Photosynthesis as an energy transformation

Photosynthesis is a metabolic process that uses light energy to make carbon compounds from simple inorganic substances. In the organisms you’ll usually meet — plants, algae and cyanobacteria — the main inorganic raw materials are carbon dioxide and water.

It isn’t just that food is “made”. Light energy is changed into chemical energy, the energy stored in the bonds of molecules. The carbon compounds made include carbohydrates, lipids, proteins and nucleic acids, so photosynthesis is one of the main ways matter and energy enter living systems.

Why ecosystems depend on it

Most ecosystems depend, in the end, on chemical energy first captured by photosynthesis. Producers use some of their new carbon compounds in respiration, keep some as biomass, and pass some on when they’re eaten. That links photosynthesis directly to primary production, food chains and the carbon cycle.

The effects on ecosystems are large: photosynthesis fixes carbon dioxide into biomass, supplies chemical energy to consumers and decomposers, and — in oxygenic photosynthesis — releases oxygen that supports aerobic respiration.

The simple word equation

Students often remember the equation, then forget where the atoms actually come from. The simple word equation for photosynthesis, with glucose named as the product, is:

carbon dioxide + water → glucose + oxygen

Because the process needs light, light is usually written above the arrow:

carbon dioxide + water —light→ glucose + oxygen

Hydrogen comes from water

Carbon dioxide is a very oxidized carbon compound. To make glucose, photosynthesis has to reduce the carbon from carbon dioxide, so hydrogen and electrons must be supplied. In photosynthesis, that hydrogen comes from splitting water.

Photolysis is a light-driven reaction that splits water molecules into protons, electrons and oxygen. Photosynthesis uses the protons and electrons; the oxygen is not needed for glucose synthesis, so it is released as a by-product.

You do not need a more detailed chemical equation for the standard word equation, but the idea matters: the oxygen released by photosynthesis comes from water, not from carbon dioxide.

Oxygenic photosynthesis

Oxygenic photosynthesis is photosynthesis that releases oxygen, because water is split and used as the hydrogen source. It occurs in plants, algae and cyanobacteria.

The oxygen made is usually waste. In a leaf, oxygen concentration builds up inside chloroplasts, so oxygen diffuses into the cytoplasm, then into the leaf air spaces, and finally out through stomata. In aquatic plants and algae, the oxygen may dissolve in the surrounding water. If the water becomes saturated, bubbles can be seen.

Why this matters beyond one leaf

Over geological time, photosynthetic organisms releasing oxygen changed Earth. Oxygen allowed aerobic respiration to become widespread, and it also caused oxidation reactions in the environment, including the oxidation of dissolved iron in ancient oceans. Biology altered geology here.

Why chromatography separates pigments

Chromatography separates dissolved substances because they move at different rates through a stationary material with a solvent. Photosynthetic pigments do not all dissolve equally well in the solvent, and they do not all stick to the paper or thin-layer coating with the same strength, so they travel different distances.

A leaf extract contains chlorophylls and accessory pigments. In paper chromatography or thin-layer chromatography, a small, concentrated spot of pigment is placed near the bottom of the strip. The solvent must start below the pigment spot. If it doesn’t, the sample simply dissolves into the solvent reservoir. As the solvent front rises, the pigments separate into coloured bands or spots.

Calculating and using Rf values

The retention factor compares the distance a substance moves with the distance moved by the solvent front in chromatography. It is calculated as:

Rf = distance moved by pigment / distance moved by solvent front, where Rf is the retention factor (unitless), distance moved by pigment is the distance from the origin to the centre of the pigment spot (m), and distance moved by solvent front is the distance from the origin to the solvent front (m).

In school practicals, distances are usually measured in millimetres. The units cancel because the value is a ratio. To identify a pigment, use both its colour and its Rf value. For example, a yellow-green band with a moderate Rf value is likely to be different from an orange band close to the solvent front.

Technique makes a real difference: use pencil for the origin line, make the initial spot small and dark, let each application dry before adding more extract, keep the chamber undisturbed, mark the solvent front immediately, and handle organic solvents in a well-ventilated space or fume cupboard.

