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Spec Mapping — OCR H420 Module 5.2.1 — Photosynthesis, content statements covering the structures and roles of photosynthetic pigments, the use of chromatography to separate pigments from a leaf, the calculation of R_f values, and the interpretation of absorption and action spectra (refer to the official OCR H420 specification document for exact wording).
Chloroplasts look green because of the photosynthetic pigments they contain — specialised molecules that absorb light energy at specific wavelengths. OCR specification module 5.2.1 and practical activity group 6 (chromatography) require you to know the main pigments, understand absorption and action spectra, and be able to use chromatography to separate pigments and calculate Rf values. This lesson links physical chemistry (absorption of photons) to biological function (exciting electrons in photosystems) and includes a required practical.
The relationship between absorption spectrum and action spectrum was made famous by the American biophysicist Robert Emerson (1957). His "red-drop" experiment (paraphrasing his findings) showed that photosynthesis efficiency dropped sharply as wavelength climbed above ~680 nm, but could be partially restored by simultaneously illuminating with shorter wavelengths. Emerson's "enhancement effect" was the first evidence that photosynthesis uses two photosystems acting in series, each absorbing different wavelengths and contributing different excitations. The result, paraphrasing Emerson's school of thought, anchored the Z-scheme architecture you will meet in the next lesson.
Key Definitions:
- Pigment — a coloured molecule that absorbs certain wavelengths of visible light and reflects or transmits others.
- Absorption spectrum — a graph showing the absorbance of different wavelengths of light by a pigment.
- Action spectrum — a graph showing the rate of photosynthesis at different wavelengths of light.
- Chromatography — a technique for separating mixtures of substances based on differences in solubility and affinity for a stationary phase.
- Rf value — the ratio of the distance travelled by a substance to the distance travelled by the solvent front.
A photon of light only becomes useful biologically when it is absorbed by a pigment molecule. When absorbed, the photon transfers its energy to an electron in the pigment, raising it to a higher energy level ("excitation"). This high-energy electron is then harvested by the photosystem and passed down the electron transport chain.
Without pigments, chloroplasts would be transparent to visible light and no photosynthesis could occur. Different pigments absorb different wavelengths — together they allow plants to capture a broad range of the visible spectrum.
OCR expects you to know four main pigments found in higher plants. They fall into two groups: primary (chlorophyll a) and accessory (chlorophyll b and carotenoids).
| Pigment | Colour | Absorbs | Role | Primary/Accessory |
|---|---|---|---|---|
| Chlorophyll a | Blue-green | Red (~670 nm) and blue-violet (~430 nm) | Direct involvement in the light reaction at the reaction centre of PSI and PSII | Primary |
| Chlorophyll b | Yellow-green | Red (~650 nm) and blue (~475 nm) | Harvests light and passes energy to chlorophyll a | Accessory |
| Carotene (β-carotene) | Orange | Blue-violet (~450 nm) | Harvests light energy; protects against photo-oxidative damage | Accessory |
| Xanthophyll | Yellow | Blue-violet (~450 nm) | Harvests light; also involved in photoprotection | Accessory |
Chlorophyll a is essential — without it no photosynthesis can occur. Accessory pigments broaden the range of wavelengths captured and funnel energy to chlorophyll a in the reaction centre. Together, hundreds of pigment molecules plus a central chlorophyll a pair form a photosystem (a light-harvesting antenna complex with a reaction centre).
An absorption spectrum tells you how much light of each wavelength a pigment absorbs. An action spectrum tells you how much photosynthesis occurs at each wavelength.
flowchart LR
W[White light] --> P[Pigments]
P -->|Absorb blue & red| E[Excited electrons]
P -->|Reflect green| G[Green colour we see]
E --> PS[Photosystem reaction centre]
PS --> ETC[Electron transport chain]
Plants reflect and transmit the wavelengths of light they do not absorb efficiently. Since chlorophylls absorb strongly in red and blue but weakly in green, green light is reflected into your eyes — hence leaves appear green. This sometimes surprises students: the green you see is the light the plant has "wasted", not the light it is using.
OCR practical activity group 11 requires you to separate the pigments in a leaf extract by paper or thin-layer chromatography (TLC) and calculate Rf values.
Rf=distance moved by solvent frontdistance moved by pigment
Rf values are always between 0 and 1, and they are characteristic of a particular pigment in a particular solvent system.
| Pigment | Colour of spot | Approximate Rf (petroleum ether:propanone) |
|---|---|---|
| Carotene | Orange-yellow | ~0.95 |
| Xanthophyll | Yellow | ~0.70 |
| Chlorophyll a | Blue-green | ~0.60 |
| Chlorophyll b | Yellow-green | ~0.45 |
Rf values depend on the solvent system — if the solvent is changed, the values will change too. Always compare values from the same chromatography run.
Each pigment has a different solubility in the solvent and a different affinity for the stationary phase (the paper or silica). Pigments that are more soluble in the solvent and less attracted to the stationary phase travel further (high Rf). Pigments that are less soluble and more attracted to the stationary phase travel less (low Rf). Carotene is non-polar, so it dissolves well in the non-polar solvent and moves near the top; chlorophyll b is the most polar, so it stays closer to the origin.
When drawing or interpreting chromatography, always remember to draw the origin line in pencil (ink would dissolve), mark the solvent front before the solvent evaporates, and measure to the centre of the spot, not the leading or trailing edge. A common OCR mark scheme point is that "Rf value has no units." If asked to identify a pigment from an Rf value, you can only do so if told the solvent system; the same pigment has different Rf values in different solvents.
Notice that the action spectrum (the rate of photosynthesis) is consistently a little higher in the green and yellow regions than chlorophyll a alone would predict. That gap is the contribution of accessory pigments. The convergence is one of the most-cited pieces of evidence in plant physiology that pigments are causal in photosynthesis — paraphrasing the standard reasoning, if any wavelength absorbed by a pigment failed to drive photosynthesis, the action and absorption spectra would diverge.
Chlorophyll a and chlorophyll b share the same core: a porphyrin ring (a flat, square structure of four pyrrole rings around a central magnesium atom) bonded to a long hydrocarbon phytol tail. The porphyrin ring absorbs photons. The phytol tail anchors the molecule in the lipid bilayer of the thylakoid membrane. Chlorophyll a has a methyl (-CH₃) group at one position; chlorophyll b has an aldehyde (-CHO) at that same position. This trivial chemical difference shifts the absorption peaks by ~20 nm, broadening the wavelengths captured by the photosystem.
Carotenoids are terpene-derived linear conjugated polyene molecules (β-carotene is a 40-carbon chain with 11 alternating double bonds). The extended π-conjugation gives them their orange–red colour. Xanthophylls are oxidised carotenoids (one or more -OH groups), shifting the spectrum slightly. Both carotenoids and xanthophylls are bound non-covalently to the photosystem proteins in the antenna complex.
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