ENERGETICS II: CELLULAR RESPIRATION

Transformation of Energy between the Environment and Organisms

Plants and other photosynthetic organisms use sunlight to synthesise carbohydrates from carbon
dioxide and water during the process of photosynthesis. The light-dependent (cyclic and non-cyclic
photophosphorylation) and light-independent stages of photosynthesis facilitate the conversion of
light energy to chemical energy in the form of carbohydrates. Carbohydrates produced from
photosynthesis can be assembled into macromolecules or broken down subsequently to fuel activities
within the plants. Carbon fixation occurs during the light-independent stage and the Calvin cycle
ultimately results in the synthesis of sugars in plants.

As heterotrophs consume plant matter, energy from the plants is transferred to them. Chemical
processes occur during aerobic respiration whereby carbohydrates are broken down to release
energy to phosphorylate ADP to ATP during aerobic respiration. The energy is transferred between
interacting molecules through the four stages of aerobic respiration when oxygen is present. In the
absence of oxygen, fermentation occurs with the release of fewer ATP molecules and the formation
of either lactate or ethanol depending on the cell type.

Learning Outcomes
Candidates should be able to:

(a) identify components of chloroplasts and mitochondria in drawings, photomicrographs and

electronmicrographs
(f) outline the process of glycolysis, highlighting the location, raw materials used and products
formed (knowledge of details of the intermediate compounds and isomerisation is not required)
(g) outline the processes of the link reaction and Krebs cycle, highlighting the location, raw materials
used and products formed (in terms of dehydrogenation and decarboxylation)
(h) outline the process of oxidative phosphorylation including the role of oxygen and the electron
transport chain in aerobic respiration (names of complexes in the ETC are not required)
(i) explain the production of a small yield of ATP from respiration in anaerobic conditions in yeast
and in mammalian muscle tissue
(j) explain the significance of the formation of ethanol in yeast and lactate in mammals in the
regeneration of NAD
(k) investigate the effect of factors such as substrate concentration, type of substrate and
temperature on the rate of respiration
(l) outline chemiosmosis in photosynthesis and respiration (names of complexes in the ETC are not
required)

Use the knowledge gained in this section in new situations or to solve related problems.

References
1. Biology: A Global Approach, 10th Ed. (2015). Campbell N. A. et al. Pearson.
2. Human Physiology: An Integrated Approach, 7th Ed. (2016). Silverthorn D. U. Pearson.
3. Biology Coursebook, 4th Ed. (2015). Jones, M et al. Cambridge University Press.
4. Biology, 2nd Ed. (2014). Clegg C. J. Hodder Education.
5. Biology for the IB Diploma, 2nd Ed. (2014). Walpole B. Cambridge University Press.
6. Biochemistry 5th Ed. (2013). Garrett R. H. and Grisham C. M. Brooks/Cole, Cengage Learning.
7. Molecular Biology of the Cell, 6th Ed (2014). Alberts B. et al. Garland Science.

HWA CHONG INSTITUTION / 2022-2023 H2 BIOLOGY / Core Idea 3 / Energetics II / Cellular Respiration 1
Learning Experiences
Further exploration could include how prokaryotes synthesise ATP when they do not possess
membrane-bound organelles.

S/N QR Code Website

Effect of factors such as substrate concentration, type of substrate and

temperature on the rate of respiration
1.
http://www.biologydiscussion.com/respiration/aerobic-respiration/factors-
affecting-aerobic-respiration-8-factors-plants/15206

Plan and conduct experiments to investigate rates of respiration using

organisms such as yeast. Factors such as temperature, type of sugar and
concentration of sugar can be investigated to determine their effect on the
rate of respiration. Simple respirometer set-ups could be used to investigate
changes in the volume of gases. Measurements of oxygen uptake or carbon
dioxide released could act as indicators of the biochemical activity and using
2.
manometers and respirometers allows quantitative measurements of gas
intake or output.

Investigating rate of Respiration

https://www.nuffieldfoundation.org/practical-biology/measuring-rate-
metabolism

This SLS lesson provides an overview of aerobic respiration with its 4 main
stages, to produce ATP for energy from carbohydrates.
3.
Aerobic Respiration SLS package

This SLS lesson provides an overview of anaerobic respiration, and how

fermentation is required to regenerate coenzyme in an oxidised state which
4. is needed for glycolysis to continue occurring.

Anaerobic Respiration SLS package

HWA CHONG INSTITUTION / 2022-2023 H2 BIOLOGY / Core Idea 3 / Energetics II / Cellular Respiration 2
OUTLINE Page
1. The Energy of Life and Metabolism 5
2. Common Reactions in Cellular Respiration 6
2.1 Energy is transferred through redox reactions 6
2.2 Role of coenzymes in redox reactions in respiration 6
3. Cellular Respiration involving Glucose 8
3.1 The effect of oxygen on glucose catabolism 8
4. Glycolysis 10
4.1 An overview of glycolysis 10
4.2 A detailed look at glycolysis 11
4.3 Importance of glycolysis 13
4.4 Regulation of glycolysis 13

CONCEPT CHECK ONE

5. Aerobic Respiration 15
5.1 Relating mitochondrial structure to its function 17
5.2 The link reaction (between glycolysis and Krebs cycle) 18
5.3 The Krebs / Citric acid cycle 19
5.3.1 An overview of the Krebs cycle 19
5.3.2 A detailed look at the Krebs cycle 21
5.3.3 Overall budget for aerobic respiration of one glucose molecule
(up till end of Krebs cycle) 22

CONCEPT CHECK TWO

5.4 Oxidative phosphorylation: Chemiosmosis couples electron transport to ATP

synthesis 24
5.4.1 An overview of the electron transport chain 24
5.4.2 The electron transport chain transfers electrons from NADH
and FADH2 to oxygen 24
5.5 ATP synthesis through oxidative phosphorylation 26
5.5.1 Chemiosmosis: The energy-coupling mechanism 26
5.5.2 ATP synthase complex 27
5.5.3 ATP yield from chemiosmosis 28
5.6 Aerobic respiration of one glucose molecule yields a maximum of 36 / 38 ATPs 29
5.7 Comparing substrate-level phosphorylation and oxidative phosphorylation 31
5.8 Respiratory Poison or Inhibitors 32

CONCEPT CHECK THREE

6. Anaerobic Respiration 34
6.1 A small yield of ATP in Anaerobic conditions 34
6.2 Formation of ethanol in yeast and lactate in mammals in regeneration of NAD 34

7. Comparing Energy Yield of Aerobic and Anaerobic Respiration 35

8. Effect of Factors on Rate of Respiration 36

8.1 Substrate concentration and type of substrate 36
8.2 Temperature 37
8.3 Investigating respiration 37

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 TOPIC OVERVIEW

Exergonic catabolic pathways Endergonic anabolic pathways

Such as breakdown / oxidation of respiratory substrates Such as biosynthesis of proteins,
(Glucose as the main respiratory substrate) lipids and polysaccharides

Key reactions in respiration:

 Redox reactions 1 Glucose (6C)
 Oxidative decarboxylation

Glycolysis
Net gain of 2 ATP & 2 NADH

2 Pyruvate (3C)

Lack of oxygen
Presence of oxygen Anaerobic respiration / fermentation
Aerobic respiration

2 Pyruvate (3C)
2 Pyruvate (3C)

Link reaction Alcoholic Lactic acid

fermentation fermentation

in yeast & in bacteria &

2 Acetyl CoA (2C) plants animals

2 Ethanol (2C) + 2 CO2 2 Lactic acid (3C)

Krebs cycle
2 ATP molecules
NADH FADH2

Oxidative
phosphorylation
Aerobic Metabolism of glucose
The energy production from one glucose molecule
can be summarised in the following two equations:

Total: 36/38 ATP molecules

&

4 CO2 + H2O

MITOCHONDRION
CYTOSOL

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 1. THE ENERGY OF LIFE AND METABOLISM

The Energy of Life

Living cells require transfusions of energy from

outside sources to perform their many tasks,
for example, assembling polymers, pumping
substances across membranes, moving and
reproducing.

