OCR A Level Biology A: Glycolysis, the link reaction and the Krebs cycle (H420)

This guide covers learning outcomes 5.2.2(a) to 5.2.2(g) of the OCR A Level Biology A (H420) specification, Version 4.1 (April 2026), first assessment 2025. It follows the first part of respiration: why cells need it, where it happens, and the stages up to the production of reduced coenzymes. Track progress on the OCR A Level Biology hub and the checklist. The revision notes and practice questions go with this page, and the free diagnostics show where to start. Earlier ideas are in nucleotides and ATP and cell structure.

What the specification asks for

Outcome	What you must be able to do
5.2.2(a)	Explain why plants, animals and microorganisms respire, with examples such as active transport and named metabolic reactions
5.2.2(b)	Describe the structure of a mitochondrion: outer and inner membranes, cristae, matrix and mitochondrial DNA
5.2.2(c)	Describe glycolysis and say where it happens, including phosphorylation, splitting and oxidation to pyruvate, with a small ATP yield and reduced NAD
5.2.2(d)	Describe the link reaction, its site, acetyl CoA formation, decarboxylation and reduction of NAD
5.2.2(e)	Describe the Krebs cycle and its site, including citrate and oxaloacetate, decarboxylation, dehydrogenation and substrate-level phosphorylation
5.2.2(f)	Explain the importance of NAD, FAD and coenzyme A
5.2.2(g)	State the site of oxidative phosphorylation and the roles of electron carriers, oxygen and cristae

The specification says that names of the intermediate compounds in the Krebs cycle are not required. You need citrate and oxaloacetate only.

5.2.2(a) Why cells need to respire

Respiration transfers energy stored in complex organic molecules, such as glucose, to ATP. ATP is the immediate source of energy for biological processes because its hydrolysis releases energy in small, usable amounts. Cells hold very little ATP, so they must keep making it.

Organism	Process that needs ATP	Detail
Animal	Active transport	Sodium–potassium pumps move ions against their gradients
Animal	Muscle contraction	Myosin heads use ATP to move along actin
Plant	Active transport	Root hair cells take up mineral ions against a concentration gradient
Plant	Synthesis of macromolecules	Building polymers such as cellulose and proteins
Microorganism	Growth and division	DNA replication and protein synthesis in yeast or bacteria
All	Metabolic reactions	Phosphorylation of glucose in glycolysis uses ATP, and joining nucleotides or amino acids needs energy

Plants respire as well as photosynthesise. Respiration goes on in all living cells, day and night, including cells with no chloroplasts such as roots. Mammals also release some of the energy from respiration as heat, which helps to maintain body temperature.

5.2.2(b) The mitochondrion

A mitochondrion is an organelle bounded by two membranes. It is roughly the size of a bacterium, so it is seen as a small oval or rod under an electron microscope, and in sections it often looks round.

Component	Structure	Function
Outer membrane	Smooth, one phospholipid bilayer	Separates the organelle from the cytoplasm and lets small molecules pass
Intermembrane space	Narrow gap between membranes	Where protons accumulate during oxidative phosphorylation
Inner membrane	Folded, more selective	Holds electron carriers and ATP synthase
Cristae	Folds of the inner membrane	Increase surface area for oxidative phosphorylation
Matrix	Fluid-filled interior	Contains enzymes for the link reaction and Krebs cycle, and DNA
Mitochondrial DNA	Short, circular DNA in the matrix	Codes for some mitochondrial proteins

Mitochondria also contain ribosomes that are similar in size to those of prokaryotes, which fits the idea that mitochondria were once free-living bacteria. Cells with high ATP demand, such as muscle fibres and the cells of the proximal convoluted tubule, have many mitochondria with extensive cristae.

