Photosynthesis: WJEC A level Biology revision notes

Lesson 1 of 17

What is photosynthesis and why are chloroplasts called transducers?

These notes cover topic 3.2, uses light energy to synthesise organic molecules, in WJEC A level Biology Unit 3: Energy, and the Environment. Each lesson follows the WJEC specification, and the exam tips are based on BioWelsh analysis of WJEC past papers.

Almost every living thing depends on . Plants, algae and some bacteria use light energy to build large organic molecules, such as glucose, from two small inorganic molecules: carbon dioxide and water. Animals, fungi and most bacteria then get their energy and carbon by eating those organic molecules, directly or indirectly.

The overall equation looks simple: 6CO₂ + 6H₂O → C₆H₁₂O₆ + 6O₂. Carbon dioxide and water go in, glucose and oxygen come out, and light supplies the energy. In reality there are dozens of reactions, each catalysed by its own , and the equation only shows the start and the end.

A is anything that changes energy from one form into another. A loudspeaker changes electrical energy into sound, and a solar panel changes light energy into electrical energy. A is a biological : it changes the energy of light into chemical energy, first in and and then in organic molecules such as glucose.

Examples of transducers
Transducer	Energy in	Energy out
Loudspeaker	Electrical	Sound
Solar panel	Light	Electrical
Muscle	Chemical ()	Kinetic and heat
	Light	Chemical (, , then glucose)

happens in two stages. The takes place on the membranes inside the . It uses light energy to split water and to make and , and it releases oxygen as a by-product. The takes place in the , the fluid around the . It uses the and to turn carbon dioxide into carbohydrate.

The diagram below shows how the two stages are .

Diagram of a chloroplast containing two boxes. The light-dependent stage on the thylakoid membranes takes in light and water and releases oxygen. An arrow labelled ATP and reduced NADP goes to the light-independent stage in the stroma, which takes in carbon dioxide and produces triose phosphate, glucose and other organic molecules. A return arrow carries ADP, inorganic phosphate and NADP back. — The two stages of photosynthesis are linked by ATP and reduced NADP. Original BioWelsh diagram.

The green box on the left is the . Light energy and water go in, and oxygen comes out.
The top arrow shows the two products that pass to the : , which supplies energy, and , which supplies hydrogen for .
The orange box on the right is the , also called the . Carbon dioxide goes in and is made.
The bottom arrow shows , () and returning, so they can be used again in the .
is then used to make glucose and all the other organic molecules the plant needs.

Lesson 2 of 17

How are leaves and chloroplasts adapted to trap light?

need light, so they are found only in the parts of a plant that light reaches: mainly the leaves, and also green stems and leaf stalks. Inside a leaf, are not spread evenly. Their position makes the most of the light that falls on the leaf.

Before looking at the leaf, it helps to picture a single . The labelled illustration below shows its structure.

Labelled cut-away illustration of an oval chloroplast showing the outer and inner membranes, the stroma, stacks of thylakoids called grana joined by lamellae, a thylakoid lumen, starch granules, lipid globules, ribosomes and a DNA ring. An enlarged circle shows water being split beside a thylakoid and hydrogen ions passing through a membrane protein to make ATP. — The structure of a chloroplast, with an enlarged view of a thylakoid. Photo: Kelvinsong, CC0.

The is surrounded by an envelope of two membranes, an outer and an inner membrane.
Inside are flattened membrane sacs called . Stacks of form (singular: ), which are joined by single called .
The membranes hold the photosynthetic , the , the and . This is where the happens.
The space inside each is the , or . It has a low pH because build up there.
The fluid around the is the . It contains the of the , starch grains, lipid droplets, and a of DNA.

How the structure of a chloroplast suits its function
Feature	How it helps photosynthesis
Many membranes stacked into	A very large surface area for , and , so more light is absorbed
held in the membranes	molecules are arranged in that pass energy to reaction centres
is a small, enclosed compartment	build up quickly, making a steep for synthesis
contains such as	The light-independent reactions happen right next to the that supply and
DNA and in the	Some of the 's own proteins can be made inside it

The photograph below is a light micrograph of a thin vertical section through a privet (Ligustrum) leaf. The stain makes the cells look green.

Light micrograph of a thin, V-shaped leaf section on a pale background. A layer of tall, tightly packed green cells runs along the top below a row of small clear cells. The lower half has loosely arranged rounded cells with large white air spaces. A round vascular bundle sits in the thickened midrib at the centre. — Light micrograph of a vertical section through a privet (*Ligustrum*) leaf, stained green. Photo: Berkshire Community College Bioscience Image Library, CC0.

The leaf is only a few cells thick, so light can reach every layer.
The single layer of flat cells along the top is the upper epidermis. It is transparent and has almost no , so light passes straight through.
Below it is of tall, tightly packed cells: the . This layer receives the most light and contains the most .
The loosely packed cells with large air spaces in the lower half are the . They also contain , but fewer.
The round structure in the middle is a vascular bundle (the midrib), which carries water to the leaf and sugars away from it.

The higher- micrograph below shows the mesophyll layers of the same kind of leaf more clearly.

Higher magnification light micrograph of a leaf section. A row of small clear epidermal cells runs along the top. Beneath it are long, narrow, closely packed cells at right angles to the surface, filled with small dots. Below these, rounded cells are separated by large white air spaces, and a row of small cells with a small gap forms the bottom surface. — Light micrograph of the mesophyll layers in a privet (*Ligustrum*) leaf. Photo: Berkshire Community College Bioscience Image Library, CC0.

Along the top is the upper epidermis, a row of small, clear cells.
The long cells arranged at right angles to the leaf surface form the . The tiny dots inside them are .
Below them, the rounded cells are separated by large air spaces, which let carbon dioxide diffuse to every cell.
The row of cells at the bottom is the lower epidermis. The small gap near the centre is a stoma, where gases enter and leave.

How leaves, cells and chloroplasts are arranged to trap light
Level	Feature	How it helps trap light
Leaf	Large, flat surface area	Intercepts as much light as possible
Leaf	Thin	Light reaches the lowest layers of cells
Leaf	Transparent cuticle and upper epidermis	Light passes through to the mesophyll below
Cell	are long and packed at right angles to the surface	Many fit into the top of the leaf, and light passes through few cell walls before reaching them
Cell	have a large vacuole	are pushed into a thin layer near the cell surface, so they do not shade each other
Cell	Many more in than in spongy cells	are concentrated where the light is brightest
	can move within the cell	In dim light they gather on the walls facing the light; in very bright light they move to the side walls, protecting the from damage
	Many membranes with	A large surface area of absorbs more light

Leaves that grow in shade can be adapted differently. Some shade leaves are thicker, with more layers of , so there are more to absorb the little light available. Others stay thin but have a larger surface area, more in each cell, or more .

Not every plant has the typical arrangement. In a grass whose leaves grow upright, light falls on both surfaces, so may be found close to both the upper and lower epidermis, and clustered around the vascular bundles. The principle is the same: sit where light reaches them and where it has few structures to pass through first.

Lesson 3 of 17

How do chlorophyll and other pigments absorb light?

White light is a mixture of , from violet and blue (about 400 nm) through green and yellow to red (about 700 nm). A is a molecule that absorbs some of light and reflects or transmits the rest. The it reflects give it its colour. A leaf looks green because its absorb blue and red light strongly but absorb little green light.

contain two main groups of photosynthetic : chlorophylls and . Each absorbs a slightly different of . Having several the can absorb a wider of than any one could on its own.

The main photosynthetic pigments in flowering plants
Group	Pigment	Colour	Wavelengths absorbed most strongly	Role
Chlorophylls		Blue-green	Blue-violet and red	Found in all plants, algae and cyanobacteria; the in the reaction centres
Chlorophylls		Yellow-green	Blue and orange-red	in the
		Orange	Blue-violet and blue-green	; also protects chlorophyll from damage by very bright light
	Xanthophylls	Yellow	Blue-violet and blue-green

and the are called . They absorb that absorbs poorly, and pass the energy on to . You will see how in the lesson on .

Two kinds of graph show how use light. An shows how much light a absorbs at each . It is measured by shining light of each through a solution of the . An shows the rate of at each , for example measured as oxygen released or carbohydrate made.

The graph below shows the of three with the drawn over them.

Line graph from 400 to 700 nm with coloured wavelength bands. Chlorophyll a peaks near 430 and 665 nm, chlorophyll b near 455 and 645 nm, and carotenoids between 440 and 490 nm only. All absorb little between 500 and 600 nm. The action spectrum on the right axis is high from 400 to 480 nm, falls to a minimum near 550 nm and peaks again near 675 nm. — Absorption spectra of chloroplast pigments with the action spectrum for photosynthesis. Original BioWelsh graph.

