What you'll learn
This required practical focuses on using paper chromatography to separate and analyse mixtures of dyes or inks. You'll learn the complete experimental method, how to calculate Rf values, and how to use chromatography to identify substances by comparing them with reference samples. This practical appears frequently in AQA GCSE Chemistry exams, both in questions about method and in data analysis tasks.
Key terms and definitions
Chromatography — a separation technique that separates mixtures into individual components based on their different solubilities in a solvent and attractions to a stationary phase.
Mobile phase — the liquid solvent (often water or ethanol) that moves up the chromatography paper, carrying dissolved substances with it.
Stationary phase — the chromatography paper itself, which remains fixed in position while the mobile phase moves through it.
Solvent front — the highest point reached by the mobile phase (solvent) on the chromatography paper.
Baseline — the starting line drawn in pencil near the bottom of the chromatography paper where samples are spotted.
Rf value — the retention factor, calculated as distance travelled by substance ÷ distance travelled by solvent front; used to identify substances.
Chromatogram — the pattern of separated spots produced on chromatography paper after the experiment.
Pure substance — a substance containing only one compound or element; produces a single spot on a chromatogram.
Core concepts
The principle of chromatography
Chromatography works because different substances have different solubilities in the mobile phase and different attractions to the stationary phase. When a mixture is placed on chromatography paper and a solvent moves through it:
- Substances more soluble in the solvent travel further up the paper
- Substances more attracted to the paper travel less far
- This causes the components of a mixture to separate into distinct spots
The balance between solubility in the mobile phase and attraction to the stationary phase determines how far each substance travels. This creates a characteristic pattern that can be used to identify substances.
Equipment and setup
The required practical uses simple, widely available equipment:
- Chromatography paper (or filter paper)
- Beaker with lid or watch glass cover
- Pencil (never pen — ink would dissolve and contaminate results)
- Ruler
- Capillary tube or fine glass rod for applying samples
- Solvent (typically water, ethanol, or a mixture)
- Sample substances (food colourings, inks, plant pigments)
The beaker must be covered to create a saturated atmosphere, preventing the solvent from evaporating too quickly and ensuring even solvent movement up the paper.
Method and procedure
Step 1: Prepare the paper Draw a horizontal baseline in pencil approximately 2 cm from the bottom of the chromatography paper. Pencil is essential because pen ink would dissolve in the solvent and contaminate your results.
Step 2: Apply samples Use a capillary tube to apply small spots of each sample along the baseline. Space spots evenly, at least 1 cm apart. Label each spot position lightly in pencil at the top of the paper. Allow spots to dry completely.
Step 3: Add solvent Pour solvent into the beaker to a depth of approximately 1 cm — crucially, the solvent level must be below the baseline. If the solvent touches the sample spots, they will dissolve directly into the solvent rather than separating.
Step 4: Develop the chromatogram Place the paper vertically in the beaker, ensuring it doesn't touch the sides. Cover immediately with a lid or watch glass. The solvent will rise up the paper by capillary action, carrying the dissolved substances with it.
Step 5: Remove and mark When the solvent front has risen about three-quarters of the way up the paper (before it reaches the top), remove the paper immediately. Quickly mark the position of the solvent front with a pencil line before it evaporates. Allow the chromatogram to dry.
Step 6: Analysis Circle any visible spots. Some substances may only become visible under UV light or after treatment with locating agents (though this is beyond GCSE requirements for this practical).
Calculating Rf values
The Rf value (retention factor) is a ratio that identifies a substance under specific conditions. It is calculated using:
Rf = distance travelled by substance ÷ distance travelled by solvent front
Key measurement points:
- Distance travelled by substance: measured from the baseline to the centre of the spot
- Distance travelled by solvent front: measured from the baseline to the solvent front line
- Both measurements must use the same units (typically cm or mm)
Important properties of Rf values:
- Rf values are always between 0 and 1 (or 0 and 100 if expressed as percentages)
- A pure substance produces the same Rf value under identical conditions (same solvent, same temperature, same paper)
- Different substances have different Rf values
- Rf values have no units (they're ratios)
- An Rf value close to 0 means the substance barely moved; close to 1 means it travelled almost as far as the solvent
Identifying substances using chromatography
Chromatography can identify unknown substances by comparing their Rf values with reference samples:
Comparing with known substances: Run the unknown sample alongside known reference substances on the same chromatography paper under identical conditions. If a spot from the unknown sample travels the same distance as a known substance (same Rf value) and appears the same colour, the unknown likely contains that substance.
Identifying pure vs impure substances:
- A pure substance produces only one spot on the chromatogram
- A mixture produces multiple spots — one for each component
- If you test a food colouring and it produces three spots, it contains at least three different dyes
Limitations: Different substances can occasionally have similar Rf values under the same conditions, so matching Rf values provides evidence but not absolute proof. Additional tests may be needed for definitive identification.
Factors affecting chromatography results
Several variables influence how substances separate:
Solvent choice: Different solvents produce different Rf values for the same substance. Water separates water-soluble substances well; ethanol works better for less polar substances. The solvent must dissolve the sample substances at least partially.
Temperature: Higher temperatures generally increase solubility and can change how far substances travel. Keeping temperature constant ensures reproducible results.
Type of paper: Different papers have different fibre structures and thicknesses, affecting how substances move. Using the same type of paper ensures consistent results.
Sample concentration: Too much sample creates large, smeared spots that are difficult to measure accurately. Too little produces faint spots that may be invisible. The optimum is a small, concentrated spot.
