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Practical Unit 5

This document provides background and methods for three core practical experiments: 1) Use a redox indicator to investigate respiration in yeast at different temperatures, measuring the time for the indicator to change color as the dependent variable. 2) Use a respirometer to determine respiration rate and respiratory quotient of organisms like germinating seeds or insects at different temperatures, measuring oxygen consumption over time. 3) Use a spirometer to investigate the effects of exercise on breathing parameters like tidal volume and rate in humans, measuring changes before and during exercise.

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Larina Mohammed
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34 views

Practical Unit 5

This document provides background and methods for three core practical experiments: 1) Use a redox indicator to investigate respiration in yeast at different temperatures, measuring the time for the indicator to change color as the dependent variable. 2) Use a respirometer to determine respiration rate and respiratory quotient of organisms like germinating seeds or insects at different temperatures, measuring oxygen consumption over time. 3) Use a spirometer to investigate the effects of exercise on breathing parameters like tidal volume and rate in humans, measuring changes before and during exercise.

Uploaded by

Larina Mohammed
Copyright
© © All Rights Reserved
We take content rights seriously. If you suspect this is your content, claim it here.
Available Formats
Download as PDF, TXT or read online on Scribd
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Practical unit 5

Core Practical 15 :Use an artificial hydrogen carrier (redox


indicator) to investigate respiration in yeast.

Equipment list
● Redox indicator - triphenyl tetrazolium chloride (TTC) solution
● Water baths at the following temperatures: 15, 20 25,30 35 °C
● Thermometer
● Yeast suspension / dried yeast, glucose and distilled water
● Distilled water
● Pipettes
● Test tubes
● Test tube rack
● Stopwatch
● Stirring rods

Background information

During oxidative phosphorylation in aerobic respiration the hydrogen


carriers, such as FAD and NAD, are regenerated by being oxidised .
In this experiment, the hydrogen carriers are oxidized by a redox indicator
(TTC) which is in turn reduced , causing a colour change to red.

Method:

Preparing the yeast : Add 10g of dried yeast and 50g of glucose to 1000
cm³ of distilled water, mix thoroughly .
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Independent variable: Temperature

Dependent variable: Time taken for the redox indicator to turn red

( can be used to indicate the rate of respiration 1/ time ).

1. Set up the first water bath at 35°C

2. Pipette 10 cm ³ of the yeast suspension into one test tube, and 1 cm³ of
TTC into a different test tube.

3. Place both test tubes into the water bath and leave them for 5 minutes to
allow them to reach the temperature of the water bath.

4. Quickly pour the TTC solution into the test tube containing the yeast,
give it a stir and start the stopwatch.

5. Stop timing once the solution has turned red and record the time
taken in a suitable table.

6. Repeat steps 1-5 with the remaining 4 temperatures.

7. Repeat several times at each temperature and calculate the mean, and
SD ., for reliability.

Controlled variables: SAME !!:

- Volume of yeast suspension, volume and concentration of TTC


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4

Core practical 16 :Use a simple respirometer to determine the


rate of respiration and RQ of an organism

❖ Describe how a respirometer works.


It is a chamber connected to a capillary tube with a drop of dye.
As the organism in the chamber respires and uses oxygen, the pressure
decreases and the liquid moves in the capillary tube. The volume of carbon
dioxide produced = the volume of oxygen used up. Soda lime/KOH is used
to absorb CO2 produced, hence allowing the measurement of the
volume of oxygen used up per unit time. Once calculated, the
rate of oxygen uptake can be calculated per gram of organism and then
compared between organisms, or under different conditions.
RQ can be calculated.
Independent variable:
- Different species of organism ( germinating seeds, insects)
- Different temperatures.
Dependent variable: Distance moved by coloured liquid in the capillary
tube per unit time!!.( used to
calculate volume of oxygen
consumed, and the
rate of respiration)
Method
1. Set up the airtight
respirometer as shown in the
diagram .
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2. Use the syringe to move the coloured fluid to the zero on the scale.
6. Close the 3-way tap to allow no more gas exchange to occur between
the apparatus set up and the outside atmosphere and start the stopwatch
immediately.
7. Measure the position of the coloured fluid from its starting point every
minute for 5 minutes, recording the distance moved in a suitable table. .
8. Open the 3-way tap connection to the outside air and use the syringe to
reset the capillary fluid.
9. Reset the drop to the zero and introduce fresh air →Repeat the steps
several times with each organism or temperature → calculate the average
and observe the variability.
Controlled variables :SAME !
1-Mass of organism
2-Temperature( if it is not the IV)
3-Mass of soda lime/ volume of KOH
Control : Use an identical apparatus , with the same conditions,
Replace the organism with an inert object of the same mass ( not size!!).
To compare , subtract ..( validity)

