Photosynthesis

In a nutshell

Photosynthesis is how plants capture light energy and use it to build sugars, storing energy in chemical bonds that nearly every other organism ultimately depends on.

It happens in two linked stages inside the chloroplast: the light-dependent reaction (in the thylakoid membranes) makes ATP and reduced NADP and splits water; the light-independent reaction (the Calvin cycle, in the stroma) uses those two products to fix carbon dioxide into a simple sugar.

Overall: 6CO₂ + 6H₂O ⟶ C₆H₁₂O₆ + 6O₂ (in the presence of light and chlorophyll).

Assumed knowledge: ATP.

Core content

Where each stage happens

The two stages are physically separated inside the chloroplast, and the products of the first stage feed straight into the second.

The ATP and reduced NADP made in the thylakoids diffuse into the stroma, where the Calvin cycle uses them. The empty ADP, P_i and NADP then return to the thylakoids to be re-used.

The light-dependent reaction

This stage needs light, and it happens on the thylakoid membranes. Four things happen, and the marks come from getting each one precise.

1. Photoionisation of chlorophyll.

• Chlorophyll absorbs light energy, which excites electrons inside it so that they are emitted from the chlorophyll.
• The chlorophyll is left positively charged. This loss of electrons on absorbing light is photoionisation.

2. ATP is made by chemiosmosis (photophosphorylation).

• The emitted electrons pass along a chain of electron carriers (the electron transfer chain) in the thylakoid membrane, releasing energy as they go.
• This energy moves protons (H⁺) across the thylakoid membrane, into the thylakoid space, building up a proton gradient.
• The protons flow back down their gradient into the stroma through ATP synthase, and the energy this releases joins ADP + P_i ⟶ ATP.
• This is chemiosmosis, and because the energy came from light it is called photophosphorylation.

Still don't get it? · chemiosmosis

Picture a hydroelectric dam. A pump pushes water uphill into a high reservoir, and that water is only allowed back down through one narrow channel, where it spins a turbine and generates electricity.

Now the chloroplast version, one step at a time:

• The electron transfer chain is the pump. As electrons drop down the chain, the energy they release is used to push protons (H⁺) uphill, from the stroma into the thylakoid space.
• The protons pile up inside the thylakoid space, just like water building up behind the dam. This store of protons is the proton gradient.
• The protons are only allowed back out through one channel: the enzyme ATP synthase. As they rush through, ATP synthase acts like the turbine, using their energy to join ADP and P_i into ATP.

Back to the exam wording: energy from the electron transfer chain moves protons across the thylakoid membrane, and the protons then pass back through ATP synthase, providing the energy to make ATP. That movement of protons down a gradient to drive ATP synthesis is chemiosmosis.

3. Photolysis of water.

• The electrons lost by chlorophyll must be replaced. Light also splits water: this is photolysis.
• Photolysis of water produces protons, electrons and oxygen.
• The electrons replace those lost from chlorophyll; the protons add to the gradient and are used to reduce NADP; the oxygen is a waste product (some is used in respiration, the rest diffuses out).

4. Reduction of NADP.

• At the end of the electron transfer chain, NADP is reduced by the electrons (together with protons) to form reduced NADP.
• Reduced NADP is not made by protons on their own: it needs the electrons.

The two products that leave for the Calvin cycle are ATP and reduced NADP.

The light-independent reaction (the Calvin cycle)

This stage happens in the stroma. It does not use light directly, but it depends completely on the ATP and reduced NADP from the light-dependent reaction, so it stops quickly in the dark.

1. Carbon fixation.

• Carbon dioxide combines with ribulose bisphosphate (RuBP), a 5-carbon compound. This reaction is catalysed by the enzyme rubisco.
• The unstable 6-carbon product immediately splits into two molecules of glycerate 3-phosphate (GP), a 3-carbon compound.

2. Reduction of GP to triose phosphate.

• GP is reduced to triose phosphate (TP).
• The reduction uses reduced NADP (which provides the hydrogen) and energy from ATP.

3. Regeneration of RuBP and making products.

• Most of the TP is used to regenerate RuBP (using more ATP), so the cycle can keep turning.
• Some of the TP is converted to useful organic substances, such as glucose (and from it starch, cellulose, amino acids and lipids).

Still don't get it? · why the Calvin cycle keeps recycling RuBP

Think of a fairground carousel that you have to keep feeding tokens into. Every full turn, you get to take one small prize off, but most of what comes round has to be put straight back in to keep the ride spinning.

