Chromatography GCSE chemistry separates the different substances in a mixture, such as dyes in ink, by how strongly each one is attracted to the paper compared with the solvent. Each substance travels a different distance up the paper, producing a chromatogram from which you calculate an Rf value to identify each spot.
What equipment do you need for paper chromatography?
The paper chromatography required practical uses simple, inexpensive apparatus:
- Chromatography paper — a special absorbent paper that draws the solvent upward.
- Pencil — used to mark the baseline and label spots, because pencil marks will not dissolve in the solvent.
- Capillary tube — used to apply small, concentrated spots of each sample.
- Solvent — often water or ethanol, chosen to dissolve the substances being separated.
- Beaker with a lid or watch glass — holds the solvent and stops it evaporating during the run.
- Ruler — for measuring distances once the chromatogram is complete.
How do you carry out paper chromatography, step by step?
- Draw a pencil baseline a few centimetres from the bottom edge of the chromatography paper.
- Add small, concentrated spots of each sample onto the baseline using a capillary tube, letting each spot dry before adding more solution on top if needed.
- Pour a shallow layer of solvent into the beaker, keeping the level below the pencil line.
- Lower the paper into the beaker so the bottom edge touches the solvent, but the baseline itself stays above the solvent surface.
- Cover the beaker with a lid or watch glass to stop the solvent evaporating and keep the atmosphere saturated with solvent vapour.
- Remove the paper just before the solvent front reaches the top, and immediately mark the solvent front in pencil before it evaporates and the mark is lost.
- Allow the chromatogram to dry, then measure and record the distances travelled by the solvent front and by each spot.
Why must you use a pencil line and not a pen?
Ink is itself a mixture of coloured dyes dissolved in a solvent, so if the baseline were drawn in pen, the ink would dissolve into the solvent during the run and separate into its own set of spots, contaminating the results. Pencil marks are made of graphite, which is insoluble in the solvents used, so the baseline and solvent-front markings stay fixed and accurate throughout the experiment.
What is a chromatogram and how do you interpret it?
A chromatogram is the finished piece of paper showing where each substance ended up after the solvent has travelled through it. Interpreting a chromatogram follows a few simple rules:
- One spot means the original sample was a single pure substance.
- Several spots at different heights mean the sample was a mixture of different substances.
- Spots at the same height and colour in different samples suggest the same substance is present in both.
- The number of spots, not their colour alone, tells you how many different substances were in the original sample.
How do you calculate an Rf value?
The retention factor, or Rf value, compares how far a substance travelled with how far the solvent travelled:
$$R_f = \frac{\text{distance travelled by substance}}{\text{distance travelled by solvent}}$$
Worked example: A student runs a paper chromatography experiment. The solvent front travels 6.0 cm from the baseline. Spot X travels 3.6 cm in the same time. Calculate the Rf value of spot X.
$$R_f = \frac{3.6}{6.0} = 0.6$$
Since Rf values have no units and are always between 0 and 1, an Rf of 0.6 means the substance travelled 60% as far as the solvent. Under identical conditions (same paper, same solvent, same temperature), a given substance always produces the same Rf value, so this number can be compared against reference chromatograms to identify unknown substances.
| Rf value | Meaning |
|---|---|
| Close to 1 | Substance is very soluble in the solvent and moves almost as far as the solvent front |
| Close to 0 | Substance is strongly attracted to the paper and barely moves from the baseline |
| Between two samples | Matching Rf values (same paper, same solvent) suggest matching substances |
What is chromatography used for beyond identifying dyes?
Paper and thin-layer chromatography are used well beyond the school laboratory. Forensic scientists use chromatography to compare ink or drug samples from a crime scene against known references. Food scientists use it to check whether food colourings are approved additives or unauthorised dyes. Environmental scientists use related chromatographic techniques to detect pollutants in water samples, matching unknown spots against Rf values from substances of known identity.
Frequently asked questions
Why is chromatography paper chosen over other materials?
Chromatography paper is made from very pure cellulose fibres that absorb the solvent evenly and consistently, without introducing extra substances that could interfere with the separation. Its uniform structure also means the solvent rises at a steady, predictable rate, giving reproducible results between repeated experiments.
What happens if the pencil baseline is drawn below the solvent level?
If the baseline sits below the surface of the solvent in the beaker, the original sample spots dissolve directly into the bulk solvent instead of being carried up the paper. This destroys the experiment, because the substances mix into the solvent rather than separating out along the paper as intended, and no usable chromatogram is produced.
Can chromatography separate colourless substances?
Yes. Many substances relevant to chromatography, such as some amino acids, are colourless, so the spots are invisible once the paper has dried. Chemists reveal them using a locating agent, a chemical spray that reacts with the substance to produce a coloured spot, or by viewing the dried paper under ultraviolet light, which makes certain compounds fluoresce.
How do you know if a substance is pure using chromatography?
A pure substance produces exactly one spot on a chromatogram, regardless of the solvent used. If a sample believed to be pure produces two or more spots, or produces a different Rf value to a genuine reference sample under the same conditions, it must actually be a mixture or a different substance rather than the pure compound claimed.
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