Next you add fresh solvent to the top of the column, trying to disturb the packing material as little as possible. Then you open the tap so that the solvent can flow down through the column, collecting it in a beaker or flask at the bottom. As the solvent runs through, you keep adding fresh solvent to the top so that the column never dries out.
The next set of diagrams shows what might happen over time. This assumes that you have read the explanation for what happens during thin layer chromatography.
If you haven't, follow the very first link at the top of the page and come back to this point afterwards. The blue compound is obviously more polar than the yellow one - it perhaps even has the ability to hydrogen bond.
You can tell this because the blue compound doesn't travel through the column very quickly. That means that it must adsorb more strongly to the silica gel or alumina than the yellow one.
The less polar yellow one spends more of its time in the solvent and therefore washes through the column much faster. The process of washing a compound through a column using a solvent is known as elution. The solvent is sometimes known as the eluent. It is going to take ages to wash the blue compound through at the rate it is travelling at the moment! This distance travelled compared with the distance travelled by the solvent is the same for a particular substance.
The R f value of a spot is calculated using:. The R f value is always the same for a particular substance using the same stationary phase and mobile phase. R f values can be used to identify unknown chemicals if they can be compared to a range of reference substances. R f values vary from 0 the substance is not attracted to the mobile phase to 1 the substance is not attracted to the stationary phase.
Paper chromatography Paper chromatography is another method that can be used to test if a substance is pure or impure. As the solvent soaks up the paper, it carries the mixtures with it. Different components of the mixture will move at different rates. This separates the mixture out. Spots of ink or plant dye are placed on a pencil line. The next diagram shows what might happen to the various spots on the original chromatogram.
The position of the second solvent front is also marked. You wouldn't, of course, see these spots in both their original and final positions - they have moved!
The final chromatogram would look like this:. Two way chromatography has completely separated out the mixture into four distinct spots. If you want to identify the spots in the mixture, you obviously can't do it with comparison substances on the same chromatogram as we looked at earlier with the pens or amino acids examples. You would end up with a meaningless mess of spots. You can, though, work out the R f values for each of the spots in both solvents, and then compare these with values that you have measured for known compounds under exactly the same conditions.
Although paper chromatography is simple to do, it is quite difficult to explain compared with thin layer chromatography. The explanation depends to some extent on what sort of solvent you are using, and many sources gloss over the problem completely.
If you haven't already done so, it would be helpful if you could read the explanation for how thin layer chromatography works link below. That will save me a lot of repetition, and I can concentrate on the problems. The key point about cellulose is that the polymer chains have -OH groups sticking out all around them.
To that extent, it presents the same sort of surface as silica gel or alumina in thin layer chromatography. It would be tempting to try to explain paper chromatography in terms of the way that different compounds are adsorbed to different extents on to the paper surface. In other words, it would be nice to be able to use the same explanation for both thin layer and paper chromatography. Unfortunately, it is more complicated than that!
The complication arises because the cellulose fibres attract water vapour from the atmosphere as well as any water that was present when the paper was made. You can therefore think of paper as being cellulose fibres with a very thin layer of water molecules bound to the surface. It is the interaction with this water which is the most important effect during paper chromatography. Non-polar molecules in the mixture that you are trying to separate will have little attraction for the water molecules attached to the cellulose, and so will spend most of their time dissolved in the moving solvent.
Molecules like this will therefore travel a long way up the paper carried by the solvent. They will have relatively high R f values. On the other hand, polar molecules will have a high attraction for the water molecules and much less for the non-polar solvent.
They will therefore tend to dissolve in the thin layer of water around the cellulose fibres much more than in the moving solvent. Because they spend more time dissolved in the stationary phase and less time in the mobile phase, they aren't going to travel very fast up the paper.
The tendency for a compound to divide its time between two immiscible solvents solvents such as hexane and water which won't mix is known as partition.
Paper chromatography using a non-polar solvent is therefore a type of partition chromatography.
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