Required Practical 9

Question 1

Describe how a respirometer can be used to measure the rate of aerobic respiration.

Answer 1

Measures O₂ uptake:

1. Add a set mass of single-celled organism e.g. yeast to a set volume / concentration of substrate e.g. glucose.
2. Add a buffer to keep pH constant.
3. Add a chemical that absorbs CO₂ e.g. sodium hydroxide.
4. Place in water bath at a set temperature and allow to equilibrate.
5. Measure distance moved by coloured liquid in a set time.

Question 2

Explain why the liquid moves in a respirometer when measuring the rate of aerobic respiration.

Answer 2

• Organisms aerobically respire -> take in O₂
• CO₂ given out but absorbed by sodium hydroxide solution
• So volume of gas and pressure in container decrease.
• So fluid in capillary tube moves down a pressure gradient towards organism.

Question 3

Explain why the respirometer apparatus is left open for 10 minutes.

Answer 3

• Allow apparatus to equilibrate.
• Allow for overall pressure expansion/change throughout.
• Allow respiration rate of organisms to stabilise.

Question 4

Explain why the apparatus used to measure the rate of aerobic respiration must be airtight.

Answer 4

• Prevent air entering or leaving.
• Would change volume and pressure, affecting movement of liquid.

Question 5

Describe a more accurate way to measure a volume of gas rather than using a respirometer.

Answer 5

• Use a gas syringe.

Question 6

Suggest a suitable control experiment when measuring aerobic respiration and explain why it is necessary.

Answer 6

• No organisms OR use inert objects OR use dead organisms
  AND all other conditions / apparatus / equipment the same.
• To show that (respiring) organisms are causing liquid to move / taking up oxygen / causing the change in volume / pressure.

Question 7

Describe how a respirometer can be used to measure the rate of anaerobic respiration.

Answer 7

Measures CO₂ uptake:

1. Add a set mass of single-celled organism e.g. yeast to a set volume / concentration of substrate e.g. glucose.
2. Add a buffer to keep pH constant.
3. Add a chemical that absorbs O₂ and a layer of oil/liquid paraffin above yeast to stop O₂ diffusing in.
4. Place in water bath at a set temperature and allow to equilibrate.
5. Leave for an hour to allow O₂ to be respired and used up.
6. Measure distance moved by coloured liquid in a set time.

Question 8

Explain why the liquid moves in a respirometer when measuring the rate of anaerobic respiration.

Answer 8

• Yeast anaerobically respire -> release CO₂
• So volume of gas and pressure in container increase.
• So fluid in capillary tube moves down a pressure gradient away from organism.

Question 9

Explain why the apparatus used to measure the rate of anaerobic respiration is left for an hour after the culture has reached a constant temperature.

Answer 9

• Allow time for oxygen to be used / respired.

Question 10

Describe how the rate of respiration can be calculated.

Answer 10

1. Calculate volume of O₂ / CO₂ consumed / released (calculate area of a cylinder).
   
   • a. Calculate cross-sectional area of capillary tube using πr²
   • b. Multiply by distance liquid has moved
2. Divide by mass of organism and time taken.
3. Units - unit for volume per unit time per unit mass e.g. cm³ min⁻¹ g⁻¹.

Question 11

Describe how redox indicator dyes such as Methylene blue can be used to measure rate of respiration.

Answer 11

• Redox indicators (eg. methylene blue) change colour when they accept electrons becoming reduced.
• Redox indicators take up hydrogens and get reduced instead of NAD / FAD -> modelling their reactions.

1. Add a set volume of organism eg. yeast and a set volume of respiratory substrate e.g. glucose to tubes.
2. Add a buffer to keep pH constant.
3. Place in water bath at a set temperature and allow to equilibrate for 5 minutes.
4. Add a set volume of methylene blue, shake for a set time (do not shake again).
5. Record time taken for colour to disappear in tube.
6. Rate of respiration (s⁻¹) = 1 / time (sec).

Question 12

Give two examples of variables that could be controlled when using redox indicator dyes to measure the rate of respiration.

Answer 12

• Volume of single-celled organism.
• Volume / concentration / type of respiratory substrate
• Temperature (with a water bath).
• pH (with a buffer).
• Volume of redox indicator (only control).

Question 13

Why leave tubes in the water bath for 5 minutes when measuring the rate of respiration using redox indicator dyes?

Answer 13

• Allow for solutions to equilibrate and reach the same temperature as the water bath.

Question 14

Suggest a suitable control experiment when measuring the rate of respiration using redox indicator dyes and explain why it is necessary.

Answer 14

• Add methylene blue to boiled / inactive / dead yeast (boiling denatures enzymes).
• All other conditions the same.
• To show change is due to respiration in organisms.

Question 15

Suggest and explain why you must not shake tubes containing methylene blue.

Answer 15

• Shaking would mix solution with oxygen.
• Which would oxidise methylene blue / cause it to lose its electrons.
• So methylene blue would turn back to its original blue colour.

Question 16

Suggest one source of error in using methylene blue. Explain how this can be reduced.

Answer 16

• Subjective as to determination of colour change / end point.
• Compare results to a colour standard (one that has already changed) OR use a colorimeter for quantitative results.