Need help with this compound's MS data

[bismuth]

This is the first time I deal with MS data as I'm a student in pharmacy doing my thesis in the organic chemistry lab of my university, so please help me, thanks a lot.

I'm trying to synthesize T3 (triiodothyronine) from tyrosine, and after 3 reactions I suppose to obtain this compound:

http://pubchem.ncbi.nlm.nih.gov/summary ... loc=ec_rcs

MW= 341.2735g/mol

I send a sample of the compound to a center to perform LC-MS (as my lab doesn't have MS equipment), and they send back the spectra for me with email:


As you can see, the peak with m/z= 338.8 is the highest peak, it's about M-2 compare to my expecting compound.
All the information about the LC-MS process I can have is present in the spectra above (name of instrument...)

So can I conclude that the compound I synthesized is the one I expected? Why is it M-2?
Please help me with this as I'm really confused now and I've synthesized a fairly large amount of this compound.

I'd really appreciate your help.

[Mattias]

The mass you should see on the MS is 340.1 (monoisotopic mass [M-H]).

If the MS was properly calibrated, you have synthesised a different compound.

[bismuth]

Thanks for your help.
Is there a possibility that the M-2H fragment is the most stable one? Maybe it lost 2H and 1e?
If the unknown compound's MW is 340 then it's really hard to understand for me

[Mattias]

The monoisotopic mass for the structure that you have provided is:
C13H15N3O8 = 341.086 Da

Since the MS was run in negative mode, you have to remove one hydrogen giving
C13H14N3O8 = 340.079 Da (this is the mass you should get on the MS)

I find it hard to believe that there can some exotic ionisations causing this. My best bet is that the MS has not been properly calibrated. Ion traps are known for not having the best mass accuracy, even when they are calibrated.

[Don_Hilton]

Do you have any other data on your reaction product, like melting point and IR? The question that comes to mind regarding the LC/MS is: Do you know if the spectrum corresponds to the main compound in the sample? Was this run by direct infusion into the MS or by an LC technique. If by an LC technique, to you know that your reaction product would have eluted from the LC column - at the time shown on the spectrum?

[bismuth]

Thanks a lot for all the replies, it'd be really bad if the MS has not been calibrated properly cos it's kinda expensive for me to have that spectra.
@Don_Hilton: Melting point is 115-116oC, and according to literature MP is 120-121 oC
I haven't had the IR data yet (I also need to send a sample to the center if I want IR data)
I'm not sure if it was run by direct infusion into the MS or by an LC technique, but I think it's LC technique, but sadly I don't know about my product's retention time.

[jonmike.reed]

The 1.6min retention time is suspect. This is pretty close to what you'd expect if your compound elutes in the void volume, in which case the "LC" component of LC-MS was essentially useless. I don't know your column dimensions, solvent system, or flow rate, and so my previous statement isn't 100% conclusive, obviously.

Are you able to get a direct infusion MS done of this compound? Also, do you have a reference standard against which you can compare retention times? If the MS analyst pulled a single full scan MS spectrum out of a 20minute LC-MS run, how did he/she know which was the correct compound? Could the analyst have run an extracted ion chromatogram of the known
[M-H] value for your compound to find it if it is present in the sample you submitted? Charge modifiers can affect this kind of data too - if the wrong one is chosen for a given analyte, it can drastically affect ionization efficiency such that it renders your molecule(s) of interest invisible in the MS.

Lastly, if this is going to be an ongoing issue, you might be able to save yourself some time and headache by working out the appropriate LC conditions (using MS-compatible mobile phases) before submitting the samples for analysis so that you can inform the analyst of the expected retention time of whatever compound you're studying.

[Don_Hilton]

Before going back to the MS lab. The melting point is a definite red flag. Review your synthetic steps and see if you can see a reason for forming the wrong product.

Are you following a literature procedure for this synthesis?

[bismuth]

Thanks for all the suggestions.
Yes, I'm following a literature procedure for this synthesis and everything seemed to go fine until now. I was worried about the melting point, that's why I thought I need MS data. If the Mass was 2 Dalton less, then maybe a double bond was formed somewhere, but 1 Dalton less??
Now I'm hoping this is just a calibration issue, and since I don't have a reference standard to do what jonmike.reed suggested, I think I will just go on with the next reaction and confirm the next compound's structure with MS and IR, if the next one's right, then most likely this one is ok.
I will post the result here latter, again, thank you a lot

[Don_Hilton]

Melting point is typically a very good characteristic for a compound. Five degrees suggests strongly that you have something else. If you can describe the reaction and the conditions you were using, perhaps somone here could ask a question or make a suggestion that would help clarify what is going on.