Alex Bouman

Aim: Investigating the effect of temperature on the membrane

permeability of beetroot cells
Introduction and science
Beetroot (Beta Vulgaris) cell membranes contain betacyanin which is a pink/red
pigment located in the vacuole of the cell. When the cell is cut or altered the pigment
will spill out of the cell however when the cells membrane is damaged the pigment
will diffuse out. High temperatures will disrupt the structure of proteins and certain
chemicals such as fat solvents dissolve in the phospholipids, damaging the
membrane and therefore increased permeability, with moderate temperatures the
rate of diffusion will be faster and the particles of the pigments will have more Kinetic
Energy which means that the particles will move faster out of the cell. The
betacyanin will diffuse from a high to low concentration. The percentage of green
light absorbance will determine the permeability of the cell as itll help gauge how
much betacyanin diffuses out of the cell. The permeability of the cell will depend on
the intact structure of the of phospholipids and proteins which contribute to its
selectively permeable nature. High temperatures will disrupt the structure of proteins
and certain chemicals such as fat solvents dissolve in the phospholipids, damaging
the membrane and therefore increased permeability.

Hypothesis: The higher the temperature the more permeable the cell will be; the
rate of diffusion will increase with temperature until the point where the cell
membranes break due to the excess heat. After that point the rate of diffusion will no
longer increase but stay constant as the membrane would leak out the betacyanin.
Equipment:
Beaker (to hold water for the beetroot to rinse in)
Timer (to time how long boiling tubes are in the water bath)
4 water baths (for the amount of temperatures you want to test)
Boiling tubes (to contain the beetroot while in the water bath)
Test tube racks (to hold the boiling tubes)
Tile (to cut the beetroot on)
Knife (to cut the beetroot into pieces)
Ruler(cm)
Dissecting needle, 4 for each temperature
Square cutter (to cut the beetroot into equal widths, 1cm by 1cm)
Colorimeter (set to green light absorbance)
Paper towels (to dab the beetroot on)
Gloves (optional- avoids staining on hands)
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Alex Bouman

Lab coat (optional- Avoids staining on clothes)

Goggles (protects eyes, safety)
Cuvette (to hold the water for the colorimeter)
Thermometer (to measure the actual temperature in the water bath)
Measuring cylinder (25 cm3)
Plan
1. Set the separate water baths to the temperature: 20,40,60 and 80 degrees
Celsius) put a thermometer inside each bath to determine the actual
temperature.
2. Using a 25 cm3 measuring cylinder, fill it with 20 cm3 of distilled water
remember to fill the cylinders at eye level to avoid parallax error.
3. Pour 20 cm3 of distilled water into 4 separate boiling tubes and leave them on
the test tube rack to avoid any spillage.
4. Cut the beet root in to rectangles using the square cutter.
5. Using a knife and ruler cut the beetroot in to cubes of 2mm thickness, cut 5
cubes per temperature and skew it on the dissecting needle, do this for each
set of temperatures therefore using 4 dissecting needles.
6. Fill a beaker with water and rinse the dissecting needles until no more colour
leaks out the beetroot cubes. Change the water once it turns too red. Repeat
this for each dissecting needle.
7. Dab each dissecting needle with the beetroot cubes on it with a paper towel to
remove the water on them from the rinse, as you dont want any impurities
from the tap water in to the distilled water.
8. Label each boiling tube for their respective temperatures to avoid mixing up
which boiling tube is for which temperature.
9. Put each boiling tube with the beetroot cubes on the dissecting needle to their
respective water baths once the water baths are at a constant temperature.
10. Immediately start the timer for 7 minutes when you put the boiling tubes into
the water baths.
11. After 7 minutes, take the boiling tubes out of the water bath and let the tubes
stand on a test tube stand for 5 minutes.
12. After 5 minutes, pour the 4 cm3 of water from the boiling tube into a cuvette,
close it with the cuvette lid and place it into the colorimeter. (make sure the
colorimeter is set to green light absorbance). Repeat this for each
temperature.
13. Repeat the whole same experiment 3 more times to get more reliable results.
With the same controlled variables.
14. Record the results (percentage of green light absorbance) into a record table
and calculate the mean for each temperature.

