AP BIOLOGY · UNIT 3 · CELLULAR ENERGETICS

Light into sugar

Almost every meal you eat, and every breath of oxygen you take, starts with a leaf catching light. This is how a plant splits water, uses the energy to make ATP and NADPH, and builds sugar from carbon dioxide, what slows it down, and how to measure it with floating leaf discs.

AP Biology 3.43 lessons13 chaptersdry lab

Start learning Run the dry lab Teach it

Zoom into a leaf and watch light split water, oxygen leave and carbon dioxide become sugar.

By the end you can tap a goal to go to its chapter

Explain how photosynthesis captures energy from sunlight and stores it in sugars, and why almost all ecosystems depend on it 3.4.A chapter 1 →
Write the equation, say where each atom of glucose comes from, and explain why the oxygen given off comes from water 3.4.A chapter 1 →
Read absorption and action spectra and explain why chlorophylls absorb some colors more than others 3.4.A chapter 1 →
Predict and test how light, carbon dioxide and temperature limit the rate, using leaf discs or pondweed 3.4.A chapter 1 →
Explain how chlorophyll boosts electrons in photosystems II and I, and how they are passed along an electron transport chain 3.4.B chapter 7 →
Explain how the proton gradient across the thylakoid membrane drives ATP synthase, and where the NADPH comes from 3.4.B chapter 7 →
Describe how ATP and NADPH power the Calvin cycle in the stroma, and why Rubisco and carbon dioxide matter 3.4.B chapter 7 →
Describe how photosynthesis in prokaryotes changed the atmosphere and led to the photosynthesis in eukaryotes 3.4.B chapter 7 →

01Learn

Thirteen chapters. Read one, play its figure, and make each prediction before you open the answer. The palisade film at the top zooms into a leaf; try the chromatography bench and the Rubisco race on the way.

1Sunlight in, chemical energy out
2Carbon from the air, hydrogen from water
3More than one green
4Why some colors and not others
5Absorbed is not the same as used
6Whatever runs short sets the pace
7Pigments in an array
8Water split, oxygen spare
9Protons downhill make ATP
10NADP+ reduced, and the thylakoid as a system
11Fixing carbon, slowly
12GP to TP, and RuBP made again
13Each half needs the other

3.4.A Light energy to chemical energy

Sunlight in, chemical energy out

Photosynthesis transforms light energy into chemical energy. In a leaf, pigments absorb light, and that energy ends up in the bonds of carbon compounds the plant builds: first sugars, then everything else it is made of. Light is not stored. It is changed into a different form, an energy transformation.

That chemical energy runs nearly every food chain. Plants, algae and cyanobacteria are producers. A rabbit eating grass, and a fox eating the rabbit, are living on energy the grass caught from the Sun. A few ecosystems, round deep-sea vents, run on chemical energy from the vent instead, but almost all the others depend on photosynthesis.

Cells need that energy for three kinds of work: building large molecules (anabolism), moving substances against their gradients (active transport, unit 2), and movement, from muscle contraction to the pulling apart of chromosomes. Respiration (topic 3.5) releases the energy from carbon compounds and holds it in ATP, which pays for the work. At every step some energy leaves as heat. Energy flows through an ecosystem and is not recycled; carbon is (unit 8).

Photosynthesis transforms light energy into the chemical energy of carbon compounds.
Producers make the carbon compounds that every consumer in a food chain depends on.
Energy flows light, then chemical energy in sugar, then ATP, then work and heat.

Watch out Photosynthesis does not make energy; it changes light energy into chemical energy. And plants respire too, in the dark and in the light: they use some of the sugar they make.

The chemical energy in a slice of cheese: where did it first come from?

Light energy from the Sun reaches a leaf.
Photosynthesis turns it into chemical energy in the bonds of sugar.
The cow lives on that energy, and stores some of it in milk.
You release it in respiration, as ATP, and every step loses some as heat.

3.4.A Carbon dioxide to glucose, with oxygen as a by-product

Carbon from the air, hydrogen from water

The word equation for photosynthesis is carbon dioxide + water → glucose + oxygen, with light energy and chlorophyll written over the arrow. In symbols it reads 6CO₂ + 6H₂O → C₆H₁₂O₆ + 6O₂, and the product that matters is glucose.

Look at the molecules. Carbon dioxide has carbon and oxygen but no hydrogen, and glucose needs plenty. The plant gets its hydrogen from water. It splits water using energy from the light that chlorophyll absorbs, a step called photolysis, and the hydrogen is added to carbon dioxide to build glucose. So glucose’s carbon and oxygen come from carbon dioxide, and its hydrogen comes from water. Most of a tree’s dry mass is carbon and oxygen taken from the air, not from the soil.

