Regenerating the column

[vishnu]

Hello guys,
how to make a whatman partisil 10 SCX column (10microns) to work long?
we are getting tired trying to regenerate, its no use.
we are not getting the same resolution even for 10 days.

Can anybody help???

[JGK]

Practical HPLC by V.R. Meyer has 2 options

At a flow rate of 0.5 - 2.0 mL/min pump:

Option 1

75 mL water while injecting 4 x 200 µL DMSO
75 mL of tetrahydrofuran

Option 2 (works for both anion and cation exchangers)

75 mL water
75 mL of a 1.0 - 0.5M solution of buffer used previously (to increase the ionic strength)
75 mL water
75 mL acetone
75 mL water
75 mL 0.1M EDTA sodium salt
75 mL water

[tom jupille]

If your samples or your mobile phase contain lots strongly bound cations (Fe+++, for example), you may have to regenerate the column frequently (e.g., every day). The way to avoid the problem is to keep those contaminants from getting on the column in the first place (better sample cleanup; use of pure buffers and salts; make sure your DI water system is working properly).

[DSP007]

Practical HPLC by V.R. Meyer has 2 options

At a flow rate of 0.5 - 2.0 mL/min pump:

Option 1

75 mL water while injecting 4 x 200 µL DMSO
75 mL of tetrahydrofuran

Option 2 (works for both anion and cation exchangers)

75 mL water
75 mL of a 1.0 - 0.5M solution of buffer used previously (to increase the ionic strength)
75 mL water
75 mL acetone
75 mL water
75 mL 0.1M EDTA sodium salt
75 mL water

Not do its do !
Its good method from silicagel RP C8,C18,Phenyl column , but dead method for alkanamine ion enchance column. For its - must not use acetone.
See old book.
http://www.nestgrp.com/pdf/colcare.pdf
Oh - its question for SCX. But also Not do its do !
See old topic
viewtopic.php?f=1&t=5999&start=0

[vishnu]

thanks for your reply all of you. i tried all you told, before soe months. but its not working. we have problem with resolution all the time.

So only i am asking for help.

[vishnu]

can anybody help me to get the resolution for long time in the above said column?
i am facing so much problems with that.

[tom jupille]

Let's back up a bit. As I read your initial post, you *can* regenerate the column, but the problem is that you have to regenerate it every ten days or so. Is this correct?

[vishnu]

Dear Tom, thanks for your response.
Not like that. i am not getting any improvement by any methods for regenerating the column.
Can you help?

[Gerhard Kratz]

Based on aromatic benzene sulfonic acid groups. Supplied in the ammonium form. Excellent for separation of nucleosides, amino acids, polyamines, drugs and other cationic species. Capable of being loaded with specific metallic cations for use in ligand exchange chromatography. Stable over pH range 1.5-7.0 when used in conjunction with a Solvecon mobile phase conditioning-column. Exceptionally stable Si-O-Si-C bond, both thermally and chemically.
That I found on the Whatman website, let us know about your mobile phase conditions, pH etc.

[tom jupille]

Not like that. i am not getting any improvement by any methods for regenerating the column.

Okay, following up on Gerhard's post, some information about your mobile phase and sample would help. Regardless, I'm afraid there is no "easy" fix.

In very general terms, decreasing retention on an ion exchange column comes about from one of two causes:
- some very strongly bound ion sticks to the ion exchange sites
- some of the bonded phase is cleaved away
In both cases, the effect is to reduce the number of available ion exchange sites.

If your problem falls in the second category (usually when operating at extremely low or extremely high pH), there is nothing you can do to fix the column. You must either change the method or simply replace the column frequently. If your problem falls in the first category, then there is hope. What you need to do is to figure out what is binding to the column (proteins? heavy metal cations? . . . ? ) and flush the column with something that will solubilize or displace those contaminants. If you look at JGK's post as an example, it would be designed to take off fatty materials and/or metals.

[vishnu]

Thanks Tom,
I think you are right. i am using ammonium dihydrogen phosphate in water with 3 pH as described in the pharmacopoeias (BP / EP / USP).
As it is based in the pharmacopoeia, i cant change the method.
I am changing columns frequently. i had no choice. so what i am searching for new methods for regeneration.

Is there any methods to Regenerate???

[tom jupille]

You know more about your samples and water than I do. What you have to do is to make an educated guess as to what kind if garbage might stick and the flush with something that will displace it. Something like EDTA for metals or perhaps ethanol for fatty materials.

My best advice would be to contact the vendor and see what *they* suggest.

[vishnu]

I am from pharmaceutical field and i am using this HPLC for anayzing metformin hcl. i am not using any fatty acids or amino acids.
This HPLC and column is dedicated for metformin only.

what to do for this?

[Sazabi]

Base on the information you've given, the column's resolution detoriate after 10 days.

Do you run this test daily? Do you wash/flush your column after each run before it detoriate? Maybe it's simply your mobile phase have been stay in the column for too long.

Otherwise, maybe you can ask other suppliers if they can provide a more robust column to replace the one you are using.