C10.2 Analysing Chromatograms

A free GCSE Combined Science: Trilogy lesson for AQA (8464) from OBA School: how paper chromatography separates substances, calculating Rf values and using chromatograms to identify substances. Watch the video, read the explanation and worked examples, then try the revision questions. You can work through the full lesson, with every video, practice quizzes and printable worksheets with mark schemes, in our free online course.

What you will learn

• Explain how paper chromatography separates substances using a stationary phase and a mobile phase
• Calculate Rf values from a chromatogram, and rearrange the equation
• Use Rf values and reference substances to identify the components of a mixture
• Explain how a chromatogram shows whether a substance is pure

Key words

• Chromatography: A method for separating the substances in a mixture so that they can be identified.
• Stationary phase: The phase that does not move. In paper chromatography it is the paper.
• Mobile phase: The phase that moves through the stationary phase. In paper chromatography it is the solvent.
• Chromatogram: The pattern of spots produced by chromatography.
• Solvent front: The furthest point the solvent reaches up the paper.
• Rf value: Distance moved by a substance ÷ distance moved by the solvent. It has no units.

How paper chromatography works

Small spots of the samples are put on a pencil baseline. The bottom of the paper is dipped into a solvent, which soaks up the paper and carries the substances with it.

Each substance is shared between the two phases. A substance that is more soluble in the solvent (mobile phase) and less attracted to the paper (stationary phase) travels further up the paper. A substance more attracted to the paper moves only a short way. Different substances move different distances, so the mixture separates.

Calculating Rf values

Rf = distance moved by substance ÷ distance moved by solvent

Measure both distances from the baseline: to the centre of the spot, and to the solvent front. Rf is always less than 1 and has no units.

Worked example: Rf of the green dye

green spot moved 6.3 cm; solvent moved 9.0 cm
Rf = 6.3 ÷ 9.0 = 0.70
The yellow spot moved 2.7 cm, so its Rf = 2.7 ÷ 9.0 = 0.30.

Answer: Rf (green) = 0.70

Worked example: rearranging

A dye has Rf = 0.35. The solvent moves 8.0 cm. How far does the dye move?
distance moved by dye = Rf × distance moved by solvent = 0.35 × 8.0 = 2.8 cm

Answer: 2.8 cm

Identifying substances

• A compound always has the same Rf value in the same solvent (with the same paper and conditions).
• Compare the spots of an unknown with reference substances run on the same paper: spots at the same height (same Rf) are likely to be the same substance.
• The Rf value of a compound changes if you change the solvent, so only compare values measured in the same solvent.
• If two substances have the same Rf in one solvent, run them again in a different solvent to tell them apart.
• A substance that does not dissolve in the solvent stays on the baseline.

• dye A; Rf in water 0.48; Rf in ethanol 0.66
• dye B; Rf in water 0.48; Rf in ethanol 0.27
• unknown; Rf in water 0.48; Rf in ethanol 0.27

In water, A and B have the same Rf, so we cannot tell which the unknown is. In ethanol, the unknown matches dye B.

Pure or mixture?

A pure compound gives one spot in every solvent. A mixture usually gives two or more spots, one for each substance that dissolves. If a sample gives a single spot in one solvent, it might still be a mixture whose components happen to have the same Rf, so test it in another solvent too.

Uses of chromatography: checking food colourings and flavourings for banned substances, comparing inks in forensic investigations, testing athletes’ urine for drugs, and checking the purity of medicines made in a laboratory.

Exam tip: The baseline is drawn in pencil because graphite is insoluble and will not run; the solvent must be below the baseline or the spots dissolve into the solvent in the beaker; a lid stops the solvent evaporating.

Common mistakes

• Measuring from the bottom of the paper instead of from the baseline.
• Measuring to the top edge of a spot instead of its centre.
• Giving an Rf value bigger than 1, or with units. Check you divided the smaller number by the larger one.
• Saying the paper is the mobile phase. The paper stays still; the solvent moves.
• Saying one spot in one solvent proves a substance is pure. It needs one spot in every solvent tried.

Revision questions

Try these, then check your answers below each one.

Question 1. A spot moves 3.0 cm and the solvent moves 7.5 cm. Calculate the Rf value.

Answer: 0.4

Working: 3.0 ÷ 7.5 = 0.40. Rf has no units.

Question 2. The chromatogram shows a mixture M and four pure dyes A, B, C and D. Which dyes are in M? (Choose from: A and C, A and B, B and D, C and D.)

Answer: A and C

Working: M has a spot at the same height as A and another at the same height as C, so M contains A and C.

Question 3. True or false? An Rf value can be greater than 1.

Answer: False

Working: False. A substance cannot move further than the solvent front, so Rf is always less than 1.

Related lessons

• Previous lesson: C10.1 Pure Substances and Mixtures
• Next lesson: C10.3 Testing for Gases
• In this chapter: C10 RP Required Practical: Chromatography

Continue in the full course

This lesson is section C10.2 (AGCSC2702) of our free GCSE Combined Science course for AQA. In the full course you get every video for this section, a self-marking practice quiz, a printable worksheet with a mark scheme, chapter tests, and your progress saved. Go to the full GCSE Combined Science course.