Lychee/Achee toxins and how to detect them with LC-MS/MS

Did you hear about this? Until the tragedy of yesterday continued to unfold in my newsfeed, I'd never once heard of these awful toxins. I think last night the death toll was up to 100 children (CNN article here).

I'm not alone. This Science article from 2015 is the earliest place I've been able to find the naming of this toxin and it honestly didn't seem super conclusive, but the article states the compound is:  methylenecyclopropylglycine, which may be a close relative of hypoglycin -- a toxin that is found in unripe achee fruit.

ChemSpider says there is an alpha and an N- form (the more recent studies below appear to focus exclusively on the alpha form)

Oh. Okay. That should be fun to detect with LC-MS/MS of any type.

mzCloud doesn't have it! I've tried every acronym in ChemSpider. Hey! Anyone with a HRAM system that has worked with this! Send your MS/MS to mzCloud!!

mzCloud is the best small molecule database in the world in terms of spectral quality. If you haven't used it, you definitely should check it out. You will need to use Microsoft Edge or Explorer because the site requires the SilverLight Extension to run it. It's worth it. If it's a shared computer, do something like this...

 You can also download the full/current mzCloud instance and use it locally through a program you might have ignored in your recent Foundation/Xcalibur/Tune updates. It's called "mzVault" and you can directly access it via FreeStyle if you have a PC that can't freely Bing the internet whenever you want. For now I've got to go somewhere else...

Bongo! PubChem has 3 papers where LC-MS/MS assays were developed for it!  Both studies focus on the Alpha-form and ignore the N- so we'll assume that's what we should be concerned with.

Here's the first one....

The other 2 studies are from the same group and the links to the rest, if you've read this far, are in the PubChem link. (I've been doing a lot of food research this year, and Elfseverer reaaaally dominates food science publications.)

Actually -- this paper might be the one that is the better reference (they used both QQQ and Q Exactive HF in this detection method...but the HF seems to be an after-thought....)

Gross. They dansylate the molecule to detect it. I'm going to be a jackass here. My experience with dansylation is that -- yes -- it increases the ionization potential of molecules -- it is not necessary for today's more sensitive instruments and -- since it is modifying all sorts of things, it often makes your job harder rather than easier. Are you using an old Varian? You've got a pre-TurboSpray 3000? Dansylate away, but if you've got something with 80% or greater transfer efficiency, a way to enrich chromatographically or in the gas phase? I'll always try that first.

If I was a betting man -- I'd say that I could just detect this ion with a SIM/PRM centered here. (On SIM I'd narrow the heck out of the isolation width -- 0.7Da on non-segmented quads (QE/Fusion 1) and 0.4Da on QEPlus/HF/Lumos -- {Correction, brain was off} I'd try the same on PRM, but wouldn't stress if I had to center a 2.2 Da window on the single C13 M+1,  but for both PRM and SIM I'd use a ridiculous fill time -- you can do 3 seconds. One target? Why not?)

(This is the "mass spec scissors" function in ACD ChemDraw -- also available in the free - to - academics ChemSketch version.)

On my Triples -- I'm going positive scan all the way unless something goes wrong and I'm setting SRMs on these fragments and just see how it goes (decimals put in because they'd actually be useful on the PRM):

113.0477
111.0466
82.0657 (the ion that can differentiate between the alpha and N-forms above -- by PRM..too close on a Triple quad)
74.0242
53.0391
44.9977 (you could track this ion on the Exploris!)

To provide confidence that I'm on the right track (...in case I'm waiting for the standard to come in....) I can check the fragmentation of similar compounds in PubChem.

The achee toxin isn't in mzCloud yet either but it is in PubChem

--and in HMDB here.

HMDB has predicted fragmentation spectra for different devices. This is the Q-TOF predicted and it appears to track with my manually calculated/predicted fragments.