it is a narrow parting.
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.
Same conclusion as the reply above, reached differently, which is mildly reassuring.
Post #27 is the version of this I will quote in future. One addition.
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.
Worth separating two things that post #27 runs together.
Genuine question rather than a rhetorical one: has anyone here actually observed 2 ppm match, as opposed to read about it? The thread is long and I cannot tell.
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.