Ammonium acetate and column rinsing

[Camisotro]

I'm running LC/MS experiments with a weak 1 mM ammonium acetate buffer (A) and methanol (B). My understanding is, ammonium acetate in methanol is soluble up to 50 mM.

Typically when I'm finished with my lab time and someone else will be using a different column after me, I run through a program that passes high aqueous through to purge any ionic junk from the column, and then sweeps up to 100% organic to purge any organic junk. To be super-careful, I usually make sure this step is carried out with non-buffered DI water rather than the 1 mM buffer.

But I was thinking, if I'm already over 50x below the solubility limit of ammonium acetate in methanol... would it just be OK to do my column rinse with the solvents I'm using anyway before storage? My thinking is that the 100% methanol would sweep out any leftover ammonium acetate.

[Alp]

I had been told that keeping silica based columns in 100% methanol might not be a good idea.
Not sure if this is correct or not, but I never argued and backflushed my columns with 90/10 or 80/20 methanol/water after use.

I think regardless of what your mobile phase was, it is good practice to flush your column out before the next guy uses it, even if that next guy would be you.

[bbkf]

there should be a product insert that came with your column. It should tell you what to clean it with and what to store it in.

to wash off the buffer with water is a good idea because of salt buildup around the LC system. Sometimes there can be a very small leak before the pump head that over time will build up a white crusty salt chunk.

[HW Mueller]

ctroster, it has been mentioned many times here that one needs to be sure that no precipitation or immiscibility is involved in changing solvents on a column. In your case I would think that getting rid of NH4+ might take quite a bit MeOH, even water, as the cation will probably be stuck on the silanolates. This might not make any difference, for instance if your next mobile phase is acidic the NH4+ will be gone in a jiffy.

Alp, it has been suggested many times, here, to store silica based columns on an aprotic solvent (ACN for RP), so what is the reason for shunning MeOH in favor of MeOH/H2O? The reason can not be storage. If protic solvents are reactive then MeOH/H2O will be more so than MeOH.
I can recall only one situation where changing to MeOH for over night was detrimental, while leaving the mobile phase or MeOH/H2O was ok over night: Some left over matrix proteins caused plugging only when pure MeOH was left over night.

[Alp]

Either ACN or Methanol is fine for short term column storage.

Long term storage they generally recommend ACN, (aprotic), but I was told by a collegue that it may be wise to have some water in the storage solvent to prevent de-hydration of the silica. It was his belief that dehydration/hydration cycles may fracture the silica particles.

Here is a link that states the water content in the ACN (long term storage) should not exceed 50%.

They way I see it, water is a protic solvent. Once you mix a protic solvent with an aprotic solvent, the result cannot be a solvent that is aprotic, can it? It is either aprotic or not. It can't have both.

http://www.nestgrp.com/pdf/colcare.pdf

Alp

[Camisotro]

How do we define long-term versus short-term column storage? Days? Weeks?

I have a C8 column I'd been storing in 100% methanol regularly, for periods ranging from a few days to 2 weeks between use. I recently went on a week's vacation and stored it in 100% methanol during that time. The column was functioning normally prior to my vacation. This week I came back and discovered on Monday that it had completely lost retention of all my compounds (phase collapse?). I tried the usual wisdom of regenerating by pumping a high concentration of ACN through at a high pressure (~250 bar) for a few hours, and tested again - no retention.

Time to order a new one... still I am not entirely sure why it went this way. My understanding is this usually happens if the column is left in 100% water and then the flow is stopped. I don't believe this has ever happened to the column in question; nobody in my group used it in my absence.

[Uwe Neue]

The observation decribed is impossible. I would look for silly errors, like the lines for the aqueous mobile phase and the organic mobile phase are mixed up etc.

[Alp]

BTW,

at $600 (Canadian) a 4L bottle ACN, how many people have tried to switch over to methanol as a replacement?
I'll be rinsing and storing my columns using methanol/water for a while yet.

Alp