Required practical 9

• Describe how a respirometer can be used to measure the rate of

  aerobic respiration (by measuring oxygen uptake)

  1. Add set mass of single-celled organism eg. yeast to set volume / conc. of substrate eg. glucose

  2. Add a buffer to keep pH constant

  3. Add a set volume / conc. of a chemical that absorbs CO2 eg. 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
• Respirometer set up:

  A) glass tubing

  B) single celled organism

  C) chemical that absorbs carbon dioxide

  D) graduated scale

  E) capillary tubing

  F) coloured liquid

  
• Explain why the liquid moves.

  (4)

  ● Organisms aerobically respire so take in O2

  ● CO2 given out but absorbed by sodium hydroxide solution

  ● So volume of gas and pressure in container decrease

  ● Fluid in tube moves down pressure gradient towards organism
• Explain why the respirometer

  apparatus is left open for 10

  minutes. (1)

  ● Allow apparatus to equilibrate

  ● Allow for overall pressure expansion/change throughout

  ● Allow respiration rate of organisms to stabilise
• Explain why the apparatus

  must be airtight. (2)

  ● Prevent air entering or leaving

  ● Would change volume and pressure, affecting movement of liquid
• Describe a more accurate way

  to measure volume of gas. (1)

  ● Use a gas syringe
• Suggest a suitable control

  experiment and explain why it

  is necessary. (2)

  ● 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
• Describe how a respirometer can be used to measure the rate of

  anaerobic respiration (by measuring carbon dioxide release)

  ● Repeat experiment as above but remove chemical that absorbs CO2

  ● Make conditions anaerobic, for example:

  ○ Layer of oil / liquid paraffin above yeast to stop O2 diffusing in

  ○ Add a chemical that absorbs O2

  ○ Leave for an hour to allow O2 to be respired and used up
• Respirometer set up: (anaerobic)

  A) glass tubing

  B) graduated scale

  C) capillary tubing

  D) coloured liquid

  E) single celled organism

  
• Explain why the liquid moves. (3)

  ● Yeast anaerobically respire so release CO2

  ● So volume of gas and pressure in container increase

  ● So fluid in capillary tube moves down a pressure

  gradient away from organism
• Explain why the apparatus is left for

  an hour after the culture has reached

  a constant temperature. (1)

  ● Allow time for oxygen to be used / respired
• Describe how rate of respiration can be calculated

  1. Calculate volume of O2 / CO2 consumed / released (calculate area of a cylinder)
  2. Calculate cross-sectional area of capillary tube using π r2

  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 eg. cm3min-1g-1
• Describe how redox indicator dyes such as Methylene blue can be used to

  measure rate of respiration

  1. Add a set volume of organism eg. yeast

  and a set volume of respiratory substrate

  eg. 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 mins

  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)
• why do redox indicators change colour?

  ● 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
• Give two examples of

  variables that could be

  controlled. (2)

  ● Volume of single-celled organism

  ● Volume / conc. / type of respiratory substrate

  ● Temperature (with a water bath)

  ● pH (with a buffer)

  ● Volume of redox indicator (only control)
• Why leave tubes in the water

  bath for 5 minutes? (1)

  ● Allow for solutions to equilibrate and reach the same temperature

  as the water bath
• Suggest a suitable control

  experiment and explain why it

  is necessary. (3)

  ● 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
• Suggest and explain why you

  must not shake tubes

  containing methylene blue.

  (3)

  ● 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
• Suggest one source of error in

  using methylene blue. Explain

  how this can be reduced. (2)

  ● 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
• explain why a log scale is used to record the number of cells

  difference between numbers is too large