Preparing a pure organic solid and a pure organic liquid
Full theory: Carboxylic acids & derivatives: esters and anhydrides · Alcohols: distillation · Organic synthesis
Aim: make an organic product, purify it, and show that it is pure. AQA name two preparations: a solid (aspirin, purified by recrystallisation and checked by melting point) and a liquid (an ester, purified by washing, drying and redistillation). The chemistry is only half the marks; the other half is the practical technique and the yield calculation.
Part A, a pure organic solid: aspirin
Aspirin is made by acylating salicylic acid (2-hydroxybenzenecarboxylic acid) with ethanoic anhydride. The –OH group on the ring becomes an ester:
HOOCC6H4OH + (CH3CO)2O → HOOCC6H4OCOCH3 + CH3COOH
Ethanoic anhydride is used rather than ethanoyl chloride because it is cheaper, less corrosive, less vigorously hydrolysed by water, and produces no HCl, a comparison AQA ask for directly.
- Weigh about 6.00 g of salicylic acid straight into a 100 cm3 conical flask, and record the exact mass.
- Add 10 cm3 of ethanoic anhydride from a measuring cylinder and swirl.
- Add 5 drops of concentrated sulfuric acid as the catalyst and swirl for a few minutes.
- Warm for about 20 minutes in a water bath at roughly 60 °C. Do not let the flask go above 65 °C.
- Cool the flask, then pour the contents into 75 cm3 of water, stirring well to precipitate the solid.
- Filter off the crude aspirin under reduced pressure on a double thickness of filter paper and leave it to dry.
- Recrystallise: dissolve the crude solid in the minimum volume of hot ethanol in a boiling tube standing in water at about 75 °C, never above ethanol’s boiling point of 78 °C.
- Pour the hot solution into about 40 cm3 of water and let it cool slowly; white needles separate. Scratching the flask with a glass rod, or cooling in ice, starts crystallisation if nothing appears.
- Filter under reduced pressure again, wash with a little cold ethanol, dry, and record the mass.
- Pack three melting-point tubes to about 0.5 cm and measure the melting point, heating slowly.
Crude aspirin carries no insoluble impurity, so this method goes straight from the hot solution to cooling. In the general recrystallisation technique (the one AQA can ask you to describe for any solid), you filter the hot solution through a warmed funnel first, which is what removes insoluble impurities; the soluble ones stay behind in the filtrate when the crystals form. The step-by-step version is on the carboxylic acids page.
Part B, a pure organic liquid: ethyl ethanoate
An alcohol and a carboxylic acid form an ester in an equilibrium catalysed by concentrated sulfuric acid:
CH3CH2OH + CH3COOH ⇌ CH3COOC2H5 + H2O
- Put a few anti-bumping granules in a pear-shaped flask, then add 10 cm3 of ethanol, 12 cm3 of glacial ethanoic acid and 15 drops of concentrated sulfuric acid in a fume cupboard.
- Clamp the flask in a beaker of water over a Bunsen, so the mixture is below the water line, and set up a condenser for heating under reflux. Do not put a stopper in the top.
- Boil gently for about 15 minutes, then turn off the heat and remove the water bath.
- Rearrange for distillation and collect the liquid that distils over.
- Shake the distillate with saturated sodium carbonate solution in a separating funnel, venting the carbon dioxide through the tap after each inversion.
- Let the layers settle. Ethyl ethanoate is less dense than water, so it is the upper layer. Run off and discard the lower aqueous layer.
- Add the ester to about 1 g of anhydrous sodium sulfate (or calcium chloride) in a dry boiling tube, and shake to take out the water.
- Decant into a dry flask, add fresh anti-bumping granules and redistil, collecting the fraction that comes over at 74–79 °C. Never distil to dryness.
- Weigh the receiver before and after to find the mass collected.
Apparatus & techniques
- heating under reflux & distillation
- filtration under reduced pressure
- recrystallisation
- using a separating funnel & a drying agent
- measuring a melting point and a boiling range
- percentage yield & atom economy
Safety. Wear eye protection throughout. Concentrated sulfuric acid is corrosive and so is ethanoic anhydride, whose vapour is a severe eye and respiratory irritant. Dispense it in a fume cupboard. Ethanol and ethyl ethanoate are highly flammable, so every heating step uses a water bath or electric heater, never a naked flame near the solvent. Aspirin is harmful if swallowed; do not taste anything you make.
