Bioenergetics: Biology unit 4 notes (FBISE class 11)

4Bioenergetics

Living things cannot grow, reproduce or show any of the characteristics of life without a ready and continuous supply of energy. Every metabolic reaction involves an energy transformation, so the study of how energy is converted and how energy relationships work in living systems has its own name.

Bioenergetics: The quantitative study of energy conversions and energy relationships in biological systems.

This chapter deals with the two most fundamental bioenergetic processes, photosynthesis and respiration. You met their outline in classes IX and X; here the detailed steps are learnt so that they can be understood and applied to real life. The chapter ends with photorespiration, a process that lowers the productivity of plants.

4.1 Photosynthesis: light energy is trapped and stored in glucose (the light dependent phase makes ATP and NADPH, the light independent phase uses them to make sugar).
4.2 Cellular respiration: the energy stored in glucose and other foods is released and trapped in ATP (glycolysis, link reaction, Krebs cycle, electron transport chain).
4.3 Photorespiration: a wasteful side reaction of RuBisCO in green cells, and C4 photosynthesis as an adaptation against it.

4.1PhotosynthesisOften examined

Chemically, photosynthesis is a redox process. Carbon dioxide (an oxidised form of carbon) is reduced into glucose (a reduced form of carbon). Water acts as the reducing agent: it is oxidised into oxygen. In terms of energy, photosynthesis is an energy conversion process in which energy-poor molecules ( $CO X_{2}$ and $H X_{2} O$ ) are changed into an energy-rich molecule, glucose. The extra energy is absorbed from sunlight by the photosynthetic pigments.

Photosynthesis: The process in which light energy absorbed by photosynthetic pigments is used to convert carbon dioxide and water into glucose (an energy-rich organic molecule), with the release of oxygen.

The overall reaction can be summarised as follows.

6 CO X_{2} + 12 H X_{2} O light chlorophyll C X_{6} H X_{12} O X_{6} + 6 O X_{2} + 6 H X_{2} O

Overall reaction of photosynthesis: carbon dioxide is reduced, water is oxidised.

The process involves the interaction of sunlight, pigments, water and carbon dioxide. The next sections take these four in turn (4.1.1 to 4.1.6) before the mechanism itself is described (4.1.7 to 4.1.10).

4.1.1Role of lightOften examined

Biological energy transformations obey the laws of thermodynamics. Sunlight is the ultimate source of energy, flowing continuously one way from the Sun to life and the ecosystem. It is an electromagnetic form of energy. Light matters most because light energy is converted to chemical energy and then to the other forms of energy needed for life processes. This trapped light energy is used to convert low-energy inorganic compounds into high-energy organic compounds.

Electromagnetic spectrum: The full range of electromagnetic radiation in the universe.

Visible light is only a small part of the spectrum, between 380 nm and 750 nm. It is not only seen by the naked eye but is also the part effective for photosynthesis.

Figure 4.1: Electromagnetic spectrum. Visible light (accent) is a very narrow band between ultraviolet and infrared; it is enlarged below (not to scale).

The effectiveness of a particular wavelength for photosynthesis depends mainly on how much of it is absorbed in the plant body. Different wavelengths (colours) of visible light are absorbed differently by the various photosynthetic pigments, so each wavelength has its own effectiveness. If a plant is lit with one colour of light at a time, the rate of photosynthesis can be measured for each colour and the data plotted on a graph.

Action spectrum: A graph which shows the effectiveness of different wavelengths of light for the process of photosynthesis (relative photosynthetic rate plotted against wavelength).

Action spectrum of photosynthesis

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430 nm (blue)

670 nm (red)

action spectrum

Figure 4.2: Action spectrum of photosynthesis. The rate is highest in blue light (about 430 nm) and red light (about 670 nm) and lowest in the green-yellow middle of the spectrum.

4.1.2Role of carbon dioxide in photosynthesis

Carbon dioxide is one of the raw materials of photosynthesis. It acts as the carbon source for the synthesis of organic compounds. Plants are therefore called autotrophs: they use a low-energy inorganic compound such as $CO X_{2}$ to make high-energy organic compounds by themselves. Carbon dioxide is used in the dark (light independent) reaction, the Calvin cycle.

Air contains about 0.03 to 0.04 percent carbon dioxide. Land plants use this atmospheric $CO X_{2}$ .
Increasing the $CO X_{2}$ concentration in the environment increases the rate of photosynthesis.
When the $CO X_{2}$ concentration rises above 1 percent, the rate of photosynthesis slows down rapidly because the stomata close.
Water contains dissolved carbon dioxide, bicarbonates and carbonates. Aquatic photosynthetic organisms use these as their carbon source.

4.1.3Role of water in photosynthesisOften examined

Water is the other raw material of photosynthesis. It acts as the hydrogen and electron donor. In the light dependent phase a water molecule is broken down into $2 H X^{+}$ , $2 e X^{-}$ and $\frac{1}{2} O X_{2}$ . Because the splitting needs light, the reaction is called photolysis of water.

H X_{2} O photolysis 2 H X^{+} + 2 e X^{-} + \frac{1}{2} O X_{2}

The electrons from water replace the electrons lost by the photo-excited chlorophyll a in the reaction centre of photosystem II (P680).
The $H X^{+}$ ions and electrons are taken up temporarily by $NADP X^{+}$ to become NADPH, which is then used to reduce $CO X_{2}$ in the light independent reaction.
The oxygen produced is released into the atmosphere.

Evidence that the oxygen comes from water

The role of water was first reported by Van Niel in 1931. He worked on photosynthesis in bacteria, which make carbohydrates from carbon dioxide but do not release oxygen. He hypothesised that plants split water as the source of hydrogen, releasing oxygen as a by-product.

The hypothesis was confirmed in 1940, when the heavy isotope $X_{18} X_{2}^{218} O$ was used in biological research for the first time.

The two experiments that proved the source of oxygen.
	Experiment 1	Experiment 2
Labelled compound supplied	Water made of $X_{18} X_{2}^{218} O$ ( $H X_{2} X_{18} X_{2}^{218} O$ ) added to an alga suspension	Carbon dioxide with $X_{18} X_{2}^{218} O$ ( $C X_{18} X_{2}^{218} O X_{2}$ )
Result	The oxygen released was radioactive (it contained $X_{18} X_{2}^{218} O$ )	The oxygen released contained none of the isotope
Conclusion	The oxygen comes from water	The oxygen does not come from carbon dioxide

6 CO X_{2} + 12 H X_{2} X_{18} X_{2}^{218} O light chlorophyll C X_{6} H X_{12} O X_{6} + 6 H X_{2} O + 6 X_{18} X_{2}^{218} O X_{2}

4.1.4Photosynthetic pigments and their major typesOften examined

Pigment: Any substance that absorbs light energy for use in the food-making (photosynthesis) process.

Photosynthetic organisms have different types of pigments for absorbing light. Algae and photosynthetic bacteria have chemically different pigments from plants. In bacteria the pigments lie in the cytoplasm, whereas in plants they are embedded in the thylakoid membranes (grana lamellae) inside the chloroplasts. Higher plants have two major groups of pigments: chlorophylls and carotenoids.

Chlorophyll

Chlorophylls absorb mainly violet, blue, orange and red wavelengths. Green and yellow are least absorbed and are reflected. The two major types are chlorophyll a and chlorophyll b. Algae also have other chlorophylls, Chl-c, Chl-d and Chl-e, while photosynthetic bacteria have bacteriochlorophyll.

Chlorophyll a and chlorophyll b.
Feature	Chlorophyll a	Chlorophyll b
Colour	Bluish green	Yellowish green
Molecular formula	$C X_{55} H X_{72} O X_{5} N X_{4} Mg$	$C X_{55} H X_{70} O X_{6} N X_{4} Mg$
Group on the 2nd pyrrole ring	Methyl group ( $- CH X_{3}$ )	Aldehyde group ( $- CHO$ )
Where found	All photosynthetic organisms except photosynthetic bacteria	All photosynthetic organisms except brown algae, red algae and photosynthetic bacteria
Role	Reaction centre pigment; converts light energy to chemical energy	Accessory pigment; passes absorbed energy to chlorophyll a

Structure of chlorophyll

A chlorophyll molecule consists of a head and two tails.

Head: a porphyrin ring with an atom of magnesium ( $Mg$ ) at the centre. The ring is made of four pyrrole rings (each pyrrole ring has four carbons and one nitrogen). The nitrogen atoms of the pyrrole rings interact with the central Mg atom, and different groups are attached around them. The head is hydrophilic, lies on the surface of the thylakoid membrane and is the light-absorbing part.
Tails: the two side chains are phytol and methyl ester. The tails are hydrophobic, are embedded in the thylakoid membrane and anchor the chlorophyll molecule there.
Only difference between chlorophyll a and b: chlorophyll a has a methyl group ( $- CH X_{3}$ ) on the 2nd pyrrole ring, whereas chlorophyll b has an aldehyde group ( $- CHO$ ) at this point.

Figure 4.3: Structure of chlorophyll. The group on the second ring (accent) is the only difference between chlorophyll a and b.

The amount of chlorophyll in a leaf is kept up for continuous photosynthesis, so plants keep making it. Its synthesis needs **sunlight, nitrogen, magnesium ions ( $Mg X^{2 +}$ ) and warm temperatures, so leaves make more chlorophyll in summer. A deficiency of chlorophyll in leaves is called chlorosis**.

Carotenoids

Carotenoids are terpenoid lipids that are yellow, orange, red or brown pigments. They absorb light strongly in the blue-violet range. They are seen in leaves before leaf fall and are present in some flowers and fruits. They act as accessory pigments along with chlorophyll b: they absorb light energy and then pass it to chlorophyll a. They also protect chlorophyll a from excess light, and they attract insects and birds for pollination and dispersal.

There are two types of carotenoids: carotenes and xanthophylls.

Carotenes and xanthophylls (the two types of carotenoids).
Feature	Carotenes	Xanthophylls
Colour	Orange-red	Yellow
Built from	Isoprenoid units	Isoprenoid units
Occurrence	All photosynthetic eukaryotes	Widely distributed in plants
Example	$β$ (beta) carotene, the most widespread and important carotene	Lutein, which gives the yellow colour of foliage in autumn

4.1.5Role of photosynthetic pigmentsOften examined

Light energy can be used by a green plant cell only when it is absorbed. Different pigments absorb particular ranges of wavelengths of visible light. All the wavelengths absorbed by a pigment disappear; a pigment shows only the wavelengths it reflects. Plants look green because chlorophyll does not absorb green light but reflects it.

The relative ability of different pigments to absorb light is measured with an instrument called the spectrophotometer.

Two roles

Absorption: the primary role of the pigment molecules is to absorb electromagnetic radiation and then transmit the energy of the absorbed photons to the reaction centre in the photosystem.
Conversion: pigments in the photosystems also take part in the photochemical conversion of light energy into chemical energy. The chief component of a photosystem is the chlorophyll a molecule, which can absorb light and directly convert it into chemical energy in the form of ATP and NADPH.

Other pigments (chlorophyll b, xanthophylls and carotenes) are called accessory pigments because they can only absorb light energy; they cannot convert it to chemical energy. They absorb wavelengths other than those absorbed by chlorophyll a and pass the energy on to chlorophyll a for conversion. In this way they help chlorophyll a in the reaction centre to produce more chemical energy than it could by absorbing light itself. Some carotenoids also protect chlorophyll from intense light by absorbing and dispersing the excess light energy instead of passing it on.

Accessory pigments pass the light energy they absorb to chlorophyll a, which converts it to chemical energy.

Primary and accessory pigments.
	Chlorophyll a (primary pigment)	Accessory pigments
Pigments	Chlorophyll a	Chlorophyll b, xanthophylls, carotenes
Absorbs light	Yes	Yes, at wavelengths other than those chlorophyll a absorbs
Converts light into chemical energy (ATP, NADPH)	Yes	No; they pass the energy to chlorophyll a
Position in photosystem	Reaction centre	Antenna complex

4.1.6Absorption spectrumOften examined

How much light of each wavelength a particular pigment absorbs is measured with a spectrophotometer, and the data are shown as a graph.

Absorption spectrum: A graph which shows the relative absorption of different wavelengths of light by a particular pigment.

Absorption spectra of different pigments

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chlorophyll a

chlorophyll b

carotenoids

Figure 4.4: Absorption spectra of chlorophyll a, chlorophyll b and the carotenoids. Chlorophylls absorb most in violet-blue and orange-red; carotenoids absorb most between about 430 and 500 nm.

The absorption spectra of different pigments show that they absorb different wavelengths of visible light, and not at the same rate.

The main photoreceptors, chlorophyll a and b, both absorb more in the violet-blue region (400 to 470 nm) and the orange-red region (630 to 660 nm).
Carotenoids absorb more at 430 to 500 nm.
The difference between the absorption spectra of chlorophyll a and b may be related to the differences in their molecular formulae, structural arrangement and physical properties, for example colour.

Absorption spectra of chlorophyll a and chlorophyll b.
Feature	Chlorophyll a	Chlorophyll b
Absorbs most in	Violet-blue and orange-red	Violet-blue and orange-red
Violet-blue peak	At the shorter wavelength end of the region (about 420 nm)	At the longer wavelength end of the region (about 460 nm)
Red peak	At the longer wavelength end (about 660 nm)	At the shorter wavelength end (about 640 nm)
Cause of the difference	Methyl group on the 2nd pyrrole ring	Aldehyde group on the 2nd pyrrole ring

Worked example: why is a healthy leaf green?

Using Figure 4.4 and the action spectrum (Figure 4.2), explain why a healthy leaf looks green even though chlorophyll is its main pigment, and which wavelengths drive photosynthesis most.

Read the absorption spectra: chlorophyll a and b absorb strongly in violet-blue (400 to 470 nm) and orange-red (630 to 660 nm). Carotenoids absorb at 430 to 500 nm.
Look at the middle of the visible range (about 500 to 600 nm, green and yellow). All three pigments absorb very little there.
A pigment shows the colours it does not absorb. The green-yellow light is reflected back to the eye, so the leaf looks green.
The action spectrum has its peaks in the same two regions, blue (430 nm) and red (670 nm), and is lowest in the green. So the wavelengths that are absorbed are the ones that drive photosynthesis.

Region	Absorbed by pigments?	Effect on photosynthesis
Violet-blue (about 430 nm)	Strongly (Chl a, Chl b, carotenoids)	High (peak)
Green-yellow (500 to 600 nm)	Very little	Low; reflected, so the leaf looks green
Orange-red (about 670 nm)	Strongly (Chl a, Chl b)	High (peak)

Absorption spectrum and action spectrum.
Absorption spectrum	Action spectrum
Graph of the relative absorption of wavelengths by a particular pigment	Graph of the effectiveness of wavelengths for photosynthesis
Plots percent of light absorbed against wavelength	Plots relative photosynthetic rate against wavelength
Measured with a spectrophotometer	Measured by lighting the plant with one colour at a time and measuring the rate of photosynthesis
Different for each pigment (chlorophyll a, b, carotenoids)	One curve for the whole plant; peaks at blue (430 nm) and red (670 nm)

4.1.7Arrangements of pigments (photosystems)Often examined

For efficient absorption and use of light energy, the photosynthetic pigments are embedded in the surface of the thylakoid membranes, where they are arranged in clusters of pigment molecules.

Photosystem: A cluster of pigment molecules on the thylakoid membrane that absorbs light and passes its energy to a reaction centre.
Antenna complex: The peripheral part of a photosystem. It consists of accessory pigments such as chlorophyll b and carotenoids.
Reaction centre: The central part of a photosystem. It contains only chlorophyll a and associated proteins.

Figure 4.5: Structure of a photosystem. Light energy absorbed by the antenna pigments is passed to the reaction centre chlorophyll a (accent), which gives up an electron to the primary electron acceptor.

Two kinds of photosystem

Chlorophyll a on its own has an optimal absorption wavelength of about 660 nm. In each type of photosystem it associates with different proteins, which slightly shift its optimal wavelength. This produces two distinct photosystems, photosystem I (PS-I) and photosystem II (PS-II).

Photosystem I and photosystem II.
Feature	Photosystem I (PS-I)	Photosystem II (PS-II)
Reaction centre chlorophyll a	Absorbs a maximum of 700 nm light, so it is called P700	Absorbs a maximum of 680 nm light, so it is called P680
Electrons are replaced from	PS-II (via the electron transport chain)	Water (photolysis of water)
Passes its excited electrons to	Ferredoxin and then NADP $^{+}$ (non-cyclic), or back to the chain (cyclic)	Plastoquinone and the cytochrome complex, towards PS-I
Used in	Both cyclic and non-cyclic photophosphorylation	Non-cyclic photophosphorylation only

How a photosystem works

A photon is absorbed by a pigment molecule of the antenna complex.
The pigments of various wavelengths pass the gained energy from one pigment to the next towards the reaction centre.
When the energy reaches chlorophyll a of the reaction centre, its electrons become so energetic that they escape and are accepted by the first molecule of a nearby electron transport chain, which is at a higher energy level than chlorophyll.
The chlorophyll molecule has lost an electron, so it is now oxidised.

Chlorophyll (reduced form) light energy Chlorophyll (oxidised form) + e^{-}

Chlorophyll a can absorb only a narrow range of wavelengths. By working with the antenna pigments it gains energy from a much larger part of the spectrum. The electron transport system then plays the key role in making ATP by changing light energy into chemical energy.

Antenna complex and reaction centre.
Antenna complex	Reaction centre
Peripheral part of the photosystem	Central part of the photosystem
Made of accessory pigments (chlorophyll b, carotenoids)	Contains only chlorophyll a with associated proteins
Absorbs light and passes the energy inwards	Receives the energy; chlorophyll a loses electrons to the primary electron acceptor
Cannot convert light energy to chemical energy	Starts the conversion of light energy to chemical energy

4.1.8Overall mechanism of photosynthesisOften examined

Photosynthesis is divided into two phases.