Pigments and electron excitation

A pigment is a chemical substance that absorbs specific wavelengths of visible light and reflects or transmits others. We see a pigment as the colour of the light it does not absorb strongly; chlorophyll looks green because much green light is reflected or transmitted.

A photon is a discrete packet of electromagnetic radiation that carries energy. When a pigment molecule absorbs a photon, that energy can excite an electron, so the electron is raised to a higher energy level. This step begins the transformation of light energy into chemical energy during photosynthesis.

Pigments absorb only some wavelengths because electron excitation needs particular energy differences. Shorter wavelengths such as blue light have higher-energy photons than longer wavelengths such as red light. If the photon energy does not match an allowed electron transition in the pigment molecule, that wavelength is not absorbed efficiently.

Absorption spectra

An absorption spectrum is a graph showing how strongly a pigment or mixture of pigments absorbs light at different wavelengths. For photosynthetic pigments, the horizontal axis should show wavelength from about 400 nm to 700 nm, with the corresponding colours marked from violet/blue through green/yellow to red. The vertical axis shows absorption, often as a percentage or absorbance.

Chlorophylls absorb strongly in the blue and red regions, but weakly in green. Accessory pigments widen the range of wavelengths that can be harvested. This is one function of pigments in living organisms: they allow light energy to be captured selectively rather than simply converted to heat.

Two spectra that answer different questions

An action spectrum is a graph showing the rate of photosynthesis at different wavelengths of light. It does not measure light absorption directly. Instead, it measures the biological effect of each wavelength.

Absorption and action spectra often peak at similar wavelengths because light absorbed well by photosynthetic pigments tends to drive photosynthesis well. The graphs still won’t be identical. An absorption spectrum may come from one pigment or from an extracted mixture, while an action spectrum measures the whole photosynthetic system in living tissue or cells.

Determining rates and plotting an action spectrum

To make an action spectrum experimentally, expose photosynthesizing material to different wavelengths while controlling other variables such as light intensity, temperature and carbon dioxide concentration. Then measure the rate of photosynthesis. Suitable rate data include oxygen production or carbon dioxide consumption.

For oxygen data, calculate the rate from the change in oxygen amount per unit time. For carbon dioxide data, calculate it from carbon dioxide removed per unit time. Plot wavelength, with colour indicated, on the x-axis and rate of photosynthesis on the y-axis. If the data are scaled to the maximum, the y-axis may show percentage of maximum rate.

Pigments have many functions in living organisms, including light harvesting in photosynthesis, signalling to other organisms, and protection from excess light. Here, the focus is on how pigment absorption translates into photosynthetic action.

Limiting factors and hypotheses

A limiting factor is an environmental factor that restricts the rate of a process because it is in shortest supply relative to demand. In photosynthesis, the usual limiting factors are carbon dioxide concentration, light intensity and temperature.

A hypothesis is a provisional explanation that makes a testable prediction. For example, a hypothesis might predict that increasing carbon dioxide concentration will increase the rate of photosynthesis until another factor becomes limiting. Hypotheses may come before an experiment, based on theory, or after early observations using evidence already collected. Either way, they need repeated testing.

The independent variable is the factor deliberately changed by the investigator. The dependent variable is the measured outcome that responds to the independent variable. In these experiments, the dependent variable is usually a rate of photosynthesis, estimated from oxygen production, carbon dioxide uptake, pH change or indicator colour change.

Varying carbon dioxide concentration

Carbon dioxide concentration can be changed by adding different concentrations of sodium hydrogen carbonate solution to water containing pondweed or immobilized algae. You can estimate oxygen production by counting bubbles, but a gas syringe or oxygen probe is more reliable because bubble size varies.

When carbon dioxide is the independent variable, keep light intensity and temperature constant. If adding more carbon dioxide no longer increases the rate, another factor has become limiting.

Varying light intensity

Light intensity can be changed by moving a lamp to different distances or by using a dimmable light source. A lux meter gives a direct measurement. If distance is used, the relationship is often treated as:

Irel = 1 / d², where Irel is relative light intensity (unitless) and d is distance from the light source (m).