The x-axis is from 400 nm (violet) to 700 nm (red). The coloured bands show the colour of light at each .
The green solid line is . It has one peak in the blue-violet, near 430 nm, and another in the red, near 665 nm.
The dashed line is . Its peaks are slightly shifted: about 455 nm in the blue and about 645 nm in the orange-red.
The dotted line is the . They absorb only in the blue and blue-green region, from 400 to about 500 nm.
All three absorb very little light between about 500 and 600 nm, which is green and yellow light.
The black line is the , read on the right-hand axis. The rate of is highest in blue and red light and lowest in green light.

The peaks of the are at the same as the peaks of the . This close match is evidence that the light absorbed by these is the light used in . The does not fall to zero in green light, because some green light is still absorbed, especially by and by chlorophyll at the edges of its peaks.

One of the first pieces of evidence came from an experiment in the 1880s by the German botanist Theodor Engelmann. He shone light split into a spectrum along a of green alga. He added motile bacteria that need oxygen, which swim towards places where oxygen concentration is highest.

The diagram below shows where the bacteria gathered.

A long alga filament lies above a spectrum from 400 to 700 nm. Many small dark bacteria cluster above and below the parts of the filament lit by blue light, around 430 to 470 nm, and red light, around 650 to 690 nm. Very few bacteria are next to the parts lit by green and yellow light. — Engelmann's experiment: aerobic bacteria gather next to an alga filament where most oxygen is released. Original BioWelsh diagram.

The green bar is the alga . Each section was lit by a different of light, shown on the spectrum below it.
The black dots are aerobic bacteria. They cluster most densely next to the parts of the lit by blue light (about 430 to 470 nm) and red light (about 650 to 690 nm).
Very few bacteria are found next to the parts lit by green and yellow light.
The bacteria gathered where most oxygen was released, so the rate of was highest in blue and red light. This matches the .

also matters for aquatic organisms. Red light is absorbed by the first few metres of water, while blue light reaches much deeper. Algae that live deep in the sea often have extra that absorb blue and blue-green light well, which lets them photosynthesise in the light that is available.

Lesson 4 of 17

How are chloroplast pigments separated by chromatography?

A leaf extract looks green, but it is a mixture of several . separates them. A small spot of extract is placed on paper or a thin layer of silica gel, and a solvent moves up through it. Each dissolves in the solvent to a different extent and is attracted to the paper to a different extent, so each one moves a different distance.

The position of each is described by its : the distance moved by the divided by the distance moved by the . Because both distances are measured on the same chromatogram, the of a stays the same for a given solvent and paper, so it can be compared with known values to identify the .

Specified practical

Separating chloroplast pigments by paper chromatography

Unit 3, section 2, specified practical 1 (BioWelsh entry number)

Aim To extract photosynthetic from leaves, separate them by paper and identify each from its .

Five panels in order: a pestle and mortar to grind leaves with sand and propanone; a capillary tube to spot the extract on a pencil origin; a stoppered boiling tube with a developing chromatogram; a pencil and ruler to mark and measure; a paper strip to calculate Rf values. — Separating chloroplast pigments at a glance. Original BioWelsh diagram.

Equipment

Fresh green leaves, such as spinach
Fresh green leaves, such as spinachabout 2 gSource of ; dark green leaves contain more.
Scissors
ScissorsCut the leaves into small pieces and remove thick veins.
Pestle and mortar with a little sand
Pestle and mortar with a little sandSand helps break open cell walls and membranes.
Propanone
Propanoneabout 5 cm³Organic solvent that dissolves the out of the leaves.
Chromatography paper strip
paper strip20 mm × 200 mm that the move along.
Pencil and ruler
Pencil and rulerDraw the and and measure distances.
Fine capillary tube
Fine capillary tubePlaces a small, concentrated spot of extract on the .
Boiling tube with a bung
Boiling tube with a bungHolds the solvent and keeps the air inside saturated with vapour.
Running solvent
Running solventpetroleum ether and propanone mixtureMoves up the paper and carries the different distances.
Hair dryer on a cool settingDries each spot quickly so it stays small.
Fume cupboard
Fume cupboardRemoves flammable, harmful solvent vapour.
Eye protection and gloves
Eye protection and glovesProtect eyes and skin from solvents.

Safety

Propanone and petroleum ether are highly flammableRiskVapour could catch fire if there is a flame or spark nearby.ControlNo naked flames; work in a fume cupboard; keep bottles closed.
Solvent vapours are harmful if breathed inRiskInhaling vapour while grinding or running the chromatogram.ControlWork in a fume cupboard and keep the tube stoppered.
Solvents irritate the eyes and remove oils from skinRiskSplashes while grinding or pouring.ControlWear eye protection and gloves.

Variables

Independent variablePigment in the leaf extract

Each separates into its own spot.

differ in how soluble they are in the solvent and how strongly they are attracted to the paper.

Dependent variableDistance moved by each pigment, as an Rf value

Distance from to centre of spot ÷ distance from to .

Using a ratio chromatograms run for slightly different times can still be compared.

Controlled variablesComposition of the running solvent, Type of chromatography paper, Temperature, Air in the tube saturated with solvent vapour

Composition of the running solvent

Use the same freshly made solvent mixture.

values change if the proportions of the solvents change.

Type of paper

Cut all strips from the same batch of paper.

Different papers attract differently.

Temperature

Run chromatograms side by side in the same room.

Temperature affects how fast the solvent evaporates and moves.

Air in the tube saturated with solvent vapour

Keep the bung in place throughout.

Evaporation from the paper would change how far the solvent travels.

Control experimentKnown Rf values for each pigment

Compare measured values with values published for the same solvent.

Identifies each spot.

Control experiment or controlled variable?

This practical has no in the usual sense. Known values act as the comparison. , such as the solvent composition and type of paper, must be kept the same for values to be compared.

Method, step by step

1)
Cut about 2 g of fresh leaves into small pieces, leaving out the thick veins.
Why?
Veins contain few , and small pieces grind more easily.
2)
In a fume cupboard, grind the leaves with a pinch of sand and about 5 cm³ of propanone until the liquid is dark green.
Why?
Grinding with sand breaks cell walls, cell membranes and membranes. The dissolve in the organic solvent but not in water.
3)
Let the solid settle and pour the green extract into a small tube.
Why?
Leaf fragments would block the capillary tube and smear on the paper.
4)
Draw a pencil line 20 mm from one end of the paper, holding the strip only at its edges.
Why?
Graphite does not dissolve in the solvent, but ink would separate into its own coloured spots. Grease from fingers affects how the solvent moves.
5)
Use a capillary tube to put a small spot of extract in the centre of the line. Dry it with cool air, then add more extract to the same spot. Repeat 10 to 20 times.
Why?
Building up a small, concentrated spot gives clear, separate spots. A large spot spreads into bands that overlap.
6)
Pour running solvent into the boiling tube to a depth of about 10 mm.
Why?
The solvent must stay below the , or the would dissolve into the solvent in the tube instead of moving up the paper.
7)
Lower the strip into the tube so its end dips into the solvent but the stays above it, without touching the sides. Fix the paper with the bung.
Why?
The bung keeps the air saturated with solvent vapour, so the solvent does not evaporate from the paper. Touching the glass makes the solvent move unevenly.
Setting up and running a paper chromatogram of leaf pigments. Original BioWelsh diagram.
Open full size
8)
Leave the tube undisturbed until the solvent is about 10 mm from the top, then remove the strip and immediately mark the in pencil.
Why?
The solvent evaporates within seconds, so the front disappears if it is not marked straight away.
9)
When the chromatogram is dry, mark the centre of each spot, measure its distance from the and calculate each .
Why?
values are compared with known values for the same solvent to identify each .

Results and analysis

Example data

Distances moved by from a spinach leaf extract ( 80 mm)

Spot	Colour	Distance from / mm
1	Orange-yellow	75	0.94
2	Yellow	55	0.69
3	Blue-green	48	0.60
4	Yellow-green	35	0.44

A chromatography strip with a dashed origin line near the bottom, a dashed solvent front near the top and a green spot part way up. A blue arrow shows the distance from origin to solvent front, and a black arrow shows the distance from origin to the centre of the spot. A box gives Rf = distance moved by pigment divided by distance moved by solvent. — Measuring the distances needed to calculate an Rf value. Original BioWelsh diagram.

Photograph of a white chromatography plate. A faint pencil line and a pale green streak mark the origin in the lower half. Above it are separated bands: a pale yellow band, a yellow-green band, a yellow band, a dark green band and a faint grey band, with a single yellow band much higher up. — Thin-layer chromatogram of pigments extracted from a fresh leaf, showing yellow, grey and green bands separated above the origin. Photo: RIT RAJARSHI, CC BY 4.0. Resized.

