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13.1.613 · Photosynthesis

Chromatography of Chloroplast Pigments — Cambridge AS & A Level Biology 9700 Revision Notes & Topical Questions

5 min read 8 parts Lesson 7 of 13 Unit quiz

📚 Cambridge AS & A Level Biology 9700 Hub / Topical Revision
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13.1.6 Chromatography of Chloroplast Pigments — CAMBRIDGE AS & A LEVEL BIOLOGY 9700 — REVISION NOTES notes: Use chromatography and Rf values to separate and identify chloroplast pigments.Technique — used to separate mixtures.Mobile phase — the solvent in which the mixture is diss…

  • Stationary phase
  • Separation
  • Unique value
  • Solvent dependence
  • Carotenoids
  • Chlorophyll a

Section 1 of 8Principle of chromatography

1Technique
  • Technique used to separate mixtures.
2Mobile phase
  • Mobile phase the solvent in which the mixture is dissolved.
3Stationary phase
  • Stationary phase the static material the mixture passes through.
4Separation
  • Separation components travel at different speeds, so they separate.

Section 2 of 8Types of chromatography

MethodStationary phaseFeatures
Paper chromatographyPaper (cellulose)Simple and cheap
Thin layer chromatographyThin layer of adsorbent such as silica gelMixture travels faster and separates more distinctly

Section 3 of 8Method for chloroplast pigments

1
Extract pigments from leaves.
2
Spot the extract on the baseline of the paper or plate.
3
Place in solvent so the solvent front rises through the stationary phase.
4
Remove before the solvent reaches the top and mark the solvent front.
5
Measure distances moved by each pigment and by the solvent.

Section 4 of 8Rf values

Rf
  • Rf retardation factor.
Rf = distance travelled by component ÷ distance travelled by solvent.
Unique value
  • Unique value each pigment has its own Rf, so pigments can be identified.
Solvent dependence
  • Solvent dependence the specific value depends on the solvent used.
Small Rf
  • Small Rf the pigment is less soluble and larger.

Section 5 of 8Relative Rf values

Carotenoids
  • Carotenoids highest Rf, usually close to 1.
Chlorophyll a
  • Chlorophyll a between carotenoids and chlorophyll b.
Chlorophyll b
  • Chlorophyll b much lower Rf.

Section 6 of 8More from the specification

1
Photophosphorylation in thylakoid membranes
  • Light energy absorbed by photosystem I and II excites electrons, driving non-cyclic and cyclic photophosphorylation to generate ATP and reduced NADP.
2
Thylakoid structure and pigment separation
  • Paper chromatography separates accessory pigments (chlorophyll a, chlorophyll b, carotenes,
  • and xanthophylls) from chloroplast extracts based on relative solubility in organic solvents.
3
Light-independent reactions in the stroma
  • The Calvin cycle in the stroma uses ATP and NADPH produced during light-dependent reactions to reduce glycerate-3-phosphate into triose phosphate.
4
Photophosphorylation in thylakoid membranes
  • Light energy absorbed by photosystem I and II excites electrons, driving non-cyclic and cyclic photophosphorylation to generate ATP and reduced NADP.
5
Thylakoid structure and pigment separation
  • Paper chromatography separates accessory pigments (chlorophyll a, chlorophyll b, carotenes,
  • and xanthophylls) from chloroplast extracts based on relative solubility in organic solvents.
6
Light-independent reactions in the stroma
  • The Calvin cycle in the stroma uses ATP and NADPH produced during light-dependent reactions to reduce glycerate-3-phosphate into triose phosphate.
7
Light-dependent stage
  • Stage in thylakoids making ATP, reduced NADP and oxygen.
8
Light-independent stage
  • Stage in the stroma where CO2 is converted to organic molecules.
9
Cyclic photophosphorylation
  • ATP formation using only PSI with electrons returning to it.
10
Electron carrier
  • A molecule such as NAD or FAD that accepts and donates electrons.
11
Non-cyclic photophosphorylation
  • ATP formation using PSII and PSI with electrons ending in NADP.
12
Triose phosphate
  • Three-carbon sugar phosphate made from GP.
13
Photosystem
  • A cluster of pigments and proteins that harvest light.
14
Photophosphorylation
  • Making ATP using light energy, an electron transport chain and chemiosmosis.
15
Thylakoid
  • Membrane sac where the light reactions occur.
16
Photosynthesis
  • The process by which plants make carbohydrates from raw materials using energy from light.
17
Stroma
  • The fluid of the chloroplast where the Calvin cycle occurs.
18
Reduced
  • Having gained electrons
19
Photolysis
  • Splitting of water by light at PSII.
20
Calvin cycle
  • Light-independent reactions that fix CO2 in the stroma.

Section 7 of 8Key terms

TermMeaning
ChromatographyTechnique for separating a mixture.
Mobile phaseSolvent that carries the mixture.
Stationary phaseMaterial the mixture moves through.
Rf valueDistance moved by a component divided by distance moved by the solvent.
Solvent frontFurthest point the solvent has reached.
Thin layer chromatographyChromatography using a thin layer of adsorbent such as silica gel.

Section 8 of 8Exam technique

1
Giving Rf units — it is a ratio with no units.
2
Measuring the solvent distance from the baseline of the pigment — measure from the baseline to the solvent front.
3
Saying a larger Rf means a larger pigment — a small Rf means less soluble and larger.
4
Letting the solvent run over the top — the front must be marked before it dries.

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