Leaf-disc experiments work well for this. Air is removed from leaf discs so they sink; as photosynthesis produces oxygen, the discs become less dense and rise. The time taken to rise can then be used as an indirect measure of photosynthesis rate. Keep carbon dioxide concentration, temperature, leaf species, disc size and number of discs constant.

Varying temperature

Temperature can be changed using a water bath, heat block or thermostatically controlled vessel. Algae can be used with an oxygen electrode or pH probe linked to a data logger. As photosynthesis removes dissolved carbon dioxide, pH may rise, so pH change can indicate photosynthetic activity.

Temperature affects enzyme-controlled stages of photosynthesis. A typical hypothesis, then, is that the rate rises with temperature up to an optimum and then falls if enzymes or membranes are disrupted.

Why enrich carbon dioxide?

In bright light and suitable temperatures, carbon dioxide often limits photosynthesis. Carbon dioxide enrichment is an experimental method where the carbon dioxide concentration is raised to test its effect on photosynthesis, growth or carbon storage.

Inside enclosed greenhouses, researchers can raise carbon dioxide concentration while keeping close control of temperature, water supply, nutrients and lighting. That makes the method useful for crop production, and for testing whether increased carbon dioxide can increase plant growth under controlled conditions.

FACE experiments

Free-air carbon dioxide enrichment is a field method that raises carbon dioxide concentration around plants growing in open-air conditions. FACE experiments are used because greenhouse results may not predict what happens in crops, forests or natural ecosystems, where wind, soil, pests, competition and seasonal conditions all matter.

In a FACE experiment, towers release carbon dioxide into a plot, and sensors monitor the concentration. Control plots are treated in a similar way but without extra carbon dioxide. A controlled variable is a factor that could affect results but is kept as similar as possible between treatments, such as plant species, soil type, water availability or nutrient status.

The NOS point is worth noticing: careful variable control is often easier in a laboratory, but some biological questions can only be answered in the field. FACE experiments give up perfect control to gain realism.

What these experiments predict

Carbon dioxide enrichment experiments help test whether rising atmospheric carbon dioxide will increase future photosynthesis and plant growth. The answer is not simply “yes”. Extra carbon dioxide may increase carbon fixation at first, but growth may later be limited by nitrogen, phosphorus, water, temperature or ecological interactions.

That connects back to the ecosystem consequence of photosynthesis: if plant biomass increases, more carbon may be stored temporarily in ecosystems; if other limiting factors prevent growth, increased atmospheric carbon dioxide will not be fully offset by photosynthesis.

What a photosystem is

A photosystem is a membrane-bound molecular array of chlorophyll, accessory pigments and proteins that absorbs light energy and emits excited electrons from a reaction centre. You find photosystems only in membranes: in the thylakoid membranes of cyanobacteria and in the chloroplast thylakoid membranes of photosynthetic eukaryotes.

A photosystem isn’t a single pigment molecule working by itself. It’s an organized array. Many pigment molecules collect light, then pass the energy through the array to a special chlorophyll molecule at the reaction centre.

Excited electrons and emission

When a pigment absorbs a suitable photon, one of its electrons becomes excited. In ordinary pigment molecules, that energy may be lost again as light or heat. Inside a photosystem, the energy is funnelled to the reaction centre, where the special chlorophyll molecule emits an excited electron to an electron acceptor.

Plants and algae have two photosystems in their chloroplasts. Photosystem II supplies excited electrons to an electron transport pathway and replaces them using electrons from water. Photosystem I emits excited electrons that are used to reduce NADP.

Why the array matters

The structured array gives three major advantages.

First, having many pigment molecules raises the chance of capturing photons. One chlorophyll molecule would intercept light too rarely to run photosynthesis at a useful rate.

Second, different pigments absorb different wavelength ranges. Chlorophylls absorb strongly in red and blue light, while accessory pigments extend the useful range. Because of that mix, the array harvests more of the visible spectrum than one pigment type could on its own.

Third, the positions and orientations of pigment molecules help excitation energy move efficiently towards the reaction centre. If the pigments were randomly arranged, much of the absorbed energy would be lost before it could generate an emitted electron.