What the results show

Four spots separated. moved furthest ( 0.94), then (0.69), (0.60) and (0.44).

that are more soluble in the solvent and less attracted to the paper travel further.

The spots show that a green leaf extract contains several . Each absorbs a different of , so together they widen the of light a can use.

A faint grey spot of phaeophytin, a breakdown product of chlorophyll that lacks , sometimes appears if the extract is old.

Worked calculations

of spot 1 () = 75 ÷ 80 = 0.94

of spot 3 () = 48 ÷ 80 = 0.60

of spot 4 () = 35 ÷ 80 = 0.44

Evaluation

A large or weak spot spreads into overlapping bands.

Effect on resultsCentres of spots are hard to judge, so values are less accurate.

ImprovementBuild up a small, concentrated spot with many applications, drying between each.

The solvent front evaporates quickly once the strip is removed.

Effect on resultsThe front may be marked too low, making every too high.

ImprovementMark the with pencil the moment the strip is removed.

Pigments fade in bright light and air.

Effect on resultsFaint spots, especially , may be missed.

ImprovementMeasure the chromatogram straight away and keep it out of bright light.

Paper chromatography gives quite broad spots.

Effect on results with similar values may not separate clearly.

ImprovementUse thin-layer on silica, which gives sharper separation.

Going further

Other ways to carry out this practical

Use thin-layer (TLC) on a plastic sheet coated with silica gel. It is faster and gives sharper spots, but values differ from those on paper.
Compare from green leaves with those from yellow autumn leaves, red leaves or brown seaweed.
Compare from sun leaves and shade leaves of the same plant.
Compare plants grown in complete with plants grown without , which make less chlorophyll.

Why Rf values differ between sources

An depends on the solvent, the paper and the temperature. Published tables only apply to the same conditions, so you may see different values for the same in different books or websites. The order of the is more reliable than the exact numbers.

Model answer: describe the method

Describe how you would separate and identify the photosynthetic in a leaf using paper .

Original practice response · 6 indicative marks

Grind fresh leaves with sand and a small volume of an organic solvent such as propanone, in a fume cupboard, to extract the .
Draw an line in pencil about 20 mm from the bottom of a strip of paper.
Use a capillary tube to apply a small spot of extract to the , drying between applications until the spot is concentrated.
Place the strip in a sealed boiling tube containing running solvent, with the solvent level below the .
Remove the strip when the solvent is near the top and immediately mark the in pencil.
Measure the distance from the to the centre of each spot and to the , calculate = distance moved by ÷ distance moved by solvent, and compare with known values to identify each .

What the answer needs to cover

extracted by grinding with sand and an organic solvent
drawn in pencil
Small, concentrated spot built up and dried between applications
Solvent level below the ; tube sealed
marked immediately
calculated and compared with known values

Exam practice

Explain why an organic solvent, rather than water, is used to extract the pigments.Show answer

Chlorophylls and are non-polar molecules held in the membranes. They dissolve in organic solvents such as propanone but are insoluble in water, so water would not remove them from the leaf.

are non-polar or lipid-soluble
They dissolve in organic solvents, not in water

Explain why the origin must be above the level of the solvent.Show answer

If the were below the solvent surface, the would dissolve into the solvent in the tube instead of being carried up the paper, so they would not separate.

would dissolve into the solvent in the tube
So they would not move up the paper and separate

A spot moved 36 mm when the solvent front moved 90 mm. Calculate its Rf value and explain why an Rf value can never be greater than 1.Show answer

= 36 ÷ 90 = 0.40. A is carried by the solvent, so it can never move further than the . The distance moved by the is therefore always less than or equal to the distance moved by the solvent.

= 0.40
A cannot move further than the

Lesson 5 of 17

What are photosystems and how is light harvested?

molecules are not scattered randomly in the membrane. They are held by proteins in groups called . Each works like a funnel: it collects light energy over a wide area and delivers it to one place where it can be used.

A has two parts. The (also called the ) contains hundreds of molecules: , and , held in place by proteins. At the centre is the , which contains a pair of special molecules.

The diagram below shows how a harvests light.

A large circle filled with small coloured circles representing chlorophyll a, chlorophyll b and carotenoid molecules around a dark green reaction centre chlorophyll a. Wavy light arrows hit pigments at the edge, and green arrows show energy passing from pigment to pigment to the centre. An orange arrow shows an excited electron leaving the reaction centre to an electron acceptor. — A photosystem: the antenna complex passes energy to chlorophyll a at the reaction centre. Original BioWelsh diagram.

The large circle is one in the membrane. The small coloured circles are molecules in the .
A of light is absorbed by one molecule. Its energy raises an electron in that molecule to a higher energy level.
The energy, not the electron, is passed from molecule to molecule, shown by the green arrows, until it reaches the .
At the , the energy excites an electron in so much that the electron leaves the molecule.
The is picked up by an . This is the start of the .

There are two types of , named in the order they were discovered, not the order in which they work. Each one is identified by the of light its chlorophyll absorbs best.

The two photosystems
Feature	Photosystem I (PSI)	Photosystem II (PSII)
name
Peak absorption of	700 nm	680 nm
Where its replacement electrons come from	, through the (or its own electrons in )	of water
Used in	Yes	No
Used in	Yes	Yes

Why have an antenna at all? in the absorbs only a narrow of . The in the antenna absorb other and pass the energy on, so the can use much more of the light that reaches it, including in shade.

Lesson 6 of 17

What happens in non-cyclic photophosphorylation?

adding a phosphate group. is making from and using energy that came from light. There are two pathways: non-cyclic, which uses both , and cyclic, which uses only. is the main one in plants, because it makes both and .

The key idea is energy levels. When a excites an electron at a , the electron is raised to a high energy level. As it passes along a chain of , it falls to lower energy levels, and the energy released at each step is used to make . To be raised high enough to reduce , the electron needs a second boost of light energy at .

The energy diagram below shows the path of the electrons. Because the path zigzags up, down and up again, it is called the .

Energy level diagram. Light excites two electrons from PSII (P680) at the bottom left up to an electron acceptor. The electrons fall through three electron carriers, where ADP and inorganic phosphate form ATP, to PSI (P700). Light excites them again up to a second, higher electron acceptor, and they pass down to NADP, forming reduced NADP. Water is split into protons, oxygen and electrons that enter PSII. — The Z scheme: the energy of electrons in non-cyclic photophosphorylation. Original BioWelsh diagram.

The vertical axis shows the energy level of the electrons. Higher on the diagram more energy.
Start at (), bottom left. Light excites two electrons, which are raised to a high energy level and accepted by an .
The electrons pass down the . At each step they lose a little energy, which is used to pump and so to make from and .
The electrons, now at a lower energy level, reach (). They replace electrons that has just lost.
Light excites the electrons in to an even higher energy level, and a second picks them up.
The electrons pass to , which also takes up two from the to become .
is now short of two electrons. It gets replacements from the splitting of water (), shown at the bottom left.

The path of electrons in

1) Light excites electrons in
2)
3) ( made)
4) , where light excites the electrons again
5)
6) is

It is called non-cyclic because the electrons travel in one direction and do not return to the they came from. They start in water and end up in .

Lesson 7 of 17

What do photolysis and the reduction of NADP do?

Two more events complete the : water is split to replace the electrons lost from , and is in the . Both also help to build the that drives synthesis.

splitting using light. An complex attached to , on the side facing the , splits water molecules: H₂O → 2H⁺ + 2e⁻ + ½O₂. It is the only known that oxidises water. Light does not split water directly; the is driven by the strongly that has just lost its electrons.

What happens to the products of photolysis
Product	Where it goes	What it does
Electrons (e⁻)	Into	Replace the electrons lost when light excited
(H⁺)	Released into the	Add to the ; then diffuse through and are used to reduce in the
Oxygen (O₂)	Diffuses out of the and the cell, into the air spaces and out through the stomata	Released as a waste product, or used by in

At the end of the chain, accepts two electrons from and two from the : + 2H⁺ + 2e⁻ → . carries hydrogen, and so reducing power, to the .

The diagram below shows the three processes that make the far more acidic than the .

Thylakoid membrane with the stroma above and the thylakoid space below. Light excites PSII and PSI. Electrons pass through carriers and a proton pump to PSI and then to NADP in the stroma, labelled as removing protons. The proton pump moves protons down into the thylakoid space. Water is split below PSII, labelled as adding protons. Protons flow up through ATP synthetase, turning ADP and inorganic phosphate into ATP in the stroma. — How the proton gradient across the thylakoid membrane is built and used. Original BioWelsh diagram.

