Using a simple respirometer: Measuring the rate of O2 uptake in organisms.

Set up the respirometer as shown in the diagram. The soda lime absorbs carbon dioxide given out
during respiration. (The volume of CO2 given out is equal to the volume of O2 taken in for
respiration. So if the CO2 is not absorbed, the colored oil drop doesn’t move. If CO2 is absorbed
the oil drop moves as the volume of air in the test tube decreases.)

water bath at known constant temperature (20 )

Method-

 Place a fixed mass of maggots or peas into the test tube and replace the bung. (the cotton
wool or gauze prevents the small organisms from coming into contact with the soda
lime). The mass of organisms must be known as different masses have different rates of
respiration.
 Place the test tube in a water bath at 20 C to maintain a constant temperature. Leave for 2
minutes for the organisms to acclimatize with the taps open & to allow air containing O2
to enter
  Close the 3 way tap (connection to the syringe and the atmosphere to ensure the
apparatus is airtight) and immediately start a stop clock
 Measure the distance moved by the colored oil drop in a fixed time, e.g. 10 minutes in the
capillary tube using the scale
 Open the 3 way tap to the syringe and depress the plunger to reset oil drop (bring it to
zero)
 Repeat the experiment 3 more times at the same temperature. Find the mean volume of
oxygen used by multiplying the mean distance moved by the cross sectional are of the
capillary tube. Divide by the time taken, 10 minutes, to find the RATE of oxygen uptake.
(and then repeat at different temperatures but not above 35 C – if the experiment asks for
effect of temp). If it asks for different species of seeds, keep the mass of seeds the same
because different masses have different rates of respiration.
 Open the connection to the outside air after each experiment so that the larvae get fresh
air having oxygen for respiration

Precautions

 The temperature and mass of organisms used must be the same for each trial (unless the
question asks you to vary these). Different masses have different rates of respiration.
Changes in temp affect enzyme activity, thereby affecting the rate of respiration because
respiration is an enzyme catalyzed reaction. By keeping the temp constant, you are
eliminating temp as a variable, increasing the validity of the expt.
 The potassium hydroxide solution or soda lime should be replaced regularly to prevent it
from getting saturated with carbon dioxide.
 All the connections when the tubes are fitted must be completely airtight.

Limitations

 The KOH may get saturated with CO2

 There is no control test tube. The KOH also absorbs carbon dioxide from the atmosphere
in addition to absorbing CO2 given out by respiration
 The oxygen supply may be insufficient for 10 minutes or the supply of glucose may be
insufficient.

To Increase Validity

A control respirometer can also be set up which doesn’t have living organisms but has
everything else the same. You would not expect the fluid to move in the control tube but it may
do so because of temperature changes, which changes the vol. of gas (if there is no water bath).
This allows you to control for this variable by subtracting the distance moved by the fluid in the
control from the distance moved in the experimental one (connected to the living organisms) to
give you the adjusted distance moved. In the control, one colored oil drop moves because the
KOH absorbs CO2 from the normal air.
e.g. distance moved in the experimental tube in 10mins = 8mm
distance moved in the control = 0.5mm
Actual distance moved = 8mm – 0.5mm = 7.5mm

The diagram below shows a more accurate respirometer.

The 2nd tube acts as a control. It compensates for any changes in pressure (or temperature) within
the apparatus. The distance moved by the manometer fluid can be recorded.

 Place both test tubes in the water bath at 20 C with the manometer tube outside the water
bath. Open the screw clip and the three way tap (to syringe and atmosphere). Leave for 5
mins for the larvae/seeds to acclimatize (and reach the respiration rate at that temperature
and also to allow for pressure changes of air in the apparatus.)
 Close the screw clip and the 3 way connection to the outside. (not the 3 way connection
to the syringe just yet)
 Use the syringe to reset the level of colored liquid in the U tube so that the level of liquid
in both arms is the same to begin with
 Close the connection to the syringe
 Record the distance moved by the colored liquid in a fixed time. (e.g. 10 minutes using
one scale in the capillary tube)
 Find the volume of oxygen used (described for the earlier respirometer) – give the
formula
 Repeat the entire experiment 3 more times and find the mean rate of oxygen uptake
(mean rate of respiration)
More Accurate Method

Also can measure distance moved by the manometer fluid or colored oil drop at 1 minute
intervals over a 10 minute period and plot distance moved on the y axis against time on the x.
The gradient of the graph will be the rate of oxygen uptake.

 At both temps the rate of O2 uptake is constant

 There is a higher rate at 30 C

The graph method is more accurate because the distance moved by the colored liquid in each
minute may not be the same

Rate of O2 uptake =( cross sectional area x mean distance moved) / time (10 mis)

Can compare rates of respiration at different temps by using a water bath

If the 3-way tap is not present, the syringe should be removed while acclimatizing and replaced
after 5 minutes to make the apparatus airtight.