Milk contains proteins which are used to make cheese. During cheese making, bacteria are added to the milk. The bacteria change the pH of the milk to acidic, causing the proteins to coagulate (clot) forming curds. The curds are then used to make the cheese. milk + bacteria --(pH more acidic)--> proteins in milk coagulate (curds) Estimating the protein concentration in milk is important when making cheese. You will need to: • make simple (proportional) dilutions of the proteins in the milk, M • carry out the biuret test on each concentration, to provide a measure of the concentration of proteins present in the milk • carry out the biuret test on milk with an unknown concentration of protein, U • estimate the concentration of milk protein in U. You are provided with the materials shown in Table 1.1. [Table 1.1] It is recommended that you wear suitable eye protection. If K comes into contact with your skin, wash it off immediately under cold water.

1. (a)You are required to make simple (proportional) dilutions of the proteins in the milk, M (1.0%). Reduce the concentration by 0.2% between each successive dilution. You will also make a 0.1% concentration. You will need to prepare 10cm³ of each concentration.
2. (a)(i)Table 1.2 shows how to make up two of the concentrations you will use, 1.0% and 0.1%. Decide which other concentrations of milk to prepare using simple (proportional) dilutions of M. Complete Table 1.2 to show how you will prepare the other concentrations. [Table 1.2][2]
3. (a)(ii)9. Compare the colour with the standard colours in Fig. 1.1. Record the colour of the mixture in (a)(ii). [Figure 1.1] (ii) Record your results in an appropriate table for the known concentrations of milk, using only the standard colours shown in Fig. 1.1.[3]
4. (a)(iii)10. Put 2 drops of U onto the spotting tile. 11. Repeat step 6 to step 9 with U. Record the colour in (a)(iii). (iii) State the colour for sample U.[1]
5. (a)(iv)Fig. 1.1 has only three colours. In (a)(ii) more than one of the known concentrations may match U. This means that an estimate of the concentration of milk protein in U may not be accurate. (iv) Using the result in (a)(iii), state the known concentrations of milk protein where the colour result for U is the same.[1]
6. (a)(v)A significant source of error in this procedure is the difficulty of matching the colour to Fig. 1.1. Complete Table 1.3 to suggest: • an improvement to this procedure of matching the colour (dependent variable) • one other significant source of error in this procedure • an improvement to reduce this error. [Table 1.3][3]
7. (b)A student suggested that another procedure to estimate the concentration of protein in milk could be to compare how much coagulation was produced in acidic conditions. The student added acid to known concentrations of milk protein and observed a range of coagulation. The concentration of milk protein in an unknown sample could then be estimated more accurately. You are provided with 1.0moldm⁻³ hydrochloric acid, labelled A. It is recommended that you wear suitable eye protection. If A comes into contact with your skin, wash it off immediately under cold water.
8. (b)(i)12. Put 10 cm³ of U into a test-tube. In step 13, you will be adding A to the milk in the test-tubes, do not stir or mix A with the milk. 13. Put 2cm³ of A into each of the test-tubes containing the known concentrations of milk protein (from step 4) and U. 14. Observe the changes in the milk in all of the test-tubes for up to 1 minute. 15. If coagulation is not visible after 1 minute, you may need to carefully tilt each test-tube to move the milk and then observe the coagulation. 16. Using the observations from step 14 and step 15 and the key in Fig. 1.2, record the results in (b)(i). [Figure 1.2] (i) Prepare the space below and record the results for the known concentrations of milk protein using Fig. 1.2.[2]
9. (b)(ii)Using the key in Fig. 1.2, record the result for U.[1]
10. (b)(iii)Complete Fig. 1.3 to show the position on the line of each of the percentage concentrations of milk decided in Table 1.2. Put the label U on Fig. 1.3 to show an estimate of the concentration of milk which provides a measure of the proteins in U, using the result in (b)(ii). [Figure 1.3][2]
11. (c)Milk proteins can also be coagulated using the enzyme rennet. A student investigated the effect of temperature on the activity of rennet, shown by the percentage coagulation of the milk. The results are shown in Table 1.4. [Table 1.4]
12. (c)(i)Plot a graph of the data in Table 1.4 on the grid in Fig. 1.4. Use a sharp pencil for drawing graphs. [Figure 1.4][4]
13. (c)(ii)Suggest explanations for the results between 35°C and 45°C.[2]
14. (c)(iii)If a student wanted to investigate the independent variable, pH, a number of experiments would need to be set up. State the pH values you would select and describe how you would change the pH. pH values description[1]

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