The top of the diagram is the , with a low concentration of . The bottom is the , with a high concentration of .
① The in the uses energy from electrons to move from the into the .
② of water in the releases more there.
③ Reducing in the removes from the .
can only return to the by diffusing through . As they flow through, and are joined to make in the . This is .

happens in the , next to .
The moves from the into the .
is in the , which removes from the .
and are both released into the , where the uses them.

The in words

1) Light energy is absorbed by
2) Water is split, releasing electrons, and oxygen
3) Electrons pass along the chain; are pumped
4) diffuse through , making
5) is by electrons and

Lesson 8 of 17

How is cyclic photophosphorylation different?

In only is involved. Light excites electrons in , and they are picked up by an , just as before. But instead of passing to , the electrons are passed back along to .

The energy diagram below shows this .

Energy level diagram summarising the light-dependent stage. Solid arrows show non-cyclic electron flow: light excites two electrons from PSII (P680) up to an electron acceptor, they pass down three electron carriers to PSI (P700), light excites them again up to a second electron acceptor, and they pass to NADP, forming reduced NADP. Photolysis of water supplies electrons to PSII. Dashed arrows beside them show cyclic electron flow: electrons excited from PSI go up to the second electron acceptor, return to the first electron carrier and pass down the carriers back to PSI. ADP and inorganic phosphate form ATP as electrons pass along the carriers. A key notes that cyclic uses PSI only and makes ATP, but no reduced NADP and no oxygen. — Cyclic (dashed) and non-cyclic (solid) electron flow in the light-dependent stage. Original BioWelsh diagram.

Light excites two electrons in (), raising them to a high energy level.
An picks up the .
The electrons pass down a series of , losing energy. The energy is used to pump , and is made by .
The electrons return to , completing the cycle, ready to be excited again.
Because the electrons are not passed to , no is made. Because gets its own electrons back, water is not split and no oxygen is released.

Why does the plant need a second pathway? The uses more than . tops up the supply of without making extra . It also happens when there is little available, for example when the is slow.

Comparing cyclic and non-cyclic photophosphorylation
Feature	Cyclic	Non-cyclic
used	only	and
Path of electrons	A back to	One direction, from water to
Source of electrons	(the same electrons are reused)	of water
	()
made	Yes	Yes
made	No	Yes
and oxygen released	No	Yes

Some bacteria that photosynthesise, such as green sulfur bacteria, have only one type of . They cannot split water, so they use hydrogen sulfide as an electron source and release sulfur rather than oxygen. Comparing them with plants is a good way to test your understanding of both pathways.

Lesson 9 of 17

How do isolated chloroplasts show the light-dependent stage?

In the 1930s, Robert Hill showed that removed from cells can still carry out the . He found that isolated in light release oxygen if they are given a substance that can accept electrons. This is known as the .

In a school laboratory, the blue dye is used as the . is blue when and colourless when . In light, electrons released by the reduce instead of , so the blue colour disappears. The faster the colour change, the faster the .

Isolating

1) Grind leaves in ice-cold, solution
2) Filter through muslin to remove cell debris
3) Centrifuge to collect the as a pellet
4) Resuspend the in ice-cold
5) Mix with and keep tubes in light or dark

Why each condition is used
Condition	Reason
Ice-cold solution	Slows activity, so released from broken cells do not damage the
solution	Has the same as the , so they do not gain or lose water by and burst or shrink
	Keeps the pH constant so proteins are not denatured

The diagram below shows the results of three tubes kept for 15 minutes.

Three pairs of test tubes showing start and after 15 minutes. Tube 1, chloroplasts and DCPIP in light, changes from blue-green to green. Tube 2, the same mixture in a dashed box labelled kept in the dark, stays blue-green. Tube 3, buffer and DCPIP with no chloroplasts in light, stays blue. — The Hill reaction: tubes of DCPIP with and without chloroplasts, in light and in the dark. Original BioWelsh diagram.

Tube 1 contains and in the light. It starts blue-green and turns green, because the has been and become colourless, leaving the green colour of the .
Tube 2 contains the same mixture kept in the dark. It stays blue-green, because without light no electrons are released, so is not .
Tube 3 contains in with no , kept in the light. It stays blue, showing that light alone does not change .

Tube 1 compared with tube 3: the cause the to be .
Tube 1 compared with tube 2: light is needed for the to release electrons.
may be present in the extract, and they respire in the dark. Tube 2 does not change colour, so are not reducing the .
Adding extra carbon dioxide makes no difference, because carbon dioxide is only used in the .

Lesson 10 of 17

What happens in the light-independent stage?

The is a cycle of -controlled reactions in the , named the after Melvin Calvin, who worked it out. It uses the two products of the : as a source of energy, and as a source of reducing power (hydrogen and electrons). It turns carbon dioxide into carbohydrate.

The diagram below shows the three main steps of the cycle.

Cycle diagram. Carbon dioxide joins ribulose bisphosphate (RuBP, 5C), catalysed by rubisco, forming two glycerate-3-phosphate (GP, 3C). GP is reduced to two triose phosphate (TP, 3C) using two ATP and two reduced NADP, which become ADP, inorganic phosphate and NADP. TP regenerates RuBP using ATP, and some TP leaves to make glucose, sucrose, starch, cellulose, amino acids and lipids. A note says for every 6 TP made, 5 regenerate RuBP. — The Calvin cycle in the stroma. Original BioWelsh diagram.

: carbon dioxide from the air combines with (), a five-carbon (5C) compound. The catalyses this reaction. The unstable six-carbon product immediately splits into two molecules of (), a three-carbon compound.
: each is to (TP), another three-carbon compound. This uses from the for energy and to supply hydrogen. The , and return to the .
Regeneration: most of the is used to remake , using more . This keeps the cycle going.
Some leaves the cycle and is used to make glucose and other organic molecules.

The steps of the Calvin cycle
Step	What happens	What is needed
	(5C) + CO₂ → unstable 6C compound → 2 (3C)
	→ (TP)	(energy) and (hydrogen)
Regeneration	→
Product synthesis	Some → glucose, starch, sucrose, amino acids and lipids	, and for amino acids

( carboxylase) is thought to be the most abundant protein on Earth. It is a slow , so leaves contain a great deal of it. Because the reaction it catalyses is the slowest step, it is often the step that limits the rate of the whole cycle.

Calvin worked out the order of the reactions using a suspension of the alga Chlorella in a flat, thin glass vessel lit from both sides, known as the lollipop apparatus. He supplied hydrogencarbonate containing radioactive carbon-14, then took samples every few seconds into hot methanol, which kills the cells and stops all reactions instantly. The compounds were separated by , and those containing carbon-14 were detected on photographic film.

How the order of the was worked out

1) Supply radioactive ¹⁴C hydrogencarbonate to algae in bright light
2) Take samples at short time intervals into hot methanol
3) Separate the compounds by
4) Detect radioactive compounds on X-ray film
5) The first compound to become radioactive is the first product:

After only a few seconds, the radioactive carbon was found in . A little later it appeared in , then in hexose sugars and . The order in which compounds became radioactive showed the order of the reactions. The experiment was carried out in bright light so that and were not in short supply.

Lesson 11 of 17

What happens to GP, TP and RuBP when light or carbon dioxide changes?

Exam questions often show a graph of how the concentrations of , TP and change when conditions change, and ask you to explain it. You can work out every answer by asking two questions: which reaction has stopped or slowed, and which reactions carry on for a while?

The graph below shows what happens in the when the light is switched off.

Line graph of concentration against time from 0 to 10 minutes. Before 5 minutes, GP is steady at 4, TP at 6 and RuBP at 5 arbitrary units. After the light is switched off at 5 minutes, GP rises to about 8.6, while TP falls to about 1.5 and RuBP falls to about 0.5. — Concentrations of GP, TP and RuBP in the stroma when the light is switched off at 5 minutes. Original BioWelsh graph.

Before 5 minutes, the plant is in the light and all three concentrations are steady, because each compound is made as fast as it is used.
At 5 minutes the light is switched off. The stops, so no more or is made.
(solid blue line) rises. that is already present still combines with carbon dioxide, so is still made. But cannot be to TP without and , so it builds up.
TP (dashed orange line) falls. No new TP is made from , while existing TP is still used to make other molecules and to regenerate .
(dotted green line) falls. It is still used up in , but it is not regenerated, because regeneration needs and TP.
Eventually levels off, because once has run out, no more can be made.

The second graph shows what happens when the carbon dioxide concentration is lowered while the light stays on.