The oxygen gas a leaf gives off is a by-product. It comes from the water that was split, not from carbon dioxide. Every organism that photosynthesizes this way gives off oxygen: plants, algae (from single cells to seaweeds) and cyanobacteria. Cyanobacteria were the first of them, as chapter 8 shows.

Adding hydrogen to a molecule is a reduction (topic 3.5). Chapters 7 to 12 follow the hydrogen from water to carbon dioxide, as electrons and protons carried by NADP⁺.

How we know In 1941 Ruben and colleagues gave algae water made with a heavy form of oxygen. The oxygen gas the algae gave off was heavy too. When the heavy oxygen was in the carbon dioxide instead, the gas was not. The oxygen had come from the water.

Carbon dioxide + water → glucose + oxygen, with light energy and chlorophyll over the arrow.
Glucose’s carbon and oxygen come from carbon dioxide; its hydrogen comes from water.
The oxygen given off comes from the water that is split, and is a by-product.

Watch out “The oxygen comes from carbon dioxide” is the commonest slip, because the equation seems to say so. Carbon dioxide’s oxygen ends up in the glucose and in the water that is made again. And light does not split water by itself: chlorophyll catches the light first (chapter 8).

Where does the oxygen gas a leaf gives off come from?

What goes in: carbon dioxide from the air, water from the soil.
Light energy splits water. The oxygen leaves as a by-product.
The hydrogen is added to carbon dioxide, building glucose.
carbon dioxide + water → glucose + oxygen. The bars show where each atom came from: green, carbon dioxide; pink, water.

Pick a card (tap it, or Tab to it and press Enter), then pick the bin it belongs in.

0 of 6 sorted

Six atoms or molecules and two bins. Which molecule going in had the atom at all, and what tells you?

Zoom into a leaf and watch light split water, oxygen leave and carbon dioxide become sugar.

3.4.A Separating the leaf pigments

More than one green

A leaf’s green is not one pigment but several, mixed. Chlorophyll a is the main one, blue-green. Chlorophyll b is yellow-green. Beside them are the yellow and orange carotenoids: carotenes and xanthophylls. All of them sit in the membranes of the chloroplast’s thylakoids.

Chromatography separates a mixture. A small, dark spot of leaf extract goes on a pencil line, the origin, near the bottom of a strip of chromatography paper (or a thin-layer plate, the stationary phase). The strip stands in a shallow layer of solvent (the mobile phase) below the spot, in a closed tube. The solvent climbs and carries the pigments with it.

Each pigment travels at its own speed. The more soluble it is in the solvent, and the less it clings to the paper, the further it goes. So the pigments end up as separate bands. The solvent front marks how far the solvent went, and the Rf value compares the two:

R_f = distance moved by the pigment ÷ distance moved by the solvent front, both from the origin, the pigment to the middle of its band.

R_f has no units and is always less than 1. You identify a pigment by its color and its R_f, read against a table made with the same solvent and the same paper or plate. Thin-layer plates run faster and separate better than paper.

Pencil, not ink, for the origin: ink is a mixture and runs too.
A small, dark spot, built up by drying between drops.
Solvent below the origin, and a lid on the tube.
Take the strip out before the front reaches the top, and mark the front at once.
Work quickly: pigments fade in the light.

Watch out Measure from the origin line, not from the bottom of the paper, and to the middle of the band. And an R_f only means something beside a table made with the same solvent.

Why it matters Chromatography separates and identifies the substances in a biological mixture. The same idea separates amino acids and sugars.

The solvent front has moved 8.0 cm from the origin and a yellow band 5.7 cm. What is the band’s R_f?

Time0 to 10 min

Slide the time. The solvent climbs and the green spot pulls apart into numbered bands. At the end, the distances are measured from the origin line.

Grind a leaf with a little sand and a few drops of acetone. Spot the extract on a pencil line 2 cm from the bottom of the strip, then stand the strip in acetone, below the line, with the lid on.
Take the strip out before the solvent reaches the top and mark the front in pencil at once. The pigments have separated into bands. The table beside it lists known values for this solvent.
Measure from the origin line to the middle of each band and to the front. Work out R_f for each band, then name each pigment from its color and its R_f.

3.4.A Absorption of light by pigments

Why some colors and not others

Visible light is the band of wavelengths from about 400 nm (violet) to about 700 nm (red). White light is every one of them together, and the shorter the wavelength, the more energy each photon carries.

A pigment absorbs a photon only if the photon’s energy matches a jump one of its electrons can make to a higher energy level. The electron is then excited. Photons with other energies are not absorbed: they pass through the pigment or bounce off it, which is reflection.

Chlorophyll’s electrons can make jumps that match blue and red photons, so it absorbs those strongly and very little green. The green is reflected or passes through. That is why leaves look green: you see the light they did not use. The energy of the excited electron is what photosynthesis turns into chemical energy.