What you should see. The aspirin appears the moment the warm mixture hits cold water, as a fine white solid; after recrystallisation the same material comes back as larger, whiter needle-shaped crystals. In the ester preparation, the sweet, fruity smell is obvious as soon as the reflux starts, and the separating funnel gives two clear layers with a sharp boundary.
Data analysis: limiting reagent, yield and purity
Both halves finish with the same calculation: work out which reagent is limiting, use it to get the theoretical yield, and compare that with what you actually collected.
| Observation | What it means |
|---|---|
| Melts sharply, at the data-book value | Pure: the sample collapses to a liquid at essentially one temperature |
| Melts lower than the book value | Impure: impurities depress the melting point |
| Melts over a range of several degrees | Impure, and the wider the range, the more impurity |
| Does not melt, or chars | The compound decomposes on heating, so this method will not work for it |
For a liquid, the equivalent test is the boiling range: a pure liquid distils over a narrow range at its literature boiling point. Ethyl ethanoate boils at 77 °C, which is why the 74–79 °C fraction is the one to keep.
Which reagent runs out first? Pick a preparation and change what you start with. The limiting reagent is recalculated every time: “in excess” is something you work out from the amounts in moles, never something you can read off the volumes.
- Reweigh the empty container. When transferring solid from a weighing boat, weigh it before and after tipping to find the mass actually transferred. Weighing directly into the reaction vessel on a tared balance is also valid.
- Heat flammable solvents on a water bath, never with a Bunsen. Say the practical step, not just “ethanol is flammable”: the mark is for the precaution.
- Recrystallise from the minimum volume of hot solvent and keep the temperature below its boiling point, so the solvent does not boil away and the product does not stay in solution when it cools.
- Wash the crystals with a little cold solvent on the Büchner funnel. Cold, so the product does not redissolve; a little, so it only rinses off the soluble impurities left in the film of solution.
- Never seal a refluxing apparatus. A bung in the top of the condenser lets pressure build until it is forced out. The top is open by design.
- Condenser water in at the bottom, out at the top. The wrong way round and the jacket never fills, so the vapour is not condensed and product escapes.
- Vent the separating funnel. Sodium carbonate neutralises the acid and produces carbon dioxide, so pressure builds up: invert, then open the tap, every time.
- Say which layer and why. Two layers form because the ester is immiscible with water; the ester is on top because it is less dense. “Lighter” does not earn the mark.
Exam questions: RP10
- Add about 6 g of salicylic acid to a weighing boat.
- Place the weighing boat on a 2 decimal place balance and record the mass.
- Tip the salicylic acid into a 100 cm3 conical flask.
- __________________
- Add 10 cm3 of ethanoic anhydride to the conical flask and swirl.
- Add 5 drops of concentrated phosphoric acid.
- Warm the flask for 20 minutes.
- Add ice-cold water to the reaction mixture and place the flask in an ice bath.
- Filter off the crude aspirin from the mixture and leave it to dry.
- Weigh the crude aspirin and calculate the yield.
Describe the instruction that is missing from step 4 of the method. Justify why this step is necessary.
Show answer
Instruction: reweigh the empty weighing boat. 1 mark
Justification: so you can calculate the exact mass of salicylic acid actually added to the reaction mixture. 1 mark
Suggest a suitable piece of apparatus to measure out the ethanoic anhydride in step 5. Identify a hazard of using concentrated phosphoric acid in step 6.
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Apparatus: a 10 cm3 measuring cylinder. A pipette, burette, graduated pipette or 10 cm3 syringe are all accepted. 1 mark
Hazard: corrosive (skin burns or permanent eye damage). 1 mark
If you name a volume it must be between 10 and 50 cm3. “Irritant” and “toxic” are both ignored for the hazard.