Line graph of concentration against time from 0 to 10 minutes. Before 5 minutes, GP is steady at 6, TP at 5 and RuBP at 3 arbitrary units. After carbon dioxide is lowered at 5 minutes, RuBP rises to about 8, GP falls to about 2 and TP falls slowly to about 3.2. — Concentrations of GP, TP and RuBP in the stroma when carbon dioxide concentration is lowered at 5 minutes. Original BioWelsh graph.

At 5 minutes the concentration of carbon dioxide falls.
(dotted green line) rises. Less combines with carbon dioxide, but is still regenerated from TP, because and are still being made in the light.
(solid blue line) falls. Less is made by , but existing is still to TP.
TP (dashed orange line) falls slowly, because less is available to make it, while TP is still used to regenerate and make other molecules.

Predicting changes in the Calvin cycle
Change	GP	TP	RuBP	Reason
Light switched off (or much dimmer)	Rises	Falls	Falls	No or : not , not regenerated
Carbon dioxide concentration lowered	Falls	Falls slowly	Rises	Less of CO₂, but still and still regenerated
that reduces is missing	Rises	Falls	Falls	No : not to TP, so less regenerated and less glucose made
inhibited	Falls	Falls	Rises	not used in

The same reasoning explains real situations. For example, some wheat plants resist a fungus that feeds on the products of . At the infection site, the for an needed to reduce is switched off. With no , is not to TP, less is regenerated, and less glucose is made, so the fungus has less food.

Lesson 12 of 17

How does a plant make other molecules from triose phosphate?

Animals must eat carbohydrates, lipids and proteins. A plant makes all of its own organic molecules, starting from the three-carbon compounds of the . You need to know what is made and where it comes from, but not the chemistry of the reactions.

What is made from the products of the Calvin cycle

Zoom 100%

Zoom in to read each branch. Swipe or scroll sideways to explore.

and (3C)
- Carbohydrates
  - Hexose sugars: glucose and fructose
    - Sucrose: transported in the phloem
    - Starch (from α-glucose): stored
    - Cellulose (from β-glucose): cell walls
- Lipids
  - : made from
  - : made from via
- Proteins
  - Amino acids: made from , with nitrogen from
- Nucleic acids: also need nitrogen and phosphate

Carbohydrates: two molecules join to make a hexose phosphate, which is converted to glucose. Glucose and fructose form sucrose, the sugar carried in the phloem. α-glucose is joined to form starch for storage, and β-glucose to form cellulose for cell walls.
Lipids: is made from . are made from , which is made from . and join by condensation reactions to form and phospholipids.
Proteins: amino acids are made from and other intermediates, but they also need an amino group containing nitrogen. The nitrogen comes from absorbed by the roots.

Lesson 13 of 17

What limits the rate of photosynthesis?

To photosynthesise, a plant needs light of suitable , carbon dioxide, water and a suitable temperature. When several factors affect a process, the rate is controlled by whichever one is in shortest supply. That factor is called the : if you increase it, the rate increases; if you increase any other factor, the rate stays the same.

Think of a sandwich production line. If there is plenty of bread but only a few slices of cheese, making more bread will not produce more sandwiches. Only more cheese will. works the same way.

The graph below shows how light intensity affects the rate of when other factors are kept constant.

Line graph of rate against light intensity. The rate rises steeply from zero, labelled light is limiting, then curves and levels off at about 25 arbitrary units from a light intensity of about 60, labelled another factor is limiting. — The effect of light intensity on the rate of photosynthesis. Original BioWelsh graph.

At low light intensity, the rate rises steeply as light intensity increases. Light is the : the cannot make and fast enough.
The curve then bends and levels off. Increasing light intensity further has no effect, so light is no longer limiting. Another factor, such as carbon dioxide concentration or temperature, is now limiting.
In a dark place, the stops, so no oxygen is released. At very high light intensity, the rate can fall because are damaged.

The next graph shows the effect of carbon dioxide concentration.

Line graph of rate against carbon dioxide concentration from 0 to 0.5%. A vertical line marks 0.04% carbon dioxide in air. The rate rises steeply at first, then levels off at about 25 arbitrary units above about 0.3%, labelled another factor is limiting. — The effect of carbon dioxide concentration on the rate of photosynthesis. Original BioWelsh graph.

At low concentrations, the rate increases as carbon dioxide concentration increases, so carbon dioxide is limiting. The rate of by depends on it.
The vertical line shows the concentration in air, about 0.04%. At this concentration carbon dioxide is usually the for land plants in good light.
At higher concentrations the curve levels off, because another factor is limiting.

Growers use this idea. In glasshouses they may raise carbon dioxide to about 0.1% using burners or gas cylinders, which increases the yield of crops such as tomatoes. Very high concentrations are not used, because they can cause stomata to close and are unsafe for workers.

The graph below shows the effect of temperature.

Line graph of rate against temperature from 0 to 50 °C. The rate rises from near zero to a peak of about 20 arbitrary units at about 30 °C, then falls steeply to near zero at 50 °C, labelled enzymes denature. — The effect of temperature on the rate of photosynthesis. Original BioWelsh graph.

As temperature rises from low values, the rate increases. Molecules have more kinetic energy, so - complexes form more often, especially in the .
The rate peaks at an optimum temperature, which depends on the . Many temperate plants have an optimum around 25 to 30 °C.
Above the optimum, the rate falls quickly because such as are denatured. Unlike the other graphs, this curve does not level off.

In nature, several factors change together. The graph below shows rate against light intensity under three different conditions.

Line graph with three curves rising from zero together. The curve for 0.1% carbon dioxide at 25 °C levels off highest at about 28 arbitrary units. The curve for 0.04% carbon dioxide at 25 °C levels off at about 17. The curve for 0.04% carbon dioxide at 15 °C levels off lowest at about 11. — Rate of photosynthesis against light intensity under three conditions. Original BioWelsh graph.

All three lines rise together at low light intensity, because light is limiting under all three conditions.
On the lowest line (0.04% carbon dioxide, 15 °C), the rate levels off first. Raising the temperature to 25 °C (middle line) increases the plateau, so temperature was limiting.
Raising carbon dioxide to 0.1% at 25 °C (top line) increases the plateau again, so carbon dioxide was limiting on the middle line.
Where a line is level, increasing light intensity has no effect. The factor that raises the plateau is the one that was limiting.

Plants respire all the time, in light and dark, releasing carbon dioxide. In dim light, can release more carbon dioxide than takes in. As light intensity increases, a point is reached where the rate of exactly equals the rate of , so there is no net exchange of carbon dioxide or oxygen. The light intensity at which this happens is the .

The graph below compares a plant adapted to bright sun with a plant adapted to shade.

Line graph of net carbon dioxide uptake against light intensity, with a horizontal line at zero. The shade plant starts at minus 2, crosses zero at about 5 and levels off at about 12. The sun plant starts at minus 5, crosses zero at about 15 and levels off at about 26. Circles mark the two compensation points. — Net carbon dioxide uptake at different light intensities for a sun plant and a shade plant. Original BioWelsh graph.

The y-axis is net carbon dioxide uptake. Below zero, is faster than and the plant releases carbon dioxide.
Each line crosses zero at that plant's . The shade plant reaches its at a lower light intensity than the sun plant.
The shade plant levels off at a low rate, so it cannot make use of bright light. The sun plant continues to increase to a much higher rate.
A plant only grows if it spends enough time above its to make more carbohydrate than it uses in .

The main limiting factors
Factor	Which stage it affects most	Effect when it is limiting
Light intensity	Light-dependent	Less and are made, so is to TP more slowly
Carbon dioxide concentration	Light-independent	Less CO₂ combines with , so less is made
Temperature	Mainly light-independent (-controlled)	work more slowly at low temperature and are denatured at high temperature
Water	Many processes	Stomata close, so less CO₂ enters; hard to study because so many processes are affected

Lesson 14 of 17

How can the rate of photosynthesis be measured?

The rate of can be measured by how fast a plant releases oxygen or takes in carbon dioxide. Aquatic plants such as pondweed are ideal, because the oxygen they release forms bubbles of gas that can be collected and measured in a .

Light intensity is changed by moving a lamp. For a small light source, light intensity is inversely proportional to the square of the distance, d, between the lamp and the plant: light intensity ∝ . Doubling the distance cuts the light intensity to a quarter.

Specified practical

Investigating the effect of light intensity on the rate of photosynthesis

Unit 3, section 2, specified practical 2 (BioWelsh entry number)

Aim To investigate the effect of light intensity on the rate of in pondweed, by measuring the volume of gas released in a at different distances from a lamp.

Five panels in order: pondweed in a boiling tube of hydrogencarbonate solution; a lamp at a measured distance; a stop clock for acclimatising and timing; a photosynthometer with a gas bubble on a scale; a metre rule to change the distance. — Measuring the rate of photosynthesis at a glance. Original BioWelsh diagram.