Now the biology, built up slowly:

• Each turn of the cycle fixes just one carbon dioxide molecule onto one RuBP, making two GP, then two TP.
• A hexose sugar like glucose has six carbons, so you need to fix six carbon dioxide molecules, which means six turns of the cycle.
• Over those six turns the cycle makes twelve TP. Only two of them are needed to build one glucose. The other ten are used to regenerate the six RuBP molecules that were used up, so the cycle can carry on.

That is why the examiner wants both fates of TP: some TP is used to regenerate RuBP, and some TP is converted to useful organic substances. Miss the regeneration and you miss a mark, because without it the cycle would run out of RuBP and stop.

Limiting factors

The rate of photosynthesis is set by whichever factor is in shortest supply. A limiting factor is a factor that, when in short supply, holds back the rate even when everything else is favourable. The three you must know are light intensity, carbon dioxide concentration and temperature.

• Light intensity: needed to excite electrons in the light-dependent reaction. More light means more ATP and reduced NADP, so more GP can be reduced to TP.
• Carbon dioxide concentration: needed in the light-independent reaction to combine with RuBP. More carbon dioxide means more GP, so more TP.
• Temperature: affects the enzymes, especially rubisco. Below the optimum there is less kinetic energy and fewer enzyme-substrate collisions; well above it the enzymes are denatured, so the rate falls sharply.

The graph below shows why you must always name which factor is limiting. Read it carefully: at low light the two curves sit on top of each other, but at high light they separate.

• Where a curve is rising, the factor on the x-axis (light intensity) is limiting.
• Where a curve has plateaued, something else has become limiting. On the lower curve the plateau at about 44 units is set by the low carbon dioxide concentration, which is why raising the carbon dioxide (upper curve) lets the rate climb higher.

Overcoming limiting factors in agriculture

Growers manipulate these factors, most obviously in a glasshouse, to keep photosynthesis (and so yield) high.

• Adding carbon dioxide (for example from a burner) raises the rate when carbon dioxide is limiting.
• Heating keeps the temperature near the optimum for the enzymes.
• Extra lighting raises light intensity so it is not limiting.

You must be able to evaluate such practices, not just list them: the extra yield has to be weighed against the cost of the fuel, lighting and equipment, so a practice is only worthwhile if the increased value of the crop is greater than the cost.

Required practical 7: chromatography of leaf pigments

This practical separates the photosynthetic pigments in a leaf, showing that a leaf contains several pigments that between them absorb a range of wavelengths of light.

Method, in outline:

• Grind leaf tissue with a small volume of solvent to extract the pigments.
• Draw the origin line in pencil near the bottom of the chromatography paper or thin-layer plate, and use a capillary tube to build up a concentrated, dry spot of extract on it.
• Stand the paper in solvent with the solvent level below the origin line, and cover the container.
• Let the solvent rise, then mark the solvent front before it dries and let the chromatogram dry.

Each pigment separates because it has a different solubility in the solvent and a different attraction to the paper (the stationary phase): more soluble pigments travel further. Identify each pigment from its colour and its R_f value:

$R_f = \frac{\text{distance moved by the pigment}}{\text{distance moved by the solvent front}}$

Required practical 8: dehydrogenase activity in chloroplasts

This practical measures the rate of the light-dependent reaction using a redox indicator, DCPIP, as an artificial electron acceptor in place of NADP. DCPIP is blue when oxidised and colourless when reduced, so the faster it loses its colour, the faster electrons are being transferred.

• Extract chloroplasts into an ice-cold, isotonic (same water potential) buffer.
• Set up tubes of chloroplast suspension plus DCPIP and time how long the DCPIP takes to go from blue to colourless in the light.
• DCPIP is reduced by the electrons released from chlorophyll in the light-dependent reaction.

Controls make the result meaningful:

Worked examples

Model 5-mark answer: "Describe how ATP and reduced NADP are produced in the light-dependent reaction."

Each numbered point is a separate marking point, in a causal order:

1. Chlorophyll absorbs light energy, which excites electrons so they are emitted from the chlorophyll.
2. The electrons pass along the electron transfer chain, releasing energy.
3. This energy moves protons across the thylakoid membrane; they pass back through ATP synthase, joining ADP and P_i to make ATP.
4. Photolysis of water produces protons, electrons and oxygen (the electrons replace those lost by chlorophyll).
5. NADP is reduced by electrons (and protons) to form reduced NADP.

Model 5-mark answer: "Describe the light-independent reaction."

