Chromatography and Rf Exam Questions

• Draw a pencil line (baseline/origin) near the bottom of a piece of chromatography paper [1]
• Place a small spot of the food colouring on the baseline using a capillary tube [1]
• Place the paper in a beaker containing a shallow layer of solvent (water), ensuring the solvent level is below the baseline/spot [1]
• Cover the beaker with a lid (to prevent evaporation of the solvent) and allow the solvent to rise up the paper until near the top, then remove and dry the paper [1]

Examiner note: Three key details candidates forget: (1) use pencil for the baseline (pen ink would dissolve), (2) the solvent must be below the baseline (or the spots dissolve into the solvent pool), and (3) cover the beaker to prevent solvent evaporation. Missing any of these typically loses 1 mark.

(a) Rf = 9.6 / 12.0 = 0.80 [1]

(b) Rf = 4.8 / 12.0 = 0.40 [1]

(c) The Rf value is characteristic of a particular substance in a particular solvent and can be used to identify unknown substances by comparing with known values [1]

(d) Spot A, because its Rf value matches the known dye (0.80) [1]

Examiner note: Rf values are always between 0 and 1. An Rf of exactly 1.0 would mean the substance moved with the solvent front (extremely soluble in the mobile phase). An Rf of 0 means the substance did not move at all (completely attracted to the stationary phase). Always measure from the centre of the spot to the baseline.

• Different substances have different relative solubilities in the solvent (mobile phase) [1]
• Substances that are more soluble in the solvent are carried further up the paper [1]
• Substances that are more strongly attracted to the paper (stationary phase) move less far [1]

Examiner note: Chromatography works because of the partitioning of substances between two phases: the stationary phase (paper) and the mobile phase (solvent). Each substance reaches an equilibrium between the two phases. The more time a substance spends dissolved in the mobile phase, the further it travels.

(a) The food colouring is a mixture of at least two dyes, which are dye X and dye Z [1]

(b) No spot from the food colouring has the same Rf value as dye Y [1]

(c) The Rf value depends on the solvent used and the conditions (temperature, type of paper). Running the known dyes on the same paper ensures the conditions are identical, making the comparison valid [1]

Examiner note: A pure substance produces one spot; a mixture produces two or more spots. Matching Rf values identifies substances only when they are run under identical conditions. If a reference dye is run on a different piece of paper or with a different solvent, the Rf values may not match even for the same substance.

• Stationary phase: the chromatography paper (cellulose fibres with water trapped between them). Substances that are more attracted to the paper stay near the baseline [1]
• Mobile phase: the solvent (e.g., water, ethanol). It carries the dissolved substances up the paper by capillary action [1]
• Separation occurs because different substances partition differently between the two phases — each substance has a different ratio of solubility in the mobile phase versus attraction to the stationary phase [1]

Examiner note: The terms “stationary phase” and “mobile phase” are required at supplement level. In paper chromatography, the stationary phase is effectively water trapped in the cellulose fibres of the paper, and substances partition between this trapped water and the moving solvent. Some questions use thin-layer chromatography (TLC), where the stationary phase is a thin layer of silica or alumina on a plate.

(a) Any two from:

• Use a different/more suitable solvent [1]
• Use a longer piece of chromatography paper so the spots travel further apart [1]
• Apply a smaller/more concentrated spot to give sharper spots [1]
• Use thin-layer chromatography for better resolution [1]

(b) Use a locating agent/UV light — some substances fluoresce under UV light, or the paper can be sprayed with a reagent (e.g., ninhydrin for amino acids) that reacts with the substances to produce coloured spots [1]

(c) Any one from: only works for small quantities / does not separate very similar substances well / Rf values can be affected by temperature changes / cannot easily be used to collect separated substances for further analysis [1]

Examiner note: Choosing the right solvent is the single most important factor in achieving good separation. If the substances are too similar in solubility, they will not separate well regardless of technique. For colourless substances (like amino acids), a locating agent is essential.

(a) The ink from the felt-tip pen dissolves in the solvent and produces additional spots on the chromatogram, making it impossible to distinguish the baseline ink from the sample spots [1]

(b) The sample spots dissolve into the solvent pool at the bottom of the beaker instead of being carried up the paper, so no separation occurs / the spots are washed off the paper [1]

(c) Any one from: cover the beaker with a lid to prevent solvent evaporation / apply small spots to avoid streaking / allow the spot to dry before placing the paper in the solvent / do not move the beaker while the experiment is running [1]

Examiner note: These are the two most common practical errors in chromatography. Pencil is insoluble in all common chromatography solvents, so it does not interfere. The solvent level below the baseline is critical — this is a safety net question that examiners expect all candidates to get right.