A 6.01 g sample of salicylic acid (Mr = 138.0) is reacted with 10.5 cm3 of ethanoic anhydride (Mr = 102.0). In the reaction the yield of aspirin is 84.1%. The density of ethanoic anhydride is 1.08 g cm−3. Show by calculation which reagent is in excess, and calculate the mass, in g, of aspirin (Mr = 180.0) produced.
Show answer
n(salicylic acid) = 6.01 ÷ 138.0 = 4.36 × 10−2 mol
1 mark
mass of (CH3CO)2O = 10.5 × 1.08 = 11.34 g
1 mark
n((CH3CO)2O) = 11.34 ÷ 102.0 = 1.11 × 10−1 mol
1 mark
The reaction is 1 : 1, and there is far more anhydride, so ethanoic anhydride is in excess. 1 mark
mass of aspirin = 4.36 × 10−2 × 0.841 × 180.0 = 6.59 g
1 mark: 2 s.f. or more.
You must both show the two amounts and state that the anhydride is in excess. Later marks carry forward from an earlier slip.
Suggest two ways in which the melting point of the crude aspirin collected in step 9 would differ from the melting point of pure aspirin.
Show answer
The value would be lower. 1 mark
It would melt over a range of temperatures rather than sharply. 1 mark
The crude aspirin can be purified by recrystallisation using hot ethanol (boiling point 78 °C) as the solvent. Describe two important precautions when heating the mixture of ethanol and crude aspirin.
Show answer
Use a water bath rather than a Bunsen burner, because ethanol is flammable. 1 mark
Heat to a temperature below its boiling point, so the ethanol does not boil away, and use the minimum volume of solvent. 1 mark
The first mark needs a practical step. Simply writing “ethanol is flammable” states the hazard without giving the precaution, and scores nothing.
The pure aspirin is filtered under reduced pressure. A small amount of cold ethanol is then poured through the Büchner funnel. Explain the purpose of adding a small amount of cold ethanol.
Show answer
To remove soluble impurities, washing away the film of ethanolic solution left on the product, while the solvent being cold and in small amount stops the aspirin itself dissolving. 1 mark
A sample of the crude aspirin is kept to compare with the purified aspirin. Describe one difference in appearance you would expect to see between these two solid samples.
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The pure product has larger, needle-like crystals, or is lighter in colour. 1 mark
Whiter, less grey, more crystalline, less powdery, shinier or a single colour are all accepted, but the answer must be tied to the pure product (or the opposite point tied to the crude one).
- A mixture of ethanol, ethanoic acid, and concentrated sulfuric acid, with anti-bumping granules, is heated under reflux for 10 minutes.
- The apparatus is rearranged for distillation.
- The mixture is heated to collect the liquid that distils between 70 and 85 °C.
- The distillate is placed in a separating funnel. Aqueous sodium carbonate is added, and a stopper is placed in the funnel. The mixture is shaken, releasing pressure as necessary.
- The lower aqueous layer is removed and the upper organic layer is placed in a small conical flask.
- Anhydrous calcium chloride is added to the sample in the conical flask. The flask is shaken well and left for a few minutes.
- The liquid from the flask is redistilled and the distillate is collected between 74 and 79 °C.
State the role of the concentrated sulfuric acid in this reaction. The reaction mixture is flammable — suggest how it should be heated in step 1.
Show answer
Role: catalyst. 1 mark: “reduces the activation energy” is allowed; “speeds up the reaction”, “provides an alternative path”, “proton donor” and “dehydrating agent” are all ignored.
Heating: an electric heater, heat mantle or hot water bath. 1 mark: a heating plate, sand bath or oil bath are fine too. Any suggestion of direct heat from a Bunsen scores nothing.
The figure below shows how a student set up the apparatus for reflux in step 1. Assume the apparatus is clamped correctly. Identify two mistakes the student made, and state the problem caused by each.
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Mistake 1: there is a bung / stopper in the end of the condenser. 1 mark
Problem: pressure builds up: the stopper could be forced out. 1 mark
Mistake 2: the water goes the wrong way through the condenser (in at the top, out at the bottom). 1 mark
Problem: the condenser does not fill with water / is not cool enough, so the vapour is not condensed effectively and reactants or products escape. 1 mark
“The glass shatters” and “it explodes” are ignored, as is “the condenser is the wrong way round” without saying which way the water flows. An unsealed neck of the flask is an equally valid pair of marks.