Accessory pigments, the carotenoids, absorb blue and blue-green light that chlorophyll a takes up less, and widen the range of light a leaf can use. In autumn the chlorophyll breaks down first, and the carotenoids show as yellow and orange. An absorption spectrum shows, wavelength by wavelength, how much of the light a pigment takes in. Draw it with both the wavelengths in nm and the colors along the horizontal axis.

The pigments are held together in arrays called photosystems (chapter 7).

Visible light runs from about 400 nm (violet) to about 700 nm (red).
A photon is absorbed only if its energy matches a jump an electron in the pigment can make.
Chlorophyll absorbs blue and red strongly and reflects green, so a leaf looks green.
The horizontal axis of an absorption spectrum carries wavelengths and colors.

Remember it The colors, longest wavelength first

RichardRedabout 700 nm, absorbed
OfOrangeabout 610 nm
YorkYellowabout 580 nm
GaveGreenabout 530 nm, mostly reflected
BattleBlueabout 450 nm, absorbed
InIndigoabout 440 nm
VainVioletabout 400 nm

Watch out A leaf is not green because chlorophyll makes green light. Green is the light it did not absorb.

Why does chlorophyll absorb red and blue light but very little green?

White light is every color together.
Chlorophyll absorbs blue and red and reflects most green.
An absorption spectrum: how much of each wavelength a pigment absorbs. These are for pigments dissolved in a solvent.
Accessory pigments absorb where chlorophyll a absorbs least.

Drag each handle up to how much you think chlorophyll absorbs at that color (or focus one and use the up and down keys). Then press Show me.

Sketch chlorophyll’s absorption spectrum: seven handles, one for each color band. Chlorophyll a is the pigment to draw.

3.4.A Absorption and action spectra

Absorbed is not the same as used

An action spectrum plots how fast photosynthesis runs against the wavelength of the light a plant is given. You measure it by lighting the plant with one color at a time, the same amount of light each time, and measuring the oxygen it gives off or the carbon dioxide it takes in. The rate of photosynthesis is the amount of oxygen produced (or carbon dioxide consumed) divided by the time taken, for example cm³ of oxygen ÷ 10 minutes, in cm³ min⁻¹. Then plot rate against wavelength.

The two spectra are alike because the light a leaf uses is the light its pigments absorb: both peak in the blue and the red and dip in the green. They are not the same. The action spectrum’s green dip is shallower, because chlorophyll b and the carotenoids absorb some of those wavelengths and pass the energy on, and because in a thick leaf green light bounces about and more of it is absorbed in the end. And absorbing light is not always using it: some of the energy that carotenoids absorb in the blue is lost as heat, so the action spectrum’s blue peak is lower than the combined absorption there.

How we know In 1882 Theodor Engelmann lit a strand of a green alga with a spectrum from a prism and added bacteria that swim towards oxygen. They crowded where red and blue light fell. It was the first action spectrum, measured with living bacteria as oxygen detectors. The film below repeats it.

Rate of photosynthesis = oxygen produced (or carbon dioxide consumed) ÷ time.
Similar: both spectra peak in blue and red and dip in green.
Different: the action spectrum is a rate, its green dip is shallower, and its blue peak is lower than the absorption there.

Watch out An action spectrum is not “the light a plant absorbs”. It is what the plant does with each color: its rate of photosynthesis.

Engelmann shone a spectrum along a strand of alga and added bacteria that swim towards oxygen. Where did the bacteria gather?

Oxygen collected from a sprig of pondweed in 10 minutes, at equal light of each wavelength
Wavelength / nm	Color	Oxygen in 10 min / cm³
425	violet	0.30
450	blue	0.30
500	blue-green	0.23
550	green	0.13
600	orange	0.24
650	red	0.35
675	deep red	0.35

Data from the dry lab’s model: a sprig in 5 % bicarbonate at 25 °C, the same number of photons at every wavelength.

Work out each rate: volume ÷ time. Then set each handle on the graph to its rate (drag it, or focus it and use the up and down keys).

Seven wavelengths of light, the oxygen a sprig gave off in 10 minutes in each, and empty axes. Rate = volume of oxygen ÷ time.

A strand of alga under a microscope, bacteria that seek oxygen, and a prism. Split the light and watch where the bacteria go.

3.4.A Light, carbon dioxide and temperature as limiting factors

Whatever runs short sets the pace

Photosynthesis needs light, carbon dioxide and a suitable temperature all at once. When several factors affect a rate, the one in shortest supply holds it back. Raising that limiting factor speeds the process up, and raising the others does nothing, until one of them becomes the shortest. Blackman stated this as the law of limiting factors in 1905.