Light dependent phase (light reaction): it can take place only in the presence of light and occurs in the thylakoid membranes. Light is used to make ATP (the assimilating power) and NADPH (the reducing power). Water is the input and oxygen the output.
Light independent phase (dark reaction): it takes place whether light is present or not, in the stroma. It needs the products of the light reaction, ATP and NADPH. Because these products are available during the day, the dark reaction also occurs in the daytime. \ce{CO2} is the input and is converted into glyceraldehyde 3-phosphate (G3P), the output of this phase.

Figure 4.6: An overview of photosynthesis in a chloroplast. The light reactions in the thylakoids supply ATP and NADPH (accent) to the Calvin cycle in the stroma, which returns ADP, Pi and NADP^+.

In the light independent phase the ATP is hydrolysed to ADP and Pi and its energy is built into the product, whereas NADPH provides energised electrons and hydrogen for the formation of G3P, which is an energy-rich molecule.

Light dependent and light independent reactions.
Light dependent reaction	Light independent reaction
Takes place only in the presence of light	Takes place in light or in darkness (but needs the products of the light reaction)
Occurs in the thylakoid membranes of the grana	Occurs in the stroma
Needs light, water, \mathrm{ADP}+\mathrm{P_i} and \mathrm{NADP^+}	Needs \ce{CO2}, ATP and NADPH
Photolysis of water; \ce{O2} is released	\ce{CO2} is fixed and reduced (Calvin cycle)
Light energy is converted into chemical energy as ATP and NADPH	ATP and NADPH energy is used to make G3P (sugar)

4.1.9Light dependent phase (light reaction)Often examined

The light dependent phase involves the absorption of light by the photosystems, excitation and flow of electrons through an electron transport chain, chemiosmotic synthesis of ATP and reduction of \ce{NADP+} to NADPH. The flow of excited electrons through the chain is of two types: non-cyclic and cyclic.

In non-cyclic flow the excited electrons that leave a photosystem do not come back. After losing their energy they are built into another molecule.
In cyclic flow the excited electrons leave a photosystem and finally return to the same photosystem.

The most important event of the light reaction is the production of ATP.

\ce{ADP + P_i ->[\text{light}] ATP}

Photophosphorylation: The production of ATP during the light reaction (ATP is made from ADP and Pi using light energy).
Chemiosmosis: The mechanism by which ATP is made in photophosphorylation: flow of electrons, pumping of protons and the generation of ATP by the thylakoid membranes.

(a) Non-cyclic photophosphorylation

This is the main pathway of the light reaction in higher plants and occurs routinely. Both photosystems (PS-I and PS-II) are used and two electron transport chains are involved. Excited electrons from PS-II flow through a chain to PS-I; the electrons liberated from PS-I are finally accepted by \ce{NADP+}. This is why it is called non-cyclic. The events are continuous but are described in steps for convenience.

Light is absorbed by PS-II and its electrons are excited. Just two photons strike the antenna complex of PS-II; the energy moves between the pigments to the reaction centre (P680), which excites two electrons. The primary electron acceptor of PS-II captures them and leaves two "electron holes", which makes chlorophyll a strong oxidising agent.
Photolysis of water fills the electron holes. Water reacts with the oxidised chlorophyll and splits into 2\ce{H+}, 2\ce{e-} and \tfrac{1}{2}\ce{O2}. The electrons fill the holes of PS-II.
Electrons flow from PS-II to PS-I. They pass from the primary electron acceptor to plastoquinone (PQ), then through the cytochrome complex (Cyt b_6 and Cyt f). The cytochrome complex is both an electron carrier and a proton pump: the flow of electrons makes it pump protons from the stroma into the thylakoid inner space (lumen).
A proton gradient forms. The energy of the flowing electrons is converted into a gradient of \ce{H+} in the thylakoid space. The gradient activates the enzyme ATP synthase in the thylakoid membrane, which lets protons flow back to the stroma and joins ADP and Pi to form ATP (photophosphorylation). The whole mechanism (electron flow, proton pumping and ATP formation) is chemiosmosis. This ATP supplies energy for sugar synthesis in the Calvin cycle.
The electrons, having lost their energy, pass from the cytochrome complex to plastocyanin (PC) and finally into PS-I.
Light is absorbed by PS-I. When P700 in the reaction centre absorbs two photons, its electrons are boosted to a higher energy level and passed to the primary electron acceptor of PS-I, creating electron holes. The holes of P700 are filled by the electrons that arrive from P680 (PS-II) through the transport chain.
Electrons flow from PS-I to \ce{NADP+}. The primary acceptor passes them to a second transport chain; they are accepted by ferredoxin (Fd), an iron-containing protein. The enzyme NADP reductase (a flavoprotein) transfers the electrons from Fd to \ce{NADP+}, which combines with electrons and hydrogen ions to form NADPH (reduced). NADPH provides the reducing power for sugar synthesis in the Calvin cycle.

\ce{NADP+ + 2e- + 2H+ -> NADPH + H+}

Figure 4.7: Chemiosmotic synthesis of ATP during the light reaction. Electron flow is shown in accent, proton movement in grey.

The path taken by the electrons through the two photosystems in non-cyclic photophosphorylation is called the Z-scheme, because of its zigzag shape when drawn against the energy level of the electrons.

Figure 4.8: Non-cyclic photophosphorylation (Z-scheme). Each photosystem raises the electrons to a higher energy level; they lose energy on the way down, which is used to make ATP.

4.1.9 (b)Cyclic photophosphorylationOften examined

A rise in NADPH and a shortage of ATP may cause the chloroplast to shift for a while from non-cyclic to cyclic electron flow, until the supply of ATP catches up with the demand. In this pathway only PS-I is used.

PS-I absorbs energy from photons. When the energy reaches the reaction centre of PS-I (P700), the electrons are boosted to a higher energy level.
The excited electrons are first captured by the primary electron acceptor of PS-I.
They then move through an electron transport chain containing **ferredoxin, the cytochrome b_6-f complex and plastocyanin**.
When electrons pass through the cytochrome b_6-f complex, ATP is generated by chemiosmosis.
Finally the electrons, having lost energy, return to P700 chlorophyll in the PS-I reaction centre.

Figure 4.9: Cyclic electron flow during photophosphorylation. The electrons (accent) return to P700, so only ATP is produced.

Non-cyclic and cyclic photophosphorylation.
Non-cyclic photophosphorylation	Cyclic photophosphorylation
Main pathway of the light reaction in higher plants	Used for a while when NADPH is high and ATP is short
Both PS-II and PS-I are used	Only PS-I is used
Two electron transport chains	One cycle through ferredoxin, cytochrome complex and plastocyanin
Electrons do not return; they end up in NADPH (Z-scheme)	Electrons return to P700 of PS-I
Water is split (photolysis); \ce{O2} is released	No photolysis of water; no \ce{O2}
Products: ATP and NADPH (and \ce{O2})	Product: ATP only

Worked example: why does cyclic flow give no oxygen and no NADPH?

Explain why cyclic photophosphorylation produces no \ce{O2} and no NADPH, but still makes ATP.

Oxygen comes only from the photolysis of water, which is needed to refill the electron holes of PS-II. In cyclic flow PS-II is not used, and the electrons return to P700, so there are no holes to refill and water is not split. No water splitting means no \ce{O2}.
NADPH is made when electrons from PS-I are handed to \ce{NADP+} by ferredoxin and NADP reductase. In cyclic flow the electrons go back to P700 instead, so they never reach \ce{NADP+} and no NADPH forms.
ATP is still made because the electrons pass through the cytochrome b_6-f complex, which pumps protons into the thylakoid space; the proton gradient drives ATP synthase (chemiosmosis).
So the plant can raise its ATP supply when ATP is short, without making more NADPH.

4.1.10Light independent phase (dark reaction)Often examined

The light independent phase (dark reaction) gets its name from the fact that light is not directly required for these reactions. It needs a supply of NADPH and ATP (the products of the light reaction) and **\ce{CO2}**.

NADPH is used to reduce carbon dioxide.
ATP is used to supply energy.
\ce{CO2} is finally converted into a phosphorylated triose sugar, glyceraldehyde 3-phosphate (G3P), which is later used to make glucose.

The dark reaction generally involves a complicated metabolic pathway, the Calvin cycle (the C3 pathway). In some plants a second pathway, the C4 pathway, is also involved (section 4.3.4). Plants in which only the Calvin cycle operates during the dark reaction are called C3 plants.

4.1.10Calvin cycleOften examined

The term Calvin cycle refers to the series of metabolic reactions in which \ce{CO2} is reduced to produce G3P. They were explored by Melvin Calvin and co-workers at the University of California, and Calvin won the Nobel Prize in 1961 for this work. The cycle has three phases: carbon fixation, reduction, and regeneration of the carbon dioxide acceptor (RuBP).

Phase 1: carbon fixation

The key substance is ribulose bisphosphate (RuBP), a five-carbon phosphorylated sugar. It is called the **\ce{CO2} acceptor because it combines with carbon dioxide with the help of the enzyme ribulose bisphosphate carboxylase/oxygenase (RuBisCO). Three intermediate molecules of six carbons are formed; they are unstable, exist for a very short time and cannot be isolated. Each six-carbon molecule breaks down into two molecules of 3-phosphoglycerate (3-PGA)**, a phosphate-containing three-carbon compound.

Since the carbon of inorganic \ce{CO2} becomes part of an organic compound (RuBP) in this phase, it is called carbon fixation. Because the first stable compound is a three-carbon compound (3-PGA), the Calvin cycle is also known as the C3 pathway.

Phase 2: reduction

Six molecules of 3-PGA react with six ATP molecules: a phosphate from each ATP is transferred to each 3-PGA, changing it into 1,3-bisphosphoglycerate. These are then reduced by the hydrogen of NADPH and a phosphate group is given off from each. The result is glyceraldehyde 3-phosphate (G3P). Six molecules of G3P are produced in this phase, but only one leaves the cycle; the other five are used to regenerate the \ce{CO2} acceptor.

Phase 3: regeneration of the \ce{CO2} acceptor

The five G3P molecules (three carbons each) undergo a series of reactions that produce three molecules of ribulose phosphate (RuP), five carbons each. Three RuP react with three ATP: a phosphate from each ATP is transferred to each RuP. The RuP are thus changed into RuBP, which again takes part in the next cycle.

Figure 4.10: Calvin cycle (for three \ce{CO2} fixed). Only one of the six G3P molecules leaves the cycle (accent).

The three phases of the Calvin cycle.
Phase	What happens	Inputs and outputs (3 \ce{CO2} fixed)
1. Carbon fixation	\ce{CO2} joins RuBP (5C) with RuBisCO; unstable 6C molecule splits	3 \ce{CO2} + 3 RuBP gives 6 3-PGA
2. Reduction	3-PGA is phosphorylated by ATP and reduced by NADPH to G3P	6 ATP and 6 NADPH used; 6 G3P made, 1 leaves
3. Regeneration of RuBP	5 G3P are rearranged to RuP; RuP is phosphorylated by ATP	3 ATP used; 3 RuBP regenerated

In all, three molecules of \ce{CO2}, six molecules of NADPH (reducing power) and nine molecules of ATP (assimilating power) are used to release just one molecule of G3P from the cycle. To make a molecule of glucose, two molecules of G3P are needed. The overall process can be written as follows.

\ce{3CO2 + 6NADPH + 9ATP -> G3P + 3H2O + 9ADP + 9P_i + 6NADP+}

Worked example: the cost of making one glucose

How many \ce{CO2}, ATP and NADPH molecules does the Calvin cycle use to make one glucose (\ce{C6H12O6})?

One turn of the cycle with 3 \ce{CO2} gives one G3P (3 carbons). Glucose has 6 carbons, so it needs 2 G3P.
So two sets of the overall reaction are needed: 6 \ce{CO2}, 12 NADPH and 18 ATP.
Check the carbon: 6 \ce{CO2} supply 6 carbons, exactly the carbons of one glucose.

	For 1 G3P	For 1 glucose (2 G3P)
\ce{CO2} fixed	3	6
NADPH used	6	12
ATP used	9	18
RuBP regenerated per cycle	3 (from 5 G3P)	6

6\ \ce{CO2},\ 12\ \text{NADPH},\ 18\ \text{ATP}

Fate of the atoms in photosynthesis

Fate of the atoms in photosynthesis. The oxygen of \ce{O2} (accent) comes from water.

Fate of atoms in \ce{6CO2 + 12H2O -> C6H12O6 + 6H2O + 6O2}.
Reactant	Where its atoms end up
6 \ce{CO2}	All 6 carbons and 6 of the 12 oxygens go into glucose; the other 6 oxygens go into the 6 new water molecules
12 \ce{H2O}	The 24 hydrogens go into glucose (12) and the 6 new water molecules (12); all 12 oxygens are released as 6 \ce{O2}

4.2, 4.2.1Cellular respiration and its kindsOften examined

In biological systems an oxidation-reduction reaction usually involves the removal of a hydrogen atom from one molecule and its gain by another molecule.

Cellular respiration: A series of complex oxidation-reduction reactions by which living cells obtain energy through the breakdown of organic matter.
Respiratory fuel: The organic molecule that generally breaks down in cellular respiration to release energy. It is usually glucose.
Glycolysis: The common first step of aerobic and anaerobic respiration, in which glucose is broken down into two molecules of pyruvate.

There are two kinds of cellular respiration.

Aerobic respiration takes place in the presence of abundant atmospheric oxygen.
Anaerobic respiration takes place in the absence of oxygen.

The initial breakdown of glucose is the same in both: glucose is broken down into two molecules of pyruvate (glycolysis). What happens to the pyruvate depends on whether oxygen is available and on the kind of organism.

Both kinds of respiration begin with glycolysis. The pyruvate then goes to one of two fates, depending on oxygen (accent: ATP yield).

Aerobic and anaerobic respiration.
	Aerobic respiration	Anaerobic respiration
Oxygen	Needed (abundant atmospheric oxygen)	Not needed
Fate of pyruvate	Broken down completely	Lactic acid, or ethanol and \ce{CO2}
Final products	\ce{CO2} and \ce{H2O}	Lactic acid, or ethanol and \ce{CO2}
Energy released	Large: 36 ATP (eukaryotes) or 38 ATP (prokaryotes)	Very small: just 2 ATP

4.2.2Mechanism of anaerobic respirationOften examined

Anaerobic respiration takes place in many microorganisms (bacteria, yeast), in the muscle cells of vertebrates and in the cells of higher plants. It is the incomplete breakdown of glucose in the absence of oxygen, and is also called fermentation. There are two pathways, depending on the final products: lactic acid fermentation and alcoholic fermentation.

Lactic acid fermentation

It consists of glycolysis followed by the reduction of pyruvate by NADH to lactic acid. The pathway can run without oxygen because, after NADH gives its electrons to pyruvate, NADH is turned back into \ce{NAD+}, which is "free" to return and pick up more electrons in the earlier reactions of glycolysis.

\ce{C6H12O6 + 2NAD+ -> 2C3H4O3 + 2NADH + 2H+}

\ce{2C3H4O3 + 2NADH + 2H+ -> 2C3H6O3 + 2NAD+}

Lactic acid fermentation occurs in anaerobic bacteria and in the muscles of mammals, including humans, during strenuous exercise when the oxygen supply is exhausted. The accumulation of lactic acid causes muscle fatigue: the muscles become unable to contract and begin to ache.

Alcoholic fermentation

It is found in yeast. It consists of glycolysis followed by the decarboxylation of pyruvate to acetaldehyde and then the reduction of acetaldehyde by NADH to ethyl alcohol (ethanol). Again NADH gives its electrons to acetaldehyde and returns as \ce{NAD+} to pick up more electrons in glycolysis, so the pathway works without oxygen.

\ce{C6H12O6 + 2NAD+ -> 2C3H4O3 + 2NADH + 2H+}

\ce{2C3H4O3 + 2NADH + 2H+ -> 2C2H5OH + 2CO2 + 2NAD+}

Figure 4.11: Anaerobic respiration. (a) Lactic acid fermentation. (b) Alcoholic fermentation. In both, NADH is used up to regenerate NAD^+ (accent).

Lactic acid fermentation and alcoholic fermentation.
Lactic acid fermentation	Alcoholic fermentation
Occurs in anaerobic bacteria and in mammalian muscles during strenuous exercise	Occurs in yeast
Pyruvate is reduced directly to lactic acid	Pyruvate is decarboxylated to acetaldehyde, which is then reduced to ethanol
End product: lactic acid (3C)	End products: ethanol (2C) and \ce{CO2}
No \ce{CO2} is released	\ce{CO2} is released (2 molecules per glucose)
Lactic acid builds up and causes muscle fatigue	Produces ethyl alcohol

Worked example: why does fermentation need the second step?

Glycolysis makes ATP without oxygen. Explain why the cell still needs to carry out lactic acid or alcoholic fermentation, and count the ATP.

In step 6 of glycolysis \ce{NAD+} is changed to NADH. A cell has only a small amount of \ce{NAD+}, and without oxygen the electron transport chain cannot turn NADH back into \ce{NAD+}.
If \ce{NAD+} ran out, step 6 and the whole of glycolysis would stop, and so would the ATP supply.
In fermentation, NADH passes its electrons to pyruvate (lactic acid) or acetaldehyde (ethanol). NADH becomes \ce{NAD+} again and glycolysis can continue.
The fermentation steps themselves make no ATP. The ATP yield is the 2 ATP net from glycolysis.

Stage	NAD^+ / NADH change	ATP
Glycolysis	2 \ce{NAD+} become 2 NADH	4 made, 2 used: net 2
Fermentation	2 NADH become 2 \ce{NAD+} again	0
Total	\ce{NAD+} is regenerated	2 ATP per glucose

4.2.3Mechanism of aerobic respirationOften examined

Aerobic respiration is a catabolic process: it is the complete oxidative breakdown of organic food (especially glucose) into carbon dioxide and water, with the release of a great deal of energy as ATP. It is the main respiratory pathway in most organisms.