State why sodium carbonate is added to the distillate in step 4. Explain why there is a build-up of pressure in the separating funnel.
Show answer
To neutralise / remove the acid (the ethanoic acid and / or the sulfuric acid). 1 mark
Carbon dioxide gas is produced: you see effervescence. 1 mark
Naming the wrong acid or the wrong gas loses the mark. “Neutralise the distillate” is ignored. Say the acid.
Give a reason why two layers form in the separating funnel. Suggest why ethyl ethanoate forms the upper layer.
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Ethyl ethanoate is immiscible with / insoluble in water, so the aqueous and organic layers do not mix. 1 mark
It is less dense than water. 1 mark
“Lighter” is not accepted, and references to polarity or intermolecular forces are ignored.
State why anhydrous calcium chloride is added in step 6.
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To remove / absorb water: it is a drying agent. 1 mark
“Dehydrates” is ignored, and “to remove soluble impurities” or “to dry the reactants” score nothing.
A student adds 10.0 cm3 of ethanol (Mr = 46.0) to 5.25 g of ethanoic acid (Mr = 60.0) and obtains 5.47 g of ethyl ethanoate (Mr = 88.0). For ethanol, density = 0.790 g cm−3. Determine the limiting reagent and calculate the percentage yield. Then suggest a reason why the percentage yield is not 100%.
Show answer
mass of ethanol = 10.0 × 0.790 = 7.90 g
1 mark
n(ethanol) = 7.90 ÷ 46.0 = 0.172 mol n(ethanoic acid) = 5.25 ÷ 60.0 = 0.0875 mol
1 mark: both amounts must be shown.
The limiting reagent is ethanoic acid. 1 mark
maximum mass = 0.0875 × 88.0 = 7.70 g
1 mark
% yield = (5.47 ÷ 7.70) × 100 = 71.0%
1 mark: 70.6 to 71.1, to at least 2 s.f.
Why not 100%: the esterification is reversible and reaches equilibrium, so it never goes to completion, and product is lost at every transfer, in the washing and on redistillation. 1 mark
Source: AQA A-Level Chemistry past papers.
Source: AQA A-Level Chemistry past papers.
Test-tube reactions to identify transition-metal ions
Full theory, every observation and equation: Reactions of aqueous ions · Transition metals
Aim: carry out simple test-tube reactions on unknown aqueous solutions of transition-metal (and other) ions, and record the observations. AQA are explicit about this one: in the task itself you are not required to identify any of the solutions or products: the practical is about seeing and describing accurately. The identifying comes in the exam.
You are given three solutions labelled Q, R and S, plus sodium hydroxide, sodium carbonate and silver nitrate solutions.
- Test 1(a). Put about 10 drops of solution Q in a test tube. Add sodium hydroxide solution dropwise, with gentle shaking, until in excess. Do not discard the mixture. Repeat with R and then S.
- Test 1(b). Half-fill a 250 cm3 beaker with freshly boiled water and stand the tubes from Test 1(a) in it for about 10 minutes. Start Test 2 while you wait.
- Test 2. Put about 10 drops of sodium carbonate solution in a fresh tube, add about 10 drops of solution Q and shake gently. Repeat with R and S.
- Test 3. Put about 10 drops of solution Q in a fresh tube, add about 10 drops of silver nitrate solution, shake gently, and let the tubes stand for about 10 minutes. Repeat with R and S.
Record what you see on dropwise addition, on addition to excess and on standing. Where nothing happens, write “no visible change”: a blank cell earns nothing.
Apparatus & techniques
- qualitative test-tube reactions
- dropwise addition, and addition to excess
- using a hot-water bath
- recording observations precisely
- safe handling of alkalis & silver nitrate
Safety. Wear eye protection. Sodium hydroxide solution is corrosive: wash any splash off immediately. Silver nitrate stains skin and clothing black and is harmful to the environment, so keep it off your hands and do not pour it down the sink. Transition-metal salts are harmful if swallowed. Use the hot water from a kettle or a beaker of previously boiled water. Never boil solutions in a test tube over a flame.