Light intensity limits the supply of energy for splitting water. The rate rises with light, then levels off when something else limits. Carbon dioxide concentration limits the fixing of carbon in the same way. Temperature acts on the enzymes (you saw why in topic 3.2): the rate rises to an optimum, about 25 to 30 °C for many temperate plants, then falls as the enzymes denature. In dim light, temperature matters little, because light is the limit. Where a curve levels off, another factor sets the height of the plateau: raise that one and the plateau rises.

To test each factor, vary it on its own. Move a lamp to change the light (the intensity falls roughly as 1 ÷ distance², so use a light meter, with a heat shield between lamp and water). Dissolve sodium bicarbonate in the water round pondweed or leaf discs to change the carbon dioxide (0 to 5 %). Use a water bath for temperature. Measure the rate by counting oxygen bubbles or collecting the gas, by the time leaf discs take to float, by the carbon dioxide taken up (bicarbonate indicator), or by the gain in dry mass, which is slow and indirect.

A hypothesis is a provisional explanation that must be tested again and again. You can write it from theory before the experiment (“if carbon dioxide is the limiting factor, the rate will rise and then level off”), or from the evidence of a first run. Either way you name the independent variable (what you change), the dependent variable (what you measure) and the controlled variables (everything else, kept the same). Then test it on the bench, in the dry lab.

Limiting factor: the factor in shortest supply, which sets the rate.
Light, carbon dioxide and temperature are the three to know; raise the limiting one and the rate rises, until another takes over.
A hypothesis names the independent, dependent and controlled variables.

Remember it The three limiting factors

LazyLight intensitythe supply of energy for splitting water
CatsCarbon dioxide concentrationthe carbon to fix
ToastTemperaturethe speed of the enzymes, up to their optimum

Watch out A limiting factor is not “a factor that stops photosynthesis”. It is the one in shortest supply, setting the rate. Raise it and the rate rises, until another one takes over.

A plant at 15 °C is given more and more light until its rate levels off. Then it is warmed to 25 °C. What happens to the rate?

Stage 1 of 3 Rate against light intensity. Draw how you think the rate changes as the light gets brighter.

Draw with a finger, a pen or the mouse (one stroke), then press Show me.

Sketch a hypothesis as a graph, then check it. Three stages: light, then temperature, then carbon dioxide at high and low light.

Sketch it: three tasks

The answers

3.4.B Photosystems and the light-dependent reactions

Pigments in an array

A photosystem is an array of a few hundred pigment molecules, chlorophylls and accessory pigments, held in fixed positions by proteins. Photosystems are always in a membrane: the thylakoid membranes of a chloroplast, or the internal membranes of a cyanobacterium. At the heart of each is the reaction center, a special chlorophyll a.

Light absorbed by any pigment in the array excites an electron in that pigment. The energy, not the electron, passes from pigment to pigment, inwards, until it reaches the reaction center. There an excited electron is emitted: it leaves the chlorophyll altogether, passing to an electron acceptor beside it in the membrane. There are two kinds of photosystem, photosystem II and photosystem I, numbered in the order they were discovered. Electrons pass through II first.

Why an array? A single chlorophyll molecule, even in sunlight, absorbs a photon only every so often, and its excitation lasts only a few billionths of a second. On its own, the energy is lost as heat or given out again as red light, and it cannot do any part of photosynthesis. In an array, hundreds of pigments collect light and funnel it to one reaction center, which keeps it busy. Different pigments widen the range of wavelengths caught. And their fixed spacing lets energy jump from one to the next before it is lost.

Seeing it Chlorophyll extracted into propanone glows deep red under a bright light. Chlorophyll in a leaf hardly glows. In the leaf the energy is used. In the tube there is no array and no acceptor, so it comes straight back out as light.

A photosystem is an array of pigments round a reaction center, always in a membrane.
Pigments pass on energy; only the reaction center emits an excited electron.
One pigment alone could not do any part of photosynthesis.

Watch out Pigments in the array pass on energy, not electrons. Only the reaction center emits an electron.

Chlorophyll dissolved in propanone glows red under a bright light. Chlorophyll in a leaf hardly glows at all. Why?

On its own, chlorophyll gives the energy straight back, as red light.
A photosystem: hundreds of pigments around a reaction center, in a membrane.
The energy hops from pigment to pigment, inwards.
The reaction center emits an excited electron.
Many pigments, many colors, all feeding one reaction center.

3.4.B Splitting water

Water split, oxygen spare

Photosystem II’s reaction center has emitted its electrons, so it needs more. It takes them from water. The splitting of water, photolysis, is done by an enzyme on the lumen side of photosystem II. Two water molecules give four electrons, four protons and one oxygen molecule.