Aerobic respiration is completed in four phases:

Glycolysis (in the cytoplasm).
Oxidation of pyruvate (the link reaction).
Krebs cycle.
Respiratory electron transport chain (ETC).

The four phases of aerobic respiration and what each yields (per glucose).

4.2.3GlycolysisOften examined

Glycolysis: The breakdown of glucose or a similar hexose sugar into two molecules of pyruvate through a series of enzyme-catalysed reactions, releasing some energy (as ATP) and reduced coenzyme (as NADH). It occurs in the cytoplasm.

Glycolysis is completed in two phases.

Preparatory phase (investment phase): two ATP are consumed. Its end products are two molecules of G3P.
Oxidative phase (pay-off phase): ATP is produced by substrate level phosphorylation, and NADH is also produced; on further oxidation in the respiratory electron transport chain this NADH yields more ATP.

The ten steps of glycolysis.
Step	Name	Enzyme	What happens
1	Phosphorylation	Hexokinase (glucokinase)	A phosphate from ATP is added to glucose: glucose 6-phosphate + ADP
2	Isomerization	Phosphohexose isomerase	Glucose 6-phosphate becomes its isomer fructose 6-phosphate
3	Phosphorylation	Phosphofructokinase	Another ATP gives a phosphate: fructose 1,6-bisphosphate + ADP
4	Splitting	Aldolase	Fructose 1,6-bisphosphate splits into two 3C compounds: G3P and dihydroxyacetone 3-phosphate
5	Isomerization	Phosphotriose isomerase	Dihydroxyacetone 3-phosphate becomes G3P (so there are two G3P; the next steps happen twice)
6	Dehydrogenation and phosphorylation	G3P dehydrogenase	G3P gives hydrogen to \ce{NAD+} (NADH formed) and takes inorganic phosphate: 1,3-bisphosphoglycerate
7	Formation of ATP	Phosphoglycerate kinase	1,3-Bisphosphoglycerate gives its phosphate to ADP: 3-phosphoglycerate + ATP (substrate level phosphorylation)
8	Isomerization	Phosphoglycerate mutase	The phosphate moves from C3 to C2: 2-phosphoglycerate
9	Dehydration	Enolase	2-Phosphoglycerate loses water: phosphoenolpyruvate (PEP)
10	Formation of ATP	Pyruvate kinase	PEP gives its phosphate to ADP: pyruvate + ATP (substrate level phosphorylation)

Figure 4.12: Glycolysis pathway. Steps 1 to 5 are the preparatory phase, steps 6 to 10 the pay-off phase. The two ATP-making steps (7 and 10) are in accent.

Energy balance of glycolysis.
	Preparatory phase (steps 1 to 5)	Pay-off phase (steps 6 to 10)
ATP	2 ATP used (steps 1 and 3)	4 ATP made (steps 7 and 10, two molecules each)
NADH	None	2 NADH made (step 6)
Starts with / ends with	1 glucose / 2 G3P	2 G3P / 2 pyruvate

4.2.4Oxidation of pyruvate to acetyl-CoAOften examined

Pyruvate is produced in the cytosol. Because it is a charged molecule, it must enter the mitochondrion by active transport with the help of a transport protein called pyruvate translocase. Once inside, pyruvate does not take part in the Krebs cycle directly; it first goes through an intermediate phase called the oxidation of pyruvate or link reaction. It is called the link reaction because it links the pathway of aerobic respiration that occurs outside the mitochondria (glycolysis) with the one inside them (Krebs cycle).

Decarboxylation: a molecule of \ce{CO2} is removed from pyruvate to form acetaldehyde.
Oxidation (dehydrogenation): \ce{NAD+} removes hydrogen from acetaldehyde. This gives a two-carbon acetyl group and NADH.
Addition of coenzyme A: the acetyl group combines with coenzyme A to form acetyl CoA.

Figure 4.13: Pathway of oxidation of pyruvate. One \ce{CO2} and one NADH are released for each pyruvate.

The whole of oxidation, decarboxylation and addition of coenzyme A is catalysed by a complex of three enzymes called the pyruvate dehydrogenase complex.

Figure 4.14: Site of oxidation of pyruvate: conversion of pyruvate to acetyl CoA, the junction between glycolysis and the citric acid cycle.

4.2.5Krebs cycleOften examined

This cycle was discovered by the British scientist Sir Hans Krebs, so it is called the Krebs cycle. It is also called the citric acid cycle or tricarboxylic acid (TCA) cycle, because the first compound formed in the cycle is citrate (citric acid), which contains three carboxylic acid groups. The cycle has nine steps.

The nine steps of the Krebs cycle.
Step	Name	Enzyme	Change	Products given off
1	Synthesis	Citrate synthetase	Acetyl CoA (2C) + oxaloacetate (4C) + \ce{H2O} give citrate (6C)	CoA
2	Dehydration	Aconitase	Citrate loses \ce{H2O}: cis-aconitate (intermediate)	\ce{H2O}
3	Hydration	Aconitase	Cis-aconitate + \ce{H2O} give isocitrate (an isomer of citrate)	none
4	Oxidative decarboxylation	Isocitrate dehydrogenase complex	Isocitrate (6C) gives \alpha-ketoglutarate (5C)	NADH, \ce{CO2}
5	Oxidative decarboxylation and addition of CoA	\alpha-Ketoglutarate dehydrogenase complex	\alpha-Ketoglutarate gives succinyl CoA (4C)	NADH, \ce{CO2}
6	Synthesis of substrate level ATP	Succinyl thiokinase	CoA removed from succinyl CoA: succinate (4C)	GTP, then ATP
7	Dehydrogenation (oxidation)	Succinate dehydrogenase	Succinate gives fumarate	\ce{FADH2}
8	Hydration	Fumarase	Fumarate + \ce{H2O} give malate	none
9	Dehydrogenation (oxidation)	Malate dehydrogenase	Malate gives oxaloacetate, which picks up the next acetyl CoA	NADH

Synthesis. Acetyl CoA (2C) and a water molecule combine with oxaloacetate (4C) to form the 6C compound citrate, with the help of citrate synthetase. It is the first product of the cycle. CoA is liberated.
Dehydration. Citrate loses a water molecule (enzyme aconitase) and becomes cis-aconitate, an intermediate in the isomerisation of citrate.
Hydration. Cis-aconitate takes up water and becomes isocitrate (same enzyme, aconitase). Citrate and isocitrate are isomers.
Oxidative decarboxylation. Isocitrate is first oxidised (dehydrogenated) and then decarboxylated to \alpha-ketoglutarate (isocitrate dehydrogenase complex). The hydrogen and electrons are taken up by \ce{NAD+} to form NADH; the carboxyl group leaves as \ce{CO2}.
Oxidative decarboxylation and addition of CoA. \alpha-Ketoglutarate again loses hydrogen and electrons (to \ce{NAD+}, forming NADH) and a carboxyl group (as \ce{CO2}), and combines with coenzyme A to give succinyl CoA (\alpha-ketoglutarate dehydrogenase complex).
Synthesis of substrate level ATP. Succinyl thiokinase removes CoA from succinyl CoA to give succinate. The energy released is used to combine GDP and Pi to form GTP; GTP reacts with ADP to form ATP and GTP becomes GDP again. One ATP is generated.
Dehydrogenation (oxidation). Succinate is oxidised by succinate dehydrogenase to fumarate. Its hydrogen and electrons are taken up by FAD to form \ce{FADH2}.
Hydration. Water is added to fumarate to form malate (enzyme fumarase).
Dehydrogenation (oxidation). Malate is oxidised by malate dehydrogenase to oxaloacetate; hydrogen and electrons go to \ce{NAD+} to form NADH. Oxaloacetate picks up another acetyl CoA and the cycle repeats.

Figure 4.15: Krebs cycle (citric acid cycle or TCA cycle). Step 6 makes ATP and step 7 makes FADH_2 (accent).

Inputs and outputs of the Krebs cycle (from Figure 4.15).
	Inputs	Outputs
For 2 acetyl groups (one glucose)	2 acetyl groups, 2 ADP + 2 Pi, 6 \ce{NAD+}, 2 FAD	4 \ce{CO2}, 2 ATP, 6 NADH, 2 \ce{FADH2}
For 1 acetyl group (one turn)	1 acetyl group, 1 ADP + 1 Pi, 3 \ce{NAD+}, 1 FAD	2 \ce{CO2}, 1 ATP, 3 NADH, 1 \ce{FADH2}

4.2.6Electron transport chain (ETC)Often examined

After the Krebs cycle most of the energy of glucose is held in the form of **NADH and \ce{FADH2}**. These molecules enter the electron transport chain, where the reduced NADH and \ce{FADH2} are oxidised and their electrons are passed along a series of oxidation-reduction reactions to the final acceptor, molecular oxygen.

Components of the respiratory electron transport chain

Components of the respiratory ETC.
No.	Component	Further composed of / note
1	NADH-Q reductase or NADH dehydrogenase complex (I)	FMN and Fe-S complex
2	FADH dehydrogenase complex (II)
3	Coenzyme Q (ubiquinone)	Mobile carrier
4	Cytochrome reductase complex (III)	Cyt b, Fe-S and Cyt c_1
5	Cytochrome c	Mobile carrier
6	Cytochrome oxidase complex (IV)	Cyt a, Cyt a_3 and copper

Passage of electron flow

The passage of electrons from high-energy NADH to the first carrier and on to the next is a downhill journey. At each step energy is released and some membrane carrier proteins use it to **pump protons (\ce{H+}) from the matrix into the intermembrane space**. The accumulation of \ce{H+} in the intermembrane space is a proton gradient, which activates ATP synthetase to make ATP in the presence of \ce{O2}. Because this ATP synthesis comes from the oxidation of NADH or \ce{FADH2} in the presence of \ce{O2}, the ETC is also called oxidative phosphorylation.

NADH is oxidised when it reacts with NADH dehydrogenase complex (I), which has two prosthetic groups, FMN and Fe-S. Electrons and protons pass from NADH to FMN; FMN moves the electrons to the Fe-S complex, which pumps protons into the intermembrane space.
\ce{FADH2} is oxidised through FADH dehydrogenase complex (II) and hands its electrons to coenzyme Q.
Electrons from coenzyme Q go to cytochrome reductase complex (III) (Cyt b, Fe-S, Cyt c_1). This complex passes its electrons to cytochrome c and also pumps protons to the intermembrane space. Like coenzyme Q, cytochrome c is a mobile carrier of electrons.
Cytochrome c delivers the electrons to cytochrome oxidase complex (IV) (Cyt a, Cyt a_3, copper), which acts as both an electron carrier and a proton pump.
Finally the electrons are transferred to oxygen, the ultimate electron acceptor. Each oxygen atom also picks up a pair of hydrogen ions from the solution and forms water.

The energy released during the downhill passage of electrons is used by specific transmembrane protein complexes to pump protons from the matrix to the intermembrane space. To avoid losing the energy as unwanted heat, it is released in steps rather than all at once. Proton pumping takes place at three sites.

Proton pumping sites of the ETC.
	Site 1	Site 2	Site 3
Proton pumping between	FMN and CoQ	Cyt b and Cyt c_1	Cyt a and Cyt a_3
Enzyme complex involved	NADH dehydrogenase complex I	Cytochrome reductase complex III	Cytochrome oxidase complex IV

Figure 4.16: Sequence of electron carriers in the respiratory ETC. Electrons (accent) lose energy at each step; protons are pumped at I, III and IV.

The electron transport chain makes no ATP directly. Its function is to ease the fall of electrons from food to oxygen, releasing energy in manageable amounts. The mitochondrion couples the ETC to ATP synthesis by a mechanism called chemiosmosis (next section).

4.2.7Chemiosmosis and oxidative phosphorylationOften examined

Oxidative phosphorylation: The synthesis of ATP molecules with the help of the energy liberated during the oxidation of the reduced coenzymes (NADH, \ce{FADH2}) produced in respiration. The enzyme required is ATP synthetase.
Chemiosmosis: The mechanism by which the energy of a proton gradient across a membrane is used to make ATP: protons pumped by the electron transport chain flow back through ATP synthase.

Figure 4.17: Mechanism of chemiosmosis in the respiratory electron transport chain. (1) The ETC pumps H^+ into the intermembrane space; (2) H^+ flows back through ATP synthase, which makes ATP.

ATP synthase (ATP synthetase) is located in the inner mitochondrial membrane. It has two parts.

**F_0** is embedded in the membrane and is the channel through which protons move from the intermembrane space to the matrix.
**F_1** (the elementary particle) is a head-like part that projects from the membrane surface into the matrix. It catalyses ATP synthesis by combining ADP and Pi.

The electron transport chain pumps protons from the matrix into the intermembrane space, so the concentration of \ce{H+} is higher on the F_0 side (intermembrane space) than on the F_1 side (matrix): a proton gradient.
The gradient makes ATP synthase active. Protons flow back down the gradient through the F_0 channel.
This flow makes the F_1 particles work as ATP synthetase: the energy of the proton gradient is used to attach a phosphate to ADP by a high-energy bond, producing ATP.

Oxidation of one molecule of NADH produces 3 ATP, while the oxidation of one molecule of \ce{FADH2} forms 2 ATP. The theory of ATP production by this mechanism is called chemiosmosis.

Oxidative phosphorylation and photophosphorylation (both use chemiosmosis).
Oxidative phosphorylation	Photophosphorylation
Occurs in respiration, in the mitochondrion (inner membrane)	Occurs in the light reaction of photosynthesis, in the chloroplast (thylakoid membrane)
Energy comes from the oxidation of NADH and \ce{FADH2}	Energy comes from light absorbed by the photosystems
Final electron acceptor is oxygen (forms water)	Final electron acceptor is \ce{NADP+} (non-cyclic) or P700 (cyclic); oxygen is released from water
Protons are pumped from the matrix into the intermembrane space	Protons are pumped from the stroma into the thylakoid space
ATP synthase: F_1 head faces the matrix	ATP synthase makes ATP on the stroma side

4.2.8Substrate level phosphorylationOften examined

The main aim of cellular respiration is to generate ATP. In aerobic respiration this is done in two ways: chemiosmosis (discussed above) and substrate level phosphorylation.

Phosphorylation: The addition of inorganic phosphate to any organic molecule.
Substrate level phosphorylation: The enzymatic transfer of a phosphate group from an organic substrate molecule to ADP, making ATP directly. It accounts for only a small percentage of the ATP a cell makes.

Substrate level phosphorylation occurs on three occasions during aerobic respiration.

Glycolysis, reaction 7: 1,3-bisphosphoglycerate is converted into 3-phosphoglycerate.
Glycolysis, reaction 10: phosphoenolpyruvate is converted into pyruvate.
Krebs cycle: succinyl CoA is converted into succinate (ATP made through GTP).

In glycolysis four ATP are made by this mechanism, but two were used in the preparatory phase, so the net gain is 2 ATP. In the Krebs cycle two ATP are made per glucose.

Because ATP can be made directly from the organic substrates of exergonic (energy-releasing) reactions such as cellular respiration, substrate level phosphorylation couples exergonic reactions with the synthesis of ATP. This ATP is then used to drive endergonic (energy-storing) reactions such as protein synthesis. So ATP is the link between catabolism and anabolism in living cells.

Figure 4.18 (a, b): (a) Substrate level phosphorylation: an enzyme moves a phosphate from an organic substrate to ADP. (b) Because ATP couples many endergonic and exergonic reactions, it is an important link between anabolism and catabolism (ATP in accent).

ATP couples exergonic and endergonic reactions (Figure 4.18 b).
Exergonic reactions	Endergonic reactions
Release energy	Need (store) energy
Example: cellular respiration	Example: protein synthesis, making new cells or cell parts
Energy is captured in ATP (ADP + Pi gives ATP)	Energy is supplied by ATP (ATP gives ADP + Pi)

ATP budget in aerobic respiration (Figure 4.18 c)

ATP yield from one glucose in a eukaryotic cell.
Stage	Substrate level ATP	Reduced coenzymes	ATP by chemiosmosis	Total ATP
Glycolysis (cytosol)	2 (net)	2 NADH	4	6
Link reaction	0	2 NADH	6	6
Krebs cycle	2	6 NADH, 2 \ce{FADH2}	18 + 4 = 22	24
Total	4		32	36

Oxidative phosphorylation and substrate level phosphorylation.
Oxidative phosphorylation	Substrate level phosphorylation
ATP is made in the electron transport chain using the proton gradient (chemiosmosis)	ATP is made by direct transfer of a phosphate group from an organic substrate to ADP
Needs oxygen as final electron acceptor	Does not need oxygen directly
Occurs on the inner mitochondrial membrane	Occurs in the cytosol (glycolysis) and the mitochondrial matrix (Krebs cycle)
Makes most of the ATP: 32 of 36	Makes a small part: 4 of 36 (net)
Needs ATP synthase	Needs only a kinase enzyme (for example pyruvate kinase)

Worked example: ATP from one glucose

Calculate the total ATP made from one glucose in a human cell, and say how much of it comes from substrate level phosphorylation.

Count the ATP made directly (substrate level): 2 net in glycolysis + 2 in the Krebs cycle = 4 ATP.
Count the reduced coenzymes: 2 NADH (glycolysis) + 2 NADH (link reaction) + 6 NADH (Krebs) + 2 \ce{FADH2} (Krebs).
Convert with the book values. The glycolysis NADH are counted as \ce{FADH2} (2 ATP each) because of the mitochondrial membrane. So: glycolysis 2 x 2 = 4; link 2 x 3 = 6; Krebs NADH 6 x 3 = 18; Krebs \ce{FADH2} 2 x 2 = 4. Total by chemiosmosis = 32.
Add: 4 + 32 = 36 ATP.

Source	Calculation	ATP
Substrate level	2 + 2	4
Glycolysis NADH	2 x 2	4
Link reaction NADH	2 x 3	6
Krebs cycle NADH	6 x 3	18
Krebs cycle \ce{FADH2}	2 x 2	4
Total		36

36\ \text{ATP}\ (4\ \text{substrate level}+32\ \text{chemiosmosis})

4.2.9Importance of G3P

Glyceraldehyde 3-phosphate (G3P) is a key molecule in both photosynthesis and respiration. Its importance can be summarised as follows.

