[shi196]

Hi there, a quick question about mass spectrometry as I can't seem to find anything clear online about the possibility of this beyond a bunch of excel spreadsheets....

I have a cyclic octapeptide which includes one lysine side chain (free amine at end), and two tryptophans (not boc protected).

https://ibb.co/g4PXG2j

Observing the course of my reaction after 4 hours which is in DMSO, I take a few drops and I use a stream of nitrogen to remove as much solvent as possible, and then dilute in methanol to run my ESI mass spec.
I can see my starting material peaks (1107.7, 1108.7,etc ) and then also peaks at 1340.7, 1342.7, 1343.6 which I thought might equate roughly to addition of 3 x DMSO (+/- protons).

Is it possible to have multiple adduct ions of DMSO? (if my terminology is incorrect here I apologise as i'm not particularly experienced in mass spectrometry at all). i.e is the dmso "sitting" at the site of the lysine+two tryptophans?

@MOTOBALL    ?

Many Thanks

[MOTOBALL]

It is indeed very likely that you are observing adducts with DMSO.
This is analogous to the moles of water of crystallization that accompany inorganic salts.
So you have [M + n DMSO + H+]+.
But the devil is, as always in science, in the details.
DMSO = 78, so 3 x 78 = 234 and you should see m/z = 1341.7.
You cannot have “+/- a few protons”.
There is a single proton to charge the CP and 3 moles of solvation by DMSO.
1) Is the mass spec calibration spot on?
2) Is the m/z 1340.7 signal strong?
3) Are the signals really at 1340.7, 1342.7, 1343.6?
Yes, 3 x S atoms at 2.2 % RA each would show that!
That tends to confirm the DMSO solvation that you propose.
You really do need to be able to answer 1).
What reference std do you use for mass calibration?
Try a dilute solution of CSI in acetonitrile.
This give [Cs+(CSI)n]+ in positive ESI over a very wide range (m/z 133 to ~2000).
Also, have you checked for doubly/triply charged signals with any of your CP type molecules? Again, would help to confirm the DMSO-adduct proposal.

Regards,
Motoball

[shi196]

It is indeed very likely that you are observing adducts with DMSO.
This is analogous to the moles of water of crystallization that accompany inorganic salts.
So you have [M + n DMSO + H+]+.
But the devil is, as always in science, in the details.
DMSO = 78, so 3 x 78 = 234 and you should see m/z = 1341.7.
You cannot have “+/- a few protons”.
There is a single proton to charge the CP and 3 moles of solvation by DMSO.
1) Is the mass spec calibration spot on?
2) Is the m/z 1340.7 signal strong?
3) Are the signals really at 1340.7, 1342.7, 1343.6?
Yes, 3 x S atoms at 2.2 % RA each would show that!
That tends to confirm the DMSO solvation that you propose.
You really do need to be able to answer 1).
What reference std do you use for mass calibration?
Try a dilute solution of CSI in acetonitrile.
This give [Cs+(CSI)n]+ in positive ESI over a very wide range (m/z 133 to ~2000).
Also, have you checked for doubly/triply charged signals with any of your CP type molecules? Again, would help to confirm the DMSO-adduct proposal.

Regards,
Motoball

Thanks for your help to answer your questions -
1. i'm not entirely sure about the mass spec reference standards (I should probably find that out next week) but I do know our open access ESI instrument had been calibrated/had maintenance the day prior to me running my samples.

2.The m/z 1340.7 is identifiable ; it has intensity of  approx 50. The 1107.7 peak has intensity approx 100.

3. Yes they are those values.

For my starting CP i've got peaks at 1107.7, 1108.7, 1109.7, 1110.7.

Probably a very stupid question, so is the fact it's 3 x DMSO adducts upon mass spec related to the fact there's 3 amine groups (lysine, 2 tryptophan)...how does that work? for example why do I not have peaks where 1x DMSO or 2 x DMSO ?

Thank You

[MOTOBALL]

Thinking about this possibly being a mass calibration problem, can you

1) name the MS instrument you are using,
2) are you the sole user, or is this a walk-up, open access system (OA)?
3) if it is OA, does the first user of the day run a mass calibration?
4) if it is OA, does every user know that you cannot retune or tweak lens voltages etc once the system has been calibrated, without having to again calibrate?

When I was analyzing samples by ESI-MS under GLP/GMP protocols, first job of the day was to tune with CsI for peak shape and sensitivity; scan m/z 125-1250, in say 3 sec and acquire a calibration file of say 5-10 scans; then repeat that acquisition and save as the calibration check file; run samples; at the end of the day, run another calibration check file to ensure that the calibration had not drifted over the work day.
If a sample came in that required a different mass range, say m/z 500-2500, I would save it for the end of the day, do every other low mass sample as described above, and then re-tune, take a new Cali file (m/z 50-2500, 3 sec), repeat this to give the Cali check file; then analyze the sample and follow with another Cali check file.

Regards,
Motoball

[MOTOBALL]

I see that you posted while I was posting my questions about open access.
So it is OA, and that is possibly at the Centre of any mass calibration issues.

I don’t know whether the 3 molecules of DMSO is correlated to the 1 x NH2 and 2 x NH of the TRP. Certainly DMSO has a S+—O- dipole which May preferentially solvate the primary sites of proton action.

You may need to have your MS person provide a solution of CSI / MeCN so that users can retune and re-Cali for their own needs.

Regards,
Motoball