Analytics · Mass spectrometry

Mass error in ppm: the arithmetic and the caveat

Analytics › Mass spectrometry LC-MS high-resolution MS MALDI-TOF counter-ion content

Solved

Solved by b.vestergaard in post #5

I read the opening post twice before replying, because I had assumed the opposite. 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).

Jump to the accepted answer →

k.chukwuTL211 Feb 2026#1

Mass error in ppm: the arithmetic and the caveat Writing it up because I had to work it out twice and would rather nobody else did.

Working through the identity arithmetic and I would like it checked.

retatrutide has a monoisotopic mass close to 4731.3 Da. On an electrospray instrument I would expect to see the multiply charged series rather than the intact singly charged ion, so for the doubly charged species I calculate (4731.3 + 2 x 1.00728) / 2, and for the triply charged the analogous expression.

The observed values in the report sit within a few ppm of those. My question is what that actually establishes, because I have seen people treat a mass match as a purity result and I do not think it is one.

17 likes 5mo

c.inglethorpeTL3Regular11 Feb 2026#2

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.

I would rather say I do not know than round it up to an answer.

2 likes 5mo

a.mwangiTL211 Feb 2026#3

The opening post and I disagree about the size of the effect, not about the direction.

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.

0 likes 5mo

cannula_driftTL3Regular11 Feb 2026#4

Taking the opening post at face value and following it one step further.

A definition problem is doing most of the work in this mass error in ppm discussion. Once the term is pinned down I suspect the disagreement mostly goes away and what is left is small.

20 likes 5mo

b.vestergaardTL2 Solution11 Feb 2026#5

k.chukwu, post #1: Mass error in ppm: the arithmetic and the caveat Writing it up because I had to work it out twice and would rather nobody else did. Working through the identity arithmetic and I would like it checked. retatrutide has a monoisotopic mass close to 4731.3 Da. On an electrospray instrument I would expect to see the multiply charged series… Go to post

I read the opening post twice before replying, because I had assumed the opposite.

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).

12 likes in reply to #1 5mo

BGiordanoTL2Member11 Feb 2026#6

b.vestergaard, post #5: I read the opening post twice before replying, because I had assumed the opposite. 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). Go to post

Adducts — sodium and potassium especially — shift the observed mass in predictable increments. A report showing an unexplained plus twenty-two is usually showing you a sodium adduct.

Where I would look next, rather than where I would stop.

0 likes in reply to #5 5mo

a.lindholmTL211 Feb 2026#7

No disagreement from me. Posting only so the question does not look ignored.

29 likes 5mo

c.delgadoTL211 Feb 2026 · edited#8

This follows post #5 rather than contradicting 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.

14 likes 5mo

p.lindqvistTL211 Feb 2026#9

a.lindholm, post #7: No disagreement from me. Posting only so the question does not look ignored. Go to post

Adding what did not work for me on mass error in ppm, since the failures never get written up and they are half the useful information.

2 likes in reply to #7 5mo

crossover_entryTL3Regular11 Feb 2026#10

Post #9 is right about the mechanism and I think understates the practical bit.

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.

0 likes 5mo

k.batistaTL212 Feb 2026#11

Adding the measurement that post #10 says would settle it.

The arithmetic for a doubly charged species is (M + 2 x 1.00728) / 2, and the analogous expression for higher charge states. Working it through once makes the reported values legible.

1 like 5mo

methods_marginTL312 Feb 2026#12

Collapsed as off-topic by two members at trust level 3 or above

Post #8 describes the usual case. This is about the unusual one.

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.

m.guerreroTL212 Feb 2026#13

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.

This is the sort of thing the wiki should carry and currently does not.

23 likes 5mo

stopper_traceTL2Member12 Feb 2026#14

m.guerrero, post #13: 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. This is the sort of thing the wiki should carry and currently does not. Go to post

On mass error in ppm, 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.

0 likes in reply to #13 5mo

n.hartmannTL212 Feb 2026#15

BGiordano, post #6: Adducts — sodium and potassium especially — shift the observed mass in predictable increments. A report showing an unexplained plus twenty-two is usually showing you a sodium adduct. Where I would look next, rather than where I would stop. Go to post

Acknowledging rather than arguing. The reasoning holds as far as I can follow it.

0 likes in reply to #6 5mo

vial_slopeTL3Regular12 Feb 2026 · edited#16

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.

This is the sort of thing that ought to be settled and apparently is not.

3 likes 5mo

m.amankwahTL212 Feb 2026#17

One caution on mass error in ppm: everything above assumes the underlying documentation is what it claims to be. That assumption is doing real work and is rarely stated.

16 likes 5mo

NLoughranTL3Regular12 Feb 2026#18

I had written a reply contradicting post #16 and deleted it. Here is what survived.

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.

32 likes 5mo

r.vukovicTL212 Feb 2026#19

c.inglethorpe, post #2: 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. I would rather say I do not know than round it up to an answer. Go to post

Mass error in ppm: I would want to see the raw numbers rather than the summary before agreeing. Summaries lose exactly the information that would settle this.

0 likes in reply to #2 5mo

r.mcalisterTL3Regular12 Feb 2026#20

The reason mass error in ppm is hard to answer is that the obvious measurement and the relevant quantity are not the same thing, and substituting one for the other is silent.

1 like 5mo

v.klausenTL3Regular12 Feb 2026 · edited#21

Post #17 and I disagree about the size of the effect, not about the direction.

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.

I would put this at better than even and not much better.

0 likes 5mo

e.coelhoTL212 Feb 2026#22

Adding a data point of agreement rather than a data point.

0 likes 5mo

GDashwoodTL3Regular12 Feb 2026#23

r.vukovic, post #19: Mass error in ppm: I would want to see the raw numbers rather than the summary before agreeing. Summaries lose exactly the information that would settle this. Go to post

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.

19 likes in reply to #19 5mo

ca.haddadTL212 Feb 2026#24

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.

The uncertainty is in the assumption, not in the calculation.

8 likes 5mo

g.ibarraTL212 Feb 2026#25

Worth separating two things that post #21 runs together.

Worth separating mass error in ppm as a question about the compound from mass error in ppm as a question about the documentation. They get answered by different people and only one of them is answerable here.

0 likes 5mo

j.teixeiraTL212 Feb 2026#26

a.mwangi, post #3: The opening post and I disagree about the size of the effect, not about the direction. 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. Go to post

This follows post #25 rather than contradicting it.

Where I have landed on mass error in ppm, 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.

27 likes in reply to #3 5mo

b.petrovTL212 Feb 2026#27

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.

13 likes 5mo

s.cardosoTL212 Feb 2026#28

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.

4 likes 5mo

n.stanescuTL212 Feb 2026#29

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.

Reading it back, the second half matters more than the first.

4 likes 5mo

j.nwosuTL212 Feb 2026#30

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.

Marking that as an opinion rather than a finding.

0 likes 5mo

Sponsored Sigma-Aldrich — certified reference materialsReference standards, analytical reagents and certified materials for method calibration. If you are validating a method, this is where the standard comes from.Reference standards →