Regenerate

[amanda]

Hi

Can anyone tell me how to regenerate silica based C-18 column?

Thanks

[unmgvar]

what did you "put" in it fist of all

[tom jupille]

unmgvar is right: the procedure depends on what's on there.

If the problem is caused by exposure to pH extremes, then the column probably cannot be regenerated.

If the problem is caused by accumulated garbage, the trick is to wash the column with something which will solubilize the garbage. You will probably have to use intermediate solvents in order to avoid miscibility/precipitation problems.

[Okkie]

reverse the column and slowly wash out the dirt.
DO NOT connect the column to your detector !!!

[amanda]

This column is new, probably used for the past 2-3months. I don't know much about what exactly has been passed through this column but my logbook indicates it was last used for plant extracts prepared in bunch of different solvents like chloroform, ethanol, dmso, water etc. The mobile phase was ACN/water with 0.1% TFA.

[james little]

in plasma analyses, we wash with about 10 column volumes of 1% formic in water then 1% formic in methanol, then 10 volumes of methanol.

Seems to regenerate the column. However, we are using a precolumn frit and a precolumn in front of the column. No success in regenerating the precolumn. Maybe the precolumn in voiding, but a sealed unit and cannot work on it.

Did you see the topic previously in this section labelled "Column rescue ideas?"

[Carlos Teixeira]

Dear Amanda,

1 - New column can to have problems with few time, too.

2 - I suggest that you flux it with the following solvent sequence:

Methanol - Acetonitrile - Tetrahydrofuran - Methylene Chloride - Tetrahydrofuran - Acetonitrile - Methanol.
Use NLT 20 volumes of your column for each solvent by step.
Soppose that your column's volume is 2.1mL (250mm x 4.6mm) you must use 42 mL MeOH, 42 mL Acn, 42mL THF, 42mL MC, 42mL THF, 42mL Acn and 42mL MerOH. Use 0.6 to 1.0mL/min. Do not conect your column to detector.

3 - To change the flux direction can damage your column. Use it like the last way.

Have good elutions!!!

Carlos Teixeira

[HW Mueller]

Not that again. Unless it´s a newfangled unsymmetric column or a 20+ year old one, reversal of the column might be very useful, some claim it even alleviates voids.
Also, with bilogical samples it´s always a good idea to start washing with a high % aqueous solvent.