We will repeat the experiment three times in order to have reasonably accurate results, but the ideal number would be around a dozen repeats, unfortunately we cannot achieve this due to time and resource limitations.
The variable in this experiment will of course be the concentration of the copper sulphate solution. There are, however, a number of constants in this experiment that need to be regulated. These include: temperature of surroundings, volume of milk solution used, volume of trypsin used and volume of copper sulphate used. The temperature needs to be kept constant as enzymes function at different rates in different temperatures and a change in temperature could invalidate our results. The volumes of the components need to be kept constant as if they are not, there would be no validity to this investigation. Another factor that must be kept constant is the PH of the trypsin, as enzymes vary in strength at different PH level, if a change in the PH level of trypsin were to occur it would reduce the reliability of our results. I am not going to test the PH of the trypsin every time though due to lack of time and resources, I will just presume that the PH remains concert as that is the best I can do under the circumstances.
METHOD
- Use syringe to draw 3cm cubed of milk solution and insert that into a test tube, 2cm cubed of trypsin and 1cm cubed of copper sulphate at a given concentration must also be to hand.
- Draw a cross on a piece of plain white paper 5cm x 5cm.
- Insert the trypsin into the test tube where the milk solution is already present. As soon as this is done insert the copper sulphate solution or distilled water and start the stopwatch straight away.
- Hold the piece of paper with the black cross on it behind the opaque test tube. The solution in the test tube should start to become translucent, as soon as it is translucent enough to be able to vaguely see the black cross stop the stopwatch and record the results.
- Repeat this three times for each concentration of copper sulphate.
- Record results and work out average time and rate of reaction (1 / time)
RESULTS
CONCLUSION
These results prove my original prediction to be correct. They prove that as the concentration of the copper sulphate solution increases, the rate of reaction decreases.
At the copper sulphate concentration of 10^-1 the milk and trypsin coagulated because the copper sulphate was so concentrated that it almost completely denatured the trypsin, causing the trypsin to have no effect on the milk but for the milk to coagulate.
The reason for the rate of reaction decreasing as the concentration of copper sulphate solution increased is because heavy metal toxins such as copper sulphate denature enzymes such as trypsin, rendering them useless, hence, the higher the concentration of copper sulphate that was present in the mixture, the more inefficient the trypsin would become.
The trypsin digests the milk protein casein causing the milk to become translucent, hence, allowing us to be able to spot the black cross behind the test time as time goes on, as more milk protein will have been digested by the trypsin.
The distilled water results were the control results and this conclusively proves to us that copper sulphate solution does decrease the rate of reaction between trypsin and milk protein. This is because the rate of reaction with only distilled water being added to the solution of trypsin and milk is 1.1 x 10^-2 min^-1, whereas as soon as a weak concentration of copper sulphate is introduced that reaction rate decreases to 9.6 x 10^-3 min^-1, and this reaction rate carries on decreasing as the copper sulphate concentration increases, coherently proving that copper sulphate solution decreases the rate of reaction between milk protein and trypsin.
The rate of reaction is decreasing as the concentration copper sulphate solution is increasing, but at a decreasing rate. That is for a given and constant change in the concentration of copper sulphate, the difference between the rates of reactions increase at an accelerating rate. This is easily spotted on the graph as the curve greatly increases in steepness as the concentration of copper sulphate solution increases by a constant value which saw no such acceleration in difference between results at the lower concentrations.
A biological explanation for this could be that trypsin has a tolerance level to which it can withstand digestion from the copper sulphate solution to a certain extent, although never fully. As the concentration increases the trypsin cannot cope with the toxin any longer as it could be working at its maximum rate, so even a small increase in the concentration of copper sulphate solution would then greatly subdue the digesting ability of the trypsin.
EVALUATION
The results I have obtained are fairly accurate which is surprising because of the method used and the conditions we operated under. My results are in the pattern they should be according to my prediction and my background scientific knowledge, hence, at first sight it would appear my results are quite accurate; however, there were many things that could have affected the reliability of my results.
This could include the PH level of the trypsin as I did not have the time or resources to test the PH of the trypsin before every use, hence, there could have been a change in the PH of the trypsin, this would reduce the accuracy of my results but I have no way of finding out if the PH of the trypsin really did change.
There would be no evidence to suggest there is an anomalous result amongst my results, furthermore strengthening the accuracy/reliability of this experiment.
The experimental procedure was not ideal as it relied a lot on human reactions and perceptions, which can vary enormously, through these methods and the fact that there were three people in our group recording results, the results had the potential to be extremely inaccurate with plenty of anomalous results.
The factors that could have seriously affected our results was the fact that the people recording the results all have different opinions on when the solution was translucent and the cross was visible. Luckily our group discussed what the appropriate level of translucency was, hence, this problem did not affect us very badly, but it definitely did affect our results in some way, it is extremely unlikely it did not.
Another factor that would have affected results, but in a much less minor way is human reaction times, i.e. the time between when the copper sulphate is added to the milk and trypsin and when the stopwatch is started and the time when the solution is translucent and the recorder pressing stop on the stopwatch. These factors certainly did affect our results but as our results were sizeable times, a second or two out did not make much of a difference.
A way we could have improved these experimental procedures would be to have one person performing all the experiments, in order to reduce inaccurate timings as one person would have a set opinion on when the cross would be visible and would have the same reaction times etc… This would improve the accuracy of our results. Unfortunately we could not do this due to limitations of time and resources.
I believe that this experiment has been a reasonably fair test. This is because there have been no major ways in which the equality of this experiment could have been breached as this was quite a simple experiment to perform. Another reason I believe this experiment is a fair test is because the results all appear to be accurate and reliable, support my background scientific knowledge and there are no anomalous results. If there were anomalous results this would mean one of the experiments has been unfair to some degree, my experiment does not contain any anomalous results.
The evidence gathered through this experiment is definitely enough to support a firm conclusion, there are no anomalous results and the results concur with my prediction. There is a definite pattern linking concentration of CuSO4 and rate of reaction between milk and trypsin, it is strikingly obvious, even more so when the graph is analysed there is a definite correlation, therefore, these results do support the form conclusion that as the concentration of CuSO4 is increased the rate of reaction between trypsin and milk protein decreases.
A way that this investigation could be extended if time allowed could be that I could test another factor that affected the rate of reaction between trypsin and milk protein. I could vary the temperature of the trypsin and record the reaction rate between trypsin and milk protein when trypsin is at different temperatures. For example:
- I would have a similar set up to this experiment but obviously, without the presence of copper sulphate solution.
- I would mark a black cross on a plain piece of paper and hold it behind a test tube with 3cm cubed of milk in it.
- I would heat the trypsin in a boiling tube and when it is at the correct temperature (I would test the effects at every 5 degrees Celsius, i.e. I would test the trypsin with the milk at 20 degrees, 25 degrees etc...) I would add the trypsin to the milk and start the stopwatch.
- I would then wait until the milk had become translucent so that the black cross behind the test tube was visible and I would then stop the stopwatch
- I would perform three repeats for every given temperature. Then record my results and analyse them in order to draw a conclusion.
I feel that if this experiment were to take place then the conclusion drawn would be that as the temperature of trypsin was round the 20 degrees mark the reaction rate would be quite low and as the temperature of trypsin increases the reaction rate would increase. I would expect the reaction rate to hit a peak at around 30 – 40 degrees Celsius and then start to decrease and eventually the effects of trypsin would be non existent after the trypsin has reached 50 degrees Celsius as after this temperature, enzymes are denatured and no longer function as they would normally.