Equipment

Pondweed, such as Elodea or Cabomba
Pondweed, such as Elodea or Cabomba5 cm lengthAquatic plant whose oxygen can be collected as gas bubbles.
Sodium hydrogencarbonate solution
Sodium hydrogencarbonate solution1%Releases carbon dioxide so it does not limit the rate.
Boiling tubeHolds the pondweed in the solution.
Photosynthometer
Collects gas in a capillary tube so its length can be measured on a scale.
Beaker of water
Beaker of waterActs as a water bath and heat shield to keep temperature constant.
Thermometer
ThermometerChecks the temperature stays constant.
LED lamp
LED lampLight source; gives off little heat.
Metre rule
Metre ruleMeasures the distance from lamp to pondweed.
Stop clock
Stop clockTimes the gas collection and acclimatisation.
Sharp scissors or scalpel
Sharp scissors or scalpelCuts the stem cleanly.
Eye protection
Eye protectionProtects eyes from splashes.

Safety

Lamp bulb and casing can become hotRiskBurns when moving the lamp.ControlUse an LED lamp and let it cool before handling.
Mains electricity near waterRiskElectric shock if water is spilt on the lamp.ControlKeep the lamp and cables away from the beaker; dry hands before touching the lamp.
Sharp scalpel or scissorsRiskCuts when trimming the pondweed.ControlCut away from the body on a board.

Variables

Independent variableLight intensity

Change the distance of the lamp from 0.10 to 0.40 m and calculate .

Light provides the energy for the .

Example: At 0.20 m, light intensity = 1 ÷ 0.20² = 25 m⁻².

Dependent variableRate of photosynthesis

Volume of gas collected in 5 minutes, divided by 5, in mm³ min⁻¹.

Oxygen from forms the gas bubble.

Controlled variablesTemperature, Carbon dioxide concentration, Piece of pondweed, Background light, Wavelength of light, Acclimatisation and collection time

Temperature

Beaker of water at 20 °C, checked with a thermometer; LED lamp.

in work faster at higher temperatures.

Carbon dioxide concentration

Same volume of 1% sodium hydrogencarbonate solution throughout.

Carbon dioxide is used in the and can be limiting.

Piece of pondweed

Use the same piece for every reading.

Different pieces have different leaf areas and numbers of .

Background light

Carry out the experiment in a darkened room.

Extra light would increase the real light intensity.

of light

Use the same lamp throughout.

Different are absorbed and used to different extents.

Acclimatisation and collection time

5 minutes each at every distance.

The plant needs time to adjust to each new light intensity.

Control experimentPondweed in the dark

Cover the tube with foil and collect gas for 5 minutes.

Little or no gas collects, showing gas production depends on light.

Control experiment or controlled variable?

Temperature and carbon dioxide concentration are : they are kept the same at every distance. Pondweed covered in foil is a : it shows that gas is only produced when light reaches the plant.

Method, step by step

1)
Keep the pondweed in bright light in sodium hydrogencarbonate solution for at least an hour before starting.
Why?
The plant is photosynthesising steadily before readings are taken.
2)
Under water, cut a 5 cm length of pondweed and place it cut end up in a boiling tube full of 1% sodium hydrogencarbonate solution.
Why?
The solution supplies carbon dioxide. Cutting under water stops air entering the stem, and gas escapes from the cut end.
3)
Stand the boiling tube in a beaker of water at 20 °C, and check the temperature with a thermometer throughout.
Why?
The water absorbs heat from the lamp and keeps the temperature constant, because temperature also affects the rate.
4)
Fill the with solution using the syringe, remove any air bubbles, and place its flared end over the cut end of the stem.
Why?
Any air left in the tube would be measured as if it were gas from the plant.
A photosynthometer set up to measure the gas released by pondweed. Original BioWelsh diagram.
Open full size
5)
Darken the room and place the LED lamp 0.10 m from the pondweed, measured with a metre rule.
Why?
Background light would change the light intensity the plant receives.
6)
Leave the pondweed for 5 minutes before collecting gas.
Why?
The rate of needs time to adjust to the new light intensity.
7)
Collect gas for 5 minutes. Then use the syringe to draw the bubble alongside the scale and measure its length in mm.
Why?
A collection time lets rates be compared. Measuring volume is more accurate than counting bubbles, which vary in size.
8)
Push the bubble out, repeat twice more at the same distance, then repeat the procedure at 0.15, 0.20, 0.25, 0.30 and 0.40 m.
Why?
Three repeats allow anomalous results to be spotted and a reliable to be calculated.
9)
Calculate light intensity as , the volume of each bubble as πr² × length, and the rate as volume ÷ 5 minutes.
Why?
Light intensity is inversely proportional to the square of the distance, and dividing by time gives a rate.

Results and analysis

Example data

Length of gas bubble collected in 5 minutes by pondweed at different distances from an LED lamp (capillary diameter 1.0 mm)

Distance from lamp / m	Light intensity () / m⁻²	Trial 1 / mm	Trial 2 / mm	Trial 3 / mm	length / mm	Volume / mm³	Rate / mm³ min⁻¹
0.40	6.2	5	6	5	5.3	4.2	0.84
0.30	11.1	9	8	10	9.0	7.1	1.41
0.25	16.0	12	13	12	12.3	9.7	1.94
0.20	25.0	18	17	19	18.0	14.1	2.83
0.15	44.4	29	31	30	30.0	23.6	4.71
0.10	100.0	36	35	37	36.0	28.3	5.65

Graph of rate in cubic millimetres per minute against light intensity from 0 to 100 per square metre. Six crosses joined by straight lines rise steadily from 0.84 at 6.2 to 2.83 at 25.0 and 4.71 at 44.4, then rise only slightly to 5.65 at 100. — Rate of gas production by pondweed at different light intensities. Original BioWelsh graph.

What the results show

Between 6.2 and 44.4 m⁻², the rate rose from 0.84 to 4.71 mm³ min⁻¹, roughly in proportion to light intensity. Light was the .

At light intensities up to about 44 m⁻², the could not make and fast enough, so each increase in light increased the rate.

From 44.4 to 100 m⁻², the rate only rose to 5.65 mm³ min⁻¹. Light was no longer limiting, and another factor, such as carbon dioxide concentration or temperature, was limiting.

The measured gas volume is an underestimate of , because some oxygen is used in and some dissolves in the water.

Worked calculations

Light intensity at 0.20 m = 1 ÷ 0.20² = 25.0 m⁻²

length at 0.20 m = (18 + 17 + 19) ÷ 3 = 18.0 mm

Volume = πr² × length = 3.14 × 0.5² × 18.0 = 14.1 mm³

Rate = 14.1 ÷ 5 = 2.83 mm³ min⁻¹

Evaluation

The lamp may warm the water.

Effect on resultsTemperature would change as well as light intensity, affecting the rate.

ImprovementUse an LED lamp, a beaker of water as a heat shield, and monitor the temperature.

Some oxygen is used in respiration or dissolves.

Effect on resultsThe rate of is underestimated.

ImprovementAccept that this measures net rate; the error is similar at each distance, so trends are still valid.

The lamp is not a true point source.

Effect on results is only an estimate of light intensity.

ImprovementMeasure light intensity at the pondweed with a light meter.

Only one piece of pondweed was used.

Effect on resultsResults may not represent the .

ImprovementRepeat with several pieces and calculate a .

Going further

Other ways to carry out this practical

Carbon dioxide concentration: keep the lamp at a distance and use sodium hydrogencarbonate solutions of 0, 0.25, 0.5, 0.75 and 1%.
Temperature: keep the lamp and stand the tube in water baths at 10, 20, 30, 40 and 50 °C.
: place coloured filters between the lamp and the plant, adjusting the distance so a light meter gives the same intensity for each colour.
Leaf discs: cut discs from a leaf, remove the air from them with a syringe so they sink in sodium hydrogencarbonate solution, then time how long they take to float as oxygen builds up.
Counting bubbles: simpler, but less accurate because bubbles vary in size.

Model answer: describe the method

Describe how you would use a to investigate the effect of light intensity on the rate of in pondweed.

Original practice response · 6 indicative marks

Place a cut piece of pondweed, cut end up, in a boiling tube of sodium hydrogencarbonate solution, which provides carbon dioxide.
Stand the tube in a beaker of water to keep the temperature constant, and check it with a thermometer.
Fit the capillary tube of the over the cut end, with no air bubbles in the tube.
Place a lamp at a measured distance, in a darkened room, and allow 5 minutes for the plant to acclimatise.
Collect gas for a time, draw the bubble alongside the scale with the syringe, and measure its length; calculate volume = πr² × length.
Repeat three times at each of at least five distances, calculate rates, and calculate light intensity as ; keep the same pondweed and solution throughout.

