In my prediction I had also included the fact that the enzymes will begin to denature so the reaction times will increase and the rate of reaction will decrease. This was shown in my graphs as well. At 70˚C the average reaction time increased to 134 seconds and the rate of reaction decreased to 0.007 seconds. These results show the beginning of the denaturing of the enzymes.
Raising the temperature had two effects. When the particles had speeded up, there were more collisions per second and these collisions would possess the activation energy needed for the reaction to occur. The trends in my graph arose because the particles in the amylase and starch obtained more kinetic energy leading them to vibrate much faster which then increases the number of collisions. (These collisions can be explained best with and by the collision theory. This theory explains that no reaction can take place without particles colliding.)
The collisions are so energetic, that the hydrogen bonds that hold the proteins together to form the enzyme begin to break. The molecules of the enzyme begin to lose its shape and activity, and only at this time is the enzyme referred to as denatured. The reactions then start to slow down as the substrate doesn’t fit well into the active site of the enzyme, but at the end the substrate finds it impossible to fit into the active site and hence a reaction can no longer occur.
Each enzyme is shaped to fit a particular substrate. This is directly related to the lock and key theory. Below is a diagram depicting this theory:
Below is another diagram showing the denaturing of an enzyme that causes the lock and key theory to not operate. This diagram explains the trends in my graphs. It explains why the reaction time increases and the rate of reaction decreases after 60˚C.
The shape of my second graph as you can see is an upside down ‘U’ outline and the shape of my first graph is a ‘U’ shape. The explanation for the rate of reaction beginning to decrease and the average reaction time beginning to increase after 60˚C is that the enzymes have reached V-max. V-max takes place when the active sites of all the enzymes are used up meaning that there is no more substrate to be broken. After this no reaction can take place of obviously and so the rate decreases. V-max can be seen as the reason for the shapes of the graphs, but it is most likely that the enzymes had reached their optimum temperature and above, started to denature, and so stopped functioning and reacting. Enzymes that are denatured are also called biologically inactive.
If you look at the graph, it is clear to see that both have anomalous results. These incorrect results could have been the cause of anything. They could have been a result of careless measurements taken out by my teacher. For example she could have accidentally put in more amylase solution that could have speeded up the breakdown of starch. It is possible that the temperature she was testing (90˚C), was altered by cooling down in room temperature. This then could have decreased the temperature to the optimum temperature allowing a quick reaction to take place.
To see if the results of the rate of reaction graph are accurate, I performed a Q10 experiment. Q10 is a theory that states that if you increase the temperature you are working with by 10˚C, the rate of reaction should double.
I have included my working out below as to whether Q10 supports the experiment or not. I would also like to add that Q10 works till the optimum temperature, thus in this case till 60˚C. I have put the rate of reaction times to four decimal places to be more accurate.
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30˚C’s Rate of Reaction → 0.0042 = 2.1
20˚C’s Rate of Reaction 0.0020
2) 40˚C’s Rate of Reaction → 0.0094 = 2.2 (1dp)
30˚C’s Rate of Reaction 0.0042
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50˚C’s Rate of Reaction → 0.0161 = 1.7 (1dp)
40˚C’s Rate of Reaction 0.0094
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60˚C’s Rate of Reaction → 0.0211 = 1.3 (1dp)
50˚C’s Rate of Reaction 0.0161
As you can see from my analysis of Q10 went considerably well. The different rates of reactions were divides by one another to see if the answer would come to near about two. If this happened the Q10 theory would have proved itself and I right. The answers were only one or two decimal places off and thus it was acceptable. My last working out was incorrect but this may be due to a systematic error. Overall Q10 demonstrate that my prediction is results are reliable to a certain extent.
In conclusion, looking at results, it is clear to see that the experiment was carried out successfully. Leaving out the anomalous outcomes, my results proved that the higher the temperature, the shorter the reaction time and the higher the rate of reaction. It also supported me when I said that when the enzymes reach their optimum temperature, then they will become denature and slow down until they are biologically inactive. As said before I did not expect the optimum temperature for this particular investigation to be as high as 60˚C and thought it the maximum it would be is 40˚C. However, I can explain this by the type of amylase that had been used. Fungal amylase was utilized instead of the human amylase found in our saliva. The fungal amylase may act differently. Analysing the evidence, it is rational to state that temperature is a key variable in the reaction between starch and amylase. Temperature is an important factor that can be used to increase the rate of reaction considering the enzyme’s optimum temperature as seen in this investigation.
