How does mobile phase organic solvent choice impact reversed-phase flash column chromatography?

Organic and medicinal chemists frequently utilize flash chromatography to purify their reaction mixtures. Normal-phase flash chromatography is most often used but may not the best methodology, especially when the compounds are quite polar and/or ionizable.

For these molecules, reversed-phase flash chromatography is preferred but often is not used due to an uncertainty regarding the best solvent choices and the reversed-phase mechanism.  In this post, I will discuss how organic solvent choice in reversed-phase chromatography can influence the chromatographic separation.

Read more here.

Published by

Bob Bickler

Technical Specialist, Biotage

3 thoughts on “How does mobile phase organic solvent choice impact reversed-phase flash column chromatography?”

  1. Thanks for the data Bob. We originally were using MeOH/water exclusively with our analytical LC-MS (and reverse phase Biotage) since it is cheaper than CH3CN, but it seems that we have had far fewer clogs when we switched to CH3CN. This may be a factor of our specific compounds, but we make a wide range of molecules. I do however recommend investigating both MeOH and CH3CN as you suggested for difficult separations. We have occasionally also seen mixtures that have significantly better resolution with one or the other solvent.

    1. Hi Chris,

      Thanks for sharing your experience with lower compound solubility in methanol-based reversed-phase chromatography. Personally, I have not encountered this phenomena though I have seen the opposite (MeOH soluble / MeCN insoluble). All the more reason to investigate chromatography with several organic solvents as you recommend.


  2. Bob
    Just a few thoughts. When it comes to choosing between normal and reverse phase I tend to favour normal phase because it’s easier to evaporate the fractions. However on occasion reverse phase is the more convenient if it means no work up. Making amides is very common. I use HATU as reagent and do the reactions in a small amount of DMF then when complete the whole reaction is injected onto my reverse phase flash column. A gradient starting with 85% water will wash off the polar reaction by products then later comes the amide. It’s very quick to purify several reactions and the product fractions go on the freeze drier overnight.
    The problem with methanol for reverse phase is that all our open access LC/MS systems use CH3CN so it’s difficult to know if MeOH would give a better result. I would have to take a chance and hope to get lucky. We really need a simple hplc machine set up for side by side comparisons.

Leave a Reply

Your email address will not be published. Required fields are marked *