Yeast cells contain enzymes that hydrolyse sucrose into reducing sugars, as shown in Fig. 1.1. enzymes in yeast cells sucrose → reducing sugars Fig. 1.1 You will investigate the activity of the enzymes in yeast cells. The yeast cells will be immobilised in sodium alginate beads. You are provided with the materials shown in Table 1.1. Table 1.1 labelled contents hazard volume/cm³ Y yeast cell suspension none 30 A sodium alginate solution harmful irritant 20 C calcium chloride solution harmful irritant 30 S sucrose solution none 100 B Benedict's solution harmful irritant 30 If any solution comes into contact with your skin, wash off immediately under cold water. It is recommended that you wear suitable eye protection. You will investigate the activity of yeast enzymes by using different numbers of beads of immobilised yeast cells in sucrose solution, S. Carry out step 1 to step 9 to immobilise the yeast cells in sodium alginate beads. step 1 Put 10 cm³ of A into a beaker. step 2 Stir the yeast cell suspension Y with a glass rod. step 3 Put 10 cm³ of Y into the beaker containing A and mix well. step 4 Put 20 cm³ of C into another beaker. step 5 Use a 10cm³ syringe to collect 10cm³ of the mixture of A and Y. step 6 Hold this syringe over the beaker containing 20 cm³ of C (step 4), as shown in Fig. 1.2. slowly press down to release one drop at a time -plunger -barrel of syringe mixture of A and Y. calcium chloride solution, C Fig. 1.2 one drop released step 7 Hold the barrel of the syringe with one hand while slowly pressing down on the plunger with the other hand so that a drop of the mixture is released into solution C. The drop will form a bead. step 8 Repeat step 7 to make at least 31 beads. The immobilised yeast beads must be left in the beaker for 5 minutes. step 9 After 5 minutes tip the beads and the solution into a Petri dish. You will test the activity of the yeast enzymes by using different numbers of beads (1, 2, 4, 8 and 16) in sucrose solution. Carry out step 10 to step 20. step 10 Label five beakers 1, 2, 4, 8 and 16 and label five test-tubes 1, 2, 4, 8 and 16. step 11 Put 1, 2, 4, 8 or 16 beads into each of the appropriately labelled beakers, as shown in Fig. 1.3. 1 2 4 8 16 yeast bead 00 0 000 Fig. 1.3 step 12 Put 10cm³ of sucrose solution, S, into each of the beakers containing the beads. step 13 Start timing and leave for 5 minutes. While you are waiting set up a water-bath ready for step 14 and step 19. In step 19 you will use the water-bath to carry out the test for reducing sugars using Benedict's solution, B.

1. (a)(i)State the temperature you will use for the water-bath. temperature ........................................................................................ [1][1]
2. (a)(ii)Record your results in an appropriate table. [5][5]
3. (a)(iii)State the independent variable in this investigation. ................................................................................................................................................................................................................................................................ [1][1]
4. (a)(iv)State one main source of error in this investigation. ................................................................................................................................................................................................................................................................ ................................................................................................................................................................................................................................................................ [1][1]
5. (a)(v)A student set up a beaker as a control experiment. The result of the control experiment showed that the sucrose was hydrolysed by an enzyme. Suggest what substances the student put in the beaker for the control experiment. ................................................................................................................................................................................................................................................................ [1][1]
6. (a)(vi)The procedure described in step 1 to step 20 investigated the effect of changing the number of yeast beads on the rate of hydrolysis of sucrose. Describe how you would modify the procedure to investigate the effect of changing the concentration of the sucrose solution on the rate of hydrolysis of sucrose. ................................................................................................................................................................................................................................................................ ................................................................................................................................................................................................................................................................ ................................................................................................................................................................................................................................................................ [2][2]
7. (b)Pectinases are enzymes that are used to increase the volume of fruit juice extracted from apples. Pectinases can be immobilised and placed inside a container with apple pulp as shown in Fig. 1.4. The fruit juice is then collected. fruit juice -apple pulp -immobilised pectinase Fig. 1.4 A scientist carried out an investigation to determine the effect of temperature on the activity of immobilised pectinase by measuring the volume of fruit juice extracted. All other variables were kept constant. The results are shown in Table 1.2.
8. (b)(i)Plot a graph of the data in Table 1.2 on the grid in Fig. 1.5. Use a sharp pencil. [Figure 1.5] [4][4]
9. (b)(ii)Use your graph in Fig. 1.5 to estimate the volume of fruit juice extracted at 55°C. Show on your graph how you obtained your answer. volume of fruit juice = ............................................................... cm³ [2][2]
10. (b)(iii)Describe the change in volume of fruit juice shown by your graph in Fig. 1.5. ................................................................................................................................................................................................................................................................ ................................................................................................................................................................................................................................................................ [2][2]
11. (b)(iv)The scientist repeated the investigation replacing the immobilised pectinase with the same amount of pectinase but not immobilised in beads (free pectinase). The results are shown in Table 1.3. Table 1.3 free pectinase (not immobilised) temperature/°C volume of fruit juice /cm³ 30 52 40 85 50 98 60 41 70 12 80 4 Suggest an explanation for why more fruit juice was obtained when using the free pectinase between the temperatures of 30°C and 50°C than when using immobilised pectinase. ................................................................................................................................................................................................................................................................ [1][1]
12. (b)(v)Suggest an explanation for why more fruit juice was obtained when using immobilised pectinase between the temperatures of 60°C and 70°C than when using the free pectinase. ................................................................................................................................................................................................................................................................ ................................................................................................................................................................................................................................................................ [2][2]

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