The electrons refill the reaction center, so it can be excited again. The protons are released into the lumen, where they add to the proton gradient (chapter 9). Both are used. The oxygen is a waste product: it diffuses out of the chloroplast, the cell and the leaf. Light does not split water directly. It excites the reaction center, and the electron-short reaction center pulls electrons from water.

The advent of oxygen from photolysis had immense consequences on Earth. Cyanobacteria were the first organisms to split water. The oxygen they released first reacted with iron dissolved in the oceans, which settled as layers of iron oxide on the sea floor: the banded iron formations. When the iron was used up, oxygen built up in the air, in the Great Oxidation Event, about 2.4 billion years ago. Oxygen poisoned many anaerobic organisms. It formed the ozone layer, which screens ultraviolet light. And it made aerobic respiration possible, with its far higher yield of ATP (topic 3.5), on which large, active organisms depend.

Photolysis at photosystem II: 2 water give 4 electrons, 4 protons and 1 oxygen.
The electrons and protons are used; the oxygen is waste.
Light excites the reaction center; the reaction center takes electrons from water.

Remember it What photolysis gives

PleaseProtonsinto the lumen, for the gradient
EatElectronsback into photosystem II’s reaction center
OrangesOxygenthrown away as waste

Watch out The protons and electrons from water are used; only the oxygen is waste. And it is photosystem II, not photosystem I, that splits water.

Cyanobacteria were splitting water for a long time before oxygen built up in the air. Where did their oxygen go at first?

Photosystem II has lost electrons.
It takes more from water.
Photolysis: electrons refill the reaction center; protons stay in the lumen.
The oxygen is a waste product.

The shape of the rise is known better than the numbers, so only 0 and today’s level are marked. Banded iron formations are mostly older than about 1.8 billion years.

3.4.B Electron transport and the proton gradient

Protons downhill make ATP

The excited electron from photosystem II passes along a chain of electron carriers in the thylakoid membrane towards photosystem I, giving up energy at each step. One carrier uses that energy for proton pumping: it moves protons across the membrane from the stroma into the thylakoid lumen.

Protons come into the lumen two ways: pumped by the chain, and released by photolysis. The lumen is small, so their concentration rises fast, and the membrane does not let them back. That difference is the proton gradient. The only way back to the stroma is through ATP synthase, a protein that spans the membrane. Protons flowing down their gradient turn part of it, and the energy joins ADP and phosphate: ATP, made on the stroma side. Making ATP with energy from a proton gradient is chemiosmosis, and when light is the first source the whole process is photophosphorylation.

You met the same machine in mitochondria (topic 3.5). The difference is direction: a thylakoid pumps protons into its lumen and makes ATP outside it, in the stroma, while a mitochondrion pumps them out of the matrix and makes ATP inside it.

In non-cyclic photophosphorylation the electrons come from photosystem II, pass along the chain to photosystem I and on to NADP⁺ (chapter 10). In cyclic photophosphorylation, electrons excited at photosystem I go back to the chain, pump more protons and return to photosystem I. It makes ATP only: no NADPH, no water split, no oxygen. It tops up the ATP, because the Calvin cycle uses more ATP than NADPH.

Electrons along the chain pump protons from the stroma into the lumen; photolysis adds more.
Protons flow back through ATP synthase, which makes ATP on the stroma side.
Non-cyclic: electrons from photosystem II to NADP⁺. Cyclic: electrons from photosystem I round the chain again, for ATP only.

Watch out Protons do not collect in an “intermembrane space”, as they do in a mitochondrion. In a chloroplast they collect in the thylakoid lumen.

In a working chloroplast, where do protons build up?

The thylakoid membrane: photosystems, a chain of carriers and ATP synthase.
Electrons give up energy along the chain, which pumps protons into the lumen.
Protons pile up in the small lumen: a proton gradient.
They flow back through ATP synthase, which makes ATP in the stroma: chemiosmosis.
Non-cyclic: on to NADP⁺. Cyclic: back round the chain, for more ATP.

The lumen is crowded with protons; the stroma has few. That gradient is stored energy, and ATP synthase is the only way down.
A proton enters the channel from the lumen side and takes a place on the rotor.
Each proton that passes pushes the rotor one step round, like water on a turbine, and leaves into the stroma.
The rotor turns the shaft inside the head. Each turn squeezes ADP and phosphate together: ADP is phosphorylated.
ATP leaves into the stroma. NADP⁺ also takes up stroma protons when it is reduced, which keeps the gradient steep.

ATP synthase as a turbine: protons flowing down their gradient turn the rotor, and the rotor drives the phosphorylation of ADP.