Interdependence inside the photosystem

A single molecule of chlorophyll, or any other pigment, cannot perform photosynthesis by itself. It can absorb light, but it cannot maintain a controlled electron flow, replace lost electrons, or connect electron emission to later chemical reactions. The photosystem works because its parts depend on each other: pigments, proteins, reaction centre and electron acceptors only make sense as a structured membrane system.

Photolysis in photosystem II

In photosystem II, light absorbed by the reaction-centre chlorophyll makes it emit excited electrons. These electrons have to be replaced. Water supplies the replacement electrons through photolysis.

Photolysis of water is a light-dependent reaction where water splits to produce electrons, protons and oxygen. The reaction can be summarized as:

2H₂O → O₂ + 4H⁺ + 4e⁻

The electrons replace those emitted by photosystem II. The protons help build the proton gradient inside the thylakoid. Oxygen is a waste product and diffuses away.

Consequences for life and Earth

Oxygen generation by photolysis had enormous consequences once it evolved. Oxygen accumulated only after reduced materials, such as dissolved iron, had been oxidized. This produced geological evidence such as banded iron formations.

When oxygen became available, aerobic respiration could evolve and later become widespread. Photosynthesis therefore did more than add oxygen to the air; it changed which metabolic pathways were possible and reshaped ecosystems.

Building the proton gradient

Chemiosmosis is the mechanism of ATP synthesis where protons diffuse through ATP synthase down an electrochemical gradient, releasing energy to phosphorylate ADP. In thylakoids, this gradient sits across the thylakoid membrane: proton concentration is high in the thylakoid space and lower in the stroma.

Two processes build the gradient. Photolysis adds protons to the thylakoid space. As electrons pass along a chain of carriers, protons are also pumped from the stroma into the thylakoid space. The thylakoid space is small, so a steep gradient can form quickly.

ATP synthase and photophosphorylation

ATP synthase is a membrane enzyme that catalyses ATP formation using the energy released when protons move through it. Protons flow back into the stroma through ATP synthase, phosphorylating ADP to ATP.

Photophosphorylation is ATP production using light-driven electron flow to generate the proton gradient. In non-cyclic photophosphorylation, electrons start at photosystem II and eventually help reduce NADP. In cyclic photophosphorylation, electrons emitted from photosystem I return through electron carriers to photosystem I; proton pumping and ATP production still occur, but reduced NADP is not produced.

NADP as the electron acceptor

NADP is a soluble coenzyme. During photosynthesis, it accepts electrons and a proton, becoming reduced NADP. Here, I’ll use the pair “NADP” and “reduced NADP” consistently.

Photosystem I absorbs light, then its reaction centre emits excited electrons. These electrons pass to an enzyme on the stroma side of the thylakoid membrane, where NADP is reduced.

NADP accepts two electrons from photosystem I and one hydrogen ion from the stroma:

NADP + 2e⁻ + H⁺ → reduced NADP

Reduced NADP carries reducing power to the Calvin cycle. There, it helps convert glycerate 3-phosphate into triose phosphate.

Keeping electron flow going

Photosystem I must replace the electrons it loses. Electrons emitted from photosystem II pass along carriers, then supply photosystem I. This connects the two photosystems in non-cyclic electron flow.

When NADP is not available because most of it has already been reduced, electrons from photosystem I can cycle back through carriers instead. This cyclic route helps generate ATP without producing more reduced NADP.

Thylakoid structure and function

A thylakoid is a membrane-bound sac containing the components for the light-dependent reactions of photosynthesis. In chloroplasts, stacks of disc-shaped thylakoids are called grana, while the unstacked connecting membranes are called stroma lamellae. Cyanobacteria have thylakoid membranes too, although these are not inside chloroplasts.

The thylakoid works as a system because its membrane, internal space and surrounding stroma form separate compartments. This compartmentalization allows electron flow, proton accumulation and ATP synthesis to happen.

Where the light-dependent reactions occur

Photolysis of water takes place at photosystem II on the thylakoid-space side of the membrane. Protons are released into the thylakoid space, and oxygen diffuses away.