The energy stored in the organic molecules

of food ultimately comes from the sun. Energy
flows into an ecosystem as sunlight and exits
as heat; in contrast, the chemical elements
essential to life are recycled (Fig. 1).

Fig. 1: Energy flow and chemical recycling in

ecosystems.

The chemical reactions that enable an organism to carry on its activities together make up its
metabolism. An organism’s metabolism consists of many intersecting series of chemical reactions,
or pathways, which are of two main types (Fig. 2):

1. Catabolism  It includes the various pathways involving the breakdown of absorbed food
substances (larger molecules) into simpler, smaller molecules.
 There is an overall release of energy i.e. exergonic.

2. Anabolism  It includes the various pathways involving the biosynthesis of complex

molecules from simpler compounds / substances.
 There is an overall energy requirement i.e. endergonic.

Fig. 2: Relationship between catabolic and anabolic pathways in metabolism.

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 2. COMMON REACTIONS IN CELLULAR RESPIRATION

2.1 Energy is transferred through Redox Reactions

In many chemical reactions, there is transfer of one or more electrons (e-) from one reactant to
another. These electron transfers are called oxidation-reduction reactions or redox reactions.

Redox reactions  The loss of electrons from one substance is oxidation, and the addition
of electrons to another substance is reduction (Fig. 3).
 Since an electron transfer requires both an electron donor and an acceptor,
oxidation and reduction always go hand in hand.

Fig. 3: Redox reactions. Xe-, the electron donor is called

the reducing agent; it reduces Y, which accepts the donated
electron. Substance Y, the electron acceptor, is the
oxidising agent; it oxidises Xe- by removing its electron.

There are three different ways in which a molecule Table 2: Changes involved in oxidation and reduction.
can be oxidised or reduced, as outlines in Table 2.
In biological oxidation reactions, addition of oxygen Oxidation Reduction
atoms is an alternative to removal of hydrogen Loss of electrons Gain of electrons
atoms.
Loss of hydrogen Gain of hydrogen
Since a hydrogen atom consists of an electron and Gain of oxygen Loss of oxygen
a proton, losing hydrogen atoms (oxidation)
involves losing one or more electrons.

Decarboxylation  Removal of carbon atoms from a compound to form carbon dioxide.

Dehydrogenation  Oxidation / breakdown of organic molecules frequently involve the

removal of electrons as well as hydrogen ions / protons (H+).
 Enzymes that catalyse oxidative reactions are called dehydrogenases.

Oxidative  Oxidation reactions whereby a carboxylate group is removed, forming

decarboxylation carbon dioxide.

2.2 Role of Coenzymes in Redox Reactions in Respiration

Coenzymes are loosely associated with the enzyme during the reaction. They act as transient
carriers of electrons, hydrogen or specific functional groups.

In biological oxidations, electrons and protons removed from oxidised substrates are transferred to
one of several coenzymes. The most common coenzymes involved in energy metabolism are
nicotinamide adenine dinucleotide (NAD) (Fig. 4a) and flavin adenine dinucleotide (FAD) (Fig.
4b).

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 Fig 4a: The Structure of NAD and NADH (FYI). Fig 4b: The Structure of FAD and FADH2 (FYI).

Coenzymes in respiration can be oxidised or reduced (Table 3).

Table 3: Representation of coenzymes in redox reactions.

Oxidised form Reduced form

NAD NADH or reduced NAD
FAD FADH2 or reduced FAD

Upon reduction, these reduced coenzymes serve as reservoirs of electrons and protons to form
ATP via oxidative phosphorylation (Section 5.4).

Fig. 5: NAD as an electron shuttle. The enzymatic transfer of two

electrons and one proton (H+) from an organic molecule in food to NAD
reduces NAD to NADH: most of the electrons removed from food are
transferred initially to NAD, forming NADH.

HWA CHONG INSTITUTION / 2022-2023 H2 BIOLOGY / Core Idea 3 / Energetics II / Cellular Respiration 7
 3. CELLULAR RESPIRATION INVOLVING GLUCOSE

Many organic molecules, such as proteins, Proteins Carbohydrate Fats

polysaccharides and lipids can serve as
respiratory substrates (Fig. 6). Amongst Amino Sugars Glycerol Fatty
acids acids
these, glucose remains as one of the most
important. GLYCOLYSIS
Glucose

Glyceraldehyde 3- P
Thinking Question:
NH
What are the advantages of using glucose 3 Pyruvate
as the main respiratory substrate?
Acetyl CoA

CITRIC
ACID OXIDATIVE
PHOSPHORYLATION
CYCLE

Fig. 6: Use of proteins, polysaccharides

and lipids as respiratory substrates.

3.1 The Effect of Oxygen on Glucose Catabolism

Organisms can be classified in terms of their need for and use of oxygen in energy metabolism. Most
organisms have an absolute requirement for oxygen and are called obligate aerobes. Obligate
anaerobes, on the other hand, cannot use oxygen under any conditions. A third class of organisms,
facultative anaerobes, can function under either aerobic or anaerobic conditions.

For obligate aerobes, access to the amount of free energy in glucose depends on the availability of
oxygen:

1. In the presence of oxygen

 This results in complete oxidation of glucose to carbon dioxide and water, and is termed
aerobic respiration.
 The process yields the maximum amount of 36 / 38 ATP molecules.

2. In the absence / scarcity of oxygen

 This results in incomplete oxidation of glucose, and is termed anaerobic respiration or
fermentation.
 The process yields a limited amount of net 2 ATP molecules.
 There are many types of fermentation, and they are identified in terms of the main end-product:
lactic acid fermentation or ethanol fermentation. (KIV Section 6)

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The harvesting of energy from glucose by cellular respiration is a cumulative function of three
metabolic stages (Fig. 7):

1. Glycolysis
2. Link reaction and Krebs cycle (or Citric acid Cycle)
3. Oxidative phosphorylation: Electron transport and chemiosmosis

Whether a cell obtains its energy via aerobic or anaerobic respiration, glucose metabolism
inadvertently begins with glycolysis.

Fig. 7: An overview of cellular respiration. During glycolysis, each glucose molecule is broken down into two molecules
of the compound pyruvate. In eukaryotic cells, as shown here, the pyruvate enters the mitochondrion. There it is oxidised
to acetyl CoA, which is further oxidised to CO2 in the citric acid cycle. NADH and a similar electron carrier, a coenzyme
called FADH2, transfer electrons derived from glucose to electron transport chains, which are built into the inner
mitochondrial membrane. During oxidative phosphorylation, electron transport chains convert the chemical energy to a form
used for ATP synthesis in the process called chemiosmosis.

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 4. GLYCOLYSIS

The process of glycolysis (glycolytic pathway) is a ten step reaction sequence, each of which is
catalysed by a specific enzyme. It is divided into two major phases:

1. Energy-investment phase: ATP utilisation

2. Energy-payoff phase: ATP formation

The process converts one molecule of glucose into 2 molecules of pyruvate, a three-carbon
compound, with the generation of 2 net ATP molecules (Fig. 8).