3.4.B NADPH, the carrier that reaches the Calvin cycle

NADP⁺ reduced, and the thylakoid as a system

Electrons arriving at photosystem I have given up energy along the chain. Light absorbed by photosystem I’s array excites them again, and from its reaction center they pass, by a short carrier on the stroma side, to NADP⁺. NADP⁺ picks up two electrons from photosystem I and one hydrogen ion (a proton) from the stroma, and becomes NADPH. This is a reduction, and taking that proton from the stroma steepens the gradient a little.

NADPH carries the hydrogen, as electrons and a proton, to the Calvin cycle in the stroma, where it reduces carbon compounds (chapter 12). This is how the hydrogen from chapter 2 gets from water to carbon dioxide: water’s electrons go through photosystem II, the chain and photosystem I, to NADP⁺.

A thylakoid is a system in which every part has its place. On the lumen side: photolysis at photosystem II, and the store of protons. In the membrane: the photosystems, the chain and ATP synthase, held in order so electrons pass from one to the next. On the stroma side: ATP is made and NADP⁺ is reduced, right where the Calvin cycle’s enzymes are. Thylakoids are stacked in grana, joined by single unstacked sheets of membrane that run through the stroma; the stacking gives a large area of membrane, and the small lumen builds a steep gradient fast. Nothing has to “move to the stroma”: ATP and NADPH are made in it.

NADP⁺ is reduced at photosystem I, by two electrons from it and a proton from the stroma.
Photolysis and the proton store are on the lumen side; ATP and NADPH are made on the stroma side.
Grana give a large area of membrane; the small lumen gives a steep gradient fast.

Remember it Two sides of the thylakoid

LoadsLumenwhere protons pile up and water is split
SpendsStromawhere ATP and NADPH are made, then spent by the Calvin cycle

Watch out NADP⁺ is reduced at photosystem I, not photosystem II; photosystem II splits water. Electrons go through II first: the numbers record the order of discovery.

On which side of the thylakoid membrane is NADP⁺ reduced, and why does that side make sense?

Tap a part of the chloroplast or the membrane, or Tab to it and press Enter.

Hide the labels, to test yourself

A chloroplast cut open, and one thylakoid membrane enlarged. Which side of the membrane does each part work on?

Light in, water split, protons pumped and ATP made. Switch the light off, or send the electrons round the cyclic path, and watch what stops.

3.4.B Fixing carbon

Fixing carbon, slowly

The light-independent reactions are a cycle in the stroma, the Calvin cycle, named after Melvin Calvin, who worked it out with radioactive carbon in algae. Its first step is carbon fixation. Carbon dioxide joins a five-carbon sugar already in the stroma, ribulose bisphosphate (RuBP). The six carbons split at once into two molecules of the three-carbon glycerate 3-phosphate (GP). The enzyme is Rubisco. So the substrates are RuBP and carbon dioxide, and the product is GP.

“Fixation” means turning carbon from a gas into part of an organic molecule: the carbon is now held. Adding carbon dioxide to a molecule is also called carboxylation, so fixation by Rubisco is the carboxylation of RuBP.

Rubisco is slow. Each active site fixes only about three carbon dioxide molecules a second, where catalase (topic 3.1) breaks down millions of hydrogen peroxide molecules a second. And it is not effective at low carbon dioxide concentrations: when carbon dioxide is scarce (on a hot day, with the stomata closed) it works far below its best. So the stroma needs a high concentration of Rubisco to fix carbon dioxide as fast as the light-dependent reactions supply ATP and NADPH. Rubisco is up to about half of the soluble protein in a leaf, and it is the most abundant enzyme on Earth.

Carbon fixation: carbon dioxide joins RuBP (5 carbons), which splits into two GP (3 carbons each).
The enzyme is Rubisco: slow, and poor at low carbon dioxide concentrations.
The stroma needs a high concentration of Rubisco for that reason.

Watch out Rubisco is an enzyme, not a product of the cycle, and it is not used up. It is abundant because it is slow, not because it is special in any other way.

A leaf cell could save a lot of protein by making less Rubisco. Why doesn’t it?

RuBP, a five-carbon sugar, waits in Rubisco’s active site.
Carbon dioxide joins it: carbon fixation.
The six carbons split into two GP.
Rubisco is slow, so a leaf makes a lot of it.

One enzyme cannot keep up.
Eleven keep pace with bright light.
With little carbon dioxide, each does less and fumbles more.
More enzyme helps until light is the limit.

3.4.B The Calvin cycle

GP to TP, and RuBP made again

NADPH and ATP from the light-dependent reactions turn GP into triose phosphate (TP), a three-carbon sugar, in the stroma. NADPH gives the hydrogen (it reduces GP) and ATP gives the energy. NADP⁺ and ADP go back to the thylakoid. TP is the first sugar and the cycle’s product: some of it leaves to make glucose and everything else.

