HILIC question

[sanks086]

Hello,

I am currently working on a project to increase the capacity factor of 3(2H)-pyridazinone. I am using a type c silica diamond hydride "hilic" column. I have been using ACN as my organic mobile phase with aqueous ammonium acetate as my aqueous buffer. In accordance with several papers written on HILIC, I have been using ACN in very high concentrations ~97% hold for the first 10 and final 10 minutes. My run time is typically 30 minutes long. Although HILIC is supposed to provide great retention times/capacity factors for polar basic compounds, my compound is showing retention times and capacity factors similar to that when it was run in reverse phase. Also, my buffer concentrations have been from 10-60 mM and none have showed any signifcant improvement of capacity factor. I have read that perhaps MeOH may be suitable as the organic. I know that this implies that I will be using 2 very strong eluents as my mobile phase, but I was wondering if I should bother using this strategy in increasing my capacity factors, or even perhaps use a different buffer.

Thanks

[sanks086]

sorry, the pH of my aqueous buffer is at 5.

[Uwe Neue]

Why do you think that the hydride diamond column is a HILIC column?

Also, HILIC pure HILIC requires a surface layer of water, which is why you start with around 95% organic.

Methanol is the wrong way to go for HILIC. It is a strong solvent, and reduces retention.

[sanks086]

Thanks for your response. Actually, I am not the one calling my column "Hilic," its actually the manufacturer Microsolve. I think because we are using this column along with an aqueous mobile phase, we are considering the method we are conducting a pseudo hilic method. Also, regarding methanol, I understand that it is a very strong eluter in hilic applications, but several papers I have read have used it as an organic as an alternative to CH3CN. I tested both neat CH3CN and MeOH on the column and surprisingly got great improvement in my capacity factor. This was very strange because we did not pre condition this diamond hydride column at all. We were also able to reproduce our results. Again, thanks for your response.

[Uwe Neue]

In general I expect greater retention with MeCN. Did you recently go back to MeCN and tried it again?

[HW Mueller]

Neat MeOH is so different from MeOH + H2O that the former has been called normal phase LC, the latter HILIC.
If the analyte gives similar rt in reverse phase compared to org.-H2O with polar stationary (HILIC), and then changing the conditions to neat MeOH or ACN, one shouldn´t be surprised at getting confusing results.

[sanks086]

I guess I should have labeled this question "aqueous normal phase" as compared to "hilic"

[Uwe Neue]

There is no difference between aqueous normal phase and HILIC. Same thing!

[zoom]

Based on my experience with type-C silica (silicon hydride surface), using methanol in place of acetonitrile is not a good idea. The -OH on the methanol combined with the small aqueous portion of the mobile phase will saturate the siloxane oxygens that allow hydrogen bonding to take place on the stationary phase. Acetonitrile can not form hydrogen bonds with the stationary phase.

[HW Mueller]

Saturation of the silane oxygens? What is that? How do you know?

[zoom]

Look at the structure of type-c silica. How could it retain a polar compound? The only way I can see is via hydrogen bonding with the oxygen atoms. There are no silanols. If both mobile phase components have polar hydrogens that can form hydrogen bonds with the oxygens, the compound being analyzed will have little or no interaction with the stationary phase.

[Uwe Neue]

All HILIC interactions that I have seen on the type-C silica are the same, or eat least very similar, to the ones on silica. The success of silica in HILIC is based on silanol groups, right? So what does that lead you to conclude about type-C silica?

[zoom]

On type c silica, the hydrogen bonding is one-way. You must have an -OH or an -NH on the compound being analyzed in order to get good retention. It works better than ordinary silica for some compounds and not as well for others.

[HW Mueller]

Even when one, strangely, opts to ignore the info given by Uwe, there is still the question of whether there is only retention via hydrogen bonding. There are obviously other methods of interaction, one should look at the old equations of dipole-dipole, dipole-induced dipole, etc., interactions.

[Uwe Neue]

-OH functions and -NH2 functions on the analytes give increased retention on silica. Again, what is the difference on type-C silica?