C6H12O6 + 2 ADP + 2 NAD 2 C3H4O3 + 2 ATP + 2 NADH + 2 H+

(Glucose) (Pyruvate)

4.1 An Overview of Glycolysis

Location  Occurs in the cytosol / cytoplasm in all

cells (plants, animals and prokaryotes).

Metabolism  Glucose, a 6-carbon (6C) sugar, is split

into two 3C sugars. Each of which
rearranged to form a 3-carbon (3C)
compound, pyruvate (ionised form of
pyruvic acid).

 Both reactions are important:

1. Substrate level phosphorylation
(ADP  ATP)
2. Dehydrogenation (NAD  NADH)

Substrates  Glucose or other hexose sugars

 ADP
 Inorganic phosphates (Pi)
 NAD

Products  Useful products formed per glucose

molecule:
1. 2 molecules of pyruvate
2. 2 net ATPs
3. 2 NADH

 Waste product formed: water

Fig. 8: An overview of glycolysis.

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 4.2 A Detailed look at Glycolysis

Steps 1 to 5: Energy-investment Phase

Steps 1 to 3:  Conversion i.e. activation of unphosphorylated glucose to a

Activation of phosphorylated fructose-1,6-bisphosphate.
glucose  Hydrolysis of 2 ATPs to provide not just the phosphate groups, but
also the energy / driving force.
 Step 3: rate-limiting step of glycolysis involving the enzyme
phosphofructokinase (see Section 4.4: Regulation of Glycolysis).

Step 4 to 5:  Cleavage of fructose-1,6-bisphopshate to two 3-carbon sugars,

Cleavage / Lysis glyceraldehyde-3-phosphate (G3P).

Steps 6 to 10: Energy-Payoff Phase

Step 6:  Each glyceraldehyde-3-phosphate (G3P) is oxidised

Reduction of NAD (dehydrogenation) and NAD is reduced to NADH.
(Dehydrogenation)  This results in a net production of 2 NADH per glucose molecule.
 One NADH supplies 2 energised electrons.
 These electrons directly drive most ATP production by oxidative
phosphorylation at the inner mitochondrial membrane.

Steps 7 and 10:  Substrate-level phosphorylation of ADP occurs at steps 7 and 10,
Substrate-level coupled to the dephosphorylation of an organic substrate.
Phosphorylation  Directly produces 4 ATPs per glucose molecule.
 Results in an overall net gain of 2 ATPs per glucose molecule.

Fig. 9: The energy input and output of glycolysis.

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 1. Phosphorylation of
glucose by ATP.

2-3. Rearrangement,
followed by a second
ATP phosphorylation.

Energy-
4-5. The six-carbon Investment
molecule is split into
Phase
two three-carbon
molecules – one G3P,
another that is
converted into G3P in
another reaction.

6. Oxidation followed by
phosphorylation
produces two NADH
molecules and two
molecules of BPG, each
with one high-energy
phosphate bond.

7. Transfer of high-energy
phosphate to two ADP
molecules produces two
ATP molecules and
leaves two 3PG
molecules.

Energy-
8-9. Removal of water yield payoff
two PEP molecules, Phase
each with a high-
energy phosphate
bond.

10. Transfer of high-energy

phosphate to two ADP
molecules produces two
ATP molecules and two
pyruvate molecules.

Fig. 10: The glycolytic pathway. The first five reactions convert a molecule of glucose into two molecules of G3P. The
second five reactions convert G3P into pyruvate.

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 4.3 Importance of Glycolysis

In most animal tissues, glycolysis is a minor, yet often vital source of energy as it directly produces
net 2 molecules of ATP by substrate-level phosphorylation.

1. Glycolysis, which occurs in most living cells,

 it is the only catabolic reaction that can be completed in the absence of oxygen.
 It hydrolyses 1 molecule of glucose into two molecules of pyruvate.

2. In the presence of oxygen,

 Pyruvate produced enters the mitochondrion and are completely oxidised to produce ATP
by oxidative phosphorylation.
 Reduced NAD and FAD supply energised electrons for ATP production.

3. It supplies cells with essential biosynthetic precursors.

 The liver carries out glycolysis to provide precursors for a whole range of molecules it
synthesises, including fats, cholesterol, bile acids and plasma proteins.
 In the well-fed animal, once liver glycogen reserves are full, carbohydrate is converted to fats.
Glycolysis is then predominantly associated with supplying the initial steps of fat biosynthesis
with substrate, rather than acting as a source of ATP.
 For microorganisms such as E. coli or yeast growing on carbohydrate sources, both energy
and necessary biosynthetic precursors are obtained from glycolysis.

4.4 Regulation of Glycolysis

Phosphofructokinase is an allosteric
enzyme.
1. As ATP accumulates, it acts as an
allosteric inhibitor by binding to
phosphofructokinase and thus slowing
down glycolysis.

2. It is stimulated by AMP which the cell

derives from ADP.

The enzyme becomes active again as

cellular work converts ATP to ADP (and
AMP) faster than ATP is being regenerated.

3. It is also sensitive to citrate, the first

product of the Krebs cycle (Section 5.3). If
citrate accumulates in the mitochondria,
some of it pass into the cytosol and inhibit
phosphofructokinase.

This mechanism helps to synchronise the

rates of glycolysis and Krebs cycle.

Fig. 11 illustrates the regulation of Fig. 11: The control of cellular respiration.

glycolysis.

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 CONCEPT CHECK ONE

The figure shows some of the details of the conversion of a molecule of glucose into two molecules
of pyruvate by glycolysis.

The reactions labeled A to J represent the different reactions of glycolysis, catalysed by different
enzymes.

Glucose
A
Glucose-6-phosphate
B
Fructose-6-phosphate

C
Fructose-1,6-bisphosphate

E
Dihydroxyacetone phosphate Glyceraldehyde-3-phosphate
F
1,3 Bisphosphoglyceric acid

G
3-Phosphoglyceric acid

H
2-Phosphoglyceric acid

I
Phosphoenolpyruvate

J
Pyruvate

(a) Where in the cell does glycolysis occur? [1]

(b) Name the processes that are occurring at A, D and F. [3]

(c) At which steps does substrate-level phosphorylation of ADP occur? [1]

HWA CHONG INSTITUTION / 2022-2023 H2 BIOLOGY / Core Idea 3 / Energetics II / Cellular Respiration 14
 5. AEROBIC RESPIRATION

In glycolysis, only 2 net molecules of ATP can be generated per glucose molecule. In order to
maximise ATP yield, aerobic respiration is employed by the cell. Aerobic respiration is generally
defined as a series of enzyme-catalysed oxidation-reduction (redox) reactions in which
respiratory substrates (usually carbohydrates or fats) are completely oxidised to CO2, and O2 (the
final electron acceptor) is reduced to H2O. This series of enzyme-controlled reactions results in a
much higher yield of ATP. Note too, that some of the energy is lost as heat.

Including glycolysis, aerobic respiration consists of 4 main stages in the following sequential order:
 Glycolysis  Link reaction  Krebs cycle and  Oxidative phosphorylation (Electron
transport and chemiomosis (Fig. 12 and Table 3).

