HILIC column for small molecules?

[smiley]

i need recommendations on HILIC column for method development of small molecules(MW<500), acidic and basic.
most reference found online is about bigger molecules like peptides. and no guide available specifically for small drug molecule.
thanks,

[MG]

The Waters HILIC Silica is suitable for small molecules, and I have used it for this purpose. I'm sure there are many others that are suitable, but that is the only one I have used.

[maris]

Big number of applications was published on web and in scientific literature. Can you specificy? And why do you prefer HILIC?
Classical RP doesn't work?

[Patrik Appelblad]

Dear Smiley,

An alternative to the Waters HILIC Silica would be our ZIC-HILIC/pHILIC columns. The ZIC-HILIC columns are silica based while the pHILIC are polymeric, and thus has a wider pH tolerability.

One important difference among bare silica HILIC and our ZIC columns, is that with a zwitterionic phase you will experience weak electrostatic interactions with the analytes in contrast to the strong electrostatic interactions obtained from the silanol groups on a bare silica phase.

More information can be retrieved at www.sequant.com

Best Regards,

Patrik Appelblad

SeQuant AB

[smiley]

is waters HILIC silica just bare silica?
my purpose is to elute small molecule at high percentage organic mobile phase, like >80% ACN, for LC/MS sensitivity.
also i can't find any prep size HILIC (silica) column at waters, they probably just have analytical?

[smiley]

to Patrik,
can you explain why the weak static interaction better than strong one on bare silica?
and if water's column is bare silica, so theoretically any silca column from whatever vendor should be able to do HILIC?
i am a little confused.

[MG]

The Waters HILIC Silica is shipped containing acetonitrile rather than hexane, so that is one difference from a normal phase column. It also comes with a test chromatogram using a HILIC method to separate polar compounds (and a nonpolar compound as dead-volume marker). I don't know if there are other differences. I have seen quite a few ASMS posters where silica columns sold for traditional normal phase were used to run HILIC methods. They would have to be flushed with isopropanol first to remove the hexane.

[Uwe Neue]

Prep-size HILIC is not as straighforward as you might think. The solubility of many samples suitable for HILIC is not very good in the standard mobile phases. This does not mean that it can't be done, it just means that it is not that simple.

[Patrik Appelblad]

Smiley,

I quote myself from a previous application note in LCGC

"The highly polar zwitterionic column provides a unique environment particularly capable of solvating polar and charged compounds, which enables high performance HILIC separations. The zwitterionic stationary phase can interact with charged analytes via weak electrostatic
interactions, as opposed to the strong electrostatic interactions
obtained with plain silica or amino HILIC phases. In practice, this
provides the chromatographer with a larger degree of freedom when
choosing between buffer salts and ionic strength in method
development.


I suggest that you study the pdf files that you may find on the following links:

http://www.sequant.com/1/040610/product ... y_2004.pdf

http://www.sequant.com/1/040610/product ... r_2700.pdf

In those documents you should be able to find the information needed to realize the difference among our columns and plain silica HILIC or amino HILIC columns.

Kind Regards,

Patrik

[HW Mueller]

Patrik,
it would still be nice to explain here, shortly, why your zwitterions give lower ionic interaction. Also, I wonder about the following: With silica you get associative ionic interaction with positive ions (this can be extremely annoying) at pH where you have SiO-, none when you have only SiOH, (I think that I have seen interactions with anions at low pH?). Also one can get anion exclusion. With zwitterions you can get interaction with both, cations and anions, or always with either one or the other ion type if you can associate something with one of the zwitter. Ion exclusion seems also possible if one ion is associated with a counterion. Isn´t that a bit complicated? Good for some specialty analyses?

[Patrik Appelblad]

Dear Hans,

I’ll try to make things as short as possible. The rationale to why our zwitterions give lower ionic interaction is that the phase contains a strong anion and strong cation functionality in close proximity. The attraction is thus counteracted by repulsive forces leading to mild ion exchange interaction hence the HILIC mechanism would be favored. From a practical point of view this means that the chromatographer will have a larger degree of freedom in choosing the mobile phase composition depending on the sample composition rather making a good match for the column.

/Patrik

[smiley]

Patrik,
thanks for the reply.
do you have any theoretical model or practical applications to support your statement?
to my understanding so far, this is pretty much a un-quantified presumption rather than rational assesment.
i have just ordered your column(only silica based available in US, not the polymeric one), but within past few days i tested on both silica column and amino column, interestingly the amino behaves better to some compound.(even the compound very hydrophobic on C18RP).
i do need help on small molecule development(drug discovery compound) on HILIC, not the biomolecules, 99% of the literature is about biomolecules.

[smiley]

i am wondering if the following is true:
as long as a molecule can be made polar/ionic in the solvent, HILIC should be able to work on it.
if this is true, then the question is how to make the molecule ionic? or say what kind of molecules are not wroking on HILIC?

[MG]

I bet naphthalene wouldn't work very well in HILIC.

[HW Mueller]

Patrick, what you say makes sense, of course, but your links didn´t furnish an obvious indication that this is functioning as believed. Also the matter is a bit confusing, because of the terms, strong ion exchangers, strong ion interaction...
On second thought:
Your silica zwitter ion columns must have some silanols as well?
Do you see ion exclusion?
Also the zwitter ion is considered to be the most hydrophilic (polar) form of amino acids (though here I seem to have seen other results), but how this looks like in immobilized zwitter ions? The charges are 6 atoms apart , still they are probably solvated rather than present in a six membered ring (with + and - charges very close), and probabply some counterions are nearby?
In short, more art than science??