Chromatography Forum

We have a compound that behaves well under ion pairing conditions using 1-octanesulfonate. The compound is not retained under RP.
I know that there are IP reagents that can be used for MS like Hexaflourobutyric acid, but I have no experience with this compound, ie. mobile phase concentration etc. and haven't really been able to find default conditions via google. Can someone post a generic starting method or some alternate ion pair reagents for basic componds for MS.

I have tried Hilic but it's difficult because the compound is only soluble in aqueous conditions and the resulting broad tailing peaks from using too strong of a diluent is not acceptable.

You can use anything volatile, but I'd warn you first: ion-pairing reagents will stick to absolutely anything associated with your LC system or your mass spec, are hard to wash off, and they ionise very strongly indeed. If you use an acidic ion-pair reagent, do not expect to be able to carry out negative mode mass spec ever again on that instrument. Well, certainly not for months.

I can't say if octane sulfonate is as bad (probably not) as my experience with nonafluoropentanoic acid, but that was truly horrendous.

Hi Mike,

Perfluoro carboxylic acids can be used as an alternative (up to pentadecafluoro octanoic acid) but as mentioned they do tend to stick around (especially in the degasser membranes) and will leach out over time. I have used succesfully ion pairing reagents in the past in an academic setting and you can clean your mass spectrometer (I can tell you what you used to do) but you will probably make a lot of your coworkers unhappy if you share the instrumentation with other people. You might want to have a look at columns with mixed chemistries on them (in this case SCX-RP or WCX-RP) as an alternative.

Mike,
What conditions did you try in the HILIC mode?

I have tried Hilic but it's difficult because the compound is only soluble in aqueous conditions and the resulting broad tailing peaks from using too strong of a diluent is not acceptable.

Do you mean the sample is dissolved I water and that is the reason for the tailing? I don’t think tailing is the expected problem/experience in such cases but rather fronting.

Best Regards

Mike,
What conditions did you try in the HILIC mode?

I have tried Hilic but it's difficult because the compound is only soluble in aqueous conditions and the resulting broad tailing peaks from using too strong of a diluent is not acceptable.

Do you mean the sample is dissolved I water and that is the reason for the tailing? I don’t think tailing is the expected problem/experience in such cases but rather fronting.

Best Regards

50 mmol Ammonium formate pH 4.0 mobile phase A
ACN Mobile phase B
Standard screening gradient.
MP-A:MP-B
10:90 0-2.5 min
step to 50:50 at 10min
and hold for 2.5 min

The compound is not soluble in ACN. I've tried to weaken sample solvent with various mixtures.
I've achieved solubility in 20% EtOH in ACN, but even though this should be weaker than initial conditions, the peaks are still broad ~1.9 USP tailing.

Column is Luna 5µ HILIC 150x4.60mmx5µm Part number 00F-4450-E0

Try another column (preferably another brand) if accessible.
As I pointed out earler you shouldn't be experiancíng tailing if strong solvent effect is the issue.

Best Regards

Unfortunately this is this only Hilic column we have. I'm not sure I described this correctly as a tailing issue. The peaks are just broad and they resembled some example chromatography of material dissolved in 100% from a troubleshooting site.

For future reference can hilic mode be used for analytes that are only soluble in h2o, meoh etoh?

Mixed-mode is the way to go, you can use all LC/MS compatible buffers you can imagine within recommended pH range, and since this is RP/cation-exchange method you will not have a problem with solubility of sample:
http://www.sielc.com/upload/file/pdf/SI ... e_2004.pdf
http://www.sielc.com/upload/file/pdf/SI ... e_2009.pdf

You can look at list of this applications and see something similar:
http://www.sielc.com/Applications_By_Compound.html

we were doing this since 2003

Hi Mike,

You wrote:

For future reference can hilic mode be used for analytes that are only soluble in h2o, meoh etoh?

That’s what HILIC is there for.

If I saw the chromatograms, perhaps I would’ve been of more help, but as I understand your description of the HILIC test/experiment, I’d say it’s not due to the sample solvent, but rather the column and potentially the chromatographic parameters – such as flow rate, temperature, sample load etc.

Best Regards

Mixed-mode is the way to go, you can use all LC/MS compatible buffers you can imagine within recommended pH range, and since this is RP/cation-exchange method you will not have a problem with solubility of sample:
http://www.sielc.com/upload/file/pdf/SI ... e_2004.pdf
http://www.sielc.com/upload/file/pdf/SI ... e_2009.pdf

You can look at list of this applications and see something similar:
http://www.sielc.com/Applications_By_Compound.html

we were doing this since 2003

We had an old mixed mode anion column that I tried and it didn't perform well. Could have just been the column though. I'll probably suggest that we buy a few of these. Ended up with a separation using 0.1% HFBA that produced adequate sepation.

Hi Mike,

You wrote:

For future reference can hilic mode be used for analytes that are only soluble in h2o, meoh etoh?

That’s what HILIC is there for.

If I saw the chromatograms, perhaps I would’ve been of more help, but as I understand your description of the HILIC test/experiment, I’d say it’s not due to the sample solvent, but rather the column and potentially the chromatographic parameters – such as flow rate, temperature, sample load etc.

Best Regards

I mean the sample dilluent. If you inject a sample in 100% water in hilic, chromatograpy is terrible. I'll remember to post a chromatogram for my next problem. Thanks for helping.

Hi Mike,

We have a compound that analysed using tetrabutylammonium hydroxide. May i change tetrabutylammonium hydroxide with tetrabutylammonium hydrogen sulfate? What is the difference between tetrabutylammonium hydroxide and tetrabutylammonium hydrogen sulfate?

Best regards

Hi Mike,

We have a compound that analysed using tetrabutylammonium hydroxide. May i change tetrabutylammonium hydroxide with tetrabutylammonium hydrogen sulfate? What is the difference between tetrabutylammonium hydroxide and tetrabutylammonium hydrogen sulfate?

Best regards

The Counter-Ion, the pH and the ion-strength?

Nevertheless, it would be interessting if the original-poster has tried a mobile phase modified with TFA in RP-Chromatography?