   

Fig. 12: The four stages of aerobic respiration.  In a eukaryotic cell, glycolysis occurs outside the mitochondria in the
cytosol. The Krebs cycle and the electron transport chains are located inside the mitochondria. During glycolysis, each
glucose molecule is broken down into two molecules of the compound pyruvate.  The pyruvate crosses the double
membrane of the mitochondrion to enter the matrix, where  the Krebs cycle decomposes it to carbon dioxide.  NADH
transfers electrons from glycolysis and the Krebs cycle to electron transport chains, which are built into the membrane of
the cristae. The electron transport chain converts the chemical energy to a form that can be used to drive oxidative
phosphorylation, which accounts for most of the ATP generated by cellular respiration. A smaller amount of ATP is formed
directly during glycolysis and the Krebs cycle by substrate-level phosphorylation.

Table 4: A summary of aerobic respiration.

Some Starting Some End

Stage Summary
Materials Products
Series of reactions in which glucose is broken down
1. Glycolysis Glucose, ATP, Pyruvate,
to pyruvate; net profit of 2 ATPs; hydrogen atoms are
(in cytosol) NAD, ADP, Pi ATP, NADH
transferred to carriers; can proceed anaerobically
Pyruvate is broken down and combined with Pyruvate,
2. Link reaction Acetyl CoA,
coenzyme A to form acetyl CoA; hydrogen atoms are coenzyme A,
(in mitochondria) CO2, NADH
transferred to carriers; CO2 is released NAD
3. Krebs cycle / Citric Series of reactions in which the acetyl portion of Acetyl CoA,
CO2, NADH,
acid cycle acetyl CoA broken down to CO2; hydrogen atoms are H2O, NAD, FAD,
FADH2, ATP
(in mitochondria) transferred to carriers; ATP is synthesised ADP, Pi
4. Oxidative Chain of several electron transport molecules;
Phosphorylation / electrons are passed along chain; released energy is
NADH, FADH2, ATP, H2O,
Electron transport used to form a proton gradient; ATP is synthesised
O2, ADP, Pi NAD, FAD
& chemiosmosis as protons diffuse down the gradient; oxygen is final
(in mitochondria) electron acceptor

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We will next focus our attention on the mitochondrion because most aerobic respiration in eukaryotic
cells takes place within this organelle (Fig. 13).

Fig. 13: The role of mitochondrion in aerobic respiration. (a) The mitochondrion plays a central role in aerobic respiration.
Most respiratory ATP production in eukaryotic cells occurs in this organelle. (b) Oxidation of glucose and other sugars
begins in the cytosol with glycolysis (stage 1), producing pyruvate. Pyruvate is transported across the inner mitochondrial
membrane and is oxidized within the matrix to acetyl CoA (stage 2), the primary substrate of the tricarboxylic acid (TCA)
cycle (stage 3). Acetyl CoA can also be formed by β oxidation of fatty acids. Electron transport is coupled to proton pumping
(stage 4), with the energy of electron transport conserved as an electrochemical proton gradient across the inner membrane
of the mitochondrion (or across the plasma membrane, in the case of prokaryotes). The energy of the proton gradient is
used in part to drive the synthesis of ATP from ADP and inorganic phosphate (stage 5).

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 5.1 Relating Mitochondrial Structure to its Function

The crucial role of the mitochondrion in meeting cellular ATP needs is often reflected in the distribution
of mitochondria within the cell. Frequently they are clustered in regions of cells with the most intense
metabolic activity and the greatest need for ATP.

Fig. 14 shows the structure of the mitochondrion. Table 5 details the relation between mitochondrial
structures and their functions.

Fig. 14: The structure of a mitochondrion.

Table 5: Relating mitochondrial structures to their functions.

Components Roles in brief

Outer membrane Freely permeable to ATP, ADP etc.

A selectively permeable membrane which is highly-folded to form

cristae which increase surface area for embedding electron transport
chain and ATP synthase complexes.
 Not permeable to NADH
 Contains proteins for transporting H+, ATP and ADP
Inner membrane
 Contains members of the ETC and ATP synthase complexes
(stalked particles)
 permeable to pyruvate

Site of oxidative phosphorylation.

Compartment enclosed by the inner membrane of the mitochondrion.
Matrix  Site of the Link reaction and Krebs cycle
 contains enzymes required for reactions in the Krebs cycle

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 5.2 The Link Reaction (between Glycolysis and Krebs cycle)

When molecular oxygen is present, the pyruvate produced by glycolysis enters the mitochondrion,
where the oxidation of glucose is completed in Krebs cycle.

Upon entering the mitochondrion via active transport, pyruvate is first converted to a 2-carbon
compound called acetyl CoA. This step is carried out by a multienzyme complex, which catalyses 3
reactions through oxidative decarboxylation:

1. Pyruvate’s carboxyl group (-COO-) that is already fully oxidised, is removed and liberated as CO2.

2. The remaining 2C fragment is oxidised, forming acetate (CH3COO-). Two electrons and a proton
removed are transferred to the coenzyme NAD, reducing it to NADH.

3. Coenzyme A is attached to acetate, forming acetyl coA.

Link reaction occurs in the mitochondrial matrix and is catalysed by pyruvate dehydrogenase (Fig.
15). It occurs twice for each glucose molecule. Thus, the overall reaction for link reaction is:

2 Pyruvate + 2 NAD + 2 CoA 2 Acetyl CoA + 2 NADH + 2 CO2

Fig. 15: Oxidation of pyruvate to acetyl coA, the step before the Krebs cycle. Pyruvate is a charged molecule, so in
eukaryotic cells it must enter the mitochondrion via active transport, with the help of a transport protein. Next, a complex of
several enzymes (the pyruvate dehydrogenase complex) catalyses the three numbered steps. The acetyl CoA will enter
the Krebs cycle. The CO2 molecule will diffuse out of the cell. By convention, coenzyme A is abbreviated S-CoA when it is
attached to a molecule, emphasising the sulfur atom (S).

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 5.3 The Krebs / Citric Acid Cycle

The Krebs cycle begins with the entry of acetate in the form of acetyl CoA. With each round of Krebs
cycle activity, 2 carbon atoms enter in organic form (as acetate) and 2 carbon atoms leave in
inorganic form (as CO2).

5.3.1 An Overview of the Krebs Cycle (Fig. 16)

Location  Occurs in the mitochondrial matrix in all cells (plants and animals).

Metabolism  A catabolic pathway that occurs twice for every glucose molecule broken
down. Completes glucose breakdown and oxidation by 8 enzyme-
controlled steps.
 It is known as a cycle because the molecule that is added to incoming
acetyl CoA (from link reaction), oxaloacetate, is regenerated at the end
of one turn of the cycle to receive more acetyl groups.
 In addition to its respiratory role, the Krebs cycle also plays a central role
in the breakdown of multiple metabolic intermediates.

Breakdown products of lipids, amino acids and carbohydrates can enter

the cycle at various steps in the pathway. The various compounds in the
cycle can also be siphoned off for the formation of chlorophyll, amino acids
etc.

Crucial 1. Oxidative decarboxylation

Reactions
2. Substrate-level phosphorylation

3. Production of reduced coenzymes

Substrates  Acetyl CoA or any of the other intermediates in the pathway

 ADP
 Inorganic phosphates (Pi)
 NAD
 FAD

Products  Useful products formed per glucose molecule, 2 turns of the cycle:
- 2 ATP
- 2 FADH2
- 6 NADH

 Waste product formed: 4 CO2

The Krebs cycle can be summarised as follows:

 The two carbon acetyl group of acetyl CoA combines with a 4C molecule called oxaloacetate
to form a six-carbon intermediate citrate.
 Citrate then goes through a sequence of electron-yielding oxidation reactions, during which 2 CO2
molecules split off, regenerating oxaloacetate.
 The oxaloacetate is then recycled to bind to another acetyl group.

