I ran my sample in LC-MS and got only one peak. But in MS, it shows three MS numbers and also indicate, these 3 ms ions came from same peak. What is happening in this experiment? Please give me some suggestions. Please see the MS data as an attach file in below.
There are two likely explanations:
1. Maybe you have singly, doubly and triply charged ions (m/z) of the same mass or
2. Maybe you have co-eluting impurities with similar chemical characteristics.
Can you provide any additional information, e.g., the m/z values of your three MS peaks, or perhaps upload the spectrum? What's the nature of the sample - peptide; small metabolite? If it's a peptide, for example, Sonja's answer #! would make sense. And yes - co-eluting peaks are always a possibility, especially at early retention times. Does your sample contain an element with a complex isotope composition? Could it be something as simple as the normal isotope distribution of the molecule?
Also, depending on your solvent system, it might be worth looking at the possibility of ammonium, sodium, and/or potassium adducts.
Thanks Sonja Hess and John Wishnok for nice suggestions.My compounds m/z are 268.15, 312.16 and 339.21 like these. Please see my LC-MS data as an attach file. Thank
May be two peaks are coeluted in same retention time.There may be chance of formation of adducts of adducts.while seeing your mass spectrum,m/z may be-339 or 268,because m/z-312 was adduct of aduct(268+22+22=312, 268+22+38=328),.Scan your sample in negative mode also, then you will get clarification of these adducts.
I also observe sodium adducts - exchange of hydrogen by sodium gives an increase of plus 22 (-1+23) - in the MS spectrum of some of my samples, but it is still one compound and not an impurity.
Mr. Siva Jyothi, you may be right. Thank for your kind suggestions. For you kind information, I upload my full LC-MS data as an attach file. Please look it again if you have time.
Dear Mohammad, please verify the spectrum, which was attached for reference to sonja. In that m/z-328 was clearly showed, that indicates the K+ adduct belongs to Na+ adduct. If these two are adducts, You mayn't get in negative mode, based on that you can confirm exact mass number.
Thank you
Did you subtract background from that mass spec? Could be ions common to the mobile phase. Looks like you're running in negative mode so they won't be Na or K adducts. In neg mode you might get formic or acetic acid adducts (-H+) if either is in your mobile phase, or even Cl- adduct. If peak is definitely due to one component then other explanation is in-source fragmentation. Since you have an ion trap why not do dependent MS to get MS/MS spectrum for additional info? Looks like you have nitrogen in your compound too, based on even m/z.
Look into the possible fragment ions for your target molecule. If we know your reaction's info, it is easy to predict the product.
Thank Dr. David Jeffery. No, my sample run in positive mode in LC-MS.We predict, it is an acidic compound. It is a growth promoting compound.
Are you sure that these ions EXACTLY co-elute, i.e., did you overlay the extracted-ion chromatograms, and expand the m/z axis? Have you tried a different column and/or solvent system? Aslo, Arumugam's suggestion is a good one, i.e., do CID on the two higher-mass ions to see whether or not either of the lower-mass ions are in-source fragments of either of the higher ones.
Jyothi, when the ion contains K+Na, how many charge you should count, 1 or 2?
You can always determine the number of charges by the difference between the C12 and C13 isotope peaks. If it's 1 Da, then the charge is 1; if it's 0.5 Da, then the charge is 2; and so forth. In general, small molecules (e.g., less than MW 600 or so,) will rarely have more than 1 charge. Larger molecules, peptides, for example, may show both singly- and multiply-charged ions.
Thanks John. my question is when a small molecule has two adduct ions like Na+K, how many charge will be counted? I haven't met such case, it's hard to check isotope peaks.
The answer is sort of contained in my previous comment. If there are, say, two sodiums, or a sodium and potassium, then each will add a + charge; essentially analogous with adding two protons. So the resulting ion will be doubly charged, and the isotope spacing will be 0.5 daltons. This is discussed nicely and in detail at: http://www.sepscience.com/Information/Archive/MS-Solutions/233-/MS-Solutions-3-Dealing-with-Metal-Adduct-Ions-in-Electrospray-Part-1.
Thanks, John. So the question about the guy's answer for this thread at beginning for adding Na+K is misleading info.
John is spot on. The Na + K adduct theory didn't make sense; it'd be doubly charged if that was the case.
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