Worked examples
Example 1: Calculating Rf values
Question: A student investigated four food colourings (A, B, C, D) using paper chromatography with water as the solvent. The solvent front travelled 8.0 cm from the baseline. Food colouring A produced one spot that travelled 2.4 cm. Food colouring B produced two spots that travelled 2.4 cm and 6.4 cm. Calculate the Rf values for all spots. Which food colouring is pure? (4 marks)
Answer:
Food colouring A: Rf = 2.4 ÷ 8.0 = 0.30 (1 mark for calculation)
Food colouring B, spot 1: Rf = 2.4 ÷ 8.0 = 0.30 (1 mark)
Food colouring B, spot 2: Rf = 6.4 ÷ 8.0 = 0.80 (1 mark)
Food colouring A is pure because it produced only one spot (1 mark for identification and correct reasoning)
Mark scheme notes: The calculation marks require correct substitution and answer to 2 significant figures. The final mark requires both identifying food colouring A and explaining that a single spot indicates purity.
Example 2: Identifying an unknown dye
Question: A student used chromatography to identify an unknown yellow dye X. Three known dyes (tartrazine, sunset yellow, and quinoline yellow) were run alongside dye X on the same chromatography paper using ethanol as solvent. The results showed:
- Tartrazine: Rf = 0.42
- Sunset yellow: Rf = 0.68
- Quinoline yellow: Rf = 0.42
- Dye X: Rf = 0.68
The student concluded that dye X is sunset yellow. Explain why this conclusion is justified and suggest one further test that could confirm the identity. (3 marks)
Answer:
Dye X has the same Rf value as sunset yellow (0.68) and they were tested under identical conditions (same solvent, same paper, same time) (1 mark for matching Rf and identical conditions)
This suggests dye X and sunset yellow have the same solubility and attraction to the paper (1 mark for explanation)
Further test: Run the chromatography again using a different solvent (such as water or propanone) to see if they still have matching Rf values / Use another analytical technique such as mass spectrometry (1 mark for appropriate suggestion)
Example 3: Experimental design and evaluation
Question: A student plans to investigate whether a brown food colouring is a pure substance or a mixture. Describe a method the student could use and explain how the results would show whether the food colouring is pure or a mixture. (6 marks)
Answer:
Method:
- Draw a baseline in pencil approximately 2 cm from the bottom of chromatography paper (1 mark)
- Use a capillary tube to place a small spot of the brown food colouring on the baseline (1 mark)
- Place the paper in a beaker containing a shallow depth of solvent (e.g., water), ensuring the solvent level is below the baseline (1 mark)
- Cover the beaker and allow the solvent to rise up the paper (1 mark)
- Remove the paper before the solvent reaches the top and mark the solvent front position (1 mark)
Interpretation: If the food colouring is pure, it will produce one spot. If it is a mixture, it will separate into multiple spots of different colours (1 mark for correct interpretation)
Common mistakes and how to avoid them
Using pen instead of pencil for the baseline: Pen ink dissolves in the solvent and contaminates your results. Always use pencil for any markings on chromatography paper, as graphite is insoluble.
Starting with the solvent level above the baseline: This causes samples to dissolve directly into the solvent without separating. Always ensure the solvent level is at least 0.5 cm below your baseline before inserting the paper.
Measuring distance travelled from the solvent front or paper bottom: The distance travelled must always be measured from the baseline to the centre of the spot, not from the bottom edge of the paper or any other reference point.
Forgetting to mark the solvent front immediately: Once removed from the beaker, the solvent evaporates quickly, making it impossible to measure accurately later. Mark it with pencil the moment you remove the paper.
Applying too much sample: Large spots spread out as they travel, creating smeared results that are difficult to measure. Apply small, concentrated spots and allow them to dry before repeating if necessary.
Not allowing spots to dry before placing in solvent: Wet spots spread when they contact solvent vapour, reducing separation quality. Always ensure spots are completely dry.
Incorrect Rf calculations: Remember Rf values are always less than 1 (unless you've made a measurement error). If you calculate an Rf greater than 1, check you haven't reversed the calculation — it's substance distance ÷ solvent distance, not the other way round.
Exam technique for "Required practical: chromatography"
Method questions: When asked to "describe a method," use numbered steps in logical order. Include specific measurements (e.g., "baseline 2 cm from bottom," "solvent depth 1 cm") and named equipment. State that pencil must be used for the baseline and explain why (marks often awarded for explanations).
Calculation questions: Show all working for Rf calculations. Write the formula, substitute values, and give your answer to 2 significant figures. Include a ruler diagram if you're calculating from a chromatogram image. Remember Rf values have no units.
Evaluation and analysis: When identifying substances, state that identical conditions are essential for valid comparison. Explain that matching Rf values suggest the same substance but aren't absolute proof. Mention limitations such as resolution of spots or measurement precision.
Variables and controls: Questions may ask about independent, dependent, and control variables. Independent variable: the substance being tested; dependent variable: the distance travelled (Rf value); control variables: solvent type, paper type, temperature, solvent volume, time allowed.
Quick revision summary
Paper chromatography separates mixtures using a mobile phase (solvent) moving through a stationary phase (paper). Substances separate based on their solubility and attraction to the paper. The baseline must be drawn in pencil above the solvent level. Pure substances produce one spot; mixtures produce multiple spots. Rf values (substance distance ÷ solvent distance) identify substances under specific conditions and are always between 0 and 1. Mark the solvent front immediately upon removal. Identical conditions are essential when comparing unknown substances with reference samples.