G3P is an important intermediate of both respiration and photosynthesis.
In the Calvin cycle of photosynthesis, G3P molecules are converted into glucose phosphate within the chloroplast. Glucose phosphate is then converted to glucose, fructose, sucrose, maltose and starch.
In respiration, G3P appears during glycolysis and acts as the connecting molecule between the preparatory phase and the pay-off (oxidative) phase. All the energy molecules, ATP or NADPH_2, can be produced only after the formation of G3P.
G3P ultimately leads to the formation of pyruvate, which is the end product of glycolysis and the raw material for anaerobic and aerobic respiration.
G3P acts as an intermediate molecule for the inter-conversion of different biological molecules, ATP and waste products.
Carbohydrates in the cell may be converted to lipids, and lipids to carbohydrates, through the formation of G3P as an intermediate.
Carbohydrates may be converted to amino acids and then proteins, and proteins to carbohydrates, through G3P as an intermediate.

Figure 4.19: The metabolic pool concept. Carbohydrates, fats and proteins enter degradative pathways at specific points; the products can be used to make other biomolecules or to release ATP. G3P (accent) is the crossing point of the pathways.

4.2.10Cellular respiration of fats and proteins

Glucose is the usual fuel, but fats and proteins can also be used as energy sources. They enter the respiratory pathway at different points (Figure 4.19).

Fats

When a fat is used as an energy source it breaks down into glycerol and three fatty acids.

Glycerol is converted to G3P, a metabolite of glycolysis.
The fatty acids are converted to acetyl-CoA, which enters the Krebs cycle. An 18-carbon fatty acid gives nine acetyl-CoA molecules.

Proteins

The hydrolysis of proteins gives amino acids. The size of the R-group of an amino acid decides whether its carbon chain is oxidised in glycolysis or in the Krebs cycle. The carbon chain is produced in the liver when an amino acid undergoes deamination, the removal of its amino group. The amino group becomes **ammonia (\ce{NH3}), which enters the urea cycle and becomes part of urea**.

Respiration of glucose and of fat (SLO 16).
Respiration of glucose	Respiration of fat
Starts with glycolysis of glucose (6C) to 2 pyruvate	Fat breaks down to glycerol and three fatty acids
Pyruvate is converted to acetyl-CoA (link reaction)	Glycerol is converted to G3P (enters glycolysis); fatty acids are converted directly to acetyl-CoA
Each glucose gives 2 acetyl-CoA	An 18-carbon fatty acid gives 9 acetyl-CoA
Acetyl-CoA enters the Krebs cycle	Acetyl-CoA enters the Krebs cycle (same pathway from here)

4.3, 4.3.1PhotorespirationOften examined

Photorespiration: The respiratory activity that occurs in green cells in the presence of light and releases carbon dioxide. It needs oxygen and produces \ce{CO2} and \ce{H2O} like aerobic respiration, but no ATP is produced.

Mechanism of photorespiration

When the \ce{CO2} level inside the leaf drops to around 50 ppm (parts per million), RuBisCO starts to combine \ce{O2} with RuBP instead of \ce{CO2}. The net result is that, instead of two three-carbon molecules of phosphoglycerate (PGA), only one PGA and a toxic two-carbon molecule called phosphoglycolate are produced. The plant must get rid of the phosphoglycolate. It first removes the phosphate group at once, converting the molecule to glycolate.

\ce{RuBP + O2 -> phosphoglycolate + PGA ->[-P_i] glycolate}

Chloroplast: RuBP + \ce{O2} give PGA and phosphoglycolate (2C); phosphoglycolate loses its phosphate (Pi) and becomes glycolate.
Peroxisome: glycolate is converted to glyoxylate (hydrogen peroxide forms and breaks down to water and oxygen) and then to glycine.
Mitochondrion: two glycine are converted into one serine, with release of \ce{CO2} (and \ce{NH3}): 2\,\text{glycine} \to \text{serine} + \ce{CO2}.
The serine is used to make other organic molecules (in Figure 4.20 it returns through the peroxisome as hydroxypyruvate and glycerate, and the glycerate is converted to PGA in the chloroplast using ATP).

Figure 4.20: Schematic representation of the pathway involved in photorespiration in the chloroplast, peroxisome and mitochondrion. The two carbons lost as \ce{CO2} and the ATP used make photorespiration wasteful.

Where the photorespiration steps occur.
Compartment	Compounds formed
Chloroplast	Phosphoglycolate (2C), glycolate (2C); at the end glycerate (3C) to PGA
Peroxisome	Glyoxylate (2C), glycine (2C); on the return path hydroxypyruvate (3C) and glycerate (3C)
Mitochondrion	Serine (3C) from two glycine; \ce{CO2} and \ce{NH3} released

4.3.2Disadvantages and evolution of photorespirationOften examined

Disadvantages

Photorespiration costs the plant energy and results in a net loss of \ce{CO2} fixation. It therefore reduces the rate of photosynthesis.
In most plants it reduces the amount of carbon fixed into carbohydrate during photosynthesis by 25 percent.
It produces no ATP.
It is not essential for the plant: if photorespiration is chemically inhibited, the plant can still grow, and some plants are naturally resistant to it.

Evolution of photorespiration

If photorespiration is wasteful and not essential, why does it exist? The common simple answer is that the active site of RuBisCO evolved to bind both carbon dioxide and oxygen. Originally this was not a problem, because there was no oxygen in the atmosphere at the time the earth was established, so the carbon dioxide binding activity was the only one used. Photorespiration started when oxygen began to accumulate in the atmosphere.

How photorespiration evolved

Early earth: almost no \ce{O2} in the air, so RuBisCO bound only \ce{CO2}.
The active site of RuBisCO was able to bind both \ce{CO2} and \ce{O2}.
Oxygen began to accumulate in the atmosphere (largely from photosynthesis itself).
RuBisCO now sometimes binds \ce{O2} instead of \ce{CO2}: photorespiration appears as a side reaction.

4.3.3Effect of temperature on the activities of RuBisCO

Photorespiration is related to the working of the enzyme ribulose bisphosphate carboxylase/oxygenase (RuBisCO). It is often called RuBisCO because it can have an oxygenase activity as well as its carboxylase activity. Which activity it shows depends on the **relative concentrations of \ce{O2} and \ce{CO2}** in the leaves. Photorespiration starts when the \ce{CO2} level inside a leaf becomes low.

If the plant continues \ce{CO2} fixation in photosynthesis when its stomata are closed, the \ce{CO2} is used up, and the \ce{O2} released by photosynthesis cannot escape from the plant body. In this way the ratio of \ce{O2} to \ce{CO2} in the leaf increases, and RuBisCO uses \ce{O2} (oxygenase activity) instead of \ce{CO2}.

Why closed stomata favour photorespiration

Stomata are closed, so no fresh \ce{CO2} enters the leaf.
Photosynthesis goes on, using up \ce{CO2} inside the leaf.
The \ce{O2} released by photosynthesis cannot get out.
The ratio \ce{O2}:\ce{CO2} in the leaf rises.
RuBisCO shows more oxygenase activity: photorespiration increases.

4.3.4C4 photosynthesis: an adaptation to the problem of photorespirationOften examined

Some plants that grow in tropical climates have an adaptation to the problem of photorespiration. They have an additional metabolic pathway in the light independent phase, besides the Calvin cycle, called the Hatch-Slack cycle or C4 pathway. In this pathway phosphoenol pyruvate (PEP) carboxylase is used instead of RuBisCO to fix \ce{CO2} to phosphoenol pyruvate (a C3 molecule), and the result is oxaloacetate, a C4 molecule. It takes place in the cytoplasm of the mesophyll cells.

\ce{PEP + CO2 ->[\text{PEP carboxylase}] oxaloacetate}

Since the first product of \ce{CO2} fixation is a four-carbon compound, oxaloacetate, these plants are called C4 plants, for example maize, sugarcane and sorghum.

Fixation in the mesophyll cell. \ce{CO2} + PEP (3C) give oxaloacetate (4C), with PEP carboxylase, in the cytoplasm of the mesophyll cell.
Oxaloacetate to malate. Oxaloacetate is transported to the chloroplasts of the mesophyll cells and converted to another 4C compound, malate, with the help of NADH produced in the photochemical phase.
Malate to the bundle sheath. Malate is transported to the chloroplasts of the bundle sheath cells.
**\ce{CO2} release. In the bundle sheath cells malate is converted to pyruvate** (3C) with release of \ce{CO2}, so the concentration of \ce{CO2} rises in the bundle sheath cells.
Calvin cycle. These cells contain the enzymes of the Calvin cycle. Because the \ce{CO2} concentration is high, RuBisCO takes part in the Calvin cycle and not in photorespiration. The sugar formed is carried into the phloem.
Regeneration of PEP. Pyruvate from the bundle sheath cells returns to the mesophyll cells and regenerates PEP, consuming one ATP.

Figure 4.21 (left): Comparison of C3 and C4 photosynthesis. In C4 plants \ce{CO2} is first fixed in the mesophyll cell (accent) and then released in the bundle sheath cell, where the Calvin cycle runs.

Figure 4.21 (right): The complete C4 photosynthetic cycle between a mesophyll cell and a bundle sheath cell.

C3 and C4 carbon fixation.
	C3 carbon fixation	C4 carbon fixation
Pathway	Calvin cycle only (C3 plants)	Hatch-Slack (C4) pathway plus the Calvin cycle
Enzyme that first fixes \ce{CO2}	RuBisCO	PEP carboxylase
\ce{CO2} acceptor	RuBP (5C)	PEP (3C)
First stable product	3-PGA (3C)	Oxaloacetate (4C)
Where fixed	Mesophyll cell chloroplasts	Cytoplasm of mesophyll cells; Calvin cycle then in bundle sheath cells
Photorespiration	Occurs (RuBisCO also binds \ce{O2})	Avoided: high \ce{CO2} in the bundle sheath keeps RuBisCO in the Calvin cycle
Examples	Most plants	Maize, sugarcane, sorghum (tropical climate)

Worked example: how does C4 photosynthesis avoid photorespiration?

Explain, in steps, how a C4 plant keeps RuBisCO from acting as an oxygenase.

In the mesophyll cell \ce{CO2} is fixed by PEP carboxylase, which does not bind \ce{O2}, so it works even when \ce{CO2} is low.
The fixed carbon travels as malate (4C) to the bundle sheath cell, which holds the Calvin cycle enzymes.
There malate is broken down to pyruvate and \ce{CO2} is released, so the \ce{CO2} concentration in the bundle sheath cell becomes high.
With plenty of \ce{CO2} the O_2 : \ce{CO2} ratio at RuBisCO is low, so RuBisCO works as a carboxylase in the Calvin cycle and does not start photorespiration.
The price is one ATP per cycle to regenerate PEP from pyruvate.

Formula sheet

Overall equation of photosynthesis

\ce{6CO2 + 12H2O ->[\text{light}][\text{chlorophyll}] C6H12O6 + 6O2 + 6H2O}

Carbon dioxide and water give glucose, oxygen and water; \ce{CO2} is reduced and \ce{H2O} is oxidised.

Photolysis of water

\ce{H2O -> 2H+ + 2e- + 1/2O2}

Water splits in PS-II into protons, electrons and oxygen.

Reduction of NADP+

\ce{NADP+ + 2e- + 2H+ -> NADPH + H+}

NADP reductase passes electrons from ferredoxin to NADP+.

Overall Calvin cycle

\ce{3CO2 + 6NADPH + 9ATP -> G3P + 3H2O + 9ADP + 9P_i + 6NADP+}

Inputs and outputs for each G3P released from the cycle.

Lactic acid fermentation

\begin{aligned}\ce{C6H12O6 + 2NAD+ &-> 2C3H4O3 + 2NADH + 2H+}\\ \ce{2C3H4O3 + 2NADH + 2H+ &-> 2C3H6O3 + 2NAD+}\end{aligned}

Glycolysis, then reduction of pyruvate by NADH to lactic acid.

Alcoholic fermentation

\ce{2C3H4O3 + 2NADH + 2H+ -> 2C2H5OH + 2CO2 + 2NAD+}

Pyruvate (from glycolysis) is decarboxylated and reduced to ethanol.

Photorespiration (RuBisCO oxygenase reaction)

\ce{RuBP + O2 -> phosphoglycolate + PGA}

At low \ce{CO2}, RuBisCO adds \ce{O2} to RuBP.

C4 carbon fixation

\ce{PEP + CO2 ->[\text{PEP carboxylase}] oxaloacetate}

First step of the C4 pathway in mesophyll cells.

Key terms

Bioenergetics: The quantitative study of energy conversions and energy relationships in biological systems.
Photosynthesis: The process in which light energy is used by photosynthetic pigments to convert carbon dioxide and water into glucose, releasing oxygen. A redox process in which \ce{CO2} is reduced and water is oxidised.
Electromagnetic spectrum: The full range of electromagnetic radiation in the universe; visible light (380 to 750 nm) is a small part of it.
Action spectrum: A graph showing the effectiveness of different wavelengths of light for photosynthesis (rate of photosynthesis against wavelength).
Photolysis of water: The splitting of a water molecule by light into 2\ce{H+}, 2\ce{e-} and \tfrac{1}{2}\ce{O2} in the light reaction.
Pigment: Any substance that absorbs light energy for use in photosynthesis.
Chlorophyll a: A bluish green pigment found in all photosynthetic organisms except photosynthetic bacteria; has a methyl group on the 2nd pyrrole ring and forms the reaction centre of a photosystem.
Chlorophyll b: A yellowish green accessory pigment with an aldehyde group on the 2nd pyrrole ring; found in all photosynthetic organisms except brown algae, red algae and photosynthetic bacteria.
Porphyrin ring: The hydrophilic, light-absorbing head of chlorophyll: four pyrrole rings around a central magnesium atom.
Phytol chain: The hydrophobic tail of chlorophyll that anchors the molecule in the thylakoid membrane.
Chlorosis: Deficiency of chlorophyll in the leaves.
Carotenoids: Terpenoid lipid pigments (yellow, orange, red or brown) that absorb blue-violet light, act as accessory pigments and protect chlorophyll a from excess light.
Carotenes: Orange-red carotenoids made of isoprenoid units, found in all photosynthetic eukaryotes; beta carotene is the most widespread.
Xanthophylls: Yellow carotenoids made of isoprenoid units; lutein gives the yellow colour of autumn foliage.
Spectrophotometer: The instrument used to measure the relative ability of pigments to absorb light of different wavelengths.
Absorption spectrum: A graph showing the relative absorption of different wavelengths of light by a particular pigment.
Accessory pigments: Pigments (chlorophyll b, xanthophylls, carotenes) that absorb light and pass the energy to chlorophyll a but cannot convert it into chemical energy.
Photosystem: A cluster of pigment molecules in the thylakoid membrane made of an antenna complex and a reaction centre.
Antenna complex: The peripheral part of a photosystem, made of accessory pigments such as chlorophyll b and carotenoids.
Reaction centre: The central part of a photosystem, containing only chlorophyll a and associated proteins.
P700 and P680: The reaction centre chlorophyll a of photosystem I (absorbs maximum 700 nm) and photosystem II (680 nm).
Light dependent phase: The first phase of photosynthesis, in the thylakoid membranes in the presence of light, making ATP and NADPH.
Photophosphorylation: The production of ATP from ADP and Pi during the light reaction, by chemiosmosis.
Non-cyclic photophosphorylation: Light reaction pathway using both photosystems and two electron transport chains; electrons end up in NADPH and oxygen is released (Z-scheme).
Cyclic photophosphorylation: Light reaction pathway using only photosystem I, in which excited electrons return to P700; only ATP is made.
Z-scheme: The zigzag path of electrons through the two photosystems in non-cyclic photophosphorylation.
Primary electron acceptor: The molecule of a photosystem that captures the excited electrons from the reaction centre chlorophyll.
Plastoquinone and plastocyanin: Mobile electron carriers (PQ and PC) of the light reaction electron transport chain.
Ferredoxin: An iron-containing protein that accepts electrons from photosystem I and passes them to NADP reductase.
NADP reductase: The flavoprotein enzyme that transfers electrons from ferredoxin to \ce{NADP+}, forming NADPH.
ATP synthase: The enzyme that makes ATP from ADP and Pi using the flow of protons down their gradient (in thylakoid and inner mitochondrial membranes).
Light independent phase: The second phase of photosynthesis (dark reaction), which uses ATP, NADPH and \ce{CO2} to make G3P.
C3 plants: Plants in which only the Calvin cycle operates during the dark reaction; the first stable product is a three-carbon compound.
Calvin cycle: The series of reactions (carbon fixation, reduction, regeneration of RuBP) that reduce \ce{CO2} to G3P; also called the C3 pathway.
RuBP (ribulose bisphosphate): The five-carbon sugar that acts as the \ce{CO2} acceptor in the Calvin cycle.
RuBisCO: Ribulose bisphosphate carboxylase/oxygenase, the enzyme that combines \ce{CO2} (or \ce{O2}) with RuBP.
Carbon fixation: The incorporation of carbon of inorganic \ce{CO2} into an organic compound (RuBP gives two 3-PGA).
G3P (glyceraldehyde 3-phosphate): A three-carbon phosphorylated sugar; the output of the Calvin cycle and an intermediate of glycolysis.
Cellular respiration: A series of oxidation-reduction reactions by which cells obtain energy through the breakdown of organic matter.
Respiratory fuel: The organic molecule, usually glucose, that is broken down in respiration to release energy.
Glycolysis: The breakdown of glucose to two pyruvate in the cytoplasm, in a preparatory phase and a pay-off phase.
Fermentation: Anaerobic respiration: incomplete breakdown of glucose without oxygen, either lactic acid or alcoholic.
Lactic acid fermentation: Glycolysis followed by reduction of pyruvate by NADH to lactic acid; occurs in anaerobic bacteria and in muscles.
Alcoholic fermentation: Glycolysis followed by decarboxylation of pyruvate to acetaldehyde and its reduction by NADH to ethanol; occurs in yeast.
Pyruvate translocase: The transport protein that carries pyruvate into the mitochondrion by active transport.
Link reaction (oxidation of pyruvate): The conversion of pyruvate to acetyl CoA by decarboxylation, oxidation by \ce{NAD+} and addition of coenzyme A.
Pyruvate dehydrogenase complex: The complex of three enzymes that catalyses the link reaction.
Krebs cycle: The cycle (citric acid cycle or TCA cycle) in which acetyl CoA is oxidised, giving \ce{CO2}, NADH, \ce{FADH2} and ATP.
Electron transport chain (ETC): The series of carriers (complexes I to IV, coenzyme Q, cytochrome c) that pass electrons from NADH and \ce{FADH2} to oxygen and pump protons.
Coenzyme Q (ubiquinone): A small lipid-soluble mobile carrier of electrons in the respiratory ETC.
Cytochromes: Iron-containing protein-pigment electron carriers; cytochrome c is a mobile carrier.
Chemiosmosis: The mechanism of ATP synthesis driven by a proton gradient: protons flow through the F_0 channel of ATP synthase.
Oxidative phosphorylation: ATP synthesis using the energy released by oxidation of NADH and \ce{FADH2} in the electron transport chain.
Substrate level phosphorylation: The direct transfer of a phosphate group from an organic substrate to ADP, making ATP.
Deamination: Removal of the amino group from an amino acid (in the liver); the amino group becomes ammonia.
Photorespiration: Respiratory activity in green cells in light in which RuBisCO combines \ce{O2} with RuBP; releases \ce{CO2}, uses \ce{O2} and makes no ATP.
Phosphoglycolate: The toxic two-carbon compound formed when RuBisCO combines \ce{O2} with RuBP.
C4 pathway (Hatch-Slack cycle): A pathway in which \ce{CO2} is first fixed to PEP by PEP carboxylase into oxaloacetate (4C) in mesophyll cells.
PEP carboxylase: The enzyme that fixes \ce{CO2} to phosphoenol pyruvate to give oxaloacetate in C4 plants.
Bundle sheath cells: Cells of C4 leaves in which malate releases \ce{CO2} for the Calvin cycle, so RuBisCO does not do photorespiration.

