A-LevelBiologyEnzymes9700/52 · Feb/Mar 2018 · Q120 Marks
Biological washing liquids are solutions that contain enzymes to help remove stains caused by biological molecules. One of these enzymes is lipase. These washing liquids are concentrated solutions and need to be diluted with water to reach their 'working' concentrations. A group of students was asked to find the 'working' concentration of the enzyme lipase for two different biological washing liquids, A and B. The recommended dilutions for washing fabrics using these biological washing liquids are: A – 20 cm³ of washing liquid in 5 dm³ water B – 35 cm³ of washing liquid in 10 dm³ water. The students found a website that lists the concentration of lipase in different biological washing liquids before they are diluted. These concentrations vary between 85 mg cm⁻³ and 155 mg cm⁻³. The students simulated fat-stained fabric by cutting 20 mm × 20 mm pieces of cotton fabric and soaking them in cooking oil. The fabric pieces were then allowed to dry. The students diluted both washing liquids according to the recommended dilutions, added a piece of fat-stained cotton fabric to each and measured the activity of lipase using a pH probe and stop-clock. The students recorded the time taken for the pH to decrease from 7.9 to 6.5. The pH decreases because fatty acids are produced from the hydrolysis of fats by lipase. [Figure 1.1] shows the apparatus the students used. Table 1.1 shows the results of two trials using each of the diluted washing liquids at the recommended temperature of 30°C. This is lower than the optimum temperature of lipase, which is 60°C. [Table 1.1]
- (a)The students used the information from the website and the recommended dilutions to calculate the concentrations of lipase that could be present in 'working' concentrations of washing liquid. The students calculated the maximum possible concentration of lipase in washing liquid A when diluted, as shown below. The maximum possible concentration of lipase in washing liquid A before dilution is: 155 mg cm⁻³ Therefore, the maximum possible concentration of lipase in diluted washing liquid A is: 155 mg cm⁻³ × 20 cm³ –––––––––––––––––––– = 0.62 mg cm⁻³ 5000 cm³ Complete the calculation below to show the minimum possible concentration of lipase in diluted washing liquid B. The minimum possible concentration of lipase in washing liquid B before dilution is: ........................................................... mg cm⁻³ Therefore, the minimum possible concentration of lipase in diluted washing liquid B is: ............................ mg cm⁻³ × ........................ cm³ –––––––––––––––––––––––––––––––––– = ........................ mg cm⁻³ ........................................................... cm³[1]
- (b)To find the 'working' concentrations of lipase in diluted washing liquids A and B, the students decided to make a range of lipase concentrations from 700 µg cm⁻³ to 200 µg cm⁻³. They were provided with a stock solution of lipase with a concentration of 1000 µg cm⁻³. Describe how the students could dilute the stock solution to make solutions of lipase with a range of concentrations from 700 µg cm⁻³ to 200 µg cm⁻³. 100 cm³ of each solution is required.[3]
- (c)The students used the apparatus shown in Fig. 1.1 to investigate the effect of lipase concentration on the activity of lipase. They used the different concentrations of lipase that they had decided to prepare. Identify the independent and dependent variables in this investigation. independent variable .................................................................................................................... dependent variable ......................................................................................................................[2]
- (d)Describe a method the students could use to find the effect of the different concentrations of lipase on the activity of lipase, using the apparatus shown in Fig. 1.1. Your method should be set out in a logical order and be detailed enough for another person to follow. It should not repeat the details from (b) of how to dilute the stock lipase solution.[7]
- (e)(i)State how the rate of release of fatty acids from fats is calculated from the students' results.[1]
- (e)(ii)Complete Fig. 1.2 to show the effect of lipase concentration on the rate of release of fatty acids from fats, by drawing the expected shape of the curve and labelling the axes, including units. [Figure 1.2][3]
- (e)(iii)Describe how the students would use this graph to find the 'working' concentrations of lipase in diluted washing liquids A and B.[2]
- (f)The students' results suggested that diluted washing liquid A had a higher concentration of lipase than diluted washing liquid B. Suggest one reason why this conclusion may not be valid.[1]
This question uses a diagram from the original paper, which isn't shown here. Open the full paper from the past papers archive to see it.
Mark scheme
Marking points from the official mark scheme (Feb/Mar 2018). The mark or mark type is shown in brackets.
- [1] 85 (mg cm³)
- 85 (mgcm³)×35 (cm³)
- 10 000 (cm³)
- = 0.30 (mg cm³) ; A 0.298/0.297
- [3] ref. to a named method of dilution or a description in words or a diagram;
- ref. to using correct proportions of stock solution and water to make 100 cm³ ;
- 700 µg cm⁻³ and 200 µg cm⁻³ plus sensible choice of at least three other intermediate concentrations described ;
- e.g. (700) 575, 450, 325 (200) µg cm⁻³
- (700) 600, 500, 400 300 (200) µg cm⁻³ A all marks on a fully labelled table
- [2] independent: enzyme / lipase, concentration ;
- dependent: time (for pH) to reach 6.5;
- [7] any six from:
- 1 ref. to using same volume of each concentration of lipase (in boiling tube) ;
- 2 ref. to method of measuring volume ; e.g. graduated pipette / measuring cylinder / syringe / burette
- 3 add, acid / alkali, to give pH 7.9; R if buffer used
- 4 idea of incubating enzyme (so it reaches the test temperature) before adding cotton fabric;
- 5 ref. to suitable incubation temperature ; A a single temp range 20-60 °C
- 6 ref. to timing until pH reaches 6.5 ;
- 7 ref. to a suitable control; e.g. boiled enzyme / water with alkali
- 8 ref. to suitable method of keeping same incubation temperature for all concentrations; e.g. incubator, water-bath, temperature-controlled room I air conditioning
- 9 ref. to using same, size / area, of stained fabric; A 20 mm × 20 mm pieces
- 10 ref. to, agitation / mixing / stirring;
- 11 ref. to low / medium risk or hazard and suitable safety precaution; e.g. allergy / sensitivity, to enzymes and wearing, gloves / eye protection
- 12 ref. to minimum of 3 replicates / replicating the whole experiment at least 3 times and finding a mean / to identify (and remove) anomalies
- plus one required marking point:
- 13 ref. to immersing the cloth and starting a, stop-clock / timer, simultaneously; A starting, stop-clock / timer, immediately after immersing the cloth A ref. to pH meter instead of cloth since both will be immersed at the same time according to Fig. 1.1
- [1] 1 / time (for pH to fall to 6.5) ;
- [3] 1 axes correctly orientated with labels ; x-axis: concentration of lipase y-axis: rate of, release of fatty acids / digestion of fats
- 2 axes have units ; x-axis unit: µg cm⁻³ y-axis unit: s⁻¹ or min⁻¹ A arbitrary units or 1 / t(ime) on y-axis
- 3 line shows increase as lipase concentration increases ; A linear curve or plateau
- [2] idea of rates calculated from times for pH to reach 6.5 for washing liquids (from Table 1.1);
- use of intercepts / described ;
- [1] any one from:
- idea that the lipase in A has a higher activity than that of B ;
- idea that lipases are of, different types / different origin ;
- idea that washing liquids and test solution are different (from each other);
- e.g. washing liquids contain other chemicals to remove fat I ref. to optimum temperature
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