Alex Bouman

Safety and risk assessment

Equipmen
t

Risk

Knife

Cutting yourself
or someone
else

Hot water
from Water
Bath or
boiling
tubes

Spillage that
will cause burns
on your skin

Beetroot

Stains on skin
or clothing.
Beetroot could
be an allergen
for some
people
Poking/stabbing
yourself or
others.

Sharp can
potentially cut
you

Dissecting
needle

Dice Cutter

Risk
assessmen
t (1-5)
4

Precaution

Make sure you pay full attention to the knife

whenever you hold it, when cutting never
loose eye contact with the knife. Avoid
walking with the knife in your hand, if you do
make sure you dont point the knife forward
while holding it and dont grip the knife too
firmly
Make sure you wear a lab coat and gloves in
case spillage does happen, the clothing will
protect your skin. Make sure you are cautious
while handling the boiling tubes with high
temperatures in them. Do not touch the
bottom of the boiling tubes if you need to
handle the boiling tubes, hold them from the
top of the boiling tube. Wash your hands with
cold water if spillage happens on your skin.
Make sure you wear a lab coat and gloves to
prevent the beetroot from staining on your
clothes and hands.

Make sure when you skew the cubes of cut

beetroot, you will poke the dissecting needle
downwards on the beetroot against the white
tile. When walking with the dissecting needle
make sure you dont face it forward as it
might stab somebody.
Keep hands and fingers away from the
blades when cutting the beetroot into
rectangles.

Variables
Independent variable: The temperatures of water the beetroot will be put in
(20,40,60 and 80 degrees Celsius)
Dependant variable: The intensity of pink/red colour that the beetroot leaks out
(green light absorbance % for each temperature the beetroot was put in).

Alex Bouman

Controlled
variable
Volume of distilled
water

Surface area of
beetroot cubes

Area of boiling
tube exposed to
the heated water

Volume of solution
in the cuvette

Time beetroot is
exposed to the
water in the water
bath

Number of
beetroot cubes per
dissecting needle

How its controlled

Why its controlled

Measure 20 cm3 of
distilled water using a
25 cm3 measuring
cylinder.
The square cutter will
make sure the width of
each cube is
constant, by using a
ruler and knife (if done
correctly) will ensure
the same 2mm
thickness of each
beetroot cube.
By using the same test
tube rack in the water
bath.

The volume of distilled water will determine

how saturated the betacyanin, water solution.
This will affect the green light absorbance the
colorimeter will measure.
The surface area of beetroot in contact with
the water will determine how much betacyanin
will move in or out the beetroot. This will alter
the colour intensity of the solution

The surface area in contact with the water in

the water bath will determine how fast and
how warm the boiling tube will get. If the
beetroot changes temperature at different
rates, it could affect how much betacyanin will
leave the cell into the distilled water therefore
effecting the colour intensity of the solution.
Every solution was
The volume in the cuvette will determine the
3
poured into a 4 cm
amount of betacyanin particles in the solution
cuvette, the cuvette
that gives off the red/pink colour. This is effect
was filled to the top.
the green light absorbance the colorimeter will
measure.
Set the timer for 7
This is effect how many betacyanin particles
minutes and start the
will leave the cell, making it stay longer or
timer once the beetroot shorter will affect how pink/red the solution will
in the boiling tube is
be.
put in to the water
bath, after 7 minutes
remove the boiling
tube from the water
bath
Skew 5 identical
This will determine how much betacyanin
beetroot cubes per
would be able to leave the cell, giving the
dissecting needle
red/pink colour of the solution.

Alex Bouman

Setting of
colorimeter

Make sure the setting

on the colorimeter is
set to green light
absorbance %.

To make the results consistent, green light is

chosen as green is on the other side of the
light spectrum from red.

