Chromatography Forum

Hello everyone,
I have a strange problem with the determination of two analytes.
I have a method on LC-MS/MS (MRM), 30 analytes - photoinitiators, which I transfer to QTOF.
All analytes, after small optimizations look excellent.
Scope of the method: 10 - 200 ng/mL
Mobile phase: pH=4 , ammonium formate / acetonitrile

I have a problem with two:
1-Hydroxycyclohexyl phenyl ketone CAS: 947-19-3

2-Hydroxy-2-methylpropiophenone CAS: 7473-98-5

These two analytes (unproblematically) are determinable by LC-MS/MS technique. I think I could even reduce the LOQ.
However, I do not see them using QTOF. What I would do, they only appear in the range of about 500 ng/mL - I don't understand why. Their monoisotopic mass is neither smaller nor larger than the others.

after creating a PCDL library, I search the analytes using the ALL IONS method (0,20,40 eV).
I have already tried with other eV, lower fragmentation energy, tried to create pseudo-MRM (Target MS/MS method).

I am out of ideas.

Hi,

If they fly through a quadrupole, I see no reason why they would not fly through a time-of-flight tube, in particular if their MW is in the same range as the other compounds which worked well. In general triple quads used in MRM mode are a bit more sensitive then Q-TOFs, but here you seem to have almost 2 orders of magnitude difference, which is a lot. The loss of signal likely occured in another part of the path. Could you tell which triple quad model and which Q-TOF model you are using?

Hi,
LC-QQQ: Agilent 6460 (AJS ESI)
LC-QTOF: Agilent 6546 (Dual AJS ESI)*

Both methods have the same source settings.
Gas Temp: 350
Drying gas: 8 l/min
Nebulizer: 55 psi
Sheath Gas Temp: 400
Sheath Gas Flow: 12 l/min
VCap 4500 V
Nozzle Volatge 300 V
Fragmentor: 60 (also I tried higher)
*Skimmer: 65
* Oct1 RF Vpp 750

On LC-QTOF, At higher concentrations, I get fragmentation spectra - just what I expect. That means in a certain way I can definitely see them.
However, I still don't understand why I lose them at lower concentrations. Could it be that they have disappeared into the background? Why only these two, when the others are visible, with similar mass and retention time.

The only difference from the others - these two compounds have only one aromatic ring. The others are analogous to benzophenone.

Hi,

If they fly through a quadrupole, I see no reason why they would not fly through a time-of-flight tube, in particular if their MW is in the same range as the other compounds which worked well. In general triple quads used in MRM mode are a bit more sensitive then Q-TOFs, but here you seem to have almost 2 orders of magnitude difference, which is a lot. The loss of signal likely occured in another part of the path. Could you tell which triple quad model and which Q-TOF model you are using?

Mmh, I am not an expert in Agilent mass specs. Maybe someone on this forum can tell if the architectures of the 6460 and 6546 (in particular the path between the source and the first quad as well as the collision cell) are identical or different. Otherwise you could ask Agilent about it.

If you run the QTOF in full scan mode, do you see nice peaks or the problem persists? This may give you hints on whether the issue comes from the collision cell or the stages before the quad.

Mmh, I am not an expert in Agilent mass specs. Maybe someone on this forum can tell if the architectures of the 6460 and 6546 (in particular the path between the source and the first quad as well as the collision cell) are identical or different. Otherwise you could ask Agilent about it.

I've worked with both of these models. Have you tried just doing a full scan with the TOF only (not using Q1 or the collision cell at all) and see if they show up? My other thought is maybe your TOF detector needs to be examined, since you said you can see the compounds on the high end but not the low end.

If you're seeing fragments correctly on the high end but not the low end ... are the fragments for those compounds significantly smaller than your other analytes' fragments? I'm assuming the collision gas for both instruments is the same.

Does your 6546 have the iFunnel technology in the front between the source and Q1? (you'll know if you do). Those were always a pain in the behind.