outline respiration in anaerobic conditions in mammals (lactate fermentation) and in yeast cells (ethanol fermentation)

Respiration – Aerobic and Anaerobic Conditions (A‑Level Biology 9700)

Learning Objective

Describe the complete pathways of aerobic respiration and the two main anaerobic fermentations (lactate in mammals and ethanol in yeast). Explain the biochemical steps, key enzymes, energy yield (ATP and NAD⁺/FADH₂), calculate the respiratory quotient (RQ), and discuss the physiological or industrial significance. Apply this knowledge in a practical investigation.

1. Overview of Cellular Respiration

• Cellular respiration is the series of catabolic reactions that convert the chemical energy of glucose (or other substrates) into usable cellular energy (ATP).
• Aerobic respiration – O₂ is the final electron acceptor; the pathway proceeds through glycolysis, the link reaction, the Krebs cycle and oxidative phosphorylation.
• Anaerobic fermentation – O₂ is absent or insufficient; NAD⁺ must be regenerated by converting pyruvate into an organic end‑product, allowing glycolysis to continue.
• Overall aerobic equation for carbohydrate substrates:
  

  \$\text{C}6\text{H}{12}\text{O}6 + 6\text{O}2 \;\longrightarrow\; 6\text{CO}2 + 6\text{H}2\text{O} + \approx30\text{ ATP}\$

• Respiratory quotient (RQ) = \(\dfrac{\text{CO}2\ \text{produced}}{\text{O}2\ \text{consumed}}\).

2. Aerobic Respiration – Step‑by‑Step Pathway

2.1 Glycolysis (Cytosol)

• Location: Cytosol.
• Overall reaction:

  \$\text{Glucose} + 2\text{ADP} + 2\text{P}i + 2\text{NAD}^+ \;\longrightarrow\; 2\text{Pyruvate} + 2\text{ATP} + 2\text{NADH} + 2\text{H}^+ + 2\text{H}2\text{O}\$

• Key enzymes (hallmarks): hexokinase, phosphofructokinase‑1 (PFK‑1), pyruvate kinase.
• Regulation (allosteric): ATP inhibits PFK‑1; AMP and fructose‑2,6‑bisphosphate activate PFK‑1; pyruvate kinase is inhibited by ATP and activated by fructose‑1,6‑bisphosphate.
• Yield per glucose: 2 ATP (substrate‑level phosphorylation) + 2 NADH.

2.2 Link Reaction (Mitochondrial Matrix)

• Location: Mitochondrial matrix.
• Overall reaction (per 2 pyruvate):
  

  \$2\text{Pyruvate} + 2\text{CoA} + 2\text{NAD}^+ \;\longrightarrow\; 2\text{Acetyl‑CoA} + 2\text{CO}_2 + 2\text{NADH} + 2\text{H}^+\$

• Enzyme complex: pyruvate‑dehydrogenase complex (PDH).
• Co‑factors: thiamine pyrophosphate (TPP), lipoic acid, FAD, NAD⁺, CoA‑SH.
• Yield: 2 NADH (≈5 ATP after oxidative phosphorylation).

2.3 Krebs (Citric Acid) Cycle (Mitochondrial Matrix)

• Location: Mitochondrial matrix.
• Each acetyl‑CoA yields:
  

  3 NADH, 1 FADH₂, 1 GTP (≈1 ATP), 2 CO₂.

• Key enzymes (one per step): citrate synthase, aconitase, isocitrate dehydrogenase, α‑ketoglutarate dehydrogenase, succinyl‑CoA synthetase, succinate dehydrogenase, fumarase, malate dehydrogenase.

2.4 Oxidative Phosphorylation (Inner Mitochondrial Membrane)

• Location: Inner mitochondrial membrane (IMM).
• Electron carriers: NADH → Complex I → Q → Complex III → cytochrome c → Complex IV → O₂.
  

  FADH₂ → Complex II → Q → Complex III → cytochrome c → Complex IV → O₂.

• Chemiosmosis (Peter Mitchell): Transfer of electrons through the ETC pumps protons from the matrix into the inter‑membrane space, creating a proton‑motive force (electrochemical gradient). ATP synthase (Complex V) allows protons to flow back into the matrix, driving synthesis of ATP from ADP + Pᵢ.
• ATP yield per carrier: ≈2.5 ATP per NADH, ≈1.5 ATP per FADH₂.

2.5 Total Aerobic ATP Yield (Cambridge value)

3. Anaerobic Fermentation

3.1 Why Fermentation Is Required

• When O₂ supply cannot meet the demand for oxidative phosphorylation, NADH accumulates.
• Fermentation provides an alternative electron acceptor, regenerating NAD⁺ so that glycolysis can continue.
• Only the 2 ATP produced by glycolysis are available – no additional ATP is generated in the fermentation steps.

3.2 Lactate Fermentation (Mammalian Skeletal Muscle)

• Location of the fermentation step: Cytosol (same compartment as glycolysis).
• Steps:

  1. Glycolysis: Glucose → 2 pyruvate + 2 ATP + 2 NADH.
  2. Reduction of pyruvate:
     

     \$\text{Pyruvate} + \text{NADH} + \text{H}^+ \xrightarrow{\text{lactate dehydrogenase (LDH)}} \text{Lactate} + \text{NAD}^+\$

• Energy yield: 2 ATP per glucose (no extra ATP from the LDH step).
• Fate of lactate:

  • Transient accumulation in muscle → lowered pH (muscle fatigue).
  • Transport to liver via the bloodstream (Cori cycle); liver converts lactate back to glucose (requires 6 ATP).