1. Carbon dioxide combines with RuBP, catalysed by rubisco.
2. This produces two molecules of GP.
3. GP is reduced to triose phosphate (TP),
4. using reduced NADP and energy from ATP.
5. Some TP is used to regenerate RuBP; some TP is converted to useful organic substances.

Notice the precision the marks depend on. "GP is converted to TP using ATP" earns nothing. You must say GP is reduced, name reduced NADP as providing the hydrogen, and say ATP provides the energy. "GP is reduced to TP using energy from ATP and reduced NADP" is the answer that scores.

Model answer: reading a limiting-factors graph.

Using the graph above: at low light intensity both curves overlap and rise together, so light intensity is the limiting factor there. The lower curve levels off at about 44 units while the upper curve keeps rising, which shows that above this point carbon dioxide has become the limiting factor for the lower curve (raising the carbon dioxide concentration removes the plateau).

Common exam mistakes

• Writing reduced NAD instead of reduced NADP. Photosynthesis uses NADP; respiration uses NAD. This is the single most common slip on this topic and it is always penalised.
• Saying "chlorophyll absorbs light" and stopping there, or writing that "chlorophyll gets excited". The marks need light energy absorbed and the electrons (not the chlorophyll) being excited and emitted.
• Giving photolysis products imprecisely. The products are protons, electrons and oxygen. Writing "hydrogen", "H" or "H₂" for the protons, "e" for electrons, or "O" for oxygen loses the mark, and students often forget oxygen altogether.
• Being vague about GP to TP. "GP is converted to TP using ATP" scores nothing; you must say GP is reduced, using reduced NADP and energy from ATP.
• Forgetting that carbon dioxide plus one RuBP makes two molecules of GP.
• Naming only ATP as a product of the light-dependent reaction and forgetting reduced NADP (or the other way round). The Calvin cycle needs both.
• Confusing the Calvin cycle with the Krebs cycle, or the photosynthesis electron transfer chain with the one in respiration. If your answer mentions the matrix, pyruvate or NAD, you have drifted into respiration.
• Putting rubisco in the wrong place. Rubisco and the whole Calvin cycle are in the stroma, not the matrix, grana or thylakoids.
• Reversing the sites: the light-dependent reaction is in the thylakoids/grana and the light-independent reaction is in the stroma.
• Writing P for phosphate. "P" is the symbol for phosphorus, so use P_i (or "phosphate") for inorganic phosphate.
• On a graph, saying the rate has plateaued without naming the new limiting factor, and explaining a control as making it "a fair test" rather than showing what the control actually tests.
• In the DCPIP practical, saying DCPIP is reduced by protons or by reduced NADP. It is reduced by the electrons from chlorophyll.

Key definitions

• Photoionisation (of chlorophyll): chlorophyll absorbs light energy, which excites electrons so that they are emitted from the chlorophyll, leaving it positively charged.
• Photolysis: the splitting of water using light energy, producing protons, electrons and oxygen.
• Photophosphorylation: the production of ATP from ADP and inorganic phosphate using energy from light, in the light-dependent reaction.
• Chemiosmosis (chemiosmotic theory): the movement of protons down their concentration gradient across a membrane, through ATP synthase, providing the energy to synthesise ATP.
• Limiting factor: a factor that, when in short supply, limits the rate of a process even when other factors are favourable.
• Rubisco: the enzyme that catalyses the reaction of carbon dioxide with ribulose bisphosphate (RuBP).

Specification

• I can state that chlorophyll absorbs light, leading to photoionisation of chlorophyll.
• I can explain how some of the energy from the electrons released in photoionisation is conserved as ATP and reduced NADP.
• I can explain how ATP is produced: electrons pass down the electron transfer chain, protons pass across the chloroplast (thylakoid) membranes, and ATP synthase catalyses ATP synthesis (chemiosmotic theory).
• I can state that photolysis of water produces protons, electrons and oxygen.
• I can describe how carbon dioxide reacts with RuBP to form two molecules of GP, catalysed by rubisco.
• I can explain how ATP and reduced NADP are used to reduce GP to triose phosphate.
• I can explain how some triose phosphate regenerates RuBP and some is converted to useful organic substances.
• I can identify the environmental factors that limit the rate of photosynthesis.
• I can evaluate data on agricultural practices used to overcome limiting factors.
• I can use chromatography to investigate the pigments isolated from leaves (Required practical 7).
• I can investigate the effect of a named factor on the rate of dehydrogenase activity in chloroplast extracts (Required practical 8).

Related notes

• ATP
• Respiration
• Energy and ecosystems