What you should see. Every one of these ions gives a precipitate with a little alkali, because the hydroxide is insoluble. What separates them is what happens next: whether the precipitate redissolves in excess, whether it changes colour on standing, and whether the carbonate gives a gas.
Data analysis: the observations that separate the ions
| Ion in solution | Colour | Precipitate with a little OH− | Excess NaOH? | With Na2CO3 |
|---|---|---|---|---|
| [Fe(H2O)6]2+ | pale green | green, Fe(H2O)4(OH)2 (darkens to brown in air) | Insoluble | Green FeCO3, no gas |
| [Fe(H2O)6]3+ | yellow-brown | red-brown, Fe(H2O)3(OH)3 | Insoluble | Red-brown precipitate and effervescence |
| [Cu(H2O)6]2+ | blue | blue, Cu(H2O)4(OH)2 | Insoluble | Blue-green CuCO3, no gas |
| [Al(H2O)6]3+ | colourless | white, Al(H2O)3(OH)3 | Dissolves → colourless [Al(OH)4]− | White precipitate and effervescence |
| [Mg(H2O)6]2+ | colourless | white, Mg(OH)2 | Insoluble | White MgCO3, no gas |
A 3+ ion is smaller and more highly charged than a 2+ ion, so it has a much higher charge density and polarises the O–H bonds in its water ligands far more strongly. That makes the 3+ aqua ion a much better proton donor, acidic enough to react with carbonate and release CO2:
2[Fe(H2O)6]3+ + 3CO32− → 2Fe(H2O)3(OH)3 + 3CO2 + 3H2O
The 2+ ions are not acidic enough. They simply swap partners and precipitate the carbonate itself, with no gas:
[Fe(H2O)6]2+ + CO32− → FeCO3 + 6H2O
So effervescence with carbonate is a 3+ test, and it is the fastest way to sort a 3+ ion from a 2+ one in an unknown.
- Every colour observation must say precipitate or solution. “Green” is not an observation; “a green precipitate” is. This is the single commonest lost mark in the whole practical.
- Add dropwise, then to excess, and record both. The aluminium precipitate only gives itself away when the excess redissolves it.
- Say “no visible change” rather than leaving a cell blank. It is a real result and it carries marks.
- Watch the iron(II) precipitate on standing. It darkens from green towards brown as it is oxidised by air, which is why AQA build a standing step into the method.
- Ammonia and sodium hydroxide give the same precipitates at first, because both provide OH−. They differ only in excess, where excess ammonia dissolves the copper precipitate to a deep blue solution but excess sodium hydroxide does not.
- Include the water ligands in A-level formulae. Fe(H2O)3(OH)3 is the expected answer, not Fe(OH)3, though the simpler form is usually allowed.
Exam questions: RP11
Aqueous aluminium sulfate is added to aqueous sodium carbonate. What are the formulas of the precipitate and the gas formed?
Cobalt(II) chloride solution changes colour when an excess of concentrated hydrochloric acid is added. What type of reaction takes place?
When anhydrous aluminium chloride reacts with water, solution Y is formed that contains a complex aluminium ion, Z, and chloride ions. Give an equation for this reaction, and give an equation to show how the complex ion Z can act as a Brønsted–Lowry acid with water.
Show answer
AlCl3 + 6H2O → [Al(H2O)6]3+ + 3Cl−
1 mark
[Al(H2O)6]3+ + H2O → [Al(H2O)5(OH)]2+ + H3O+
1 mark: an equation going on to [Al(H2O)4(OH)2]+ is equally acceptable.
Describe two observations you would make when an excess of sodium carbonate solution is added to solution Y. Give an equation for the reaction, including the formula of each complex aluminium species.
Show answer
Observation 1: a white precipitate forms. 1 mark
Observation 2: effervescence / bubbles / fizzing. 1 mark
2[Al(H2O)6]3+ + 3CO32− → 2[Al(H2O)3(OH)3] + 3CO2 + 3H2O
1 mark: multiples are accepted; spectator ions only if the equation still balances.
The two observations can be given in either order, but both are needed. “White” on its own does not score: it has to be a white precipitate.