Worked example: size from an electron micrograph

A mitochondrion measures 27 mm long on an electron micrograph. The magnification is ×15 000. Find its real length in micrometres.

actual size = image size / magnification
            = 27 mm / 15 000
            = 0.0018 mm
            = 0.0018 × 1000 = 1.8 µm

Convert units at the end, and keep the unit with the answer. Image sizes measured in mm are easiest to handle by dividing, then multiplying by 1000 for micrometres.

5.2.2(c) Glycolysis

Glycolysis happens in the cytoplasm. It does not need oxygen, so it is part of both aerobic and anaerobic respiration. It turns one six-carbon glucose into two three-carbon pyruvate molecules.

1. Phosphorylation. Two ATP each transfer a phosphate group, first to glucose and then to the intermediate, producing hexose bisphosphate. This makes the sugar reactive and traps it in the cell. Two ATP are used per glucose.
2. Splitting. Hexose bisphosphate (6C) splits into two molecules of triose phosphate (3C each).
3. Oxidation. Each triose phosphate is oxidised by removing hydrogen, which is accepted by NAD to form reduced NAD. Each is converted in further steps to pyruvate. Four ATP are made by substrate-level phosphorylation, two from each triose phosphate.

The net result for each glucose is 2 ATP, 2 reduced NAD and 2 pyruvate. Four ATP are made in total but two were spent in stage 1, leaving 4 − 2 = 2.

Worked example: glycolysis in moles

A sample of cells uses 0.25 mol of glucose by glycolysis. Find the net ATP, reduced NAD and pyruvate, and the mass of glucose (Mr 180).

ATP used            = 0.25 × 2 = 0.50 mol
ATP made            = 0.25 × 4 = 1.00 mol
net ATP             = 1.00 − 0.50 = 0.50 mol
reduced NAD         = 0.25 × 2 = 0.50 mol
pyruvate            = 0.25 × 2 = 0.50 mol
mass of glucose     = 0.25 × 180 = 45 g

Substrate-level phosphorylation means ATP is made by transferring a phosphate group directly from a phosphorylated substrate to ADP. It does not use an electron transport chain.

5.2.2(d) The link reaction

If oxygen is available, pyruvate leaves the cytoplasm and enters the mitochondrial matrix through the two membranes using transport proteins. The link reaction takes place in the matrix. Each pyruvate (3C) is:

• decarboxylated: a carbon dioxide molecule is removed, leaving a two-carbon acetyl group
• dehydrogenated: hydrogen is removed and accepted by NAD, which becomes reduced NAD
• combined with coenzyme A to form acetyl CoA

pyruvate + NAD + CoA  →  acetyl CoA + reduced NAD + CO₂

No ATP is made. One glucose gives two pyruvate, so the link reaction runs twice per glucose, giving 2 CO₂, 2 reduced NAD and 2 acetyl CoA. Acetyl CoA carries the acetyl group to the Krebs cycle.

5.2.2(e) The Krebs cycle

The Krebs cycle also takes place in the matrix. It is a cycle because the starting compound is regenerated at the end.

1. The two-carbon acetyl group of acetyl CoA joins oxaloacetate (4C) to form citrate (6C). Coenzyme A is released and can pick up another acetyl group.
2. Citrate is decarboxylated and dehydrogenated in a series of enzyme-controlled steps, losing two CO₂ molecules and several pairs of hydrogen atoms.
3. One molecule of ATP is made by substrate-level phosphorylation.
4. The 4C compound that remains is converted back to oxaloacetate, ready to accept another acetyl group.

Each turn of the cycle releases two molecules of CO₂ and forms three of reduced NAD, one of reduced FAD and one of ATP. One glucose gives two acetyl CoA, so the cycle turns twice per glucose and the numbers double.