HWA CHONG INSTITUTION / 2022-2023 H2 BIOLOGY / Core Idea 3 / Energetics II / Cellular Respiration 19
Fig. 16: An overview of pyruvate oxidation and Krebs cycle. The inputs and outputs per pyruvate molecule are shown.
To calculate on a per-glucose basis, multiply by 2, because each glucose molecule is split during glycolysis into two pyruvate
molecules.

HWA CHONG INSTITUTION / 2022-2023 H2 BIOLOGY / Core Idea 3 / Energetics II / Cellular Respiration 20
 5.3.2 A Detailed Look at the Krebs Cycle

 Condensation
Acetyl CoA (from
oxidation of pyruvate)
adds its 2C acetyl group
 NAD to oxaloacetate,
Reduction producing citrate.
 Citrate is
The substrate is
converted to its
oxidised,
isomer, isocitrate, by
reducing NAD
removal of 1 water
to NADH and
molecule and
regenerating
addition of another.
oxaloacetate.

 Oxidative
decarboxylatio
n
 Addition NAD
of a water Reduction
molecule Isocitrate is
rearranges oxidised,
bonds in the reducing NAD
substrate. to NADH. Then
the resulting
compound loses
a CO2 molecule.

 Oxidative
decarboxylation
NAD Reduction
Another CO2 is lost,
and the resulting
 FAD Reduction compound is
Two hydrogens are oxidised, reducing
transferred to FAD, NAD to NADH. The
forming FADH2 and remaining molecule
oxidising succinate. is then attached to
coenzyme A by an
unstable bond.

 Substrate Level
Phosphorylation
CoA is displaced by a
phosphate group, which is
transferred to GDP, forming
GTP, a molecule with
functions similar to ATP.
GTP can also be used, as
shown, to generate ATP.

Fig. 17: A closer look at the Krebs cycle. In the chemical structures, red type traces the fate of the two carbon atoms that
enter the cycle via acetyl CoA (step 1), and blue type indicates the two carbons that exit the cycle as CO2 in steps 3 and 4.
(The red type goes only through step 5 because the succinate molecule is symmetrical; the two ends cannot be distinguished
from each other.) Notice that the carbon atoms that enter the cycle from acetyl CoA do not leave the cycle in the same turn.
They remain in the cycle, occupying a different location in the molecules on their next turn, after another acetyl group is
added. Therefore, the oxaloacetate regenerated at step 8 is made up of different carbon atoms each time around. In
eukaryotic cells, all the citric acid cycle enzymes are located in the mitochondrial matrix except for the enzyme that catalyses
step 6, which resides in the inner mitochondrial membrane. Carboxylic acids are represented in their ionised forms, as –
COO-, because the ionised forms prevail at the pH within the mitochondrion.

HWA CHONG INSTITUTION / 2022-2023 H2 BIOLOGY / Core Idea 3 / Energetics II / Cellular Respiration 21
We can summarise what has been accomplished by noting the following steps of the Krebs cycle in
Fig. 17:

 Oxidative Decarboxylation occurs at steps 3 and 4 in the cycle, so that the input of 2 carbons
as acetate is balanced by the loss of 2 carbons as CO2.

 Substrate-Level Phosphorylation where ATP is generated at step 5, with GTP as an

intermediate.

 Production of reduced coenzymes by dehydrogenation, with NAD as the electron acceptor at

steps 3, 4 and 8 and FAD as the electron acceptor at step 6.

One turn of the cycle is completed upon regeneration of oxaloacetate (step 8), the original 4C
acceptor. Note that the cycle must occur twice to metabolise both of the acetyl CoA molecules
derived from a single molecule of glucose.

5.3.3 Overall budget for aerobic respiration of 1 glucose molecule

(up till end of Krebs cycle)

ATP
Process CO2 NADH FADH2
(Substrate Level Phosphorylation)

Glycolysis - 2 2 -

Link Reaction 2 - 2 -

Krebs Cycle
4 2 6 2
(2 turns of the cycle)

TOTAL 6 4 10 2

Thus far, the ATP yield has been modest. However, it is important to bear in mind that the reduced
coenzymes are high-energy compounds.

This means that the transfer of electrons from these coenzymes to oxygen is highly exergonic, and
thus accounts for the synthesis of most of the 36 / 38 ATP molecules produced during the complete
oxidation of a glucose molecule.

We will now proceed to examine oxidative phosphorylation (electron transport and chemiosmosis),
the remaining stage of aerobic respiration, for the release of that energy.

HWA CHONG INSTITUTION / 2022-2023 H2 BIOLOGY / Core Idea 3 / Energetics II / Cellular Respiration 22
 CONCEPT CHECK TWO

The figure below shows an overview of the Krebs cycle.

(a) Where do the reactions of the Krebs cycle occur? [1]

(b) What is the major function of the Krebs cycle? [2]

(c) Fill in the product formed in each blank in the figure above. [7]

(d) What are the products of the cycle following the breakdown of one glucose molecule? [1]

(e) Why is this series of reactions known as a cycle? [1]

HWA CHONG INSTITUTION / 2022-2023 H2 BIOLOGY / Core Idea 3 / Energetics II / Cellular Respiration 23
 5.4 Oxidative Phosphorylation:
Chemiosmosis couples Electron Transport to ATP Synthesis

Recall that the electrons removed from a molecule of glucose during glycolysis, link reaction and the
Krebs cycle are transferred as pairs of hydrogen atoms to the acceptors NAD and FAD, forming
NADH and FADH2.

These reduced coenzymes then transfer the electrons to oxygen in an electron transport chain
(ETC). The exergonic transport of electrons is coupled to the endergonic process of ATP synthesis.
These two processes (electron transport and ATP synthesis) are functionally linked by the
electrochemical proton gradient that is the result of electron transport as well as the source of the
energy that drives ATP synthesis.

5.4.1 An Overview of the Electron Transport Chain

Location  Occurs in the inner mitochondrial membrane, which is highly folded to form
cristae in all cells (plants and animals).

Crucial  The ETC is made up of a sequence of electron carriers.

reaction  They undergo temporary reduction and oxidation as electrons (from NADH
and FADH2) are passed down to the final electron acceptor, molecular
oxygen.

ATP  No ATP is directly generated.

generation  Instead, energy from the electrons carried by NADH and FADH2 are used to
pump protons across the inner mitochondrial membrane to the intermembrane
space, generating a proton gradient across the inner mitochondrial membrane.
 The flow of these protons back into the matrix along their concentration gradient
through the ATP synthase complex, allows for ATP synthesis by oxidative
phosphorylation.

5.4.2 The ETC transfers Electrons from NADH and FADH2 to Oxygen

The ETC, embedded in the inner mitochondrial membrane, comprises a sequence of electron
carriers that have the ability to be reversibly reduced and oxidised (i.e. temporarily accommodate
either one or two electrons within their molecular structure) as electrons from NADH and FADH2 are
passed down the mitochondrial ETC.

Several of the coenzymes and cytochromes in the ETC are bound to enzymes and other essential
factors to form functional groups known as complexes. There are 4 main complexes, Complexes I-
IV in the ETC (Fig. 18).

Each subsequent member of the ETC has a higher affinity for electrons than its predecessor, but
a lower affinity than its successor. This ensures a one-way transport of electrons down the ETC.
The electrons thus move along the sequence of electron carriers in order of increasing electron
affinity.