Aqueous potassium hydroxide is added, until in excess, to solution Y. Describe two observations you would make, and for each give an equation for the reaction that occurs, including the formula of each complex aluminium species.
Show answer
Observation 1: a white precipitate forms. 1 mark
[Al(H2O)6]3+ + 3OH− → [Al(H2O)3(OH)3] + 3H2O
1 mark
Observation 2: the precipitate dissolves: a colourless solution forms. 1 mark
[Al(H2O)3(OH)3] + OH− → [Al(H2O)2(OH)4]− + H2O
1 mark: the 4-coordinate form [Al(OH)4]− + 3H2O is equally accepted; only 6- or 4-coordination is allowed.
| Test | Observations with L | Observations with M |
|---|---|---|
| Add ammonia solution slowly until in excess. | A red-brown precipitate forms that is insoluble in excess. | A green precipitate forms that is insoluble in excess. |
| Add sodium carbonate solution. | A red-brown precipitate forms. Effervescence is seen. | A green precipitate forms. |
| Add dilute nitric acid, then divide into two portions. | No change is seen. | No change is seen. |
| Add barium chloride solution to the first portion. | No change is seen. | A white precipitate forms. |
| Add silver nitrate solution to the second portion. | A white precipitate forms. | No change is seen. |
Identify L and M using the results in the table. In your answer: identify all precipitates; explain why effervescence is seen in the reaction of sodium carbonate with L but not with M; and give ionic equations for all reactions.
Show answer
Marked by levels of response across three stages.
Stage 1: the identifications. Both red-brown precipitates from L are Fe(H2O)3(OH)3. The green precipitate from M is Fe(H2O)4(OH)2 with ammonia and FeCO3 with carbonate. The white precipitate with L is AgCl and with M is BaSO4. So L is FeCl3 and M is FeSO4.
Stage 2: the effervescence. The gas from L is carbon dioxide. [Fe(H2O)6]3+ is more acidic than [Fe(H2O)6]2+, because Fe3+ is smaller and more highly charged: a greater charge density, so it polarises the O–H bonds more and is a better proton donor.
Stage 3: the equations.
[Fe(H2O)6]3+ + 3NH3 → Fe(H2O)3(OH)3 + 3NH4+
2[Fe(H2O)6]3+ + 3CO32− → 2Fe(H2O)3(OH)3 + 3CO2 + 3H2O
[Fe(H2O)6]2+ + 2NH3 → Fe(H2O)4(OH)2 + 2NH4+
[Fe(H2O)6]2+ + CO32− → FeCO3 + 6H2O
Ag+ + Cl− → AgCl Ba2+ + SO42− → BaSO4
Level 3 — 5–6 marks all three stages covered, correct and virtually complete, communicated coherently with the equations.
Level 2 — 3–4 marks all three stages covered but incomplete or containing inaccuracies, or two stages covered well.
Level 1 — 1–2 marks two stages covered but incomplete, or one stage covered well.
Precipitates count as identified if they appear as products in your equations or in a table. Names are allowed provided the oxidation state of the iron is given.
Source: AQA A-Level Chemistry past papers.
Source: AQA A-Level Chemistry past papers.
Separation of species by thin-layer chromatography
Full theory, Rf and the interactive plate: Chromatography · Amino acids: two-way TLC
Aim: separate a mixture by thin-layer chromatography and identify its components from their Rf values. AQA’s exemplar analyses the active ingredients in common painkillers (aspirin, paracetamol, ibuprofen and caffeine) and asks you to work out what a combination tablet contains.
- Crush an aspirin tablet with a pestle and mortar and transfer it to a weighing boat.
- Dissolve about 0.1 g of the powder in 0.5 cm3 of ethanol.
- Repeat for the ibuprofen and paracetamol tablets.
- Crush the caffeine tablet and dissolve about 0.1 g in 7.0 cm3 of ethanol, a much more dilute solution, because a caffeine tablet is nearly all caffeine.
- Repeat for the combination tablet (one containing aspirin, paracetamol and caffeine together).
- Draw a faint pencil line 1 cm above the bottom of a TLC plate and mark five equally spaced spots along it.
- Use a capillary tube to put a tiny drop of each solution on a different spot, and let the plate air dry.