3.3 Ethanol Fermentation (Yeast – Saccharomyces cerevisiae)

• Location of the fermentation steps: Cytosol.
• Steps:

  1. Glycolysis: Glucose → 2 pyruvate + 2 ATP + 2 NADH.
  2. Decarboxylation of pyruvate (pyruvate decarboxylase, co‑factor TPP):
     

     \$\text{Pyruvate} \xrightarrow{\text{PDH (yeast)}} \text{Acetaldehyde} + \text{CO}_2\$

  3. Reduction of acetaldehyde (alcohol dehydrogenase):
     

     \$\text{Acetaldehyde} + \text{NADH} + \text{H}^+ \xrightarrow{\text{ADH}} \text{Ethanol} + \text{NAD}^+\$

• Overall reaction:

  \$\text{C}6\text{H}{12}\text{O}6 + 2\text{ADP} + 2\text{P}i \;\longrightarrow\; 2\text{C}2\text{H}5\text{OH} + 2\text{CO}_2 + 2\text{ATP}\$

• Energy yield: 2 ATP per glucose (same as lactate fermentation).
• Industrial relevance: production of alcoholic beverages, bio‑ethanol fuel, and CO₂ for baking/leavening.

4. Respiratory Quotient (RQ)

RQ is calculated from the stoichiometric equations for the oxidation of a substrate:

4.1 Worked Example – Glucose (Carbohydrate)

• Equation: \$\text{C}6\text{H}{12}\text{O}6 + 6\text{O}2 \rightarrow 6\text{CO}2 + 6\text{H}2\text{O}\$
• CO₂ produced = 6 mol, O₂ consumed = 6 mol → RQ = 6/6 = 1.0.

4.2 Worked Example – Palmitic Acid (Fat)

• Equation (simplified): \$\text{C}{16}\text{H}{32}\text{O}2 + 23\text{O}2 \rightarrow 16\text{CO}2 + 16\text{H}2\text{O}\$
• RQ = 16/23 ≈ 0.70.

4.3 Worked Example – Average Protein (e.g., alanine)

• Equation (simplified for an average amino acid, C₅H₁₀N₂O₃): \$\text{C}5\text{H}{10}\text{N}2\text{O}3 + 5\text{O}2 \rightarrow 5\text{CO}2 + 5\text{H}2\text{O} + \text{NH}3\$
• RQ = 5/5 = 1.0 for the carbon skeleton; when nitrogen is considered, the effective RQ for mixed protein metabolism is ≈0.8 (Cambridge value).

5. Practical Investigation – Measuring Fermentation Activity

1. Objective: Compare the rate of lactate fermentation in mammalian muscle tissue with ethanol fermentation in active yeast.
2. Materials: sealed fermentation tubes or gas‑tight syringes, 0.5 M glucose solution, fresh baker’s yeast, freshly obtained muscle biopsy (or homogenate), pH meter, spectrophotometric lactate assay kit, alcohol oxidase‑peroxidase kit for ethanol, ice‑bath, water bath (37 °C).
3. Method (outline):

   • Flush each tube with nitrogen to ensure anaerobiosis.
   • Add 5 mL glucose solution + 0.5 g tissue (muscle) in one tube; add 5 mL glucose + 0.5 g yeast in a second tube.
   • Incubate at 37 °C; record the volume of gas displaced at 0, 5, 10, 15 min.
   • Withdraw 0.5 mL aliquots at the same time points:

     • Muscle sample – determine lactate concentration (colourimetric assay).
     • Yeast sample – determine ethanol concentration (alcohol oxidase assay).

4. Data analysis:

   • Plot CO₂ volume (or gas pressure) versus time; the initial slope gives the fermentation rate (mmol CO₂ min⁻¹).
   • Check stoichiometry:

     • Lactate fermentation – no CO₂ expected; any gas measured indicates contamination.
     • Ethanol fermentation – 2 mol CO₂ per mol glucose; compare measured CO₂ with ethanol produced.

5. Evaluation (AO3):

   • Potential errors – gas leakage, incomplete removal of O₂, temperature drift.
   • Improvements – use a gas‑tight syringe, maintain constant temperature with a circulating water bath, verify anaerobiosis with a redox indicator.
   • Link to theory – discuss how the rate of NAD⁺ regeneration limits the overall ATP production in each system.

6. Comparison of Lactate vs. Ethanol Fermentation

7. Summary

• Aerobic respiration extracts ~30 ATP per glucose through glycolysis, the link reaction, the Krebs cycle and oxidative phosphorylation (chemiosmosis).
• In the absence of O₂, cells switch to fermentation to recycle NADH to NAD⁺, allowing glycolysis to continue at a modest 2 ATP per glucose.

  • In mammals, pyruvate is reduced to lactate (no CO₂ released); lactate is later reconverted to glucose in the liver (Cori cycle).
  • In yeast, pyruvate is decarboxylated to acetaldehyde (producing CO₂) and then reduced to ethanol.

• Both fermentations are essential: lactate fermentation for short‑term muscular activity, ethanol fermentation for industrial production of alcohols and bio‑fuels.
• Understanding the enzymes, regulation, energy yield and RQ calculations fulfills the core requirements of the Cambridge AS & A‑Level Biology syllabus.

8. Suggested Diagram

A flowchart illustrating the two branches from pyruvate:

• Glucose → Glycolysis → Pyruvate → (a) Lactate (LDH) –> muscle ↔ liver (Cori cycle)
• Glucose → Glycolysis → Pyruvate → (b) Acetaldehyde (pyruvate decarboxylase) → Ethanol (ADH) + CO₂
• Parallel aerobic route: Pyruvate → Acetyl‑CoA → Krebs cycle → ETC → ATP (≈30 ATP).

Label each step with its cellular compartment, key enzyme(s) and ATP/NADH yield.