The enzyme urease catalyses the breakdown of urea into carbonate ions and ammonium ions as shown in Fig. 1.1. urease urea + water --[urease]--> carbonate ion + ammonium ion CO(NH₂)₂ 2H₂O CO₃²⁻ 2NH₄⁺ Fig. 1.1 A solution of urea does not conduct electricity, but a solution of the two ions does conduct electricity. An increase in ions increases conductivity, so the rate of reaction is proportional to the conductivity. A meter measures conductivity in microsiemens per centimetre (µScm⁻¹). The conductivity meter also records temperature. K_m shows the affinity of an enzyme for its substrate. The lower the K_m the greater the affinity. A student carried out an investigation to find the K_m of the enzyme urease at different temperatures. The student: • made 500 cm³ of 0.2 mol dm⁻³ solution of urea, molar mass 60.1 g mol⁻¹ • used serial dilution to make a total of five urea solutions from 0.2 moldm⁻³ stock solution • used 1 g per 10 cm³ urease solution • used a conductivity meter to measure the initial rate of reaction at each temperature. Fig. 1.2 shows the experimental set-up during the course of a reaction. [Figure 1.2]

Mark scheme

Marking points from the official mark scheme (Oct/Nov 2017). The mark or mark type is shown in brackets.

1. [2] weigh / measure (out) / take / put / add / use/ AW, 6 (.01) g urea ;
2. [2] add 500 cm³ (distilled / deionised) water (and stir until dissolved) ;
3. [2] idea of removing (a known and) same volume of urea solution (with second dilution taken from first dilution, etc.) at each stage of dilution ;
4. [2] idea of adding (a known and) same volume of (distilled) water at each stage of dilution ;
5. [2] independent temperature;
6. [2] dependent conductivity (of enzyme and substrate / ions / solution); A in conductivity units
7. [1] substituting, the active enzyme / urease, by an unreactive substance (at all temperatures) A e.g. boiled or denatured enzyme / water in place of enzyme / 0.0M enzyme / (solution of) urea without urease
8. [1] or substituting, urea / substrate, with water; I 'use (distilled) water' unqualified A e.g. water in place of substrate / 0.0M substrate / (solution of) urease without urea
9. [6] ref. to a suitable range of at least 5 temperatures ; A any 5 in the range 10-70 °C
10. [6] ref. to using suitable apparatus (to incubate enzyme and urea solutions at constant temperature(s)); e.g. (thermostatically controlled) water baths / incubators / thermostatically or temperature controlled room / (magnetic stirrer) hotplate A beaker plus hot water as water-bath I air conditioning
11. [6] ref. to using same volume of urease each time ; total volume must not exceed 30 cm³
12. [6] ref. to using same volume of (each) urea concentration ; total volume must not exceed 30 cm³
13. [6] ref. to using (same volume of) buffer to maintain a constant pH ; A use buffer to control pH
14. [6] ref. to incubating urease and urea concentrations separately;
15. [6] ref. to mixing urea and urease solutions (on the magnetic stirrer) and immersing (conductivity) probe ; I 'probe is used' unqualified
16. [6] ref. to taking reading (from meter) at same time (for each solution / temperature); A any stated time from 0s to 5 min A take reading immediately / AW A 'use meter to measure rate of reaction' if time context correct
17. [6] ref. to testing each of the concentrations (of urea) at each temperature ;
18. [6] ref. to a min. of 3 replicates / repeats and a mean / find anomalies ;
19. [6] ref. to suitable hazard and precaution; A low risk experiment I medium risk experiment R no risk / high risk A urease / enzyme is allergen or irritant and wear gloves / goggles A Ammonia (given off at high temperature) is irritant/mask or gloves
20. [4] one curve drawn correctly; does not need to go through origin A straight line up then level without curving
21. [4] Vmax shown;
22. [4] ½Vmax shown;
23. [4] Km shown; A Michaelis-Menten (constant)
24. [2] D;
25. [2] (temperature showing) lowest Km or shows, greater / greatest / higher / highest, affinity (of the, enzyme / urease) for its substrate / AW ;