Chromatography Forum

How do you regenerate columns? IS THERE any body to share views with me

About 20 years ago there was a popular song in the US by Paul Simon titled "Fifty Ways to Leave Your Lover". There are at least that many ways to regenerate a column, depending on what's wrong with it in the first place.

We regenerate our C18 columns two ways.

We wash the monochrom C18 columns with 1% formic in 95/5 water methanol, then 5/95 water methanol, then pure methanol. We reverse the flow of the column during the washing. Not all columns can be reversed. Even though these have arrows for flow, don't mind being reversed. Mainly use these columns for analysis of drugs in protein precipitate plasma. We clean them when the peakwidth of the basic drugs of interest start to broaden and tail.

We use Polaris C18 columns for routine survey runs of a wide variety of compounds. The failure mode is different for these columns. The main problem is that peaks begin to split for early eluting peaks. We take the front nut and filter off the column, and there is always a void. We take some C18 stationary phase (doesn't always match that in the column) and slurry in isopropanol to make a paste. Take a spatula and spread a little on head of column, place nut back on column with filter, and flush some methanol through the column. Usually fixes the column for a few more weeks. Often do this many times to a column before tossing it. Sometimes, dig out a little bit of material on the front of column before topping it off again if the material at front of column is particulary dirty looking.

Amazing what cheap people will do to save a little money..

Probably better just to get a new column, but old habits from saving money in graduate school are hard to break..

Also, look at the top hits on a Google search of "regenerate HPLC columns."

About 20 years ago there was a popular song in the US by Paul Simon titled "Fifty Ways to Leave Your Lover". There are at least that many ways to regenerate a column, depending on what's wrong with it in the first place.

Jupille, you have to undestand my question in a different way. How do you ensure that a regenerated column works with the same capacity as a new one ?

Your chances that a regenerated column works exactly the same way as a new column are slim to none...

(Of course, I represent a column manufacturer who spends a lot of time and money to make sure that all columns of a particular brand are the same as they go out the door...)

Hello

I've got a couple of powerpoint slides on how to regenerate a column and restore its original performance. I've evolved and tested these regeneration methods myself and I'm quite sure they work. These are part of my regular HPLC troubleshooting and maintenance program that I've conducted in various pharma companies in Mumbai and Bangalore.

Columns are expensive and I can appreciate your efforts in trying to save your columns.

Do you want me to email these to you? If so, let me know your email ID.

Thanks and rgds,

S.K. Srinivas

we use various test mixtures to check the performance after we regenerate the columns and the peformance is acceptable. If not, we toss the column.

How do you regenerate columns? IS THERE any body to share views with me

No method is available as such that ensures a column of same efficiency after regeneration.

If you are doing it for 1st time and as Sir Jupille said 50 ways I would suggest you follow the procedure in your accompanying column pamphlet. The manufacturer has better idea how to save column.

One good LCGC artilce over Column regeneration...nice one check it.

Regards,

Amaryl.

Hello

I've got a couple of powerpoint slides on how to regenerate a column and restore its original performance. I've evolved and tested these regeneration methods myself and I'm quite sure they work. These are part of my regular HPLC troubleshooting and maintenance program that I've conducted in various pharma companies in Mumbai and Bangalore.

Columns are expensive and I can appreciate your efforts in trying to save your columns.

Do you want me to email these to you? If so, let me know your email ID.

Thanks and rgds,

S.K. Srinivas

SRINIVAS , would you please e-mail those articles and slides which may be useful for me. My id is rc_12321@yahoo.com

Hello

Do you have a corporate email ID I can write to?

rc_12321, James Little already gave the same answer that I would have. I'll elaborate a bit and comment that any method you run should have an associated system suitability test and specification (if it is a validated regulatory method, such a specification is required, but I recommend that it be done for any method). If your regenerated column passes the test, it is suitable for that method.

Hello

Do you have a corporate email ID I can write to?

I do not have any corporate ID.
Do you have any problems to mail to my personnel id? If you send the details we will try to put it into practice in our lab.

rc_12321, James Little already gave the same answer that I would have. I'll elaborate a bit and comment that any method you run should have an associated system suitability test and specification (if it is a validated regulatory method, such a specification is required, but I recommend that it be done for any method). If your regenerated column passes the test, it is suitable for that method.

Jupille, your suggestion may hold good for single peak analysis i.e dissolution, assay etc , BUT for related substances analysis espcially of
stability samples one cannot easily rely on regenerated columns because of expected and unexoested degradation processes which are likely to take place over a period of storage.Is it possible to identify those unexpected peaks with regenerated columns?How to confirm that regenerated columns can also be useful in this situation?

Here are my 5c worth...

We test columns with a very sophisticated test that tests for various contributions to retention at pH 7 (published in J Chrom A 849). If a used or refurbished column gives identical results to a new column with this rather sophisticated test, I will place very serious bets that the column will show the same results under most real-life circumstances.

That said, I also know that there are limitations. The test does not test for the complexing properties of a column, and a column that has been exposed for some period of time to a stainless steel instrument will show different properties for complexing analytes. We test for this, but I have no clue how I can fix this in a world of HPLC instrumentation made from stainless steel.

We also test columns internally under other test conditions.

rc_12321: If your method (for related substances or anything else) has been validated according to ICH recommendations, then it has an associated system suitability test. If you pass system suitability, then your system (column, instrument, batch of mobile phase, etc.) is, by definition, suitable for the intended purpose. As far as I know, no distinction is made regarding the provenance of the column (new, old, used, regenerated, whatever).

The argument is no different for a new column from a different lot, or for a column which has been in use for 6 months, or for a regenerated column.

If your system suitability test cannot weed out unsuitable columns, then the validation needs to be re-done and meaningful system suitability criteria established.

If the question is "can I be sure that a regenerated column will be able to separate new degradants which may appear?", the answer is "NO". But that same answer applies to new columns as well.

That said, my personal feeling is that columns are inexpensive compared to the cost of botched assays. I would rather minimize the risk and use new columns from established sources (but still run the system suitability! )