Product, per glucose	Glycolysis	Link reaction (two runs)	Krebs cycle (two turns)	Total
Carbon dioxide	none	2	4	6
Reduced NAD	2	2	6	10
Reduced FAD	none	none	2	2
ATP made directly	2 net	none	2	4

Decarboxylation matters because it removes carbon from the cycle so citrate can be converted back to oxaloacetate, and it is how the carbon of glucose leaves as CO₂. Dehydrogenation matters because the hydrogen is accepted by NAD and FAD, which carry it to oxidative phosphorylation. Substrate-level phosphorylation gives a small direct ATP yield. All six carbons of glucose have left as CO₂ by the end of the Krebs cycle. Most of the energy is still held in the reduced coenzymes, which is why oxidative phosphorylation matters.

Worked example: tracking 12 pyruvate molecules

Twelve pyruvate molecules enter the matrix. Find the totals from the link reaction and Krebs cycle.

Link reaction:  12 acetyl CoA, 12 CO₂, 12 reduced NAD
Krebs cycle:    12 turns
                CO₂         = 12 × 2 = 24
                reduced NAD = 12 × 3 = 36
                reduced FAD = 12 × 1 = 12
                ATP         = 12 × 1 = 12
Totals:         CO₂ = 12 + 24 = 36
                reduced NAD = 12 + 36 = 48
                reduced FAD = 12, ATP = 12

Check: 12 pyruvate come from 6 glucose, and 6 × 6 = 36 CO₂ matches.

5.2.2(f) The importance of coenzymes

A coenzyme is a non-protein organic molecule that helps an enzyme to work. In respiration, coenzymes act as carriers that move chemical groups or hydrogen between reactions, and are recycled.

Coenzyme	What it carries	Where it is used
NAD	Hydrogen (as hydrogen atoms, or protons and electrons)	Glycolysis, link reaction and Krebs cycle; the reduced form delivers to the electron carriers
FAD	Hydrogen	Krebs cycle; the reduced form delivers to the electron carriers
Coenzyme A	The acetyl group	Link reaction (forms acetyl CoA) and delivery to the Krebs cycle

NAD and FAD are reduced when they accept hydrogen and oxidised when they give it up. Dehydrogenase enzymes catalyse the removal of hydrogen. The supply of oxidised NAD and FAD must be renewed for glycolysis, the link reaction and the Krebs cycle to carry on, which is why the electron carriers matter.

5.2.2(g) Oxidative phosphorylation: site and key roles

Oxidative phosphorylation takes place on the inner mitochondrial membrane, including the cristae. Reduced NAD and reduced FAD from the earlier stages give up their hydrogen, and electrons pass along a chain of electron carriers. Energy is released step by step as the electrons move.

• The electron carriers are fixed in the inner membrane, so the cristae, with their large surface area, hold many chains and ATP synthase enzymes.
• Oxygen sits at the end of the chain as the final electron acceptor: it takes up the electrons and, with protons, forms water.
• Without oxygen the chain stops, reduced NAD and FAD stay reduced. Oxidised NAD and FAD then run short, so the link reaction and Krebs cycle slow down.

How the released energy builds ATP is a separate outcome (chemiosmosis), covered in the chemiosmosis and anaerobic respiration guide, so only the site and roles are needed here.

Common errors

• Saying glycolysis happens in the mitochondrion. It is in the cytoplasm.
• Writing that the link reaction or Krebs cycle makes lots of ATP. They make little or none directly; they mainly make reduced coenzymes.
• Forgetting that everything after glycolysis is doubled for each glucose, because glucose gives two pyruvate.
• Calling the cycle “an oxygen-using stage” without explaining that oxygen is needed only for reoxidation of the carriers.
• Mixing “decarboxylation” (CO₂ removed) with “dehydrogenation” (hydrogen removed).
• Giving 2 ATP as the glycolysis yield without distinguishing gross from net.

Next steps

Condense the content with the revision notes, then test yourself with the practice questions. The Foundations in biology guide covers microscopy and magnification calculations.

Official syllabus

OCR, A Level Biology A (H420) specification, Version 4.1 (April 2026), first assessment 2025: section 5.2.2, learning outcomes (a) to (g).