Assessment of ethicality: No assessment was needed as the only biomass used

for the experiment was beetroot. Beetroot is part of the plant kingdom, not the animal
kingdom. Due to the conservation status of Beta Vulgaris, its a plentiful root that is
extensively grown for agriculture.
Table of results
Temperature (Celsius)

colorimeter (absorbance%)

mean absorbance (%)

23.5

0.04

0.05

0.04

0.04

40

0.04

0.04

0.07

0.05

52.5

1.44

0.97

1.65

1.35

68

1.27

1.53

1.56

1.45

77

1.55

1.08

1.32

1.32

Alex Bouman

= anomaly

Secondary evidence
As I could not find any secondary results with the same procedures and variables for
this experiment, my results may be unreliable. However, the experiment I have done
has 3 results per temperature making a mean result.

Conclusion
I have come to the conclusion with the evaluation of my results that the permeability
of the beetroot cell membrane will increase with an increase of temperature until the
point where the protein structures in the cell will be disrupted and the betacyanin will
leak out. From the temperature 40 Celsius to 52.5 Celsius the permeability increased
uniformly but from 52.5 Celsius to 77 Celsius the permeability stayed around the
same percentage of green light absorbance with a 0.1% range. This proves my
hypothesis right as the true values would lie around that range however, this leads
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Alex Bouman

me to my next point where there is some degree of inaccuracy due to the relatively
large range in comparison to my other results. Due to the inaccuracy I wont be able
to give the conclusion on which temperature the proteins of the beetroot cell
membranes structure would get damage and leak out betacyanin.

Evaluation
Accuracy: When pouring the 20cm3 of distilled water and 4cm3 of solution into the
cuvette make sure youre at eye level with the measuring equipment to avoid
parallax error. I also further increased my accuracy by taking my readings from the
bottom of the meniscus. When cutting the beetroot, a dice/square cutter was used
which was 1cm x 1cm, this made sure the width of the beetroot was uniform
throughout the experiment. When measuring the actual temperature of the water
bath, a glass thermometer was used to determine the actual temperature of the bath.
However, to improve accuracy a digital thermometer with 2 to 3 decimal place
accuracy would increase the accuracy. The water bath shouldve been covered
completely to stop the heat of the water bath from radiating into the environment
therefore affecting the temperature. The beetroot thickness was not consistent as we
used a knife to cut the beetroot, the beet root was cut at an angle making certain
parts of the beetroot cube thicker than the other, affecting surface area. Callipers
would be used to determine the width of each beetroot slice.
Precision: The appropriate measuring cylinder was used as a 25cm 3 measuring
cylinder to measure 20cm3. An appropriate ruler was used to measure the width of
the beet root as it displayed cm and mm, a sharp knife was used to cut the beetroot.
The colorimeter was appropriate as it displayed 2 decimal places. The same units
were used throughout the experiment, cm3 used throughout the experiment.
Reliability: The experiment was repeated 3 times per temperature however, the
experiment was done by a different group. One group would do one temperature 3
times and all the results from each group would be put into a single recording sheet.
The procedures were discussed between the groups to make sure close to identical
procedure were carried out to avoid in unreliability. Each dissecting needle with the
beetroot were blotted onto a paper towel to make sure no betacyanin solution was
on the surface of the beetroot. To improve each group should do all the temperatures
3 times and compare the results with the other group as the experiment I carried out,
I couldnt compare my results with my colleagues. Alternatively, I could carry out the
experiment and collect more results per temperature (7 results) and find the mean
and on top of that, to get secondary results from me colleagues. Error bars were
included in the graph to show the range of results per temperature, anomalies were
also identified in the table of results.
Validity: All the beetroot cubes were from the same root or the same plant and
species. All the distilled water in the boiling tubes were the same volume and all the
boiling tubes were put into the same depth of water in the water bath by using the
same type of test tube racks. All the colorimeters were set to green light absorbance
and the colorimeter was calibrated with distilled water before the experiment. The
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Alex Bouman

beetroots were all left in the water baths for 7 minutes. The same volume of solution
was inside the cuvette(4cm3)