- Put about 10 cm3 of ethyl acetate in the developing chamber.
- Stand the plate in the chamber with the solvent level below the spotting line, and replace the lid so it seals.
- When the solvent reaches about 1 cm from the top, take the plate out and mark the solvent front in pencil straight away. Dry it in the fume cupboard.
- Visualise the spots under a UV lamp and draw round them lightly in pencil.
- Calculate the Rf value of every spot.
Apparatus & techniques
- thin-layer chromatography
- spotting with a capillary tube
- using a sealed developing chamber
- visualising with UV or a locating agent
- measuring and comparing Rf values
Safety. Wear eye protection. Ethanol and ethyl acetate are highly flammable and their vapours are irritating. Keep them away from flames and work in a fume cupboard or a well-ventilated room. Never look directly at the UV lamp, and keep the plate flat under it rather than holding it. Handle plates by the edges only: fingerprints leave their own spots and grease ruins the run.
What you should see. Nothing at all, until the plate goes under the lamp: all five compounds are colourless. Under UV the silica fluoresces and the spots show up as dark patches. The combination tablet gives several spots, one lining up with each pure substance it contains, which is exactly how you read it.
Data analysis: Rf and what it can and cannot tell you
Rf = distance moved by the spot ÷ distance moved by the solvent front
Both distances are measured from the pencil origin line, and the spot is measured to its centre. Rf is a ratio, so it has no units and is always between 0 and 1.
| What you see | What it tells you |
|---|---|
| A spot in the mixture lane at the same height as a pure reference | That substance is probably present (same Rf, same conditions) |
| A high Rf | More soluble in the moving phase, and less strongly adsorbed by the polar silica |
| A low Rf | Held tightly by the stationary phase: a more polar substance on silica |
| One spot that will not separate | Two substances may have the same Rf in this solvent: run it again in a different one |
| A streak rather than a spot | The spot was too concentrated, or the solvent was above the pencil line and washed it off |
Two different compounds can share an Rf value, so a match is evidence, not proof. It only means anything at all when the reference and the unknown are run on the same plate, in the same solvent, at the same temperature, which is why the pure references go on the plate beside the mixture rather than being looked up in a book. To be certain you either run a second solvent (two-way TLC) or take the separated substance away for NMR or mass spectrometry.
- Pencil, not pen. Ink is a mixture and would run up the plate with everything else.
- The solvent must start below the pencil line. If it covers the spots they dissolve straight into the solvent in the tank and never travel up the plate.
- Lid on the chamber. It keeps the atmosphere saturated with solvent vapour, so the solvent does not evaporate off the plate as it rises and the front stays level.
- Mark the solvent front the moment the plate comes out. Once the solvent evaporates you cannot tell where it got to, and every Rf is lost.
- Tiny, dry spots. A big wet spot streaks and the components overlap; let each application dry before adding the next.
- Measure to the centre of the spot, from the pencil origin line, not from the bottom edge of the plate.
- Colourless compounds need visualising: a UV lamp for anything that fluoresces or quenches, ninhydrin for amino acids, or iodine vapour.
Exam questions: RP12
Suggest a suitable reagent for the hydrolysis of a protein.
Show answer
Concentrated hydrochloric acid. 1 mark
Any concentration of 5 mol dm−3 or higher is allowed, and concentrated sulfuric acid or a concentrated strong alkali are both accepted.
Suggest how the positions of the amino acids on the TLC plate were located.
Show answer
Using ninhydrin or ultraviolet light. 1 mark
Iodine vapour is also allowed.
Deduce the minimum number of amino acids present in the original mixture.
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Seven. 1 mark
“Minimum” matters: two amino acids could still be sitting under one another in a single spot even after two solvents, so seven spots means at least seven amino acids.
Suggest why it was necessary to use two different solvents.
Show answer
Some of the amino acids did not separate in the first solvent: they had the same Rf value, or the same affinity for that solvent. 1 mark
Simply saying “amino acids have different Rf values in different solvents” is not accepted: the mark is for explaining that the first solvent left some of them unseparated.
Source: AQA A-Level Chemistry past papers.
Source: AQA A-Level Chemistry past papers.