HWA CHONG INSTITUTION / 2022-2023 H2 BIOLOGY / Core Idea 3 / Energetics II / Cellular Respiration 24
The electrons are eventually passed to the final electron acceptor in the ETC, molecular oxygen
(O2) that has diffused into the mitochondrion. Oxygen is reduced in the mitochondrial matrix to
produce a molecule of water.

The energy released as the electrons flow through the ETC is sufficient to power the pumping of H+
ions across the inner mitochondrial membrane into the intermembrane space, establishing an
electrochemical proton gradient.

As members of the ETC accept electrons from Question:

NADH and FADH2, the oxidised coenzymes What is the significance of the regeneration of oxidised
coenzymes NAD and FAD?
NAD and FAD are regenerated.

Fig. 18: Chemiosmosis couples the electron transport chain to ATP synthesis.  NADH and FADH2 shuttle high-
energy electrons extracted from food during glycolysis and the Krebs cycle into an electron transport chain built into the
inner mitochondrial membrane. The gold arrows trace the transport of electrons, which are finally passed to a terminal
acceptor (O2, in the case of aerobic respiration) at the ‘downhill’ end of the chain, forming water. Most of the electron carriers
of the chain are grouped into four complexes (I-IV). Two mobile carriers, ubiquinone (Q) and cytochrome c (Cyt c), move
rapidly, ferrying electrons between the large complexes. As the complexes shuttle electrons, they pump protons from the
mitochondrial matrix into the intermembrane space. FADH 2 deposits its electrons via complex II – at a lower energy level
than complex I, where NADH deposits its electrons and so results in fewer protons being pumped into the intermembrane
space than occurs with NADH. Chemical energy that was originally harvested from food is transformed into a proton-motive
force, a gradient of H+ across the membrane.  During chemiosmosis, the protons flow back down their gradient via ATP
synthase, which is built into the membrane nearby. The ATP synthase harnesses the proton-motive force to phosphorylate
ADP, forming ATP. Together, electron transport and chemiosmosis make up oxidative phosphorylation.

HWA CHONG INSTITUTION / 2022-2023 H2 BIOLOGY / Core Idea 3 / Energetics II / Cellular Respiration 25
 5.5 ATP Synthesis through Oxidative Phosphorylation

So far we have learnt that reduced coenzymes are reoxidised (regenerated) by the exergonic transfer
of electrons to oxygen via a system of reversibly oxidisable intermediates located within the inner
mitochondrial membrane.

Now we will consider how the energy released during electron transport is used to generate a proton
gradient and how the energy of the gradient is then used to drive ATP synthesis. The answer is
a mechanism called chemiosmosis i.e. energy stored in the form of a hydrogen ion gradient across
a membrane is used to drive the synthesis of ATP.

ATP synthesis involves phosphorylation events that are linked to oxygen-dependent electron
transport, hence the process is called oxidative phosphorylation (4th stage of aerobic respiration).

5.5.1 Chemiosmosis: The Energy-Coupling Mechanism

In 1961, British biochemist Peter Mitchell proposed a revolutionary model, the Chemiosmotic
Coupling Model, or Chemiosmosis, to link electron transport to ATP synthesis.
As the electrons are transferred along the ETC, energy is released. This energy drives proton pumps
(located at the inner mitochondrial membrane) to actively pump H+ unidirectionally across the inner
mitochondrial membrane from the mitochondrial matrix to the intermembrane space (Fig. 19).

Protons are pumped across this

membrane as electrons flow Outer mitochondrial
through the electron transport chain membrane

Inner mitochondrial
High [H+] H+ membrane

Matrix

Low [H+]
Intermembrane
space

Fig. 19: Proton pumping creates a proton gradient across the inner mitochondrial membrane.

This proton pumping generates a proton gradient with two components:

1. Concentration gradient of H+: chemical or pH gradient.
2. Electrical gradient: voltage across the membrane due to the higher concentration of positively
charged protons on one side of the membrane.

With the build-up of H+ concentration in the intermembrane space, there is, thus, a tendency for
H+ to diffuse back into the matrix. This creates a proton motive force.

However, as the inner mitochondrial membrane is impermeable to H+, protons can only re-enter the
matrix via an ATP-synthesizing complex that spans the inner mitochondrial membrane. This complex
is known as the ATP synthase complex.

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 5.5.2 ATP Synthase Complex

The ATP synthase complex or stalk particle functions to couple the exergonic passage of H+ to the
endergonic phosphorylation of ADP to form ATP (Fig. 20).

Multiple copies of ATP synthase stud

the inner mitochondrial membrane.
ATP synthase is a multisubunit
complex with four main parts, each
made up of multiple polypeptides.

Protons move one by one into binding

sites on one of the parts (the rotor),
causing it to spin in a way that
catalyses ATP production from ADP
and Pi. The flow of protons thus
behaves somewhat like a rushing
stream that turns a water wheel.

Fig. 20: ATP synthase, a molecular mill. The ATP synthase protein complex functions as a mill, powered by the flow of
hydrogen ions. Multiple ATP synthases reside in eukaryotic mitochondrial and chloroplast membranes and in prokaryotic
plasma membranes. Each part of the complex consists of a number of polypeptide subunits. ATP synthase is the smallest
molecular rotary motor known in nature.

Fig. 21: The electron transport chain includes three proton pumps that are located in three of the four electron
transport complexes.

HWA CHONG INSTITUTION / 2022-2023 H2 BIOLOGY / Core Idea 3 / Energetics II / Cellular Respiration 27
 5.5.3 ATP yield from Chemiosmosis

The amount of ATP produced from chemiosmosis depends on the number of H+ pumped into the
intermembrane space as electrons are passed down the ETC. This is in turn dependent on the
number of electrons contributed by the reduced coenzymes NADH and FADH2 that originated from
glycolysis and the Krebs cycle.

For each pair of electrons stripped from For each pair of electrons stripped from
NADH or reduced NAD FADH2 or reduced FAD

 5 pairs / 10 H+ are pumped across the inner  3 pairs / 6 H+ are pumped across the inner
mitochondrial membrane into the mitochondrial membrane into the
intermembrane space. intermembrane space.

 Re-entry of H+ into the matrix provides the  Re-entry of H+ into the matrix provides the
energy to generate 3 ATP molecules. energy to generate 2 ATP molecules.

The dynamics of the chemiosmotic model is summarised in Fig. 22.

Fig. 22: Dynamics of the electrochemical proton gradient. Respiratory complexes I through IV are integral components
of the inner mitochondrial membrane. Complexes I, III and IV (but not complex II) couple the exergonic flow of electrons
(red lines) through the complex with the outward pumping of protons (blue) across the membrane. The proton motive force
of the resulting electrochemical proton gradient drives ATP synthesis by F1 as protons are translocated back across the
membrane by the Fo complex, which is also embedded in the inner membrane. ATP synthesis also consumes protons in
the mitochondrial matrix, a depletion that contributes to the transmembrane proton gradient driving ATP synthesis.

HWA CHONG INSTITUTION / 2022-2023 H2 BIOLOGY / Core Idea 3 / Energetics II / Cellular Respiration 28
 5.6 Aerobic Respiration of 1 Glucose molecule yields a Maximum of 36 / 38 ATPs

Fig. 23: Energy yield from the complete oxidation of glucose by aerobic respiration.

