5 Tips on extending reversed-phase flash chromatography cartridge life

With reversed-phase flash column chromatography becoming increasingly popular for routine purification, understanding how to make the cartridges last (since they cost more) is important to know. 

In this post I will mention a few tips to prolong reversed-phase cartridge life.

Reversed-phase flash chromatography is an extremely useful purification technology for a wide array of compound types.  Since reversed-phase chromatography uses partitioning as its separation mechanism hydrophobic compounds will bind to the lipophilic stationary phase and elute as solvent polarity decreases.

Strongly lipophilic compounds/impurities can bind strongly and build up with repeated injections.  Over time this build-up can cause changes in compound retention and even selectivity so keeping the cartridge clean is important for a long, useful life.

To achieve this goal here are the steps I try to use…

1.       At the end of each purification run flush the cartridge with 100% of the strong solvent (typically methanol or acetonitrile).  You can wash with stronger solvents as well (acetone, THF, EtOAc) but be sure to convert back to your main organic solvent (methanol or acetonitrile)

2.       Clean up crude mixtures to remove possible strong-binding compounds

a.       Liquid-liquid extraction – this can be done without separatory funnels using disposable hydrophobic frit based vessels (Figure 1)

Figure 1. Simple phase separator with a hydrophobic frit allows dense organic solvents such as DCM to elute while water is retained.

b.       Sample clean-up (SPE), Figure 2

Figure 2. Solid-phase extraction is an excellent tool to remove strongly binding lipophilic compounds prior to reversed-phase flash chromatography.

3.       Use dry loading (reversed-phase media) to capture the strongly lipophilic impurities, Figure 3

Figure 3. External dry loading using reversed-phase media as the dry load sorbent is a very efficient way to protect your reversed-phase flash cartridge from contamination.

4.       Flush the cartridge with aqueous solvent to remove pH modifiers and buffers

5.       Always store reversed-phase cartridges in your method’s strong solvent (without pH modifiers or buffers) and keep them sealed to prevent evaporation

What do you use to maximize your reversed-phase flash cartridge useful life?

 

Published by

Bob Bickler

Technical Specialist, Biotage

2 thoughts on “5 Tips on extending reversed-phase flash chromatography cartridge life”

  1. I worry about particles blocking the column much more than I do about accumulated lipophilics.
    I have reverse phase columns which I have used for years and are as good as new, though perhaps a bit discoloured. I put this down to dry loading as a way of introducing the sample and like to think of this as the ultimate guard cartridge. With very black reaction mixtures, a lot of coloured does pass through onto the column but usually elutes before 100% CH3CN and if needed an occasional wash with THF usually cleans of any staining. We have some C18AQ columns which need replacing due to increasing back pressure. With these people have been introducing their water soluble samples as a liquid injection and despite syringe filtering, particles have built up on the inlet frit and eventually killed the column.

    1. Hi Derek,

      Particulate build up can be a problem, especially with HPLC, but I have not seen that issue with flash. Like you, I find dry loading to be a major help keeping the main cartridge clean. I have also found that in some cases I can use a normal-phase clean-up to remove most of the colored materials which allows for a simpler and cleaner reversed-phase run.

      Bob

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