Multiple choice questionsp. 128–129

1

Removal of the source of carbon dioxide from photosynthesizing chloroplasts results in rapid changes in the concentration of certain chemicals. Which one of the following represents the correct combination of concentration changes?

AATP decreases; ribulose bisphosphate decreases; phosphoglyceric acid (PGA) increases
BATP decreases; ribulose bisphosphate increases; PGA shows no change
CATP increases; ribulose bisphosphate increases; PGA decreasesAnswer
DATP increases; ribulose bisphosphate shows no change; PGA decreases

Why

Without \ce{CO2}, the Calvin cycle cannot fix carbon. RuBP is no longer used up, so it accumulates, and 3-PGA is no longer made, so it falls. The light reactions continue, but their ATP is not used by the Calvin cycle, so ATP rises.

A and B show ATP falling, which would happen only if the light reactions stopped. D keeps RuBP unchanged, but RuBP is not consumed, so it must rise.

2

What are the products of the light reactions in photosynthesis?

AATP and NADP
BATP, NADPH_2 and oxygenAnswer
CATP, PGA and NADH_2
DATP, PGA and oxygen

Why

The light reactions make ATP (photophosphorylation), NADPH (NADP^+ is reduced by electrons from PS I) and oxygen (from the photolysis of water). The book writes the reduced coenzyme as NADPH_2, which means NADPH.

A lists NADP, which is a reactant (the electron acceptor), not a product. C and D contain PGA, a product of the Calvin cycle, and C also contains NADH_2, which comes from respiration, not from the light reactions.

3

During the light dependent stage of photosynthesis, the photoactivated pigment removes an electron from the hydroxylation derived from the water molecule. The fate of the free hydroxyl radical is that it

Ais broken down into oxygen and a free radical of hydrogenAnswer
Bis used to raise the activation level of chlorophyll by donating a positive charge
Cis used to produce adenosine triphosphate from adenosine diphosphate
Dreduces carbon dioxide to sugar

Why

The photoactivated P680 takes an electron from water. The hydroxyl radical (OH) left behind is broken down into oxygen and a free hydrogen radical; this is the photolysis of water that the book describes, and the hydrogen is then taken up by NADP^+.

B is wrong because the pigment loses electrons and does not gain a charge from the hydroxyl. C is wrong because ATP is made by chemiosmosis from the proton gradient, not from the radical. D is wrong because sugar is made in the Calvin cycle with ATP and NADPH, not directly from the radical.

Note. The question prints 'hydroxylation' where the sense is the splitting (photolysis) of water; the intended reading is the hydroxyl produced when water is split.

4

Carbon dioxide labeled with ^{14}C has been used to identify the intermediate compounds in the Calvin cycle, the light-independent stage in photosynthesis. Which compound would be the first to contain the ^{14}C?

Aglucose
BPGAAnswer
CRuBP
Dstarch

Why

Labelled \ce{CO2} is fixed onto RuBP, forming an unstable 6-carbon compound that at once splits into two molecules of 3-phosphoglycerate (PGA). PGA is the first stable compound of the cycle, so it is the first to carry the label.

RuBP is the acceptor that receives the \ce{CO2}; it does not carry the label until it is regenerated, later in the cycle. Glucose and starch are made only after G3P has been formed, so they come much later.

5

The rate of photosynthesis of a freshwater plant is measured using five spectral colours. Which sequence of colours would give an increasing photosynthetic response? (Smallest response on the left, largest on the right.)

Ablue, green, yellow, orange, red
Bgreen, yellow, orange, red, blueAnswer
Cred, orange, yellow, green, blue
Dyellow, green, orange, blue, red

Why

Fig 4.2 (the action spectrum) shows the lowest effectiveness in the green region and a steady rise through yellow and orange to red, with blue and red the most effective. Of the two peaks, the blue one (about 430 nm) is slightly the higher, so blue gives the largest response.

A puts blue at the bottom although it is one of the two most effective colours. C puts red at the bottom and green above orange, both wrong. D places yellow below green and reverses the order of blue and red.

Note. The book prints no key. The answer is read from the action spectrum in Fig 4.2; the blue and red peaks are close, so the order of the top two depends on reading the figure.

6

During dark reactions the three carbon atoms of 3-PGA are derived from

ARuBP only
BCO_2 only
CRuBP + CO_2Answer
DRuBP + CO_2 + PEP

Why

RuBP (5 carbons) takes up one carbon from \ce{CO2} to form a 6-carbon compound, which splits into two 3-PGA molecules. Each 3-PGA therefore has one carbon from \ce{CO2} and two from RuBP.

A and B each name only one source. D adds PEP, which belongs to the C4 pathway, not to the Calvin cycle.

7

Chlorophyll is soluble in

Awater
Borganic solventAnswer
Cwater and organic solvent
Dnot in any solvent

Why

The two tails of chlorophyll, the phytol chain and the methyl ester, are hydrophobic, so the molecule dissolves in organic solvents (such as alcohol, acetone or ether) and not in water.

C is tempting because the book calls the head hydrophilic, but the whole molecule is dominated by its hydrophobic tails. D is wrong because chlorophyll is extracted with solvents in the laboratory.

8

Photorespiration takes place only in

Aroot
Bmitochondria
Cgreen parts of the plantAnswer
Dall cells of the plant

Why

Photorespiration needs light, oxygen and chloroplasts, so it occurs in the green cells (leaves and green stems) that carry out photosynthesis. The book describes it as respiratory activity in green cells in the presence of light.

A: roots have no light and no chlorophyll. B: mitochondria take part in photorespiration, but they are found in all cells, so they do not show where it happens. D: cells without chloroplasts do not carry it out.

9

In C_4 plants, fixation of CO_2 occurs in

Apalisade tissue
Bcortex of stem
Cspongy mesophyll and bundle sheathAnswer
Dphloem tissue

Why

In C4 plants the first fixation, by PEP carboxylase, takes place in the mesophyll cells, which include the spongy mesophyll. The second fixation, by RuBisCO in the Calvin cycle, takes place in the bundle sheath cells. Option C names both sites.

A is partly right, since palisade cells are also mesophyll, but it names only one of the two fixation sites and misses the bundle sheath. B and D do not fix CO_2 in C4 photosynthesis.

10

ATP synthesis during light reactions is

Aoxidative
Bphotolysis
Csubstrate phosphorylation
DphotophosphorylationAnswer

Why

ATP made in the light reactions, using energy from light to drive a proton gradient across the thylakoid membrane, is photophosphorylation. The book states that this is the most important event in the light reaction.

A is oxidative phosphorylation, which takes place in mitochondria in respiration. B, photolysis, is the splitting of water, which releases O_2 and electrons, not ATP. C, substrate level phosphorylation, occurs in glycolysis and the Krebs cycle.

11

In C_3 plants first stable product of photosynthesis during dark reaction is

APGAAnswer
BG3P
CRuBP
Doxaloacetate

Why

The first stable compound of the Calvin cycle is the three-carbon 3-phosphoglycerate (PGA), which is why the cycle is also called the C3 pathway. The book says so directly.

B, G3P, is formed later, by reduction of PGA. C, RuBP, is the acceptor that is used up and regenerated. D, oxaloacetate, is the first product of the C4 pathway, not of C3 plants.

12

The diagram shows the Krebs cycle. At which numbered stages does decarboxylation take place?

Diagram as printed: pyruvate, with arrow 1, gives acetylcoenzyme A; citrate, with arrow 2, gives a 5C compound; the 5C compound, with arrow 3, gives a 4C compound; the 4C compound, with arrow 4, gives oxaloacetate. A coenzyme feeds into the cycle at the start.

A1 and 2
B1, 2 and 3Answer
C1, 3 and 4
D1, 2, 3 and 4

Why

Decarboxylation removes a carbon as CO_2. Stage 1 turns pyruvate (3C) into acetyl CoA (2C), stage 2 turns citrate (6C) into a 5-carbon compound, and stage 3 turns the 5-carbon compound into a 4-carbon compound (succinyl CoA). Each of these releases one CO_2.

Stage 4 turns the 4-carbon compound (malate) into oxaloacetate by dehydrogenation (oxidation). No carbon is lost, so it is not decarboxylation. Option A misses stage 3, and C and D wrongly include stage 4.

Short answer questionsp. 130

13 marks

What is electromagnetic spectrum?

Solution

The electromagnetic spectrum is the full range of electromagnetic radiation in the universe, arranged by wavelength (or frequency). It runs from very short gamma rays and X-rays, through ultraviolet, visible light, infrared and microwaves, to very long radio waves.

Visible light is only a small part of it, between about 380 nm and 750 nm.
It is the part that the eye sees and that photosynthetic pigments absorb (about 400 to 700 nm), so it drives photosynthesis.
All these forms are energy travelling as waves; light energy is the ultimate energy source for living things.

23 marks

Explain 'action spectrum' of photosynthesis.

Solution

The action spectrum is a graph of the rate of photosynthesis (relative photosynthetic rate) plotted against the wavelength of light. The plant is lit with one colour at a time and its rate of photosynthesis is measured for each colour.

The graph shows how effective each wavelength is for photosynthesis.
Blue (about 430 nm) and red (about 670 nm) give the highest rates; green (around 500 to 550 nm) gives the lowest.
Its shape follows the absorption of the pigments in the plant, so it is a measure of the effect of light, not of the light alone.

33 marks

What are the types of chlorophyll?

Solution

Higher plants and algae have chlorophyll-a and chlorophyll-b, which are the two major types; other types occur in algae and bacteria.

Chlorophyll-a is bluish green and is found in all photosynthetic organisms except photosynthetic bacteria. It is the only pigment in the reaction centre.
Chlorophyll-b is yellowish green and is found in all photosynthetic organisms except brown and red algae. It is an accessory pigment.
Algae have chlorophyll-c, chlorophyll-d and chlorophyll-e, and photosynthetic bacteria have bacteriochlorophyll.

43 marks

What is the importance of carotene?

Solution

Carotene is an accessory pigment. It absorbs blue and violet light strongly and transfers the energy to chlorophyll-a, so it widens the range of light used in photosynthesis.
It protects chlorophyll-a from excess light: some carotenoids absorb and disperse surplus light energy instead of passing it to chlorophyll-a, which prevents damage.
Carotene gives the orange and red colour of some flowers and fruits, and attracts insects and birds for pollination and seed dispersal.

53 marks

Describe 'absorption spectrum' in photosynthesis.

Solution

The absorption spectrum of a pigment is a graph of the percentage of light absorbed at each wavelength, obtained with a spectrophotometer. The graph is drawn for one pigment at a time.

Chlorophyll-a and chlorophyll-b absorb most in the violet-blue region (400 to 470 nm) and the orange-red region (630 to 660 nm).
Carotenoids absorb most at about 430 to 500 nm, in the blue region.
Green and yellow light are absorbed least, so they are reflected, which is why plants look green.

63 marks

What is photosystem? Explain.

Solution

A photosystem is a cluster of pigment molecules, with associated proteins, embedded in the thylakoid membrane of the chloroplast. It absorbs light and converts it to chemical energy.

The antenna complex is the outer part. It contains accessory pigments (chlorophyll-b and carotenoids) that absorb light and pass the energy to the centre.
The reaction centre is the central part. It contains only chlorophyll-a and proteins; when its chlorophyll receives the energy, it loses electrons to the primary electron acceptor.
There are two types: PS I (reaction centre P700) and PS II (reaction centre P680), named in the order they were discovered.

73 marks

What is the role of carbon dioxide in photosynthesis?

Solution

Carbon dioxide is a raw material of photosynthesis. It is the carbon source from which plants make organic compounds, so plants are called autotrophs.
In the light independent (Calvin) cycle, CO_2 combines with RuBP, with the help of RuBisCO, and is reduced to 3-phosphoglycerate and then to G3P, which is used to make glucose and other organic compounds.
The rate of photosynthesis depends on CO_2 concentration; it rises as CO_2 rises, but falls when CO_2 is above about 1 percent because the stomata close.

83 marks

How it was confirmed that 'plants split water as a source of hydrogen releasing hydrogen as a byproduct'?

Solution

Van Niel (1931) studied photosynthetic bacteria that make carbohydrate from CO_2 but release no oxygen. He hypothesized that plants split water as the source of hydrogen, releasing oxygen as a by-product.
In 1940, ^{18}O, a heavy isotope of oxygen, was first used in biological research. Water labelled with ^{18}O was added to an alga suspension, and the oxygen evolved was found to be radioactive.
In a second experiment, CO_2 labelled with ^{18}O gave oxygen with no label. So the evolved oxygen came from water, not from carbon dioxide, which confirmed the hypothesis.

93 marks

What is the importance of G3P?

Solution

G3P (glyceraldehyde 3-phosphate) is the first sugar made in photosynthesis. In the Calvin cycle it is converted into glucose phosphate, which then gives glucose, fructose, sucrose, maltose and starch.
In respiration, G3P links the preparatory phase and the payoff phase of glycolysis. It is then converted to pyruvate, the end product of glycolysis and raw material for both aerobic and anaerobic respiration.
G3P is an intermediate for converting one biological molecule into another: carbohydrates can become lipids, and lipids can be converted into carbohydrates through G3P. Proteins can also be converted into carbohydrates through G3P.

103 marks

What is the effect of temperature on the activities of RuBisCO?

Solution

RuBisCO can act as a carboxylase (fixing CO_2 in the Calvin cycle) and as an oxygenase (fixing O_2 in photorespiration). Its activity depends on the relative concentrations of O_2 and CO_2 in the leaf.
At high temperature, plants close their stomata to save water. CO_2 is then used up inside the leaf while O_2 from photosynthesis cannot escape, so the O_2 to CO_2 ratio rises.
With more O_2 and less CO_2, the oxygenase activity of RuBisCO increases and photorespiration increases, so less carbon is fixed.

Note. The book's text on section 4.3.3 gives the O_2 to CO_2 argument but does not state the temperature link explicitly. The answer adds the stomatal closure step, which is the reason temperature matters.

113 marks

What are the disadvantages of photorespiration?

Solution

It reduces the rate of photosynthesis. In most plants it lowers the amount of carbon fixed into carbohydrate by about 25 percent, because CO_2 fixed by photosynthesis is released again.
It wastes energy. Photorespiration needs O_2 and produces CO_2 and water, but no ATP is made, so the plant spends energy on a pathway that gains nothing.
In hot tropical climates it lowers dry matter production and crop yield, because up to 5 percent of the photosynthetic potential may be lost.

123 marks

How photorespiration evolved?

Solution

The active site of RuBisCO evolved to bind both CO_2 and O_2. Photorespiration is a side effect of this double affinity.
Originally there was almost no oxygen in the atmosphere when the earth was established, so RuBisCO used only its carboxylase activity to bind CO_2, and photorespiration was not a problem.
Photorespiration began when O_2 started to accumulate in the atmosphere, because RuBisCO then began to bind O_2 as well.

