Having the water to cold though will make the enzyme inactive. This does not mean that it is denatured just will not work or do anything. When temperature has got to a higher temperature the enzyme will become active again.
- Label the test tubes 1-6, and make a note how much enzyme is in each test tube.
- Prepare the solutions of Trypsin and water needed, and place them also within the water bath until they have reached 30˚C.
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Test Tube 1: 5ml of water 0ml of Trypsin
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Test Tube 2: 4ml of water 1ml of Trypsin
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Test Tube 3: 3ml of water 2ml of Trypsin
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Test Tube 4: 2ml of water 3ml of Trypsin
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Test Tube 5: 1ml of Water 2ml of Trypsin
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Test Tube 6: 0ml of Water 1ml of Trypsin
- When both the substrate and enzyme have reached 30˚C. Transfer the enzyme into the each of the 6 test tubes, using a pipette. As soon as the Trypsin has been added start the stopwatch.
- Time the amount of time it takes for the trypsin and water solution to turn colourless. Record the results in seconds.
- Repeat steps 1-4 for each of the test tubes.
Independent Variable
To control the independent variable we will be measuring out the amount of Trypsin and water needed, using a pipette. This way it’s very precise and accurate.
Dependant Variable
To find the transparency of the milk, we will be using a calorimeter; this will be precise and very accurate to the percentage of light able to travel through the solution.
Controlled Variable
The temperature will be fixed at 30˚C by using a water bath that will remain that temperature throughout.
We will always use 5ml of milk powder, this and the Trypsin solution will then add up to 10ml, so will show how accurate the results are. Also I will be using a pipette, which will create more precision.
The pH of the enzyme will remain the same as will be using a buffer to stop a change in the pH. I will combine the water with the buffer; this will be the buffer I’ll be using to concentrate the Trypsin solution.
Accuracy
The equipment will bring precise and accurate results as the calorimeter will be within 0.01% and my pipettes, used to transfer my enzyme and substrate into the test tubes, will be within 0.1ml. Also as the water bath will be at a controlled temperature there won’t be any chance of the temperature of the enzyme to decrease of increase, therefore creating a fair result.
Safety
- Wear goggles, in case the Trypsin or Casein solution goes in your eyes.
- Make sure the pipette tip doesn’t touch the desk as could contaminate it.
Ethics
No ethical concerns.
Results
Conclusion
The higher the concentration of the enzyme trypsin, the less time was taken for the protein casein to turn colourless. The lower the concentration of the enzyme trypsin, so the more time was taken for the casein to turn colourless.
This is because when the concentration of the enzyme is increased, there are more active sites available on the enzyme for the substrate (protein) to attach onto. This creates enzyme-substrate complexes; this is part of the lock-and-key theory, where the substrate (the protein casein) forms temporary bonds with the active site upon the enzyme (trypsin). The enzyme holds the substrate molecule in such a way that the reaction can happen more easily. Upon both the substrate and enzyme there are charged groups on the surface, when joined in an enzyme-substrate complex the charged groups are attracted to one another, this distorts the substrate and helps to break bonds within it causing it to break up into separate products. The products are what we can see, and show when the reaction is taking place, or finished.
If we kept increasing the concentration of the enzyme we should notice that there would become a point when the line will start to level off. This is when all the active sites on the enzyme are full, so increasing the concentration even more wouldn’t affect the rate of reaction.
Errors
There weren’t any random errors within my data. My results came out in a steady trend, which is what I was expecting. However a systematical error could be that I couldn’t put all the enzyme solution into each test tube at exactly the same time. This meant that some of the solutions had a few more seconds to react so is a little more unreliable.
Reliability
I don’t think my results were as reliable as they could have been. I feel I needed to repeat the experiment, for each concentration about 5-10 times instead of just the once. That way I can take an average of my results and see if my first set of results are reliable and near a true value. If they are, they should be showing around the same results with each experiment. If I were to do the experiment again I would repeat the experiment 5-10 times for each concentration. I would also make sure I have enough time to get all results, this is because after 40mins I still did not get a result for the 1:4 concentration and this may make a difference to my results. If I got an error that was completely different again then I would repeat that experiment straight away, as it would have been an error I had made.
Accuracy
I don’t think it is possible to say for definite whether my results were close enough to the true value. However I think that they were fairly accurate, as they appeared to go in the pattern, which I had predicted as it agrees with my hypothesis.
Validity
The systematical error has not made any significance to the results as they are still in a pattern, and the values are not to far out of range.
