Professor Davis briefly reviews the anatomy of a chromatography column and how packing and loading a column can affect the quality of the subsequent separation.
I would agree with other comments -- This is the best presentation on column chromatography that I've seen or read. Thank you very much for preparing and posting it. Your efforts are much appreciated.
WOW this is by far the best video on chromatography. What would usually take someone hours of reading is all condensed in to one 10 min video. Chromatography was very confusing for me initially took hours to figure out. If I would have seen this video first I could have saved my self alot of time.
Best video on internet on chromatography!Every single detailed explained well!TY sir! My teachers were not able to give answers to my questions.By u surely took away all my headache!Ty once again!and keep uploading more!
Nice video! I am quite a cowboy when it comes to "gently" adding my mobile phase so I prefer putting a royal layer of sand on top as well, especially for large columns.
Yes, a little sand on top can be a column-saver sometimes. In my teaching labs it proves to be more of a liability, though. Many students manage to disturb the top of their columns when adding the sand intended to protect it. Ha!
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What you are referring to is a slightly more complicated method known as a 'gradient' run: one in which the eluting power (read: polarity) of the solvent is increased through the course of the run by mixing solvents in different proportions as you proceed. effective? Yes! Suitable for an ten minute introduction? I say no. Thanks for the comment!!!!
Beautiful! Very nicely done video. It's really helpful. How to know when to collect our samples if we're running a column that doesn't show color (e.g protein desalting)
Hi, thank you for nice and clear sumarise of column chromatography, but there is one more thing I would like to know. Do you have any rule to determine, how much silica gel you will need for individual separations? P.S. Sorry about my english, but I am not from country, which english is spoken in.
Question: Why does the more polar layer (purple) of the sample move slower then the polar layer (yellow)? As you said, your yellow layer is less polar, but moves more quickly in the stationary phase. The stationary phase is in this case a silica slurry which is also polar, so why doesn't the more polar purple layer move faster than the less polar yellow layer? If the stationary phase was Alumina, would the movement of the layers be reversed? Thank you for the very informational video, much appreciated.
+Kaitlin Allerton Hi Kaitlyn. The more polar compound does indeed have a greater affinity for the silica. But remember, the silica ISN'T MOVING. It is the stationary phase. Sticking to a stationary object better means you are less likely to move! In contrast, the less polar compounds spends less time on the stationary phase and more time in the liquid mobile phase being carried along, so it moves faster. ru-vid.com/video/%D0%B2%D0%B8%D0%B4%D0%B5%D0%BE-kMIWNUrAx0k.html This is a link to one of my very first RU-vid videos. It explains the situation you are asking about. Hope it helps! P.S. Alumina is also very polar, and would likely produce a similar result to that of silica.
+ChemSurvival thank you! yes I got it now. We are just now performing a lab like this, but dry?, for my organic chemistry course. In addittion we are writing the procedure ourselves. Needless to say I'm referencing your RU-vid video, thanks so much! Will be sharing this with my classmates.
Hi Sasha. No, you do not! As the name implies, a 'gradient' elution involves a slow, controlled (though not necessarily linear) change in the mobile phase concentration over time to increase its eluting power. The column should never be allowed to drain completely of mobile phase, since this would cause cracks and channels to form, ruining the column. Thanks for the question!
because the stationary phase must always be saturated by the mobile phase to ensure that it does not dry out. Silica swells somewhat to varying degrees in different organic solvents, so allowing it to dry out causes the silica particles to contract, potentially forming channels and voids in the column. Think of a bucket of wet sponges, all packed together very well. If the sponges begin to dry out and contract, voids and channels will naturally open up. inside of the pile of sponges. Even pouring more water back in won't necessarily cause the voids and channels to close up completely.
That depends on what you mean by "ordinary". The sands needs to be of uniform and sufficiently small grain size to prevent silica gel from penetrating the bed and flowing through it. Naturally, you also want to be sure you're using sand that has been chemically cleaned so that you don't risk contaminating your materials as they pass through the column. Many chemical suppliers sell so-called "chromatography sand" or "sand for chromatography," which is guaranteed to meet both of those requirements right out of the bottle.
Short answer - You don't. This is a tertiary resource (general knowledge). If you are citing a resource for technical writing it is because you want your reader to have access to an original account of peer-reviewed research (which this is not).
There are those who believe in placing sand on top of the silica slurry as well. I do not teach it to my introductory students for two reasons. First, my students have never had trouble maintaining a level surface at the top of the silica slurry without a sand layer on top. Second, and probably more importantly, adding the sand to the top of a column without disturbing the bed takes slight of hand that most introductory students cannot hope to achieve in an afternoon. It is easy to embed the sand within the gel at the top of the column, creating irregularities that actually cause more harm than good.