-To calculate the volume of oxygen taken in since the start (cm³) you
calculate the volume of the manometer tube that has been filled with
oxygen: V = 兀r² l

Where r is the radius of the tube (half its diameter), l is the distance moved.
E.g. If the fluid moved 1.5 cm in the first minute and the radius of the tube
was 0.5 mm then the volume produced would be:
V = 兀 x (0.5)² x 1.5 = 1.178 cm
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The rate of oxygen uptake per minute can be calculated by:


Dividing the VOLUME of oxygen produced by the TIME since starting
the stopwatch.
( MAY BE REQUIRED PER GRAM BODY MASS OF THE ORGANISM-→
DIVIDE BY THE MASS!
How can the volume of carbon dioxide produced be measured?
Perform two set-ups, one with soda lime (A) and onewithout (B).
Find the volume of gas used in the given time.
Volume of carbon dioxide: volume of A - volume of B

Formula for the respiratory quotient


RQ = Volume of carbon dioxide produced / Volume
of oxygen absorbed

Alternatively : 2 tubes can be used , connected to a U shaped


manometer, as shown in the diagram below.
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w.pmt
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Core practical 17 :Investigate the effects of exercise on tidal


volume, breathing rate, respiratory minute ventilation, and oxygen
consumption using data from spirometer traces.

Independent variable: Whether the test subject is resting or exercising.


Dependent variable: Change* in: -Tidal volume and breathing rate
- Volume or rate of oxygen consumption.
Method
A spirometer is closed chamber of air/oxygen floating over water,
connected to a tubing and mouth piece. The person breaths in and out ,
the carbon dioxide in the exhaled air is absorbed by soda lime, allowing us
to measure the volume and rate of oxygen uptake .
To use it, it must first be calibrated as follows:
● Empty the spirometer so no air remains in it, use the pen to make a mark
on the graph paper trace on the drum.
● Now add 1 dm3 volume of air to the spirometer, again let the pen mark on
the graph paper. From this you can use the vertical scale to calculate how
many squares/mm on the graph are equivalent to the volume of air just
added to the spirometer.
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You can now begin the main part of the experiment with the calibrated
spirometer.
1. While at rest, place a nose clip , so they are only breathing through their
mouth ( validity).
2. Put the disinfected mouthpiece of the spirometer into the person’s mouth
and allow them to breathe normally until they’re are accustomed to
breathing into the apparatus.
3. Set the kymograph to a rate of 1 mm per second and turn it on so it
starts rotating.
4. Record their normal, resting breathing for a minute and then ask them to
breathe deeply for one breath ( to show the vital capacity), then returning to
normal breathing.
5. The effects of exercise can then be observed by recording the
breathing pattern while asking the test subject to exercise for 1 minute
(such as running on the spot or star jumps).
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-Tidal volume – the volume of air that we breathe in and then out during one normal
breath. This is usually about 500 cm3.

From the trace= Vertical distance between the peak and trough of the wave
( take the mean of several waves)

-Breathing rate: number of breaths per minute

From the trace= Number of full* waves per minute.

- Inspiratory reserve volume – the maximum volume (in excess of the tidal volume)
we can breathe in during one forced breath.
- Expiratory reserve volume – the maximum volume (in excess of the tidal volume) we
can breathe out in one forced breath.

- Vital capacity – the maximum volume of air that can be breathed during a forced
breath in and then out.
-Residual volume is the volume of air left in the lungs after a forced breath out.

mt Minute volume (VE) (Ventilation Rate)


It is a function of the breathing rate X tidal volume. Unit dm3 min-1 (or cm3 min-1)

VE = breathing rate X Tidal volume


Ex . At rest = 12 X 0.5 dm3 = 6 dm3 min-1
-In exercise, both breathing rate and tidal volume increase
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Oxygen consumption:
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Core practical 18 : Investigate the production of amylase in


germinating cereal grains

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