Year 12 Diploma Biology

THE THERMAL DEATH POINT OF CYTOPLASM

Membrane biology (Syllabus 1.4)
Beetroot contains an intensely red, water-soluble pigment, Betacyanin, which is found in the cell
vacuoles. If the plasma membrane is damaged, the pigment may escape in sufficient quantities to be
detected in the water around the tissue. One effect of heat on cell membranes is to denature the
proteins of the membrane irreversibly. The proteins will no longer be able to control the flow out of the
red pigment, so the pigment will be found in the surrounding water.

EXPLORATION
GENERAL QUESTION, WHICH DRIVES THIS INVESTIGATION
At which temperature would you expect the proteins in the plasma membrane of beetroot cells to be
denatured by heat?
HYPOTHESIS
You should propose a hypothesis, based upon your knowledge of proteins, to suggest at which
temperature the proteins will be denatured.
A Write your hypothesis.
B Justify your hypothesis.
TESTING THE HYPOTHESIS
Carry out an investigation to determine the approximate temperature at which membrane protein is
denatured, using replicate (identical) cylinders from well-washed beetroot tissue. Much of the
information needed to design the investigation is given below:
1
Identical cylinders of tissue, cut with a cork borer to a size which fits easily into the available
test tubes which contain water, should be subjected to a fixed period (10 minutes?) in water at a
range of temperatures between 0c and boiling. Use the same number of equally-sized
cylinders in, perhaps, 6 different temperatures. (At this stage, the volume of water is not
important it is simply a liquid medium which surrounds the cylinders, so that they are
properly heated at the necessary temperature.)
2
After heating, the cylinders should be removed and placed into clean test tubes, containing the
same volume of distilled water in each tube. They should be left for a fixed period say 15
minutes - at room temperature. The pieces of beetroot should then be discarded. The remaining
water sample will now contain any pigment that has leaked out of the tissue, through the
denatured proteins.
3
The intensity of the colour of the solutions reflects how much pigment is present. This can be
measured accurately (ie. quantitatively) with a colorimeter, using a blue filter (complementary
to red). But, without the service of a colorimeter, a quantitative scale must be devised.

ANALYSIS
DATA PRESENTATION
C Make a table of the quantitative results obtained.
D Plot the results against temperature in a graph.
E List uncertainties of measurement as well as degrees of precision (+/-) of measuring apparatus
You should obtain a curve from which you can deduce the approximate temperature at which the
escape of betacyanin was accelerated. At the point where the proteins are denatured, a greater density
of the pigment should be apparent.

EVALUATION
CONCLUSIONS
F Comment upon the results and the graph which you obtained. What does the graph show? What
happened? What is the explanation? If the value for the denaturing of the membrane proteins is
different from the one proposed in your hypothesis, comment on the possible explanations by
analysing your experimental method, or reviewing the hypothesis.
DISCUSSION
G Review the investigation, commenting upon possibilities for improvement and follow-up.

Technician: each group probably requires:

Beetroot
Scalpel
Cork borer
Scissors
12 boiling (test) tubes
Forceps
Equipment for water baths, incl. ice & boiling water
Graduated, measuring cylinder
Distilled water
Cutting surface

John Osborne
November 2014