133 marks

Write the differences between:

Solution

(a) chlorophyll a and chlorophyll b

Feature	Chlorophyll a	Chlorophyll b
Colour	Bluish green	Yellowish green
Group on ring II	Methyl (CH_3)	Aldehyde (CHO)
Molecular formula	C_{55}H_{72}O_5N_4Mg	C_{55}H_{70}O_6N_4Mg
Found in	All photosynthetic organisms except photosynthetic bacteria	All except brown and red algae and photosynthetic bacteria
Role	Main pigment of the reaction centre	Accessory pigment that passes energy to chlorophyll-a

(b) carotene and xanthophyll

Feature	Carotene	Xanthophyll
Colour	Orange or red	Yellow
Most important example	Beta (B) carotene, the most widespread carotene	Lutein, the most widely distributed xanthophyll
Where it is seen	In all photosynthetic eukaryotes; in flowers and fruits	In leaves; it gives the yellow colour of foliage in autumn
Role	Accessory pigment that passes energy to chlorophyll-a and protects it from excess light	Accessory pigment; it gives the yellow colour of leaves in autumn

(c) action spectrum and absorption spectrum

Feature	Action spectrum	Absorption spectrum
What is plotted	Rate of photosynthesis against wavelength	Percentage of light absorbed by one pigment against wavelength
Measured with	The whole plant, one colour at a time	A spectrophotometer, for a single pigment
Shows	How effective each colour is for photosynthesis	Which wavelengths a pigment can absorb

(d) absorption spectrum of chlorophyll a and b

Feature	Chlorophyll a	Chlorophyll b
Blue absorption peak	About 430 nm (read from Fig 4.4)	About 450 nm (slightly longer wavelength)
Red absorption peak	About 660 nm; the taller peak	About 645 nm; lower peak
Cause of difference	Molecular formula, structure and physical properties, as the book explains	Same cause: the two molecules differ in formula, structure and physical properties

The peak positions are read approximately from Fig 4.4 of the book.

(e) antenna complex and reaction centre

Feature	Antenna complex	Reaction centre
Position in photosystem	Peripheral part	Central part
Pigments	Accessory pigments: chlorophyll-b and carotenoids	Only chlorophyll-a and associated proteins
Function	Absorbs light and passes the energy to the reaction centre	Converts light energy to chemical energy by losing excited electrons

(f) photosystem I and photosystem II

Feature	Photosystem I (PS I)	Photosystem II (PS II)
Reaction centre	P700	P680
Optimal absorption	700 nm	680 nm
Electron source and fate	Receives electrons from PS II via the chain; electrons go to ferredoxin and NADP^+	Gets its electrons from the photolysis of water; releases O_2

Both are used in non-cyclic photophosphorylation; PS I alone is used in cyclic photophosphorylation.

(g) light dependent reaction and light independent reaction of photosynthesis

Feature	Light dependent reaction	Light independent reaction
Site	Thylakoid membranes (grana)	Stroma
Needs light	Yes	No, but it needs the products of the light reaction
Inputs	Water, light, NADP^+, ADP and Pi	CO_2, ATP and NADPH
Outputs	ATP, NADPH and O_2	G3P, which becomes sugar; ADP, Pi and NADP^+ are returned

(h) oxidative phosphorylation and photophosphorylation

Feature	Oxidative phosphorylation	Photophosphorylation
Energy source	Energy released by oxidation of NADH and FADH_2 in the electron transport chain	Energy of light absorbed by the photosystems
Site	Inner mitochondrial membrane	Thylakoid membrane of the chloroplast
Occurs in	Aerobic respiration	Light dependent reaction of photosynthesis

(i) cyclic photophosphorylation and non-cyclic photophosphorylation

Feature	Cyclic	Non-cyclic
Photosystems used	PS I only	PS II and PS I
Electron path	Electrons return to P700	Electrons leave the system and reach NADP^+
Products	ATP only	ATP, NADPH and O_2
Photolysis of water	Does not occur	Occurs, replacing electrons lost by PS II

(j) C_4 carbon fixation and C_3 carbon fixation

Feature	C4 fixation	C3 fixation
First product	Oxaloacetate (4C), from PEP and CO_2	3-phosphoglycerate (3C), from RuBP and CO_2
Enzyme	PEP carboxylase in mesophyll cells	RuBisCO in the Calvin cycle
Where the Calvin cycle occurs	Bundle sheath cells, where CO_2 is concentrated	Mesophyll cells
Example plants	Maize, sugarcane, sorghum	Plants that use only the Calvin cycle (C3 plants)

(k) lactic acid fermentation and alcoholic fermentation

Feature	Lactic acid fermentation	Alcoholic fermentation
Products	Lactic acid	Ethanol and CO_2
Key step	Pyruvate is reduced by NADH to lactic acid	Pyruvate loses CO_2 to acetaldehyde, which is reduced by NADH to ethanol
Organisms	Some bacteria and muscle cells of animals	Yeast

(l) Calvin cycle and Krebs cycle

Feature	Calvin cycle	Krebs cycle
Process	Light independent reaction of photosynthesis	Aerobic respiration
Site	Stroma of the chloroplast	Matrix of the mitochondrion
Starting compound	RuBP (5C) with CO_2	Acetyl CoA (2C) with oxaloacetate (4C)
Main products	G3P, which forms sugars	CO_2, NADH, FADH_2 and ATP (as GTP)
Energy and coenzymes	Uses ATP and NADPH	Produces NADH and FADH_2

(m) oxidative phosphorylation and substrate level phosphorylation

Feature	Oxidative phosphorylation	Substrate level phosphorylation
Mechanism	Chemiosmosis: ATP synthase uses a proton gradient	Phosphate is transferred directly from an organic substrate to ADP
Enzyme	ATP synthase	Kinases, such as phosphoglycerate kinase and pyruvate kinase
ATP made per glucose	32 ATP (of the 36)	4 ATP (2 net from glycolysis, 2 from the Krebs cycle)

Extensive answer questionsp. 130–131

1

What is photosynthesis? Explain the role of light in photosynthesis.

Solution

Definition

Photosynthesis is a redox process in which carbon dioxide (an oxidized form of carbon) is reduced to glucose (a reduced form of carbon), and water is oxidized to oxygen. It is an energy conversion process: light energy is trapped and stored in the energy-rich glucose made from energy-poor \ce{CO2} and \ce{H2O}.

\ce{6CO2 + 12H2O ->[light][chlorophyll] C6H12O6 + 6O2 + 6H2O}

Role of light

Light is the ultimate source of energy for photosynthesis. Pigments absorb it and use it to make the high-energy compounds ATP and NADPH, which then drive the synthesis of sugar in the Calvin cycle. Light has five roles:

Excitation of pigments: photons are absorbed by chlorophyll-a and the accessory pigments of the antenna complex, and the energy is passed to the reaction centre (P680 in PS II, P700 in PS I).
Electron excitation: the energy excites the electrons of the reaction-centre chlorophyll-a, which leave it and enter electron transport chains.
Photolysis of water: light splits water (in PS II), giving electrons that replace those lost, protons and oxygen.
Production of ATP and NADPH: the energy of the electron flow drives photophosphorylation (ATP) and the reduction of NADP^+ (NADPH).
Colour of light: only some wavelengths are effective. The action spectrum shows that blue (about 430 nm) and red (about 670 nm) light are the most effective.

Light energy is trapped by the photosystems and stored in ATP and NADPH, which drive the Calvin cycle that makes sugar.

2

Describe the structure of chlorophyll.

Solution

Types and formula

Higher plants have two major types of chlorophyll: chlorophyll-a, a bluish green pigment found in all photosynthetic organisms except photosynthetic bacteria, and chlorophyll-b, a yellowish green pigment found in all of them except brown and red algae. Their molecular formulas are \mathrm{C_{55}H_{72}O_5N_4Mg} (chlorophyll-a) and \mathrm{C_{55}H_{70}O_6N_4Mg} (chlorophyll-b).

Structure of a chlorophyll molecule

A chlorophyll molecule consists of a head and two tails.

Head: a porphyrin ring with a magnesium ion (Mg) at its centre. The porphyrin ring has four pyrrole rings, each with four carbon atoms and one nitrogen atom, and the four nitrogen atoms hold the Mg. The head absorbs light and is exposed on the surface of the thylakoid membrane; it is hydrophilic.
Ring II: the second pyrrole ring carries a methyl group (CH_3) in chlorophyll-a and an aldehyde group (CHO) in chlorophyll-b. This is the only chemical difference between the two.
Tails: two side chains, the phytol chain and the methyl ester. They are hydrophobic and are embedded in the thylakoid membrane, anchoring the molecule.

Chlorophyll-a: the Mg-porphyrin head absorbs light and the hydrophobic tails anchor the molecule in the thylakoid membrane.

Chlorophyll-a and chlorophyll-b compared

Feature	Chlorophyll-a	Chlorophyll-b
Colour	Bluish green	Yellowish green
Group on ring II	Methyl (CH_3)	Aldehyde (CHO)
Found in	All photosynthetic organisms except photosynthetic bacteria	All except brown and red algae and photosynthetic bacteria
Role	Reaction-centre pigment that converts light to chemical energy	Accessory pigment that passes absorbed energy to chlorophyll-a

3

Write a note on the photosynthetic pigment carotene.

Solution

What carotenoids are

Carotenes and xanthophylls together form the carotenoids, which are terpenoid lipids made of isoprenoid units. They are yellow, orange, red or brown pigments. They absorb light strongly in the blue-violet range (about 430 to 500 nm), and they are seen in leaves before leaf fall, in some flowers and in fruits.

Carotene

Carotenes are orange or red pigments found in all photosynthetic eukaryotes. The most widespread and important carotene is beta (B) carotene.

Carotene and xanthophyll compared

Feature	Carotene	Xanthophyll
Colour	Orange or red	Yellow
Example	Beta (B) carotene, the most widespread	Lutein, widely distributed
Visible effect	Orange and red colour of some flowers and fruits	Yellow colour of foliage in autumn

Functions of carotene

Accessory pigment: carotene absorbs light that chlorophyll-a cannot absorb well, then transfers the energy to chlorophyll-b and chlorophyll-a. It cannot itself convert light into chemical energy.
Photoprotection: carotenoids protect chlorophyll-a from excess light. They absorb and disperse surplus light energy instead of passing it on, which prevents damage.
Attraction: the bright colour attracts insects and birds for pollination and seed dispersal.
Human eye (science tidbit in the book): lutein, zeaxanthin and meso-zeaxanthin in the macula absorb up to 90 percent of blue and violet light, which protects the retina from oxidative damage.

Carotene passes the energy it absorbs to chlorophyll-a, the pigment that converts light energy into chemical energy.

4

Explain the arrangement of photosystems.

Solution

Photosystems

For efficient absorption and use of light, the photosynthetic pigments are embedded in the surface of the thylakoid membrane in clusters called photosystems. Each photosystem has two parts: the peripheral antenna complex and the central reaction centre.

Antenna complex: accessory pigments (chlorophyll-b and carotenoids) that absorb light of many wavelengths and pass the energy to the reaction centre.
Reaction centre: only chlorophyll-a and associated proteins. Its chlorophyll-a has an optimal absorption near 660 nm, and it converts light energy into chemical energy by losing excited electrons to a primary electron acceptor.

Path of energy

A photon is absorbed by an antenna pigment.
The energy is passed from pigment to pigment until it reaches the reaction centre.
The chlorophyll-a of the reaction centre is excited and loses its electrons to the primary electron acceptor, leaving an electron hole.
The electrons pass into an electron transport chain, and the hole is later filled by electrons from water (PS II) or from the chain (PS I).

A photosystem: many antenna pigments funnel light energy to the reaction centre of chlorophyll-a in the thylakoid membrane.

Photosystem I and photosystem II

Feature	Photosystem I	Photosystem II
Reaction centre	P700	P680
Optimal absorption	700 nm	680 nm
Electron fate	Electrons go to ferredoxin and then NADP^+	Electrons pass to plastoquinone; holes are filled from water

5

Describe the role of water in photosynthesis.

Solution

Water as donor

Water is a raw material of photosynthesis and is the hydrogen and electron donor. In the light dependent phase it is split by photolysis. Its electrons replace those lost by P680, its protons build a gradient for ATP synthesis, and its oxygen is released to the air.

\text{H}_2\text{O} \xrightarrow{\text{light}} 2\text{H}^+ + 2e^- + \tfrac12\,\text{O}_2

Steps of photolysis

Light energy excites P680 in PS II. Its electrons are captured by the primary acceptor, and P680 is left with electron holes, making it a strong oxidizing agent.
The water-splitting (oxygen-evolving) enzyme at PS II splits water into 2\mathrm{H}^+, 2e^- and \tfrac12\mathrm{O_2}.
The two electrons fill the holes of P680, so that PS II can absorb light again.
The protons are released into the thylakoid lumen, adding to the proton gradient that drives ATP synthesis.
The electrons and protons are taken up temporarily by NADP^+ to form NADPH, which reduces \ce{CO2} in the Calvin cycle.
The oxygen is released into the atmosphere.

Photolysis of water supplies electrons to PS II, protons to the thylakoid lumen and oxygen to the air.

Evidence that water is the source of oxygen

Van Niel (1931) studied bacteria that make carbohydrate from \ce{CO2} without releasing oxygen, and suggested that plants split water as the source of hydrogen. In 1940, labelled oxygen-18 showed the source:

Experiment	Labelled molecule	Oxygen released
1	Water with ^{18}O added to algae	Radioactive (labelled), so it came from water
2	Carbon dioxide with ^{18}O added	Not labelled, so it did not come from carbon dioxide

6

Describe the mechanism of photosynthesis.

Solution

Two phases

Photosynthesis takes place in two phases. The light dependent phase (light reaction) takes place in the thylakoid membranes and needs light. It makes ATP and NADPH, and releases oxygen from water. The light independent phase (dark reaction) takes place in the stroma; it does not need light directly, but it needs the ATP and NADPH from the light reaction to fix carbon dioxide into G3P.

The two phases of photosynthesis: the light dependent phase makes ATP and NADPH; the light independent phase uses them to make G3P from CO_2.

Light dependent phase

Absorption of light by PS II, excitation of P680 and photolysis of water.
Electron transport from PS II to PS I through plastoquinone, the cytochrome b_6-f complex and plastocyanin, with the pumping of protons into the thylakoid lumen.
Chemiosmosis: the proton gradient drives ATP synthase, which makes ATP from ADP and Pi (photophosphorylation).
Absorption of light by PS I and reduction of NADP^+ to NADPH through ferredoxin and NADP reductase.
Cyclic flow, which uses PS I only and makes ATP without NADPH or oxygen, can occur when ATP is needed.

Light independent phase

Carbon fixation: RuBP (5C) combines with CO_2 under RuBisCO to form an unstable 6-carbon compound, which splits into two molecules of 3-phosphoglycerate (PGA).
Reduction: PGA is phosphorylated by ATP and then reduced by NADPH to glyceraldehyde 3-phosphate (G3P). One G3P leaves the cycle to make sugars.
Regeneration: the remaining G3P regenerates RuBP with the help of ATP, so that the cycle can continue.

Inputs and outputs

Phase	Location	Inputs	Outputs
Light dependent	Thylakoid membrane	Light, H_2O, ADP, Pi, NADP^+	ATP, NADPH, O_2
Light independent	Stroma	CO_2, ATP, NADPH	G3P (then glucose and other compounds), ADP, Pi, NADP^+

Overall equation

\ce{6CO2 + 12H2O ->[light][chlorophyll] C6H12O6 + 6O2 + 6H2O}

72026 Annual I, Set B, Q.62026 Annual I, Set D, Q.6 OR2025 Annual I, Set D, Q.2 (vii) OR

Explain in detail the light dependent phase of photosynthesis?

Solution

Overview

The light dependent phase takes place on the thylakoid membranes. Light energy is used to split water, to move electrons through two photosystems and to make ATP (by chemiosmosis) and NADPH. Oxygen is released as a by-product. The main route is non-cyclic electron flow, which is shown as a zigzag (Z) scheme.

Steps of non-cyclic electron flow

Absorption of light by PS II. Antenna pigments pass energy to P680, which loses two excited electrons to the primary acceptor of PS II and becomes a strong oxidant.
Photolysis of water. The water-splitting enzyme gives electrons to refill the holes in P680, releases protons into the thylakoid lumen, and releases oxygen.
Electron transport from PS II to PS I. The electrons pass to plastoquinone (PQ), then to the cytochrome b_6-f complex, which pumps protons from the stroma into the lumen, and then to plastocyanin (PC).
Chemiosmosis. The proton gradient is high in the lumen. Protons flow back to the stroma through ATP synthase, which joins ADP and Pi to make ATP (photophosphorylation).
Absorption of light by PS I. P700 is excited and its electrons are captured by the primary acceptor of PS I; its holes are filled by electrons from PC.
Reduction of NADP^+. The electrons pass to ferredoxin (Fd), an iron protein, and then to NADP reductase, which transfers them with H^+ to NADP^+ to form NADPH.

The Z scheme of non-cyclic photophosphorylation: electrons rise in PS II, fall through the chain, rise in PS I and are used to make NADPH.

Products

ATP, made by photophosphorylation (chemiosmosis) as protons flow through ATP synthase.
NADPH, which provides the reducing power for the Calvin cycle.
O_2, released from photolysis of water.

8

Explain in detail the light independent phase of photosynthesis?

Solution

Overview

The light independent phase (dark reaction) takes place in the stroma of the chloroplast. It is called the Calvin cycle or C3 pathway. It fixes \ce{CO2} into a three-carbon compound and reduces it to G3P. It does not need light directly, but it uses the ATP and NADPH made in the light dependent phase.

The light independent phase: CO_2 is fixed, reduced with NADPH and ATP, and RuBP is regenerated with ATP.

Phase 1: carbon fixation

RuBP, a five-carbon sugar, combines with one molecule of \ce{CO2} in a reaction catalysed by RuBisCO. The product is an unstable six-carbon compound that splits into two molecules of 3-phosphoglycerate (PGA). Because the carbon of \ce{CO2} becomes part of an organic compound, this is called carbon fixation. PGA is the first stable compound, which is why this is the C3 pathway.

Phase 2: reduction

Six molecules of PGA are phosphorylated by six ATP, giving six molecules of 1,3-bisphosphoglycerate and six ADP.
Each 1,3-bisphosphoglycerate is reduced by NADPH. A phosphate is released, and six molecules of G3P are formed (six NADPH oxidized to NADP^+).
One G3P leaves the cycle to be used for sugars; the other five stay in the cycle to regenerate RuBP.

Phase 3: regeneration of RuBP

Five G3P molecules (each with three carbons) are converted, using ATP, into three molecules of ribulose phosphate (RuP, five carbons). Each RuP receives a phosphate group from an ATP to become RuBP, so the acceptor is ready for the next turn.

