Question 1 (60 minutes]

You are required to investigate some reactions carried out by live yeast cells after they
have been incorporated into alginate 'beads'. First make up two sets of yeast-alginate
beads.
Proceed as follows:
Place 5 cm3 of well-stirred yeast suspension in a small beaker. Add to it an equal volume
of glucose solution and stir. Place 3 cm3 of this yeast-glucose mixture into a beaker or
tube. Add 3 cm3 of the alginate to the yeast-glucose mixture and stir very thoroughly
using a glass rod. In a further small beaker, place enough calcium chloride solution to
give a depth of about 3 cm. Use the glass rod, by holding it level with the rim of the
beaker as shown in Fig. 1.1, to drop some of the yeast-glucose-alginate mixture into the
calcium chloride solution.

Figure 1.1
A bead of about 5 mm in diameter should be produced which lies at the bottom of the
beaker. Stirring the yeast-glucose-alginate mixture continuously with the glass rod,
repeat this procedure to produce 15 beads. Remove and discard any which are obviously
different in size, distorted in shape, or which float. The beads can be picked up gently
using a pair of forceps.
Label this beaker yeast-glucose beads and put it to one side.
Using a container with fresh calcium chloride solution, follow the same procedure to
make a further set of 15 beads but use 5 cm3 of distilled water with the yeast suspension
instead of glucose solution. Label these yeast beads.
Bromothymol blue is an indicator which changes colour as follows:
pH 6 yellow
pH 7 green
pH 7.6 blue

Label three test-tubes A, B and C respectively. To A add 5 yeast-glucose beads, to B add

5 yeast beads. Do not add any beads to C. Place 2 cm3 of bromothymol blue solution in
each of the three test-tubes. Add a cork or a bung and put the tubes to one side for 15
minutes. During this time continue with the rest of the question.
(a) (i) After 15 minutes note the colours of the solutions in the three tubes.
Tube A ...
Tube B ...
Tube C ...

(ii)

Carefully pour off and discard the liquid in tubes A and B but retain and
observe the beads.
Note any changes which have taken place in the appearance of the beads.

(iii)

Account as fully as possible for the various colour changes which you have
noted in (a) (i) and (a) (ii).

One-third fill a test-tube with 1.0 mol dm-3 hydrogen peroxide solution with which you
have been supplied.
Hydrogen peroxide is corrosive. If any should come into contact with your skin wash
immediately under cold water.
Drop one of your yeast beads into the test-tube and observe it for the next minute or so
and repeat this procedure using a fresh bead and a test-tube one-third filled with distilled
water.
(b)

(i)

Record your observations on the behaviour and appearance of the bead.

Observations

(ii)

Explain your observations in (b) (i) as fully as possible.

Repeat your procedure in (b) (i) using hydrogen peroxide solution. This time, as soon as
the bead touches the bottom of the test-tube, start a stopwatch or stopclock. Stop the
timing as soon as the bead rises from the bottom of the test-tube.
(iii) Record this time interval in seconds: .
Repeat this measurement twice more using yeast beads and then take three
measurements using yeast-glucose beads. Use fresh hydrogen peroxide
solution each time and a new bead.
Before you start, prepare a table in the space below in which to record your
readings. Include your result in (b) (iii).

(iv)

What deductions can you make from your results about the effect of glucose
on the process which you have been measuring?

Plan and carry out a further procedure to find out if the reaction you have been observing
and measuring in (b) is due to the presence of an enzyme in the yeast cells.
(c) Record briefly your method, results and conclusions.

(d)

Sketch a graph of the results that you would expect from investigating the
effect of hydrogen peroxide on time that the yeast beads take to rise.

(e)

Suggest three ways in which the independent variable might be varied.

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