Required practical 9 - Investigation into the effect of a named variable on the rate of respiration

Using a respirometer to measure rate of respiration

add set amount of single celled organism with set concentration of substrate

add pH buffer

add a chemical that absorbs CO2

place in water bath at a set temperature and allow to equilibrate

measure distance moved by coloured liquid in set time

why the liquid moves in a respirometer

organisms respire → take in O₂ and release CO₂ which is absorbed by sodium hydroxide

So volume of gas and pressure in container decrease

so fluid in capillary tube moves down a pressure gradient towards organism

Why respirometer is left open for 10 mins

allow apparatus to equilibrate

allow overall pressure expansion/change over time

allow respiration rate of organisms to stabilise

why the apparatus must be air tight

prevents air from entering or leaving

would change volume and pressure, affecting movement of liquid

more accurate way to measure volume of gas

use a gas syringe

Calculating respiration rate

distance liquid has moved x cross sectional area divided by mass of organism and time taken

how a respirometer could be used to measure the rate of anaerobic respiration

layer of oil above yeast to stop O₂ diffusing in

add a chemical that absorbs O₂

Leave for an hour to allow O₂ to be used up

why redox indicators (methylene blue) can be used to measure rate of respiration

change colour when they are reduced by electrons

Methylene blue method

set volume of organism and respiratory substrate

add a buffer to keep pH constant

place in water bath at a set temperature and allow to equilibrate for 5 mins

add a set volume of methylene blue, shake for a set time

record time take for colour to disappear in tube

variables which should controlled in methylene blue method

volume of single celled organism

volume/ conc of respiratory substrate

Temperature

pH

Volume of redox indicator

control experiment for methylene blue

add methylene blue to inactive yeast

maintain all other conditions

shows change is due to respiration in organisms

why you can’t shake tubes containing methylene blue

shaking would mix solution with oxygen

which would oxidise methylene blue

so methylene blue would turn back into its original blue colour

problem with using methylene blue

subjective when colour change happens/end point

compare results to a colour standard

or use a colorimeter for a quantitative result