Summary

Phase	Process	Energy used	Key product
1 Fixation	CO_2 + RuBP, by RuBisCO	None	2 PGA (3C each)
2 Reduction	PGA to G3P	6 ATP and 6 NADPH	6 G3P (1 leaves)
3 Regeneration	5 G3P to 3 RuBP	3 ATP	RuBP for the next turn

Overall, three molecules of \ce{CO2}, six NADPH and nine ATP are needed to release one molecule of G3P.

92025 Annual I, Set B, Q.2 (v)

Describe cyclic photophosphorylation.

Solution

Definition

Cyclic photophosphorylation is the production of ATP by cyclic electron flow, in which electrons leave PS I, pass through an electron transport chain and return to the chlorophyll of PS I. It uses only PS I. The cycle is switched on temporarily when the cell has too much NADPH and too little ATP for the Calvin cycle.

Cyclic photophosphorylation: electrons leave P700, pass through ferredoxin, the cytochrome b6-f complex and plastocyanin, and return to P700.

Steps

Light excites the electrons of P700 in the PS I reaction centre, which are captured by its primary acceptor.
The electrons move through ferredoxin to the cytochrome b_6-f complex, which pumps protons into the thylakoid lumen, and then to plastocyanin.
The proton gradient drives ATP synthase, so ATP is made by chemiosmosis.
The electrons, having lost their energy, return to P700 and the cycle repeats.

Cyclic and non-cyclic flow compared

Feature	Cyclic	Non-cyclic
Photosystems used	PS I only	PS II and PS I
Products	ATP only	ATP, NADPH and O_2
Photolysis of water	Does not occur	Occurs
Electron path	Returns to P700	Leaves the system and reaches NADP^+

10

Describe Calvin cycle.

Solution

Overview

The Calvin cycle is the series of reactions in which \ce{CO2} is reduced to G3P. It was worked out by Melvin Calvin and co-workers at the University of California. It takes place in the stroma and has three phases: carbon fixation, reduction and regeneration of the \ce{CO2} acceptor (RuBP).

The Calvin cycle: CO_2 is fixed onto RuBP, PGA is reduced to G3P, and five of six G3P regenerate RuBP while one leaves as sugar.

Phases

Carbon fixation: RuBP combines with CO_2 under RuBisCO to give two molecules of 3-PGA.
Reduction: six 3-PGA take phosphate from six ATP to form 1,3-bisphosphoglycerate, which is reduced by six NADPH to six G3P. One G3P leaves the cycle.
Regeneration: five G3P (15 carbons) form three RuP (15 carbons), using three ATP. RuP is phosphorylated to RuBP, which is the CO_2 acceptor again.

Net equation

The book gives the overall equation as shown below. Its phosphate count needs a correction: G3P keeps one phosphate group, and nine ATP give nine ADP, so the phosphorus balance needs 8 Pi, not 9. The water term is copied as printed.

3\,\text{CO}_2 + 6\,\text{NADPH} + 9\,\text{ATP} \rightarrow \text{G3P} + 3\,\text{H}_2\text{O} + 9\,\text{ADP} + 8\,\text{Pi} + 6\,\text{NADP}^+

Note. The book's overall Calvin cycle equation prints 9Pi; the phosphate balance gives 8 Pi because G3P carries one phosphate.

11

Describe the kinds of cellular respiration.

Solution

Definition

Cellular respiration is a series of complex oxidation-reduction reactions by which living cells obtain energy from the breakdown of organic matter, usually glucose, which is the respiratory fuel. Its first stage, glycolysis, is common to aerobic and anaerobic respiration.

Kinds of respiration

Feature	Aerobic respiration	Anaerobic respiration
Oxygen	Needed (abundant atmospheric O_2)	Absent
Final products	CO_2 and H_2O	Lactic acid, or ethanol and CO_2
ATP per glucose	36 (eukaryotes) or 38 (prokaryotes)	2
Organisms	Most organisms	Anaerobic bacteria, yeast, muscle cells of vertebrates, cells of higher plants

Kinds of cellular respiration: glycolysis is common to all; in the absence of oxygen the pyruvate is fermented, and in its presence it enters the aerobic stages.

Anaerobic respiration (fermentation)

Lactic acid fermentation: glycolysis is followed by reduction of pyruvate by NADH to lactic acid. It occurs in some bacteria and in muscles during strenuous exercise; the lactic acid causes fatigue.
Alcoholic fermentation: glycolysis is followed by decarboxylation of pyruvate to acetaldehyde and reduction of acetaldehyde by NADH to ethanol. It occurs in yeast.

\begin{aligned}\ce{C6H12O6 &-> 2C3H6O3}\quad\text{(lactic acid, 2 ATP)}\\ \ce{C6H12O6 &-> 2C2H5OH + 2CO2}\quad\text{(ethanol, 2 ATP)}\end{aligned}

Aerobic respiration

Aerobic respiration oxidizes glucose completely to carbon dioxide and water. It has four stages: glycolysis, oxidation of pyruvate (link reaction), the Krebs cycle, and the electron transport chain with chemiosmosis.

\ce{C6H12O6 + 6O2 -> 6CO2 + 6H2O}\quad (\text{about 36 or 38 ATP})

122026 Annual I, Set B, Q.2 (ii) OR2026 Annual I, Set D, Q.2 (ii) OR

Give an account of 'Glycolysis'.

Solution

Definition and location

Glycolysis is the breakdown of glucose (or a similar hexose sugar) into two molecules of pyruvate, through a series of enzyme-catalysed reactions that release some energy as ATP and reduced coenzymes (NADH). It takes place in the cytoplasm and is the first stage of both aerobic and anaerobic respiration.

Two phases

Glycolysis has a preparatory phase, which uses 2 ATP, and a payoff phase, which makes ATP and NADH.

The ten steps of glycolysis: the preparatory phase (steps 1 to 5) uses ATP; the payoff phase (steps 6 to 10) makes ATP and NADH.

Steps

Phosphorylation: hexokinase (or glucokinase) transfers a phosphate from ATP to glucose, giving glucose 6-phosphate; ATP becomes ADP.
Isomerization: phosphohexose isomerase converts glucose 6-phosphate to fructose 6-phosphate.
Phosphorylation: phosphofructokinase transfers a second phosphate from ATP, giving fructose 1,6-bisphosphate.
Splitting: aldolase splits fructose 1,6-bisphosphate into G3P and dihydroxyacetone phosphate (DHAP).
Isomerization: phosphotriose isomerase converts DHAP into G3P, so there are now two molecules of G3P.
Dehydrogenation and phosphorylation: G3P dehydrogenase oxidizes each G3P, reducing NAD^+ to NADH, and adds inorganic phosphate to form 1,3-bisphosphoglycerate.
Substrate level phosphorylation: phosphoglycerate kinase transfers a phosphate from 1,3-bisphosphoglycerate to ADP, giving 3-phosphoglycerate and ATP.
Isomerization: phosphoglycerate mutase moves the phosphate from C3 to C2, giving 2-phosphoglycerate.
Dehydration: enolase removes water from 2-phosphoglycerate, giving phosphoenolpyruvate (PEP).
Substrate level phosphorylation: pyruvate kinase transfers the phosphate from PEP to ADP, giving pyruvate and ATP.

Net yield per glucose

Item	Produced	Used	Net
ATP	4	2	2
NADH	2	0	2
Pyruvate	2	0	2

132025 Annual I, Set B, Q.2 (xi)

Explain oxidation of pyruvate.

Solution

Transport into the mitochondrion

Pyruvate is produced in the cytosol. Because it is charged, it enters the mitochondrion by active transport through the transport protein pyruvate translocase. Inside the matrix it does not enter the Krebs cycle directly; it first undergoes the oxidation of pyruvate, also called the link reaction, which links glycolysis to the Krebs cycle.

Oxidation of pyruvate: pyruvate loses CO2, is oxidized to an acetyl group, and is joined to coenzyme A to form acetyl CoA.

Three steps

Decarboxylation: pyruvate loses one molecule of CO_2 and becomes acetaldehyde (2 carbons).
Dehydrogenation (oxidation): NAD^+ removes hydrogen from acetaldehyde. This produces NADH and a two-carbon acetyl group.
Addition of coenzyme A: the acetyl group combines with coenzyme A to form acetyl CoA, which enters the Krebs cycle.

\ce{C3H4O3 + NAD+ + CoA-SH -> CH3CO-SCoA + CO2 + NADH + H+}

Summary

The three steps are catalysed by the pyruvate dehydrogenase complex, which consists of three enzymes.
Per glucose, two pyruvate give two acetyl CoA, two CO_2 and two NADH.
No ATP is made in this stage.

14

Explain Krebs cycle.

Solution

Overview

The Krebs cycle, also called the citric acid cycle or tricarboxylic acid (TCA) cycle, was discovered by Sir Hans Krebs. It takes place in the matrix of the mitochondrion. Acetyl CoA (2C) combines with oxaloacetate (4C) to form citrate (6C), which is then oxidized step by step, releasing CO_2, and regenerating oxaloacetate.

The Krebs cycle: acetyl CoA enters at step 1; CO2 is released at steps 4 and 5; GTP, NADH and FADH2 are made; oxaloacetate is regenerated.

The nine steps

Synthesis: acetyl CoA and water combine with oxaloacetate to form citrate; citrate synthetase catalyses it and CoA is released.
Dehydration: aconitase removes a water molecule from citrate to give cis-aconitate.
Hydration: aconitase adds water to cis-aconitate to give isocitrate.
Oxidative decarboxylation: isocitrate dehydrogenase oxidizes isocitrate, reducing NAD^+ to NADH, and releases CO_2, forming alpha-ketoglutarate (5C).
Oxidative decarboxylation and addition of CoA: the alpha-ketoglutarate dehydrogenase complex oxidizes alpha-ketoglutarate, releases CO_2, makes NADH and joins CoA, forming succinyl CoA (4C).
Substrate level phosphorylation: succinyl thiokinase removes CoA and forms succinate. GDP and Pi form GTP, which gives ATP with ADP.
Dehydrogenation: succinate dehydrogenase oxidizes succinate to fumarate, and FAD is reduced to FADH_2.
Hydration: fumarase adds water to fumarate, giving malate.
Dehydrogenation: malate dehydrogenase oxidizes malate to oxaloacetate, reducing NAD^+ to NADH. Oxaloacetate is ready to take up another acetyl CoA.

Inputs and outputs

Inputs (per glucose, 2 turns)	Outputs (per glucose, 2 turns)
2 acetyl groups; 2 ADP and 2 Pi; 6 NAD^+; 2 FAD	4 CO_2; 2 ATP (as GTP); 6 NADH; 2 FADH_2

152025 Annual I, Set B, Q.4 OR

Explain electron transport chain.

Solution

Components

NADH-Q reductase (NADH dehydrogenase complex, complex I), containing FMN and an Fe-S complex.
FADH_2 dehydrogenase complex (complex II).
Coenzyme Q (ubiquinone), a mobile carrier.
Cytochrome reductase complex (complex III), containing cytochrome b, Fe-S and cytochrome c_1.
Cytochrome c, a mobile carrier.
Cytochrome oxidase complex (complex IV), containing cytochrome a, cytochrome a_3 and copper.

Passage of electrons

NADH is oxidized at complex I, which passes its electrons and protons to FMN and then to the Fe-S complex. Electrons then move to coenzyme Q. FADH_2 gives its electrons to complex II, which also passes them to coenzyme Q. From Q, the electrons go to complex III, then to cytochrome c, then to complex IV, and finally to oxygen. Oxygen, the final acceptor, combines with the electrons and protons to form water.

The electron transport chain in the inner mitochondrial membrane: electrons pass from NADH or FADH2 through complexes I to IV to oxygen, and protons are pumped at complexes I, III and IV.

Energy and proton pumping

The electrons lose energy at each step (the chain is a downhill journey). Part of this energy pumps protons from the matrix to the intermembrane space at three sites:

Site	Carriers	Enzyme
Site 1	FMN to CoQ	NADH-dehydrogenase complex I
Site 2	Cytochrome b to cytochrome c_1	Cytochrome reductase complex III
Site 3	Cytochrome a to cytochrome a_3	Cytochrome oxidase complex IV

162026 Annual I, Set B, Q.2 (xiii) OR2026 Annual I, Set D, Q.2 (xiii) OR2025 Annual I, Set D, Q.3 OR

Explain chemiosmosis and oxidative phosphorylation.

Solution

Definitions

Oxidative phosphorylation is the synthesis of ATP using the energy released when NADH and FADH_2 are oxidized in the electron transport chain. Chemiosmosis is the mechanism: the energy of electron flow is used to pump protons, which build a gradient that drives ATP synthase.

Chemiosmosis: protons pumped into the intermembrane space flow back through the F0 channel, and F1 uses the flow to make ATP from ADP and Pi.

ATP synthase

F_0 is embedded in the membrane and forms a channel for protons from the intermembrane space to the matrix.
F_1 is the head-like particle that projects into the matrix. It catalyses the joining of ADP and Pi to make ATP.

Steps

The electron transport chain pumps protons from the matrix to the intermembrane space at three sites.
A high proton concentration builds up in the intermembrane space. This is a gradient of protons (and an electrical charge) across the inner membrane.
Protons flow back into the matrix through the F_0 channel, down the gradient.
The flow turns F_0, which causes F_1 to join ADP and Pi, so ATP is made by a high-energy bond.

Yield

Oxidation of one NADH produces 3 ATP, and oxidation of one FADH_2 produces 2 ATP. The total from aerobic respiration is 32 ATP by chemiosmosis, out of 36 ATP per glucose in eukaryotes.

17

Describe substrate level phosphorylation.

Solution

Definition

Substrate level phosphorylation is the synthesis of ATP by the direct transfer of an inorganic phosphate from an organic (phosphorylated) substrate to ADP, in a reaction catalysed by an enzyme. It is a coupling of exergonic reactions to the synthesis of ATP, and it accounts for only a small percentage of the ATP made by a cell.

Substrate level phosphorylation: an enzyme transfers a phosphate from a substrate directly to ADP, forming ATP without a proton gradient.

The three sites in aerobic respiration

Stage	Reaction	Enzyme	ATP
Glycolysis, step 7	1,3-bisphosphoglycerate to 3-phosphoglycerate	Phosphoglycerate kinase	2 (per glucose)
Glycolysis, step 10	Phosphoenolpyruvate to pyruvate	Pyruvate kinase	2 (per glucose)
Krebs cycle, step 6	Succinyl CoA to succinate (GTP, then ATP)	Succinyl thiokinase	2 (per glucose)

Net yield

Glycolysis produces four ATP by substrate level phosphorylation, but two are used in the preparatory phase, so the net is 2 ATP. The Krebs cycle adds 2 ATP. Substrate level phosphorylation therefore makes 4 ATP per glucose, and the other 32 of the 36 ATP come from chemiosmosis.

Coupling

ATP links the reactions that release energy (exergonic, such as cellular respiration) to those that need energy (endergonic, such as protein synthesis). Substrate level phosphorylation is one of the links between catabolism and anabolism.

FBISE 2025 and 2026 questions

Questions from the FBISE HSSC-I 2026 (1st Annual) and FBISE HSSC-I 2025 (1st Annual) papers that belong to this unit, answered the way an examiner expects.

2026 Annual I, Set B, MCQ 31 mark

During anaerobic respiration in human muscles, pyruvate is converted into:

AEthanol
BCarbon dioxide
CLactic acidAnswer
DGlucose

Why

In muscle during strenuous exercise, oxygen runs short and the cell uses lactic acid fermentation: pyruvate is reduced by NADH to lactic acid, and NADH returns as \ce{NAD+} so that glycolysis can continue.

Ethanol (A) and \ce{CO2} (B) are the products of alcoholic fermentation in yeast, where pyruvate is first decarboxylated. Glucose (D) is the starting material of glycolysis, not a product.

2025 Annual I, Set B, MCQ 51 mark

Which one of the following sequence of reactions is correct regarding Calvin cycle?

A3PGA \to 1-3BPG \to G3PAnswer
B3PGA \to G3P \to RuP
CG3P \to RuBP \to 3PGA
DG3P \to RuP \to 3PGA

Why

In the reduction phase of the Calvin cycle, 3-PGA takes a phosphate from ATP to become 1,3-bisphosphoglycerate, which NADPH then reduces (losing a phosphate) to G3P. So the sequence is 3PGA, 1,3-BPG, G3P (option A).

B leaves out 1,3-BPG; G3P to RuP is also only part of the regeneration, and 3PGA does not become RuP directly. C and D start from G3P and reverse the cycle: RuBP is the acceptor that combines with \ce{CO2} to make 3PGA, and G3P is not changed into RuBP in one step.

2026 Annual I, Set B, Q.2 (ii) OR3 marks

Outline steps of preparatory phase of glycolysis.

Solution

What the phase is (1 mark)

The preparatory (investment) phase is steps 1 to 5 of glycolysis in the cytoplasm. It uses 2 ATP and changes one glucose (6C) into two molecules of G3P (3C). No ATP is made and no NADH is formed in this phase.

The five steps (2 marks)

Step	Name	Enzyme	What happens
1	Phosphorylation	Hexokinase	ATP gives a phosphate to glucose: glucose 6-phosphate + ADP
2	Isomerization	Phosphohexose isomerase	Glucose 6-phosphate becomes its isomer fructose 6-phosphate
3	Phosphorylation	Phosphofructokinase	A second ATP gives a phosphate: fructose 1,6-bisphosphate + ADP
4	Splitting	Aldolase	Fructose 1,6-bisphosphate (6C) splits into two 3C sugars: G3P and dihydroxyacetone 3-phosphate
5	Isomerization	Phosphotriose isomerase	Dihydroxyacetone 3-phosphate becomes G3P, so there are two G3P

Because step 5 gives a second G3P, everything in the pay-off phase happens twice for each glucose.

