Chromatography Forum

Hi everybody,
My question is about a column lifetime at higher pH. I need to separate some analytes on a reversed phase at really high pH.

I use 20mM ammonium acetate pH 10 vs. MeOH on a Gemini NX column. I also use gradient elution (2-95% MeOH) which probably even makes thigs worse. I am really dissapointed with its lifetime. <200 separations were performed on this column and it went wrong (some peaks which were pretty focused (30s) are over 3 minutes wide at the moment).

Does Waters have some C18 columns with extended pH range ? (we have one Xbridge coulmn, but it is too short to repeat the separation) Do you have any experience with the lifetime of Xbridge in higher pH ? ( or any suggestions about an alternative coulmn from Waters for this type of separation, Uwe?)

Thanks.

I have been very impressed with xbridge at high pH. I have experienced excellent column lifetimes with gradient methods at pH 10 with ACN as the organic.

thanks floppy, that´s the thing I wanted to hear. btw using ACN shortens the lifetime more than MeOH.

I am sorry. flappytango of course.

Though they use older column technology, XTerra columns are good for wide pH range applications as well.

I can not speak for the XBridge columns as I have not used them, but I do know that a Gemini column will give nearly the same selectivity as a XTerra will.

i was thinking about it some more

We have investigated the use of Gemini NX as a backup to Xbridge also. My coworker has done this. I do not know all of the details but i believe he found them to be robust.

So if you try Xbridge and it is significantly better in your solvent system that will be interesting. It my understanding is that xbridge and gemini nx utilize similar chemistry (ethane cross linking). The waters is a full hybrid particle and NX is silica core with hybrid surface.

Also, In our method Xbridge performed much better than Xterra and standard gemini from a lifetime standpoint.

good luck

If you want to go outside the Waters line, there are many options for high pH use. All of the other major manufacturers offer high pH options, and all "claim" to be stable.

But you have to realize that in all of these silica based columns you are just holding off the flood, not preventing it. Silica dissolves at this pH, and you can do a pretty good job of covering it up, but the thermodynamics are still there.

The polymeric columns are quite stable, although efficiency is usually less. Zirconia phases are available (Zirchrom) that will tolerate this pH even at high temperatures. Several different reversed phase options are available, although selectivity is different than with silica based columns. And I think there is a titania phase sold by someone, but can't remember who at the moment.

thanks, I will try it. I was satisfied with the separation on the gemini NX.
I could achieve baseline separation of cis- trans- isomers.
I am a little bit worried though. The gemini NX has 3 um particles, while the XBridge 3,5.

One additional thought:
Quartz is a form of silica, and also may have a reduced lifetime at pH 10. So, watch the windows on your flow cell for changes.

To get the same separation using a different particle size, all you need to do is make sure you keep the linear velocity of the solvent system the same.

In a nutshell:

F2 = F1*(dc2^2/dc1^2)*(dp1/dp2), where:

F1 = original flow rate (mL/min)
dc1 = original column diameter (mm)
dp1 = original column particle size (um)
dc2 = new column diameter (mm)
dp2 = new column particle size (um)

During the development of the XBridge product, we did a lot of comparisons with other packings. At alkaline pH, the XBridge column is at least 10 times more stable than XTerra, at least 20 times more stable than Gemini, and at least 50 times more stable than the best silica alternatives.

If you contact me, I can send you the test results.

For further comparison, according to Phenomemex's literature, the Gemini-NX is approximately five times more durable than the Gemini at high pH.

The durability graphs for Prednisolone and Amitriptyline in a 7 minute gradient of 100% pH 10.5 ammonium bicarbonate to 90% CH3CN and hold for 7 mins, indicate a four times increase in durability for the NX to 80%N, and five times durability to 50%N, reflecting the more gradual decline of the NX.

They don't offer a durability comparision with the Xbridge.

Bruce Hamilton

Thank you for reply, Uwe. I would appreciate some more information about different column lifetime comparison. I have sent you an email. Thanks once more.

Phobius: the material has been sent to you...

Thank you all for your replies, suggestions and tips. I got the information that I needed.