1) Temperature
2) pH
3) Concentration of the enzyme (sucrase)
4) Concentration of the substrate (sucrose)
Enzymes has different environmental factors to deal with, the temperature and the pH could vary to give a different set of results.
In temperature, there is an optimum temperature and there is also a denaturing range where the enzymes just stop working. The pH can affect the enzyme because if it were too low or too high, it wouldn’t work at all. The pH disrupts the hydrogen bonds so it needs to perfect for the rate of reaction to be optimal.
The concentration of the enzyme and substrate needs to be taken into account. The concentration of the enzyme and the substrate will be the same throughout the experiment so that it would be a fair test.
In this experiment, the temperature will be the variable and the rest will be constant.
The temperature can be varied via a water bath, which can either increase or lower the temperature.
The pH will be kept constant by a buffer solution, which minimizes the pH changes.
The amount of substrate and enzyme in each of the experiments will be kept the same by using the same concentration and volume.
Temperature
Enzymes work better in higher temperature; this is due to the kinetic theory that when there is more energy (in this case heat) molecules will speed up. In this experiment the enzyme molecule will speed up and will make more frequent contact with the substrate molecule meaning greater reactivity. However enzymes don’t work very well when they get too hot, this is when it begins to denature. When it gets even hotter the enzymes get permanently destroyed. Heat destroys the hydrogen bonds in the enzymes so that the shape and the flexibility are changed. So the active site has been modified.
Before the temperature gets to the level that it is dangerous for the enzyme there is an optimum point of reaction. So the rate of reaction works best. The optimum temperature is normally about 40 °C.
My prediction about sucrase’s optimum point is at 37.5°C because that is the temperature in our body, and surcrase works at optimum levels in our body breaking down sucrose into energy for respiration.
Q10 is the rate of reaction increasing due to temperature. The rate of reaction normally doubles every 10 °C rise until it reaches the optimum rate. The equation to work this out is:
Q 10 ≈ 2 for most reactions
Method
Firstly, 3 test tubes will be filled with exactly the same amount of sucrase for instance 10ml using a measuring cylinder or a burette.
This is heated or cooled to the temperature, which it is needed at in an electronic water bath. It is in a electronic water bath because it is able to keep the temperature for some time. A thermometer would come in handy to check if the water bath is at the right temperature.
A Clinistix is put into the test tube; the colour should still be pink.
Pick up the stopwatch, and be ready to time as soon as you put 10 ml of sucrose into the test tube of sucrase.
Stop the stopwatch as soon as you see the Clinistix turn from pink to blue. The time recorded will show how quick the rate of reaction was to turn sucrose into glucose.
Repeat this test with the same amount of substrate and enzyme, with the same temperature two more times to show that the results are correct.
This test needs to be done at different temperature, it is suggested to do them at:
5 °C 15 °C 25 °C 35 °C 37°C 40 °C 45 °C 55 °C 65 °C
|Q10| |Optimum| |Denature|
The optimum temperature that has been predicted by myself has been added to prove at that temperature is where the rate of reaction is highest.
These temperatures are to see how quick the rate increase, where is the optimum and when does it denature.
The results from the experiment will be plotted down in a table showing the time taken in the 3 test tubes in the first set of readings of 5 °C, then 10°C etc.
With an average at the end of each
What should happen is that, when the temperature is at 37.5°C, which is the fastest rate of reaction (so sucrose is broken down into glucose very quickly) and any temperature after that will make that reaction very slow. Before the optimum rate, the rate of reaction should be increasing steadily. This is because of the kinetic theory, and it denatures because too much heat change’s their active site.
Equipment list
A Measuring cylinder- electronic water bath- Burette- Clinistix- test tubes- thermometer- stopwatch- pipette - buffer solution
Bibliography- Advanced biology, principles and applications - CJ Clegg with DG Mackean
-Biology for advanced level- Glenn and Susan Toole
Internet- www.advanced-biology.co.uk
To help in this experiment I was taught the basics in enzymes, as in what the optimum temperature was, and what factors could have affected enzymes working.
This was all taught in the fourth form.