The course asks for what the cycle does: it takes carbon from carbon dioxide into sugar, spending ATP and NADPH. You do not need to memorize the steps, the molecules or the names of the enzymes (ATP synthase is the one to know).

The rest has to remake RuBP, using more ATP. Five TP (fifteen carbons) are rearranged into three RuBP (fifteen carbons), so the cycle can fix carbon dioxide again. This is regeneration. You do not need the steps in between. Counting a round: three RuBP fix three carbon dioxide to give six GP; six GP give six TP; one TP leaves; five remake three RuBP. Three carbons in, three carbons out as one TP.

To make one glucose (six carbons, two TP), six carbon dioxide must be fixed, and five-sixths of all the TP made goes back to RuBP. Without regenerating RuBP, fixation would stop within moments. One turn of the cycle fixes one carbon dioxide, so six turns make one glucose. The figure runs three turns at a time (a “round”) only because three is the smallest number that gives a whole TP out.

Why a cycle? The molecule that accepts carbon dioxide is remade every turn, so a small stock of RuBP keeps fixing carbon as long as ATP and NADPH keep coming. The tally in the corner counts ATP and NADPH as they are used; it is there to show the cost, not to be learned.

GP is reduced to TP, using NADPH and ATP.
Five TP of every six made are remade into three RuBP, using ATP; one in six leaves.
Six turns of the cycle fix six carbon dioxide and make one glucose.

Remember it The Calvin cycle’s three stages

FreshFixRubisco joins carbon dioxide to RuBP, making GP
RollsReduceATP and NADPH turn GP into TP
RiseRegeneratefive TP in six remade into RuBP, using ATP

Watch out It is GP (glycerate 3-phosphate) that is reduced, to TP. “Glyceraldehyde 3-phosphate” is another name for a triose phosphate, so “the reduction of glyceraldehyde 3-phosphate” has the stages muddled.

Of every six TP the Calvin cycle makes, how many must be used to remake RuBP?

Three RuBP wait for three carbon dioxide.
Rubisco fixes them: six GP.
NADPH and ATP turn GP into TP.
One TP in six leaves the cycle.
Five TP are remade into three RuBP, using ATP.
Six turns, six carbon dioxide: one glucose.

3.4.B The two halves depend on each other

Each half needs the other

The light-dependent reactions in the thylakoids make ATP and NADPH. The light-independent reactions, the Calvin cycle in the stroma, use them and hand back ADP, phosphate and NADP⁺. Each half needs what the other makes: that is their interdependence.

No light. The photosystems are not excited, so no ATP or NADPH is made. The Calvin cycle runs on for a few seconds on what is left, then stops.

No carbon dioxide. The Calvin cycle cannot fix carbon, so it stops using ATP and NADPH, and no NADP⁺ comes back. Once all the stroma’s NADP⁺ is reduced, photosystem I has nothing to pass its electrons to. The chain backs up, and photosystem II cannot pass its electrons on, so it stops, and so does photolysis.

So a leaf’s two halves run at the pace of the slower one, which links back to limiting factors (chapter 6). In bright light, carbon dioxide or Rubisco limits the rate; in dim light, the light-dependent reactions do. And a plant in the dark still respires. Everything in Unit 4 so far is on one map below, the metabolism map, where you can see both.

Light-dependent reactions make ATP and NADPH; the Calvin cycle uses them and returns ADP, phosphate and NADP⁺.
No light: the Calvin cycle stops a few seconds later.
No carbon dioxide: NADP⁺ is not returned, so photosystem I fills, then the chain, then photosystem II stops.

Watch out “Light-independent” does not mean “happens in the dark”. The Calvin cycle needs the products of the light reactions, so in a leaf it runs only in the light.

A leaf in bright light is suddenly given no carbon dioxide. What happens to photosystem II?

Choose a state, once you have made your prediction.

The thylakoid makes ATP and NADPH for the Calvin cycle, and the Calvin cycle hands back ADP, phosphate and NADP⁺. Switch off the light, or the carbon dioxide, and watch what stops.

The light-dependent reactions and the Calvin cycle at work. Then switch off the light, or the carbon dioxide, and see what stops.

Glycolysis, the Krebs cycle and the Calvin cycle in one leaf cell. Follow a molecule to see where it is made and where it is used.

02Dry lab: Leaf discs and pondweed

Leaf discs sink in bicarbonate solution, then float as the oxygen they make collects inside them. Move the lamp, change the carbon dioxide or the temperature, and time how long half the discs take to float. Or switch to a sprig of pondweed and count bubbles.

Your questionHow does one limiting factor change the rate of photosynthesis? Start with the lamp: how does the rate change as the lamp moves away, and is it the distance or the light that matters?

