How paper chromatography separates a mixture
A rising solvent (the mobile phase) carries soluble substances up the chromatography paper (the stationary phase). Spot the mixture onto a pencil baseline. Stand the paper in solvent with the baseline above the solvent surface, in a sealed container. Components more soluble and less strongly held by the paper travel further, so the mixture separates. Remove the paper and mark the solvent front in pencil. A pure substance gives one spot; a mixture gives two or more.
Locating agents reveal colourless spots
Amino acids and other colourless substances leave invisible spots, so a locating agent makes them visible. Ninhydrin is sprayed on and the paper warmed gently, turning amino-acid spots purple. Under a UV lamp, paper treated with a fluorescent dye shows the spots as dark patches on a bright background. Iodine vapour turns many organic spots yellow-brown, though these fade on removal. In an answer, name the agent, say how it is applied, and state the colour it produces.
(Extended) Retention factor Rf
(Extended only.) Rf = distance moved by the spot ÷ distance moved by the solvent front, both measured from the baseline, taking the spot to its centre. Rf always lies between 0 and 1: near 0 the substance is held strongly by the paper; near 1 it moves almost with the solvent front. Rf is fixed under set conditions (same solvent, paper, temperature), so an unknown with the same Rf as a reference is likely the same substance. Example: 3.6 ÷ 9.0 = 0.40 (no units).
Drawn from real examiner reports.
Drawing the baseline in pen, not pencil
Draw the baseline in pencil. Pen ink contains soluble dyes that dissolve in the solvent and travel up the paper, adding false spots and ruining the chromatogram. Pencil is graphite, which is insoluble in the solvent and stays put on the baseline. "Draw a pencil line near the bottom of the paper" is the mark-scheme wording; just "draw a line" is not enough.
Starting the baseline below the solvent
Position the paper so the baseline sits above the solvent surface. If the spots are submerged, they dissolve straight into the solvent reservoir instead of being carried up the paper by capillary action, and no separation occurs. The solvent must start below the baseline and rise through it. "Baseline above the solvent surface" is the required statement.
(Extended) Inverting the Rf fraction
Rf = spot ÷ solvent front, not solvent front ÷ spot. Inverting the fraction gives a value greater than 1, which is impossible because a spot can never travel past the solvent front. If your Rf comes out above 1 you have divided the wrong way round. Keep the spot distance on top and the solvent-front distance underneath, both measured from the baseline.
Naming a locating agent without the result
Writing just "ninhydrin" or "UV" scores little. The marks need the agent named, how it is applied, and the observable result — for example, "spray ninhydrin and warm gently; amino-acid spots turn purple". Stating the visible change is what earns credit, the same "observations not product names" rule that runs through the whole paper.
Section 2 — observable changes replaced by product names: S23 P31 Q4(d)(ii); W22 P31 Q6(a).
Calling UV light a chemical that stains
UV light does not react with or stain the sample. The paper carries a fluorescent dye and glows under the lamp; spots that absorb UV quench the glow and show up as dark patches on the bright background. Describing UV as "a chemical that stains the spots" is wrong — it is a physical method of locating them, unlike ninhydrin or iodine.
Saying a pure substance gives no spots
A pure substance gives exactly one spot on a chromatogram, not "no spots"; a mixture gives two or more. When judging purity, count the spots and quote the rule, rather than commenting on how far a spot travelled. Two spots at the same position as a reference suggest the same substance, but purity is decided by the number of spots.
(Extended) Rf calculation scaffold
Measure both distances from the pencil baseline: the solvent front to its leading edge, the spot to its centre. Then Rf = spot ÷ solvent front. Check it lies between 0 and 1; a value above 1 means the fraction is inverted. Rf is a ratio, so it has no units.
Identify a spot by comparison
To identify a spot, compare its position (or Rf) with a reference on the same paper under the same conditions. The same position suggests the same substance; a different one proves they differ. Confirm a match by co-chromatography: spotting both together gives a single spot.
Describe the chromatogram precisely
Describe what is seen, not just what happens. Write "the solvent front moved 9.0 cm and spot A moved 3.6 cm, so Rf = 0.40", and "two spots show the sample is a mixture". Avoid bare statements like "the substance moved 3.6 cm" without saying which substance is meant.
Paper chromatography is a technique used to separate a mixture of soluble substances. The mixture is dissolved and spotted onto a piece of chromatography paper (the stationary phase). A solvent (the mobile phase) rises up the paper by capillary action, carrying components of the mixture with it at different rates. Substances that dissolve more readily in the solvent travel further; substances adsorbed more strongly to the paper travel less far.
Stationary phase: the chromatography paper.
Mobile phase: the solvent, which moves up the paper.
Locating agent: a chemical or physical method used to reveal colourless spots on a chromatogram, making them visible. Examples: ninhydrin solution (reveals amino acids as purple spots), iodine vapour (reveals organic spots as yellow-brown), UV light with a fluorescent indicator paper (spots appear dark on a bright background).
(Extended) Retention factor (): a dimensionless ratio that describes how far a substance travels relative to the solvent front, under fixed conditions. It is always between 0 and 1 (inclusive).
(Extended) A spot travels 4.5 cm from the baseline. The solvent front travels 6.0 cm from the baseline. Calculate the Rf value.
(Extended) A student runs paper chromatography on an unknown dye mixture. After the experiment, the following measurements are recorded:
Reference data:
| Dye | Reference value |
|---|---|
| Blue dye | 0.40 |
| Red dye | 0.80 |
| Green dye | 0.60 |
(a) Calculate the values for spots A and B. Show your working.
(b) Identify the dye corresponding to each spot using the reference data.
(c) State whether the original unknown dye is a pure substance or a mixture. Justify your answer.