What the answer needs to cover

Sodium hydrogencarbonate as carbon dioxide source
Temperature kept constant with a water bath and monitored
Light intensity varied by distance, with
Background light excluded and acclimatisation time
Gas volume measured over a time; volume = πr² × length
Repeats at each distance and calculated

Exam practice

Explain why the measured volume of gas underestimates the rate of photosynthesis.Show answer

Some of the oxygen made in is used by the pondweed's own in , and some dissolves in the water before it forms a bubble. The gas collected therefore shows the net rate, which is lower than the true rate.

Oxygen used in
Oxygen dissolves in the water
So the measured rate is lower than the true rate

The rate at 0.10 m was lower than predicted from a straight line through the other points. Explain why.Show answer

At high light intensity, light is no longer the . Another factor, such as carbon dioxide concentration or temperature, is limiting, so increasing light intensity has less effect on the rate.

Light is not limiting at high intensity
Another named factor, carbon dioxide or temperature, is limiting

A student counted bubbles instead of measuring gas volume. Explain why this is less accurate.Show answer

Bubbles are not all the same size, so the same number of bubbles can contain different volumes of gas. Small bubbles may also be missed. Measuring the length of gas in a capillary tube gives the actual volume.

Bubbles vary in size
Volume measured directly is more accurate

Lesson 15 of 17

Why do plants need nitrogen and magnesium?

supplies carbon, hydrogen and oxygen, but plants also need inorganic nutrients, sometimes called mineral ions. They are absorbed from the soil by root hair cells, by or , and carried to the leaves in the xylem. You need to know the roles of two: nitrogen and .

Inorganic nutrients have three broad roles: some are part of structures (calcium in cell walls), some are needed to make important molecules (nitrogen in amino acids, in chlorophyll), and some activate ( activates such as ATPase). If a nutrient is in short supply, it can limit growth just as light or carbon dioxide can limit .

Plants absorb nitrogen mainly as (NO₃⁻), and sometimes as ammonium ions (NH₄⁺). They cannot use nitrogen gas from the air directly. In the plant, is converted to ammonium, which provides the amino group (-NH₂) of amino acids.

The roles of nitrogen and magnesium
Nutrient	Absorbed as	Used to make or do	Deficiency symptoms
Nitrogen	(NO₃⁻), or ammonium ions (NH₄⁺)	Amino acids and proteins (including ), nucleotides, DNA, RNA, and chlorophyll	Poor, stunted growth; (yellowing), which appears first in older leaves
	(Mg²⁺)	Part of every chlorophyll molecule; activates such as ATPase	between the veins, starting in older leaves, which look mottled; less

yellowing of leaves because not enough chlorophyll is made. Without enough chlorophyll, less light is absorbed, less happens, and the plant grows less.

The photograph below shows maize leaves from a plant growing in soil short of .

Close-up photograph of long maize leaves in a field. Narrow green strips run along the parallel veins, with pale yellow stripes of tissue between them, giving a striped appearance. — Maize (*Zea mays*) leaves showing magnesium deficiency symptoms, in acid soil in KwaZulu-Natal, South Africa. Photo: Alandmanson, CC BY 4.0. Resized.

The long leaves have many parallel veins, as is typical of grasses such as maize.
The narrow strips along the veins stay green, but the tissue between the veins has turned pale yellow. This striped yellowing between veins is interveinal .
The yellow areas contain less chlorophyll, because is needed to make each chlorophyll molecule.
can be moved from older leaves to growing leaves, so older leaves usually show the symptoms first.

Why do older leaves turn yellow first? When supply is short, the plant breaks down some chlorophyll and proteins in older leaves and moves the nitrogen and in the phloem to young, growing leaves, where they are most useful.

Nitrogen supply shapes where plants can live. Insect-eating plants such as sundews grow in boggy soils where bacteria keep levels very low. By digesting insects, they obtain nitrogen-containing compounds such as proteins, and use the nitrogen to make their own amino acids and nucleotides.

Lesson 16 of 17

How can the effect of nitrogen and magnesium on growth be investigated?

The roles of mineral ions can be tested by growing plants in solutions that contain every ion they need except one. (Lemna) is a good choice: it is a tiny floating flowering plant made of small leaf-like fronds with roots hanging into the water. It reproduces quickly by budding off new fronds, so its growth can be measured by counting fronds over two weeks.

Three are compared. A complete solution contains all the essential ions, including and , and acts as the . The other two are identical except that one lacks and one lacks . Any difference in growth or colour can then be to the missing ion.

Specified practical

Investigating the role of nitrogen and magnesium in plant growth

Unit 3, section 2, specified practical 3 (BioWelsh entry number)

Aim To investigate the effect of a lack of or on the growth and colour of over two weeks.

Four panels in order: three bottles of culture solution labelled all, minus N and minus Mg; forceps to add duckweed; a lamp to give the same conditions; a Petri dish of duckweed to count and observe. — Investigating nitrogen and magnesium in plant growth at a glance. Original BioWelsh diagram.

Equipment

Duckweed (Lemna)
(Lemna)Small floating plant that grows quickly by producing new fronds.
Three culture solutions
Three 20 cm³ per dishComplete, lacking and lacking .
Nine Petri dishes with lidsThree dishes for each solution; lids reduce evaporation.
Forceps
ForcepsLift plants gently from underneath without damaging them.
Syringe
Syringe20 cm³Measures the same volume of solution into each dish.
Dilute sodium hypochlorite and distilled water
Dilute sodium hypochlorite and distilled water1%Surface-sterilise the plants, then rinse them.
Labels or a marker pen
Labels or a marker penIdentify each dish.
Lamp or light bank
Lamp or light bankGives all dishes the same light.
Hand lensHelps count small new fronds.
Gloves and eye protection
Gloves and eye protectionProtect against sodium hypochlorite.

Safety

Sodium hypochlorite is an irritant and corrosiveRiskSkin or eye damage when dipping or rinsing plants.ControlWear gloves and eye protection; rinse any splashes with water.
may irritate skin and eyesRiskSplashes when filling dishes.ControlWear eye protection and wash hands after handling.

Variables

Independent variableMineral ions available

Complete solution, solution lacking , solution lacking .

and are needed to make proteins, nucleic acids and chlorophyll.

Dependent variableGrowth of duckweed

Number of fronds every two days, plus colour of fronds.

More fronds more growth.

Example: In complete solution the rose from 10 to 60 fronds.

Controlled variablesVolume of culture solution, Starting number of fronds, Light intensity and duration, Temperature, Concentration of all other ions

Volume of

20 cm³ in every dish.

Method, step by step

1)
Label three Petri dishes for each : complete, lacking and lacking .
Why?
Three dishes per solution allow a to be calculated and anomalies spotted.
2)
Dip the in 1% sodium hypochlorite for about 30 seconds, then rinse it twice in distilled water.
Why?
This kills bacteria and algae on the surface, which would compete for nutrients. Rinsing removes the hypochlorite, which would damage the plants.
3)
Use a syringe to add 20 cm³ of the correct to each dish.
Why?
The same volume the same amount of each ion is available.
4)
Use forceps to transfer healthy green plants with a total of 10 fronds into each dish, lifting them from underneath.
Why?
Every dish starts with the same number of fronds. Lifting from below avoids crushing the fronds.
5)
Put lids on the dishes and place them all under the same light at room temperature, about 20 °C.
Why?
Light and temperature affect growth, so they must be the same for every dish.
6)
Every two days for 14 days, count the fronds in each dish and record their colour.
Why?
Counting over time shows the pattern of growth, and colour shows .
7)
Calculate the number of fronds for each solution on each day, and the percentage increase after 14 days.
Why?
and percentage increases allow fair comparison between solutions.

Results and analysis

Example data

number of fronds in three (three dishes per solution)

Time / days	Complete solution	Lacking	Lacking
0	10	10	10
2	13	11	12
4	17	12	14
6	22	13	17
8	29	13	19
10	37	14	22
12	47	14	24
14	60	15	27

Three Petri dishes seen from above. The complete solution dish has many green clusters of fronds. The dish lacking nitrate has only four clusters of small, pale yellow-green fronds. The dish lacking magnesium has eight clusters of green fronds with yellow patches. — Duckweed after two weeks in three culture solutions. Original BioWelsh diagram.

Line graph of mean number of fronds against time from 0 to 14 days. All start at 10. Complete solution rises fastest to 60. Lacking magnesium rises to 27. Lacking nitrate rises only to 15. — Mean number of duckweed fronds over 14 days in three culture solutions. Original BioWelsh graph.

