CHROMATOGRAPHY separates mixtures of soluble substances based on how they distribute between a STATIONARY PHASE and a MOBILE PHASE.
STATIONARY PHASE: doesn't move — e.g. chromatography paper.
MOBILE PHASE: moves — e.g. solvent (water, ethanol etc.).
PRINCIPLE:
Each substance in the mixture has different SOLUBILITY in the solvent and different AFFINITY for the paper.
Substances that are more soluble in the solvent and less attracted to paper TRAVEL FURTHER.
Substances that are less soluble or more attracted to paper TRAVEL LESS FAR.
This separates the mixture into distinct SPOTS.
PAPER CHROMATOGRAPHY:
1. Draw a pencil baseline near the bottom of chromatography paper (pencil — ink would run).
2. Spot the mixture onto the baseline.
3. Place in a small depth of solvent — below the baseline (so the spot isn't dissolved directly).
4. Solvent rises up the paper by capillary action, carrying dissolved substances.
5. Remove when solvent front is near the top.
6. Mark the solvent front immediately (it evaporates).
7. Measure distances moved by each spot and the solvent front.
Rf Values
The RF VALUE (retention factor) is used to identify substances — each substance has a characteristic Rf in a given solvent.
FORMULA:
Rf = distance moved by substance ÷ distance moved by solvent front
Rf values are always between 0 and 1.
Rf = 0: substance doesn't move at all (stays at baseline).
Rf = 1: substance moves exactly as far as the solvent front.
EXAMPLE:
Substance moves 4.5 cm. Solvent front moves 9.0 cm.
Rf = 4.5 ÷ 9.0 = 0.50
IDENTIFICATION:
Compare Rf values to known reference data or reference spots run alongside the unknown.
If Rf matches a known substance in the same solvent → likely the same substance.
CHROMATOGRAM:
The paper with spots after development.
Each spot represents a different component of the mixture.
A pure substance produces ONE spot.
A mixture produces MULTIPLE spots (one per component).
Applications of Chromatography
FOOD TESTING:
Identify artificial colourings in food — compare to known reference spots.
Check for undeclared additives.
FORENSIC SCIENCE:
Analyse ink from documents to detect forgeries — different pens use different inks.
Drug testing — identify substances in blood or urine samples.
SPORTS DOPING:
Identify banned substances in athlete samples.
Chromatography separates the many compounds in blood/urine.
PHARMACEUTICAL QUALITY CONTROL:
Verify purity of medicines — check for impurities.
Confirm active ingredient is present in correct amount.
LIMITATIONS:
Some substances have similar Rf values — different substances can overlap.
Better to use TWO different solvents — if Rf values match in both, identification is more reliable.
More advanced techniques (HPLC, GC-MS) give greater precision.
⚠️ Common Mistake
Rf values are ALWAYS between 0 and 1. If you get a value greater than 1, you've divided the WRONG WAY — always divide substance distance BY solvent distance. Also: the baseline and spots must be drawn in PENCIL not pen — pen ink contains dyes that would run with the solvent and confuse results.
📐 Variables
RfRetention factor (Rf) is measured in ()
📐 Key Equations
Rf = distance moved by substance ÷ distance moved by solvent front
📌 Key Note
Chromatography: separates mixtures by how far substances travel in solvent on paper. Rf = substance distance ÷ solvent front distance (always 0–1). Same Rf in same solvent = same substance. Pure substance = 1 spot. Mixture = multiple spots. Use pencil for baseline.
🎯 Matching Activity — Chromatography Concepts
Match each term to its correct description. — drag the symbols on the right to match the component names on the left.
Rf value
Drop here
Pure substance
Drop here
Mixture
Drop here
Mobile phase
Drop here
Stationary phase
Drop here
Produces only one spot on the chromatogram
The paper — does not move
Produces multiple spots — one for each component
The solvent that moves up the chromatography paper
Distance moved by substance ÷ distance moved by solvent — always between 0 and 1
⚽ FIFA Worked Examples
Rf Calculation
In a chromatography experiment, a substance moves 7.2 cm. The solvent front moves 9.6 cm. Calculate the Rf value.
F
Rf = distance moved by substance ÷ distance moved by solvent front
I
Rf = 7.2 ÷ 9.6
F
No unit conversion needed — Rf has no units
A
Rf = 0.75
⭐ Higher Tier Only
Explain Rf in terms of relative solubility: substances more soluble in mobile phase travel further. Two-way chromatography: run in one direction, rotate, run again with different solvent → better identification. High-performance liquid chromatography (HPLC) and gas chromatography (GC) as more advanced methods.
🧪 Required Practical
🔬 RP1 (Chemistry) — Investigate paper chromatography to separate and identify colours in inks or food dyes. Measure Rf values and compare to known reference substances.
Know the method, variables, equipment and how to analyse results.
🎯 Test Yourself
Question 1 of 2
1. A student runs chromatography on a food colouring and gets three spots. What does this tell them?
2. In chromatography, why must the initial spot be placed ABOVE the solvent level?
⭐ How Well Do You Understand This Topic?
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