What you can changeOne independent variable at a time: the distance of the lamp, the concentration of sodium bicarbonate, the temperature of the water, or the color of a filter. Everything else stays fixed unless you change it on purpose, and some of it changes when you are not looking.

What to recordThe number of discs floating each minute, then the time for half of them to float; or bubbles counted in each of three minutes. Three trials at each value, a mean and a standard deviation, and a graph of the means with error bars.

Leaf disks. Spinach disks sink in sodium bicarbonate solution; count how many float each minute for 20 minutes, and find the time for half of them to float.

Find out

How does the rate change with distance? Move the lamp from 5 to 100 cm: at least six distances, three trials at each, after the sprig has settled. Graph the rate against distance, then against 1 ÷ distance². Where is the graph a straight line, and where not?
Light is not the only thing changing. Read the thermometer before and after a run with the lamp 5 cm away, with and without the heat shield.
Carbon dioxide. Run bicarbonate from 0 to 5 % with the lamp at 10 cm, then again at 40 cm. Where does each level off?
Temperature. Run the bath from 5 to 45 °C, lamp at 10 cm, 1 % bicarbonate. Then cool a sprig you ran at 45 °C back to 25 °C and run it again.
Color. Run white light and the four filters with the lamp at 10 cm, reading the light meter each time. Then improve your method so the comparison is fair. Comparing colors, the lamp may sit at a different distance for each filter: the table writes it down with each trial.
Leaf discs. Switch the method to leaf discs and run bicarbonate at 0, 0.1, 0.2, 0.5 and 1 % with the lamp at 10 cm. Find the time for half the discs to float.

Record

Every run's settings: what you changed, and everything you kept the same.
Change one thing at a time, and let the sprig settle after each change before you count.
The bubbles in each of three minutes, or the gas bubble's two ends on the scale, or the discs floating each minute.
Three or more trials at each value, then a mean and its standard deviation.
Use the graph under the lab, or download the CSV and plot it yourself.
When you are done, choose one investigation and write it up: the question, the method, a table, a graph with error bars, and a conclusion that cites your data. Improve your method, under the graph, opens better apparatus as you think of it.

With the lamp 80 cm away you count no bubbles at all. Has the sprig stopped photosynthesizing?

Without the heat shield, your rates with the lamp close to the tube sit above the trend of the others. Why?

The rate plotted against 1 ÷ distance² is a straight line for distant lamps but bends over close up. Why?

Under the green filter the light meter reads more lux than under the red one, yet the sprig gives fewer bubbles. Why?

03Check

A fresh paper from the exam bank on this guide's topics each time: past-paper style questions, marked as you go, and saved to your Exam practice progress when you are signed in. It opens on biologybybradford.com, so it needs the internet.

Exam practice

Exam questions on 3.4, marked

Start 15 marks of exam questions Your exam practice

04Downloads

This file is the guide, saved to run without the internet.

Saved from biologybybradford.com/guides/ap-biology-unit-3-photosynthesis, where the guide is kept up to date with the notes, the revision guide and, for the teacher, the lesson plans and slides.

05Teacher resources

The guide as three one-hour lessons. Each has a plan to print (the timings, what to project and where to stop, the activities with what they need, what students get wrong) and slides to teach it from, in the same order, with the teacher's moves in the notes.

Lesson	Covers	Focus	Chapters
1Capturing light	3.4.A60 min	The equation, the pigments and their spectra, and the first limiting factor with floating leaf discs	1, 2, 3, 4, 5, dry lab
2The light reactions	3.4.B60 min	The light reactions: photosystems, the chain, the proton gradient and NADPH, built with a class model	7, 8, 9, 10, 13
3Building sugar	3.4.B60 min	The Calvin cycle, how the two halves depend on each other, and a timed free response with data	6, 8, 9, 11, 12, 13

The note template, the revision guide and the offline copy are in Downloads, for you and the class.

PRACTISE AND REVISE

Make it for your class, or set it

Build for your class

Tarsia puzzle, dominoes and picture reveal on Unit 3, built from its key terms and ready to print
A pub quiz with rounds on Unit 3, to present or run with no projector
The Kelp Light Habitat escape room, or an escape room built from the question bank on Unit 3

Already on the site for Unit 3

Lesson: Photosynthesis
Quiz: Photosynthesis
Exam questions: Unit 3 in the exam maker, a paper with scoring guidelines

Card sets on splycr

Open a set to host it as a class game, or make a copy to edit.

How this was made. The course structure, the activities and the teaching choices come from my own classroom, my presentations and my years of teaching IB and AP Biology. I use Claude, an AI, to help write and build the interactive guides, lesson plans, slides and practice questions, and to draw the diagrams and code the interactives. I direct the work and review it against the IB and AP specifications before it goes live.