2025 Annual I, Set B, Q.2 (v)1+2 = 3 marks

Why and how cyclic Photophosphorylation occurs in light reactions?

Solution

Why it occurs (1 mark)

Non-cyclic flow makes NADPH and ATP, but the Calvin cycle needs more ATP than NADPH. When NADPH rises and ATP is short, the chloroplast shifts for a while to cyclic flow, which makes only ATP, until the supply of ATP catches up with the demand.

How it occurs (2 marks)

Only PS-I is used. P700 absorbs light and its electrons are boosted to a higher energy level.
The excited electrons are captured by the primary electron acceptor of PS-I.
They pass through a chain of **ferredoxin, cytochrome b_6f complex and plastocyanin**.
As electrons pass through the cytochrome complex, protons are pumped and ATP is made by chemiosmosis.
The electrons, having lost energy, return to P700.

Figure 4.9: Cyclic electron flow during photophosphorylation. The electrons (accent) return to P700, so only ATP is produced.

2025 Annual I, Set B, Q.2 (xi)3 marks

Illustrate the conversion of pyruvic acid to acetyl CoA as link reaction between glycolysis and Krebs cycle.

Solution

Pyruvate made in the cytosol by glycolysis enters the mitochondrion by active transport through pyruvate translocase. There the pyruvate dehydrogenase complex (three enzymes) converts it to acetyl CoA, which enters the Krebs cycle. This is why it is the link reaction.

Figure 4.13: Pathway of oxidation of pyruvate. One \ce{CO2} and one NADH are released for each pyruvate.

Decarboxylation: a \ce{CO2} is removed from pyruvate (3C), forming acetaldehyde (2C).
Oxidation (dehydrogenation): \ce{NAD+} removes hydrogen from acetaldehyde, giving a two-carbon acetyl group and NADH.
Addition of coenzyme A: the acetyl group joins CoA to form acetyl CoA.

\ce{Pyruvate + NAD+ + CoA -> Acetyl CoA + NADH + CO2}

For one glucose (2 pyruvate) this gives 2 acetyl CoA, 2 NADH and 2 \ce{CO2}. No ATP is made directly.

2026 Annual I, Set B, Q.2 (xiii) OR1x3 = 3 marks

State three roles of ATP-Synthetase during chemiosmosis.

Solution

ATP synthetase (ATP synthase) sits in the inner mitochondrial membrane (and in the thylakoid membrane in chloroplasts). In chemiosmosis it does three jobs.

Proton channel: its F_0 part, embedded in the membrane, lets protons flow back down the gradient from the intermembrane space (high \ce{H+}) to the matrix (low \ce{H+}).
Uses the proton gradient: it is activated by the gradient built by the electron transport chain, and changes the energy of the flowing protons into chemical energy.
Makes ATP: its F_1 head, projecting into the matrix, joins ADP and inorganic phosphate (Pi) by a high-energy bond to form ATP.

Chemiosmosis: protons pumped into the intermembrane space flow back through the F0 channel, and F1 uses the flow to make ATP from ADP and Pi.

2025 Annual I, Set B, Q.4 OR5+2 = 7 marks

How electrons move through electron transport chain till they reach final acceptor? Explain. Draw diagram of electron transport chain.

Solution

Passage of electrons (5 marks)

After glycolysis, the link reaction and the Krebs cycle, most of the energy of glucose is held in **NADH and \ce{FADH2}. They are oxidised in the electron transport chain (ETC) in the inner mitochondrial membrane, and their electrons fall down a series of carriers to the final acceptor, oxygen. The passage is a downhill journey**: energy is released in small steps instead of all at once.

No.	Component	Further composed of
1	NADH dehydrogenase complex (I)	FMN and Fe-S
2	FADH dehydrogenase complex (II)
3	Coenzyme Q (ubiquinone)	mobile carrier
4	Cytochrome reductase complex (III)	Cyt b, Fe-S, Cyt c_1
5	Cytochrome c	mobile carrier
6	Cytochrome oxidase complex (IV)	Cyt a, Cyt a_3, copper

NADH is oxidised at complex I. Electrons and protons pass to FMN, then to the Fe-S complex, which pumps protons into the intermembrane space.
**\ce{FADH2}** is oxidised at complex II and hands its electrons to coenzyme Q (NADH electrons also reach Q).
Electrons go from Q to complex III, which passes them to cytochrome c and pumps protons.
Cytochrome c carries the electrons to complex IV (Cyt a, a_3, copper), which is an electron carrier and a proton pump.
Finally the electrons reach oxygen, the final acceptor. Oxygen combines with them and with protons to form water.

\ce{1/2 O2 + 2H+ + 2e- -> H2O}

The energy released pumps protons from the matrix into the intermembrane space at three sites: between FMN and CoQ (complex I), between Cyt b and Cyt c_1 (complex III) and between Cyt a and Cyt a_3 (complex IV). The resulting gradient is used by ATP synthetase to make ATP, so the chain is also called oxidative phosphorylation.

Diagram (2 marks)

Figure 4.16: Sequence of electron carriers in the respiratory ETC. Electrons (accent) lose energy at each step; protons are pumped at I, III and IV.

2026 Annual I, Set B, Q.62+2+2 = 6 marks

Draw events of non-cyclic photophosphorylation and explain absorption of light by PS-II and chemiosmotic synthesis of ATP during light reaction.

Solution

Events of non-cyclic photophosphorylation (2 marks)

Non-cyclic photophosphorylation is the main pathway of the light reaction. Both photosystems and two electron transport chains are used, and the electrons do not return: they leave PS-II, pass to PS-I and end up in NADPH. The zigzag path of the electrons is called the Z-scheme.

Figure 4.8: Non-cyclic photophosphorylation (Z-scheme). Each photosystem raises the electrons to a higher energy level; they lose energy on the way down, which is used to make ATP.

Products: **ATP, NADPH and \ce{O2}**. Order of carriers: PS-II (P680), primary acceptor, PQ, cytochrome b_6f, PC, PS-I (P700), primary acceptor, Fd, NADP reductase, \ce{NADP+}.

Absorption of light by PS-II (2 marks)

Light strikes the antenna complex of PS-II. Two photons are enough for one cycle of the reaction centre.
The pigments pass the energy from one to the next until it reaches the reaction centre, the special chlorophyll a called P680.
P680 absorbs the energy and two electrons are excited to a higher energy level.
The primary electron acceptor of PS-II captures the two electrons. This leaves two "electron holes" in P680, which becomes a strong oxidising agent.
Photolysis of water fills the holes: \ce{H2O} splits into 2\ce{H+}, 2\ce{e-} and \tfrac12\ce{O2}. The electrons refill P680; the protons stay in the thylakoid space and oxygen is released.

Chemiosmotic synthesis of ATP (2 marks)

Excited electrons pass from the primary acceptor to plastoquinone (PQ) and then through the **cytochrome b_6f complex**.
The cytochrome complex is a proton pump: the energy of the falling electrons pumps \ce{H+} from the stroma into the thylakoid space (lumen), so a proton gradient builds up (high \ce{H+} inside the lumen).
The gradient activates ATP synthase in the thylakoid membrane. Protons flow back to the stroma through its F_0 channel.
The flow drives F_1 to join ADP and Pi into ATP (photophosphorylation). This ATP supplies the Calvin cycle.

Figure 4.7: Chemiosmotic synthesis of ATP during the light reaction. Electron flow is shown in accent, proton movement in grey.

2026 Annual I, Set D, MCQ 31 mark

End product of anaerobic respiration in yeast is:

ALactic acid
BWater
COxygen
DEthanolAnswer

Why

Yeast carries out alcoholic fermentation: pyruvate is decarboxylated to acetaldehyde (releasing \ce{CO2}), and acetaldehyde is reduced by NADH to ethanol. Ethanol (with \ce{CO2}) is the end product.

Lactic acid (A) is the end product in muscles and some bacteria. Water (B) and oxygen (C) are not products of fermentation: water is formed only when oxygen accepts electrons in aerobic respiration, and oxygen is not made at all.

2026 Annual I, Set D, Q.2 (ii) OR3 marks

Outline steps of payoff phase of Glycolysis.

Solution

What the phase is (1 mark)

The pay-off (oxidative) phase is steps 6 to 10. It starts with 2 G3P and ends with 2 pyruvate. ATP is made by substrate level phosphorylation and NADH is made. Each step happens twice for one glucose.

The five steps (2 marks)

Step	Name	Enzyme	What happens
6	Dehydrogenation and phosphorylation	G3P dehydrogenase	G3P gives hydrogen to \ce{NAD+} (NADH forms) and takes inorganic phosphate: 1,3-bisphosphoglycerate
7	Formation of ATP	Phosphoglycerate kinase	1,3-Bisphosphoglycerate gives its phosphate to ADP: 3-phosphoglycerate + ATP
8	Isomerization	Phosphoglycerate mutase	The phosphate moves from C3 to C2: 2-phosphoglycerate
9	Dehydration	Enolase	2-Phosphoglycerate loses water: phosphoenolpyruvate (PEP)
10	Formation of ATP	Pyruvate kinase	PEP gives its phosphate to ADP: pyruvate + ATP

	Pay-off phase (steps 6 to 10)
ATP	4 ATP made (steps 7 and 10, two molecules each)
NADH	2 NADH made (step 6)
End product	2 pyruvate

2025 Annual I, Set D, Q.2 (vii) OR3 marks

Outline the process of non-cyclic photophosphorylation with the help of a labelled flow sheet.

Solution

Non-cyclic photophosphorylation uses both photosystems and two electron transport chains. Electrons flow from water to \ce{NADP+} and do not return.

The Z scheme of non-cyclic photophosphorylation: electrons rise in PS II, fall through the chain, rise in PS I and are used to make NADPH.

PS-II absorbs light. P680 loses two excited electrons to the primary acceptor.
Photolysis of water fills the holes of PS-II and releases \ce{O2} and \ce{H+}.
Electrons pass through **PQ, the cytochrome b_6f complex and PC; protons are pumped into the thylakoid space and ATP is made by chemiosmosis**.
PS-I absorbs light. P700 gives its electrons to the primary acceptor; its holes are filled by electrons from PC.
Electrons go through ferredoxin and NADP reductase to \ce{NADP+}, forming NADPH.

Products: ATP, NADPH and oxygen.

2025 Annual I, Set D, Q.2 (ix)3 marks

Differentiate between absorption spectrum of Chlorophyll- a, b and Carotenoids.

Solution

An absorption spectrum is a graph of the relative absorption of different wavelengths of light by a particular pigment. The three pigments absorb differently.

Feature	Chlorophyll a	Chlorophyll b	Carotenoids
Regions of main absorption	Violet-blue (400 to 470 nm) and orange-red (630 to 660 nm)	Violet-blue and orange-red	Blue-green, 430 to 500 nm only
Violet-blue peak	At the shorter wavelength end (about 420 nm)	At the longer wavelength end (about 460 nm)	Broad peak around 430 to 500 nm
Red peak	At the longer wavelength end (about 660 nm)	At the shorter wavelength end (about 640 nm)	None; absorbs little or no red
Reason for the difference	Methyl group on the 2nd pyrrole ring	Aldehyde group on the 2nd pyrrole ring	Different structure; accessory pigment

Absorption spectra of different pigments

Figure 4.4: Absorption spectra of chlorophyll a, chlorophyll b and the carotenoids. Chlorophylls absorb most in violet-blue and orange-red; carotenoids absorb most between about 430 and 500 nm.

2026 Annual I, Set D, Q.2 (xiii) OR1+2 = 3 marks

Write down composition and roles of parts of ATP synthetase.

Solution

Composition

ATP synthetase (ATP synthase) is a membrane enzyme complex found in the inner mitochondrial membrane (thylakoid membrane in chloroplasts). It has two parts: F_0 and F_1.

Part	Composition and position	Role
F_0	Embedded in the inner membrane; forms a channel across it	Channel through which protons flow from the intermembrane space (high \ce{H+}) to the matrix (low \ce{H+})
F_1	Head-like particle (elementary particle) that projects from the membrane into the matrix	Catalyses the joining of ADP and inorganic phosphate (Pi) into ATP, using the energy of the proton flow

Chemiosmosis: protons pumped into the intermembrane space flow back through the F0 channel, and F1 uses the flow to make ATP from ADP and Pi.

2025 Annual I, Set D, Q.3 OR5+2 = 7 marks

Explain how the movement of protons across mitochondrial membrane is involved in ATP synthesis. Also draw diagram showing movement of electrons.

Solution

Protons and ATP synthesis (5 marks)

ATP is made in the electron transport chain by chemiosmosis (oxidative phosphorylation). The electron transport chain itself makes no ATP; it builds a proton gradient that ATP synthetase then uses.

NADH and \ce{FADH2} give their electrons to the chain. As the electrons fall from carrier to carrier, energy is released.
Complexes I, III and IV use this energy to **pump protons (\ce{H+}) from the matrix into the intermembrane space**. Protons cannot cross the inner membrane by themselves.
The intermembrane space now has a higher \ce{H+} concentration than the matrix: a proton gradient across the inner membrane.
The gradient makes ATP synthetase active. Protons flow back down the gradient into the matrix through the **F_0 channel**.
This flow makes the **F_1 head work: it joins ADP and Pi with a high-energy bond to form ATP**.

One NADH gives 3 ATP and one \ce{FADH2} gives 2 ATP by this mechanism.

Figure 4.17: Mechanism of chemiosmosis in the respiratory electron transport chain. (1) The ETC pumps H^+ into the intermembrane space; (2) H^+ flows back through ATP synthase, which makes ATP.

Diagram showing movement of electrons (2 marks)

Figure 4.16: Sequence of electron carriers in the respiratory ETC. Electrons (accent) lose energy at each step; protons are pumped at I, III and IV.

2026 Annual I, Set D, Q.6 OR2+2+2 = 6 marks

Draw events of non-cyclic photophosphorylation and explain absorption of light by PS-I and electron flow from PS-I to \mathrm{NADP^+}.

Solution

Events of non-cyclic photophosphorylation (2 marks)

Both PS-II and PS-I work, with two electron transport chains. Electrons excited in PS-II (replaced from water) pass down a chain to PS-I and are finally accepted by \ce{NADP+}. The path is a zigzag, the Z-scheme. Products: ATP, NADPH and \ce{O2}.

Figure 4.8: Non-cyclic photophosphorylation (Z-scheme). Each photosystem raises the electrons to a higher energy level; they lose energy on the way down, which is used to make ATP.

Figure 4.7: Chemiosmotic synthesis of ATP during the light reaction. Electron flow is shown in accent, proton movement in grey.

Absorption of light by PS-I (2 marks)

Electrons arriving from PS-II reach plastocyanin (PC), which carries them into PS-I and fills the electron holes of P700.
The antenna pigments of PS-I absorb light and pass the energy to the reaction centre, the chlorophyll a called P700.
P700 absorbs two photons and its electrons are boosted to a higher energy level.
The primary electron acceptor of PS-I captures the electrons and leaves electron holes in P700. These holes are filled by the electrons that came from P680 (PS-II) through the transport chain.

Electron flow from PS-I to NADP^+ (2 marks)

The primary acceptor of PS-I passes the electrons to a second electron transport chain.
They are accepted by ferredoxin (Fd), an iron-containing protein.
The enzyme NADP reductase (a flavoprotein) transfers the electrons from Fd to \ce{NADP+}.
\ce{NADP+} combines with the electrons and hydrogen ions to form NADPH.

\ce{NADP+ + 2e- + 2H+ -> NADPH + H+}

NADPH carries the reducing power to the Calvin cycle in the stroma.

Exam focus

Non-cyclic photophosphorylation: draw the Z-scheme or flow sheet and explain light absorption by PS-II or PS-I, photolysis and electron flow to \ce{NADP+}. (long)
The most repeated question: a 6-mark Section C question in both 2026 sets (Set B Q.6, Set D Q.6 OR) and a 3-mark flow sheet in 2025 (Set D Q.2 (vii) OR). Book long question 7.
Go to answer
Electron transport chain: order of carriers, where protons are pumped, and a labelled diagram. (long)
A 7-mark question in 2025 (Set B Q.4 OR) and the diagram part of 2025 Set D Q.3 OR; book long question 15.
Go to answer
Chemiosmosis and ATP synthetase: how the proton gradient drives ATP synthesis; composition (F_0, F_1) and roles. (long)
Asked in 2025 (Set D Q.3 OR, 7 marks) and both 2026 sets (Set B Q.2 (xiii) OR, Set D Q.2 (xiii) OR); also the chemiosmosis part of 2026 Set B Q.6. Book long question 16.
Go to answer
Glycolysis: the preparatory (investment) and pay-off phases, with enzymes, ATP used and made, and net yield. (short)
Each phase was a 3-mark question in 2026 (Set B and Set D Q.2 (ii) OR); the complete pathway is book long question 12.
Go to answer
Cyclic photophosphorylation: why and how it occurs, and how it differs from non-cyclic. (short)
Asked in 2025 (Set B Q.2 (v)), and the Z-scheme questions expect the contrast. Book long question 9.
Go to answer
Link reaction: conversion of pyruvate to acetyl CoA with a diagram. (short)
A 3-mark question in 2025 (Set B Q.2 (xi)); book long question 13.
Go to answer
Anaerobic respiration: lactic acid and alcoholic fermentation, end products and where each occurs. (mcq)
One MCQ in each 2026 paper (Set B MCQ 3 muscles, Set D MCQ 3 yeast). Learn both end products and the role of NADH.
Go to answer
Absorption spectrum of chlorophyll a, chlorophyll b and carotenoids: compare the peaks. (short)
A 3-mark differentiate question in 2025 (Set D Q.2 (ix)); book short questions 2 and 5.
Go to answer
Calvin cycle: three phases, the reduction sequence 3-PGA, 1,3-BPG, G3P, and the inputs for one G3P (3 \ce{CO2}, 9 ATP, 6 NADPH). (long)
2025 Set B MCQ 5 asked the reduction sequence; the cycle is book long question 10 and an SLO.
Go to answer