Location 4 Stages of Aerobic Respiration NADH FADH2 ATP

Cytoplasm  Glycolysis +2 0 +2

Mitochondrial
 Link reaction +2 0 0
matrix

 Krebs cycle +6 +2 +2

 Oxidative phosphorylation -10 -2 +34 / +32*

Total 0 0 +38 / +36*

*Whether a cell produces 36 or 38 molecules of ATP via aerobic respiration, it depends on the shuttle
system used in mitochondrion (See Shuttle Systems).

HWA CHONG INSTITUTION / 2022-2023 H2 BIOLOGY / Core Idea 3 / Energetics II / Cellular Respiration 29
 Aerobic Respiration – Shuttle Systems

The inner mitochondrial membrane lacks a NADH transport protein, and NADH produced in the
cytosol cannot penetrate directly into the mitochondria.

Thus, the electrons and protons from the 2 NADH produced in the cytosol during glycolysis are passed
to a shuttle system, which in turn passes these electrons and protons to either NAD or flavoproteins
within the mitochondrial matrix.

There are two known shuttle systems:

1. Glycerol phosphate shuttle
 Found in muscle and nerve cells.
 Cytosolic glycerol-3-phosphate dehydrogenase converts dihydroxyacetone phosphate
(DHAP) to glycerol 3-phosphate by oxidising one molecule of NADH to NAD.
 2 ATP will be formed per molecule of cytosolic NADH oxidised.
 This accounts for a total of 36 ATP molecules produced per glucose molecule.

Fig 24: The Glycerol Phosphate Shuttle. The inner mitochondrial membrane is impermeable to NADH so electrons from cytosolic NADH
are moved onto the mitochondrion by one of two shuttle mechanisms. Cells of skeletal muscle, brain, and other tissues use the glycerol
phosphate shuttle for this purpose. The red line traces the path of electrons from cytosolic NADH to oxygen.  The cytosolic enzyme
glycerol-3-phosphate dehydrogenase (G3PDH) uses electrons (and protons) from NADH to reduce dihydroxyacetone phosphate (DHAP)
to glycerol-3-phosphate (glycerol-3-P), which  can cross the outer mitochondrial membrane by diffusing through a porin channel. 
Glycerol-3-P is then reoxidised to DHAP by an FAD-linked G3PDH in the inner membrane, with concomitant reduction of FAD to FADH 2.
Because NADH is more energy-rich coenzyme than FADH2, the inward transport of electrons is exergonic, driven by the difference in the
reduction potentials of the two coenzymes. The cost of this inward transport is decreased ATP yield because  electrons from the FADH2
generated by the mitochondrial G3PDH bypass complex I, thereby reducing the number of protons pumped across the inner membrane.

2. Malate-aspartate shuttle
 Found in heart and liver cells.
 The electrons and protons stripped from cytosolic NADH are passed to matrix NAD.
 3 ATP will be formed per molecule of cytosolic NADH oxidised.
 This accounts for a total of 38 ATP molecules produced per glucose molecule.

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5.7 Comparing Substrate-Level Phosphorylation and Oxidative Phosphorylation

Table 5: Two processes by which ATP is formed during cellular respiration.

Substrate-level phosphorylation Oxidative phosphorylation

Process

 The enzymatic endergonic

 The enzymatic endergonic
phosphorylation of ADP to form
phosphorylation of ADP with a 5’
ATP by ATP synthase, coupled to
monophosphate, is coupled to
exergonic electron transport from a
exergonic dephosphorylation of an
substrate to the final electron
organic substrate.
acceptor, oxygen.
 Exergonic passage of protons along
a proton gradient.

 During glycolysis in the cytoplasm  During electron transport chain

Occurrence  During Krebs cycle in the (ETC) in the inner mitochondrial
mitochondrial matrix membrane

 Produces almost 90% of ATP

ATP  Produces only a small amount of
generated in respiration.
production ATP.

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 5.8 Respiratory Poison or Inhibitors

Oxidative phosphorylation is susceptible to inhibition at all stages of the process (Fig. 25). Specific
inhibitors of electron transport were invaluable in revealing the sequence of electron carriers in the
respiratory chain. For example, rotenone and amytal block electron transfer in NADH-Q
oxidoreductase (Complex I) and thereby prevent the utilisation of NADH as a substrate

The effects of three classes of respiratory poisons provided evidence for chemiosmosis and its
dependence on the structural organisation of the mitochondrial membrane.

1. Poisons that block electron flow

 Various poisons block the movement of electrons down the ETC.
 For examples, carbon monoxide, cyanide and hydrogen sulfide block the transport of
electrons from cytochrome a3 to oxygen.
 Hence, ATP production is completely inhibited.

2. Poisons that inhibit ATP synthase

 This class of poisons, which includes the antibiotic oligomycin, directly inhibits ATP
synthase by preventing the influx of protons through ATP synthase.
 Although the proton gradient becomes larger than normal, its potential energy cannot be
tapped to make ATP.

3. Poisons that make the inner mitochondrial membrane leaky to protons

 These poisons, such as 2, 4-dinitrophenol and certain other acidic aromatic compounds, are
known as uncoupling agents. These substances carry protons across the inner
mitochondrial membrane.
 As the protons leak back across the membrane, the proton gradient dissipates, i.e. the ETC
and proton gradient build-up are uncoupled.
 Since no proton gradient is formed, no ATP can be made by oxidative phosphorylation.
Rather, the energy derived from electron transport is released as heat.

Fig. 25: Poisons blocking electron flow through the ETC.

HWA CHONG INSTITUTION / 2022-2023 H2 BIOLOGY / Core Idea 3 / Energetics II / Cellular Respiration 32
 CONCEPT CHECK THREE

(a) Where does oxidative phosphorylation take place? [1]

(b) What are the major functions of the oxidative phosphorylation? [2]

Whiteflies feed on the crops and pose a major problem. Diafenthiuron inhibits the ATP synthase
complex in insects and is used in the insecticide spray.

(c) Describe what happens when the ATP synthase complex is inhibited. [2]

HWA CHONG INSTITUTION / 2022-2023 H2 BIOLOGY / Core Idea 3 / Energetics II / Cellular Respiration 33
 6. ANAEROBIC RESPIRATION

6.1 A Small yield of ATP in Anaerobic conditions

As mentioned in Section 3.1, glycolysis can take place in the absence of oxygen. Under such
anaerobic conditions, no further oxidation of pyruvate occurs, no acetyl CoA is formed and no
additional ATP can be generated.

Instead, the energy needs of the cell are met by the modest yield of 2 ATP molecules per glucose
in the glycolytic pathway. As a result, cells must consume glucose more rapidly in order to maintain
steady-state cellular ATP levels.

6.2 Formation of Ethanol in Yeast and Lactate in Mammals in Regeneration of NAD

Anaerobic respiration or fermentation is thus an oxygen-independent process, occurring in the

cytosol that uses organic molecules as terminal electron acceptors for the regeneration of NAD.

There are two common types of fermentation, where pyruvate is used as an electron acceptor:

1. Lactic acid fermentation (Fig. 26):pyruvate is converted to lactate (ionised form of lactic acid)
2. Alcoholic fermentation (Fig. 27): pyruvate is converted to ethanol and CO2

Lactic Acid Fermentation Alcoholic Fermentation

Equation
NADH + C3H4O3  NAD + C3H6O3 NADH + C3H4O3  NAD + C2H5OH + CO2
(pyruvate) (lactate) (pyruvate) (ethanol)
Steps involved
 Pyruvate is reduced directly by Pyruvate is converted to ethanol in 2
NADH to form lactate as a waste steps:
product, with no release of CO2  CO2 released from pyruvate, which
 Lactate is the ionised form of is itself converted to acetaldehyde.
lactic acid.  Acetaldehyde is reduced by NADH to
ethanol.
Occurrence  Occurs in animals and in certain In fungi e.g. yeast
fungi and bacteria.
In most plant tissues
In animals:
 In the short term, the reaction
satisfies the greater priority of
regenerating NAD.
 In the long term, lactic acid is
toxic and must ultimately be
removed.

