Yeast cells contain the enzyme catalase which catalyses the breakdown of hydrogen peroxide, releasing oxygen. You will investigate the effect of pH on the activity of catalase in an extract from yeast cells. You will need to immobilise the yeast cells in sodium alginate beads. When a bead containing yeast cells is dropped into hydrogen peroxide solution the bead will sink. As oxygen is released the bead will rise. The more oxygen released, the faster the bead will rise. You are provided with the materials shown in Table 1.1. Table 1.1 labelled | contents | hazard | volume/cm³ ---|---|---|--- Y | yeast cell suspension | none | 15 H | 3.0% hydrogen peroxide solution | harmful irritant | 30 S | sodium alginate solution | none | 30 C | calcium chloride solution | none | 30 B3 | buffer pH 3 | none | 10 B4 | buffer pH 4 | none | 10 B6 | buffer pH 6 | none | 10 B7 | buffer pH 7 | none | 10 B8 | buffer pH 8 | none | 10 If any solution comes into contact with your skin, wash off immediately under cold water. It is recommended that you wear suitable eye protection. Carry out step 1 to step 19. step 1 Put 10cm³ of C into a large test-tube. step 2 Put 5 cm³ of S into a small beaker. step 3 Stir Y and put 3 cm³ of Y into the beaker used in step 2. Mix well. step 4 Use a 5 cm³ syringe to collect 2cm³ of the mixture of S and Y (prepared in step 3). step 5 Position the 5 cm³ syringe over the large test-tube containing C as shown in [Figure 1.1]. gently press down with thumb to release one drop at a time 5 cm³ syringe barrel resting on the top of a large test-tube 2 cm³ of a mixture of S and Y 10 cm³ of C large test-tube [Figure 1.1] step 6 Gently press down on the plunger of the 5cm³ syringe with your thumb to release one drop into solution C. The drop should form a bead. step 7 Repeat step 6 until you have used all 2cm³ of the mixture. Leave the beads in the solution C for 1 minute. step 8 Tip the contents of the large test-tube from step 7 into a Petri dish. step 9 Put two beads into each of the beakers containing pH buffers B3, B4, B6, B7 and B8. step 10 Label a small test-tube B3. step 11 Put 3 cm³ of the pH buffer B3 into the test-tube labelled B3. step 12 Put 3cm³ of hydrogen peroxide solution, H, into this test-tube and shake to mix. Leave this test-tube in a test-tube rack. step 13 Pick up a bead from the pH buffer B3 using blunt forceps. step 14 Drop the bead into the test-tube from step 12. Start timing when the bead reaches the bottom of the test-tube. step 15 Time how long it takes for the bead to reach the surface of the liquid. If the bead does not reach the surface after 60 seconds, stop timing and record as 'more than 60'. step 16 Record the result from step 15 in (a)(i). step 17 Pick up the second bead from the pH buffer B3 using blunt forceps. step 18 Repeat step 14 to step 16. step 19 Repeat step 10 to step 18 with the remaining pH buffers instead of B3.