top of page
  • Liz

Improved Chromatography Column Packing Method: The Best of 2 Worlds

Updated: Mar 17


To run column chromatography, properly packing the column is essential to achieving good separation of compounds. When preparing a manual silica column, there are 2 common ways to do this:

1.Dry-packing. Powder silica is poured into the column, followed by wetting of the entire column by adding solvent. Advantage: This allows easier handling of the silica, that one can easily transfer a desired amount or height of silica onto the column. Disadvantage: the column could be poorly packed, non-homogeneous and have bubbles inside affecting separation performance.

2. Wet-packing. Powder silica is first poured into a flask, added solvent to form a slurry, and then transferred onto the column. Advantage: the column will achieve the best packing quality of a manual column. Disadvantage: pre-forming a slurry, and transferring of the slurry silica onto the column are extra steps, and which the slurry needs multiple rounds of extra rinsing to be fully onto the column. Furthermore, it is not straightforward to see how much silica should be added into the flask.

Can we do better? We should, and yes we can. To achieve the advantages and avoid the disadvantages of both ways, here is an improved method. In this method, powder silica is directly added to the column, with wet-shaking to achieve good quality homogeneous packing. Detailed steps below.

Step #1 - Stuff the bottom of the column with cotton.

A thin narrow swirl of cotton should be applied to the outlet end of the column. The cotton should not be laid flat on the bottom, but gently sticked into the exit stem. This works better with a removable flow control valve column, but will work all fine with the common design column. Once the cotton is added, wet it with solvent. To avoid unnecessary back pressure, the cotton should not be too thick or tight. Wetting the cotton will expand it and help the seal. If there is a frit at the bottom of the column, then this entire step is to be happily skipped.

Step #2 - Adding powder silica.

Add powder silica into the column. Try to add silica up to no more than 2/3 of the height of the column. The mass of your silica can be estimated from the volume (which the ConciseColumns by AAdvance Instruments will provide you with convenient volume marks, pending launch!) Then, add a measured volume amount of solvent to your column. To know the volume is often neglected in current practices but is very useful-it will tell you the amount of solvent needed to fully wet the silica, i.e. your column volume. The column volume will help you estimate how much solvent needs to flow out before your first compound shows up. For example, if the Rf of your compound on your column is 0.5, then it should start coming out at double the column volume amount of solvent. To note, the flow valve should be open.

Step #3 - Thorough shaking.

This is the step that allows the silica to achieve good homogeneous packing. First, cap the column with a glass stopper, and then use a plastic clamp to secure it. Next, in a way similar to shaking a separatory funnel, shake the column along its longitudinal axis as vigorously as is needed, so that all silica including parts all the way at the bottom of the column is well shaken. Release the pressure from the valve time to time the same way as a with a separatory funnel. Once all silica is loose, the shaking should be changed to a gentle side-to-side rocking.

To note, if your cotton is stuffed from the top down, make sure it is not sticking out from the stem, otherwise the sticking out part could become the anchor point that will get dragged out during shaking.

Step #4 - Settling and rinsing down.

After all the silica is wetted and properly shaken, set up the column in its ready-to-operate pose, allowing the in situ formed slurry to settle. On the column walls there will be remaining silica, which a small amount of solvent will be able to rinse it down. Gently remove the glass stopper, and then open the valve to allow the solvent to flow out. Collect this solvent for rinsing or later use.


Say goodbye to making a slurry in an intermediate flask, doing hassle-filled transfers and washes. At the same time avoid a poorly packed column. If you have a fritted column, follow steps 2-4, otherwise stuff the column with cotton following step 1.


Post: Blog2_Post
bottom of page