The end of the paper, nearest the spot, is then dipped into the solvent without submerging the spot itself. In ascending chromatography, the solvent is in a pool at the bottom and moves up by capillarity. In descending chromatography it is in a trough at the top and flows down by capillarity and gravity. The solvent flows along the paper through the spots and on, carrying the substances from the spot.
Each of these will, if the solvent mixture has been well chosen, move at a different rate from the others. After a time the paper is taken out and dried: the substances can be seen at once if coloured, or located by treating with a suitable locating agent.
The distance a substance travels depends upon the resultant between propelling and retarding forces. Propellors a Solvent flow Usually the more soluble a substance is in the solvent, the more rapidly it will move along the paper.
Solvents are chosen for the greatest differential solubilities of the substances concerned. They could equally well, of course, both have been the same color - in which case you couldn't tell whether there was one or more dye present in that spot. It is very unlikely that the two confusing spots will have the same R f values in the second solvent as well as the first, and so the spots will move by a different amount.
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. How to Separate the Components of Ink. Food Coloring Experiments. How Does Ink Diffuse in Water? How to Separate Ink From Water. Easy Minute Science Projects. How Ink Is Made. Cool Science Experiments for Teens.
Plant Pigments Found in Spinach. How to Make Ozone Test Strips. Materials Two white coffee filters Scissors Ruler Drawing markers not permanent : brown, yellow and any other colors you would like to test At least two pencils one for each color you will be testing At least two tall water glasses one for each color you will be testing , four inches or taller Water Two binder clips or clothespins Drying rack or at least two additional tall water glasses one for each color you will be testing Pencil or pen and paper for taking notes Preparation Carefully cut the coffee filters into strips that are each about one inch wide and at least four inches long.
Cut at least two strips, one to test brown and one to test yellow. Cut an extra strip for each additional color you would like to test. How do you expect each of the different colors to behave when you test it with the paper strip?
Draw a pencil line across the width of each paper strip, about one centimeter from the bottom end. Take the brown marker and a paper strip and draw a short line about one centimeter on the middle section of the pencil line. Your marker line should not touch the sides of your strip.
Use a pencil to write the color of the marker you just used on the top end of the strip. Note: Do not use the colored marker or pen to write on the strips, as the color or ink will run during the test. Repeat the previous three steps with a yellow marker and then all the additional colors you would like to test. Hold a paper strip next to one of the tall glasses on the outside of it , aligning the top of the strip with the rim of the glass, then slowly add water to the glass until the level just reaches the bottom end of the paper strip.
Repeat with the other glass es , keeping the strips still on the outside and away from the water. What role do you think the water will play? Procedure Fasten the top of a strip the side farthest from the marker line to the pencil with a binder clip or clothespin.
Pause for a moment. Do you expect this color to be the result of a mixture of colors or the result of one color molecule? If you like, you can make a note of your prediction now.
Hang the strip in one of the glasses that is partially filled with water by letting the pencil rest on the glass rim. The bottom end of the strip should just touch the water level. If needed, add water to the glass until it is just touching the paper. Note: It is important that the water level stays below the marker line on the strip.
Leave the first strip in its glass as you repeat the previous two steps with the second strip and the second glass.
Repeat with any additional colors you are testing. Watch as the water rises up the strips. What happens to the colored lines on the strips? Does the color run up as well? Do you see any color separation? When the water level reaches about one centimeter from the top this may take up to 10 minutes , remove the pencils with the strips attached from the glasses.
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