In certain fungi and bacteria

 Lactic acid fermentation is used
in the dairy industry to make
cheese and yoghurt.

Products Lactic acid, NAD Ethanol, NAD

HWA CHONG INSTITUTION / 2022-2023 H2 BIOLOGY / Core Idea 3 / Energetics II / Cellular Respiration 34
 Fig. 26: Lactic acid fermentation. Fig. 27: Alcoholic fermentation.

7. COMPARING ENERGY YIELD OF AEROBIC AND ANAEROBIC RESPIRATION

Aerobic Respiration Anaerobic Respiration

(Alcoholic and lactic acid fermentation)

Maximum no. of ATP produced per glucose No. of ATP produced per glucose molecule
molecule oxidised = 38 molecules oxidised = 2 molecules

Amount of energy captured as ATP during aerobic respiration is maximally 19x as much as during
anaerobic respiration. From this viewpoint, therefore, aerobic respiration is much more efficient
than anaerobic respiration.

A great deal of energy remains locked with ethanol and lactic acid.

1. The energy in ethanol is permanently unavailable to yeast as 1 carbon has been lost as CO2,
hence ethanol cannot be regenerated to pyruvate which clearly indicates that alcoholic
fermentation is an inefficient energy-producing process.

2. Lactic acid, however, can be reconverted to pyruvate (which will enter the Krebs cycle) when
oxygen becomes available again.

HWA CHONG INSTITUTION / 2022-2023 H2 BIOLOGY / Core Idea 3 / Energetics II / Cellular Respiration 35
 8. EFFECT OF FACTORS ON RATE OF RESPIRATION

Aerobic respiration requires a steady input of fuel molecules and oxygen. Under normal conditions
these materials are adequately provided and do not affect the rate of respiration. Instead, the rate of
aerobic respiration is regulated by how much ADP and phosphate are available, with ATP
synthesis continuing until most of the ADP has been converted to ATP. At this point oxidative
phosphorylation slows considerably, which in turn slows down the citric acid cycle.

8.1 Substrate Concentration and Type of Substrate

The concentration and type of substrate present in the cells greatly affect the rate and course of
respiration. It has been shown that plants respire more rapidly after having been exposed to
conditions favourable for photosynthesis during which carbohydrates are synthesised. Increase in
respiration has also been observed to be associated with increase in soluble sugars.

Many organisms, including humans, depend on nutrients other than glucose as a source of energy.
In fact, you usually obtain more of your energy by oxidising fatty acids than by oxidising glucose.
Amino acids derived from protein digestion are also used as fuel molecules. Such nutrients are
transformed into one of the metabolic intermediates that are fed into glycolysis or the citric acid cycle
(Fig. 28).

Fig. 28: Energy from proteins, carbohydrates and fats.

HWA CHONG INSTITUTION / 2022-2023 H2 BIOLOGY / Core Idea 3 / Energetics II / Cellular Respiration 36
 8.2 Temperature

Like most chemical reactions, the rate of respiration is greatly influenced by temperature. Estimation
of temperature coefficient Q10 of the process for a rise in temperature from 8 to 18°C gives a Q10 of
2, indicating a chemical reaction. If the rise is at a much higher starting temperature, say between
20° and 30°C, then the Q10 may fall below 2. Different plants or plant parts may show considerable
variation in regard to optimum temperature for respiration.

In some cases the rate of respiration increases at lower temperature. For example, in white
potatoes, rate of respiration increases if the temperature in lowered to just above freezing point. This
increase in the rate of respiration is primarily due to increase in the quantity of substrate (such as
soluble carbohydrates) which tend to accumulate in Irish potato at temperature slightly above 0°C.

8.3 Investigating Respiration

8.3.1 Method I

The rate of respiration of an organism is an indication of its demand for energy. Respiration rate,
the uptake of oxygen per unit time, may be measured by means of a respirometer (Fig. 29). The
manometer in this apparatus detects change in pressure or volume of a gas. Respiration by tiny
organisms (germinating seeds or fly maggots are ideal) that are trapped in the chamber of the
respirometer alters the composition of the gas there, once the screw clip has been closed.

Fig. 29: A respirometer to measure respiration rate.

Carbon dioxide produced in respiration is absorbed by a suitable chemical such as soda lime or a
concentrated solution of potassium hydroxide (KOH) or sodium hydroxide (NaOH). Any decrease in
the volume of air surrounding the organisms results from their oxygen consumption.

HWA CHONG INSTITUTION / 2022-2023 H2 BIOLOGY / Core Idea 3 / Energetics II / Cellular Respiration 37
Oxygen consumption in unit time can be measured by reading the level of manometer fluid against
the scale.

Alternatively, the volume of air from the syringe that must be injected back into the respirometer tube
to make the manometer fluid level in the two arms equal again is equivalent to the volume of oxygen
taken up by the respiring organisms.

8.3.2 Compensation Tube

Changes in temperature and pressure alter the volume of air in the apparatus, and so the temperature
of the surroundings must be kept constant while readings are taken, by using a thermostatically
controlled water bath. The presence of a control tube containing an equal volume of inert materials
to the volume of organisms used helps to compensate for changes in atmospheric pressure.

Once measurements have been taken at a series of temperatures, a graph can be plotted of oxygen
consumption against temperature.

8.3.3 Respiratory Quotient

The same apparatus can be used to measure the Respiratory Quotient (RQ) of an organism. First,
oxygen consumption at a particular temperature is found (xcm3min-1). Then the respirometer is set
up with the same organism at the same temperature, but with no chemical to absorb carbon dioxide.
The manometer scale will show whether the volumes of oxygen absorbed and carbon dioxide
produced are the same.

When the volumes are the same, the level of the manometer fluid will not change and the RQ = 1.
When more carbon dioxide is produced than oxygen absorbed, the scale will show an increase in the
volume of air in the respirometer (by ycm3min-1).

The RQ can then be calculated:

CO2 x+y
RQ = 
O2 x

Conversely, when less carbon dioxide is produced than oxygen absorbed, the volume of air in the
respirometer will decrease (by zcm3min-1) and the calculation will be:

CO2 x-z
RQ = 
O2 x

8.3.4 Method II

Another way of investigating the rate of respiration of yeast is to use a redox dye such as a solution
of dichlorophenolindophenol (DCPIP) or of methylene blue. These dyes do not damage cells and so
can be added to a suspension of yeast cells. When reduced, these blue dyes become colourless.
The rate of change from blue to colourless is a measure of the rate of respiration of the yeast.

This technique can be used to investigate the effect of various factors on yeast respiration, such as
substrate concentration, types of substrates and temperature.

HWA CHONG INSTITUTION / 2022-2023 H2 BIOLOGY / Core Idea 3 / Energetics II / Cellular Respiration 38
 Fig. 30: Using the respirometer to derive respiratory quotient value.

HWA CHONG INSTITUTION / 2022-2023 H2 BIOLOGY / Core Idea 3 / Energetics II / Cellular Respiration 39