What the results show

In complete solution, the number of fronds rose from 10 to 60, an increase of 500%, and all fronds stayed green.

Without , the number only rose to 15 (50%), and the fronds were small and pale yellow-green. Nitrogen is needed to make amino acids, proteins, nucleotides and chlorophyll, so very little new tissue could be made.

Without , fronds rose to 27 (170%), but they developed yellow patches. is part of every chlorophyll molecule, so less chlorophyll was made, less light was absorbed and less happened.

Growth without was slower to fall, because already in the older fronds could be moved to new ones for a while.

Worked calculations

Percentage increase in complete solution = (60 − 10) ÷ 10 × 100 = 500%

Percentage increase without = (15 − 10) ÷ 10 × 100 = 50%

Percentage increase without = (27 − 10) ÷ 10 × 100 = 170%

Evaluation

Deciding what counts as a new frond is subjective.

Effect on resultsCounts may differ between people or days.

ImprovementAgree a rule, for example count a frond once it is visible beyond its parent, and use the same person with a hand lens.

Frond number does not show how large the fronds are.

Effect on resultsGrowth may be underestimated or overestimated.

ImprovementAlso measure the of the plants, or the area covered using a grid or photograph.

Water evaporates from the dishes.

Effect on resultsIon concentrations increase over two weeks.

ImprovementKeep lids on and top up to the mark with distilled water.

Algae or bacteria may grow in the dishes.

Effect on resultsThey use the ions and shade the .

ImprovementSurface-sterilise the plants and use sterile dishes and solutions.

Going further

Other ways to carry out this practical

Measure growth as the of plants at the end, by drying them at 80 °C until the mass is constant.
Measure the area of water covered by fronds using a transparent grid or a photograph analysed with software.
Grow seedlings, such as wheat, in the same and measure height, leaf number or .
Extract from the fronds and compare them by .

Model answer: describe the method

Describe how you would investigate the effect of a lack of and a lack of on the growth of .

Original practice response · 6 indicative marks

Prepare three that are identical except that one is complete, one lacks and one lacks ; the complete solution is the .
Surface-sterilise the briefly in dilute sodium hypochlorite and rinse in distilled water.
Put the same volume, for example 20 cm³, of each solution into three dishes each.
Add plants with the same number of fronds, for example 10, to every dish.
Keep all the dishes at the same light intensity and temperature, with lids on, for two weeks.
Count the fronds and record their colour every two days, then calculate the and percentage increase for each solution.

What the answer needs to cover

Complete solution as ; solutions lacking and lacking
Plants surface-sterilised and rinsed
Same volume of solution; repeats for each solution
Same starting number of fronds
Same light and temperature; lids to reduce evaporation
Fronds counted and colour recorded over time; or percentage increase calculated

Exam practice

Explain why the fronds in the solution lacking nitrate were pale and few.Show answer

provides the nitrogen needed to make amino acids and proteins, including , as well as nucleotides for DNA and RNA, and chlorophyll. Without , the plants could not make new cells and tissues, so few fronds grew, and little chlorophyll was made, so the fronds were pale.

Nitrogen needed for amino acids and proteins
Nitrogen needed for nucleic acids and chlorophyll
Fewer new cells, so few fronds; less chlorophyll, so pale

Explain why older leaves show magnesium deficiency before younger leaves.Show answer

When is in short supply, the plant breaks down chlorophyll in older leaves and moves the in the phloem to young, growing leaves. The older leaves lose chlorophyll and turn yellow first.

is moved from older to younger leaves
Older leaves lose chlorophyll first

Explain why the duckweed was rinsed in dilute sodium hypochlorite at the start.Show answer

Sodium hypochlorite kills , such as bacteria and algae, on the surface of the plants. These would otherwise grow in the dishes, use the mineral ions and shade the , affecting its growth.

Kills on the plants
They would compete for ions or light

Lesson 17 of 17

How do the ideas in photosynthesis fit together?

is a chain of ideas. Light is trapped by in placed where light reaches them. Its energy drives electrons from water to , making on the way. and then drive the , which fixes carbon dioxide and makes the used for everything else. The tree below shows how the topics connect.

How the ideas in this topic connect

Zoom 100%

Zoom in to read each branch. Swipe or scroll sideways to explore.

in
- Trapping light
  - Leaf and
  - , absorption and action spectra
    - of (practical)
  - : antenna and
- ()
  - : and
  - : only
  - and the
  - with
- ()
  - : , , regeneration
  - Products: carbohydrates, lipids, amino acids
- Rate and nutrients
  - and
    - Measuring rate with a (practical)
  - Nitrogen and (practical)

Oxygen comes from water, not from carbon dioxide.
happens in the ; is in the .
is made as electrons pass along the chain between and , not at the .
uses ; uses .
Name both and and give a use for each in the .
In the dark, rises while TP and fall.
When a rate graph levels off, another factor is limiting.
Plants absorb and , not 'nitrogen'.
Compare spectra at the same , and draw a conclusion.

Photosynthesis uses light energy to synthesise organic molecules

What is photosynthesis and why are chloroplasts called transducers?

How are leaves and chloroplasts adapted to trap light?BWRBioWelsh recommendedWe recommend revising this because it has appeared regularly in past papers. Asked in 2 of 7 WJEC exam series (2017 to 2025), 9 marks in total. This is a revision priority, not a prediction of your exam.

How do chlorophyll and other pigments absorb light?

How are chloroplast pigments separated by chromatography?

Equipment

Safety

Variables

Method, step by step

Results and analysis

Distances moved by pigments from a spinach leaf extract (solvent front 80 mm)

Measuring the distances needed to calculate an Rf value. Original BioWelsh diagram.

Thin-layer chromatogram of pigments extracted from a fresh leaf, showing yellow, grey and green bands separated above the origin. Photo: RIT RAJARSHI, CC BY 4.0. Resized.

What the results show

Worked calculations

Evaluation

Going further

Model answer: describe the method

Exam practice

What are photosystems and how is light harvested?

What happens in non-cyclic photophosphorylation?BWRBioWelsh recommendedWe recommend revising this because it has appeared regularly in past papers. Asked in 3 of 7 WJEC exam series (2017 to 2025), 16 marks in total. This is a revision priority, not a prediction of your exam.

What do photolysis and the reduction of NADP do?

How is cyclic photophosphorylation different?

How do isolated chloroplasts show the light-dependent stage?

What happens in the light-independent stage?BWRBioWelsh recommendedWe recommend revising this because it has appeared regularly in past papers. Asked in 3 of 7 WJEC exam series (2017 to 2025), 6 marks in total. This is a revision priority, not a prediction of your exam.

What happens to GP, TP and RuBP when light or carbon dioxide changes?BWRBioWelsh recommendedWe recommend revising this because it has appeared regularly in past papers. Asked in 3 of 7 WJEC exam series (2017 to 2025), 12 marks in total. This is a revision priority, not a prediction of your exam.

How does a plant make other molecules from triose phosphate?

What limits the rate of photosynthesis?

How can the rate of photosynthesis be measured?

Equipment

Safety

Variables

Method, step by step

Results and analysis

Length of gas bubble collected in 5 minutes by pondweed at different distances from an LED lamp (capillary diameter 1.0 mm)

Rate of gas production by pondweed at different light intensities. Original BioWelsh graph.

What the results show

Worked calculations

Evaluation

Going further

Model answer: describe the method

Exam practice

Why do plants need nitrogen and magnesium?BWRBioWelsh recommendedWe recommend revising this because it has appeared regularly in past papers. Asked in 3 of 7 WJEC exam series (2017 to 2025), 11 marks in total. This is a revision priority, not a prediction of your exam.

How can the effect of nitrogen and magnesium on growth be investigated?

Equipment

Safety

Variables

Method, step by step

Results and analysis

Mean number of duckweed fronds in three culture solutions (three dishes per solution)

Duckweed after two weeks in three culture solutions. Original BioWelsh diagram.

Mean number of duckweed fronds over 14 days in three culture solutions. Original BioWelsh graph.

What the results show

Worked calculations

Evaluation

Going further

Model answer: describe the method

Exam practice

How do the ideas in photosynthesis fit together?

Worksheets

Worksheet 1: 3.2 Photosynthesis (154 marks)

Check your understanding.

How are leaves and chloroplasts adapted to trap light?

Distances moved by from a spinach leaf extract ( 80 mm)

What happens in non-cyclic photophosphorylation?

What happens in the light-independent stage?

What happens to GP, TP and RuBP when light or carbon dioxide changes?

Why do plants need nitrogen and magnesium?

number of fronds in three (three dishes per solution)