The apparatus that was used for the experiment were:
- Pipettes - Starch
- Measuring cylinder - ‘Fungal’ Amylase
- Thermometer - Water bath
- Iodine - Beaker
- Stopwatch - Test tubes
I think it is fair to say that the results are accurate but not as reliable as I had hoped for unfortunately. I know the results are correct as my prediction was proved correct and I analysed my results after with the Q10 theory. It is not evident however from my personal knowledge, that the results are reliable as there were many flaws.
My teacher tried to make reliability precautions for the investigation to be fair test by using pipettes, a measuring cylinder and thermometers. However despite these advantages, there are a lot of improvements I could make if I was to redo the experiment. Some of the improvements I could make are to use a syringe rather than measuring cylinder for more accuracy in measuring the concentrations of amylase and starch. I can also used Gilson’s pipettes rather than normal ones as this has a knob which can be tuned to a certain measurement I wanted in a precise degree of accuracy. These methods of improvement could have been initially carried out by the teacher but unfortunately wasn’t. If the amounts of the measurements had even the slightest flaw, it could have jeopardised and manipulated the real result.
I think it would have been wiser if my teacher had used an electric water bath as it would have given me more accurate reading of the temperatures she was experimenting with. Since she didn’t use an electric bath this could have caused a systematic error. I recall an error of contamination where she did not wash out the used equipments such as the measuring cylinder and the test tubes which could have slightly altered my results. For example, if she had not washed out the solution in the test tube of the previous experiment, she could have had unequal amounts on the starch and amylase she experimented with next, which would have changed the reaction state possibly.
Technological improvements could be made by my teacher with the experiment also. For example, she could use a colorimeter to test whether the break down of amylase had completely finished. The colorimeter is a scientific device which accurately specifies certain points in a reaction. It is also an instrument which compares the amount of light getting through a solution with the amount which can get through a sample of pure solvent. It would also be able to tell us exactly how much of the sun’s light was able to pass though. This could have helped us decide when the reaction had actually finished in case she had made any early or late incorrect timing in the results. As she was working alone, her timing could have been also wrong because she only had two hands to pour the enzyme and substrate in with each other and the iodine and also start the stopwatch to time it.
It is possible that she had also made a parallax error. This name is given to an error such as if she had not correctly read the needle on the measuring cylinder or pipette accurately as the naked eye isn’t precisely accurate and I might have read my readings from an angle instead of head on.
Any of these flaws I have mentioned may have been the cause of the anomalous result and if any of these improvements were made, they could have been the prevention of my anomalous results. There is an extent of accuracy in the investigation but not much accuracy involved due to careless and imprecise actions taken out.
There are a few limitations I realised after the experiment. If I were to do the experiment once again, I would carry out all the improvements I have mentioned. I would also do the experimentation at every 5˚C interval to have more results which may have given me further insight on my findings. This is also a good idea to carry out, as to be fairly honest, I have no idea what happens between the intervals of 0-20, 20-30, 30-40 etc. In redoing the experimentation, I would also carry on the test the results after 90˚C as I am unaware if anything could have changed to maybe prove my prediction wrong.
Further work, which I could possibly carry out to add to this investigation, is to carry out another experiment on the affect of temperature between another enzyme and substrate, like carbohydrase and carbohydrates. From this I can compare the results I get from that experimentation to the one with amylase and starch to configure whether there is any relationship with their trends and optimum temperatures. This could help enable the further reliability of my initial investigation.
Overall, I am proud to say that the investigation has followed out to be a successful experimentation. Besides the anomalous results, the rest of the results are quite close to each other fitting a trend which proves the experiment to be fairly reliable. I have productively managed to establish my prediction and provide evidence from scientific knowledge as well as my own.