ATP synthesis by chemiosmosis happens at ATP synthase in the thylakoid membrane. Protons move from the thylakoid space to the stroma, and ATP is released into the stroma.

Reduction of NADP occurs on the stroma side of the thylakoid membrane after electrons are emitted from photosystem I. The location fits the job: reduced NADP is needed in the stroma for the Calvin cycle.

The first step of the Calvin cycle

Carbon fixation converts inorganic carbon dioxide into an organic carbon compound. In chloroplasts, it takes place in the stroma during the Calvin cycle.

The enzyme involved is Rubisco. It catalyses the reaction between carbon dioxide and ribulose bisphosphate. Ribulose bisphosphate, usually shortened to RuBP, is a five-carbon sugar phosphate that accepts carbon dioxide in the Calvin cycle.

The substrates are RuBP and CO₂. The product is glycerate 3-phosphate, often shortened to GP, a three-carbon phosphorylated organic acid. One carbon dioxide molecule combines with one RuBP molecule, and the six-carbon intermediate formed then immediately produces two GP molecules.

Why so much Rubisco is needed

Rubisco is usually described as the most abundant enzyme on Earth. Not because it’s especially fast. It is abundant because it works relatively slowly and is not very effective when carbon dioxide concentration is low. For that reason, chloroplast stroma contains high concentrations of Rubisco, keeping carbon fixation moving at a useful rate.

This is one reason carbon dioxide can limit photosynthesis, especially when light and temperature are favourable.

Reducing GP to TP

Triose phosphate, often shortened to TP, is a three-carbon sugar phosphate made in the Calvin cycle. GP is converted into TP using ATP and reduced NADP from the light-dependent reactions.

ATP provides the energy for this conversion. Reduced NADP provides the electrons and hydrogen needed to reduce GP. Chemically, GP has a carboxyl group, while TP has a more reduced aldehyde-type sugar structure.

This stage takes place in the stroma. It is called light-independent because the reaction does not use light directly, but the name can be misleading: in darkness, it stops quickly because ATP and reduced NADP run out.

Why RuBP must be regenerated

The Calvin cycle keeps going only if RuBP is replaced after it reacts with carbon dioxide. If every TP molecule left the cycle to form glucose or starch, RuBP would run out and carbon fixation would stop.

The stoichiometry is simple but worth remembering: five molecules of TP are converted into three molecules of RuBP. ATP is needed for this regeneration. You don’t need to know the individual enzyme steps.

The five-sixths rule

Treat glucose as the product of photosynthesis, and six carbon atoms have to be fixed. Each turn of the Calvin cycle fixes one carbon atom from carbon dioxide. To keep the cycle running, five-sixths of the TP produced must be recycled to regenerate RuBP. That leaves only one-sixth as a net gain for making glucose and other carbon compounds.

That’s why the Calvin cycle is a cycle, not a straight pathway: RuBP is consumed and then regenerated.

Calvin cycle products feed metabolism

Simple photosynthesis equations often put glucose as the final product. In real photosynthesizing cells, though, the output goes well beyond glucose. All carbon in the carbon compounds of photosynthesizing organisms can be traced back to carbon fixed in the Calvin cycle.

TP and GP act as starting points for metabolic pathways. TP can be used to make hexose phosphates, which can then form glucose, sucrose, starch or cellulose. Some carbon compounds leave the chloroplast; others are kept there temporarily, for example as starch grains.

Beyond carbohydrates

Photosynthesizing organisms also produce lipids, amino acids, nucleotides and many other carbon compounds. You don’t need these pathways in detail, but the logic is worth keeping straight: carbon skeletons come from Calvin cycle intermediates, or from respiratory pathway intermediates that ultimately trace back to Calvin cycle carbon.

Mineral nutrients are inorganic ions absorbed by organisms that supply elements needed for metabolism. Nitrate or ammonium ions provide nitrogen for amino acids. Phosphate is needed for nucleotides, ATP and phosphorylated intermediates. Sulfate supplies sulfur for some amino acids. Photosynthesis provides the carbon framework; mineral nutrition provides elements that carbon dioxide and water cannot.