An experiment to show how protease enzymes are affected by temperature

Enzymes only work in specific conditions, and this experiment should also demonstrate that the correct temperature is required for the digestion of food in the body. This temperature is 37ºC (body temperature)

Enzymes must have the correct pH to work. They do react in acidic, alkaline and neutral conditions but enzymes will only work if the conditions are correct e.g. a protease enzyme in the stomach will only work in acidic conditions because the stomach contains a strong hydrochloric acid.

Enzyme molecules connect like pieces of a jigsaw puzzle to form these reactions. When the enzyme molecule and the reactant molecule fit together, a reaction can take place. (see diagram below)

Diagram:

List of apparatus:

* Two beakers

* 3 boiling tubes(washed after testing each temperature)

* 3 test tubes(washed after testing each temperature)

* Thermometer

* Stopwatch

* Kettle

* Measuring cylinder

* Safety goggles

Method:

The apparatus above where used, to keep the tests fair and as accurate as possible.

To start with as a safety precaution safety goggles where worn throughout the experiment. Then 2 beakers, 3 boiling tubes, 3 test tubes and a thermometer were set up as shown in the diagram on the previous page. Next using a measuring cylinder, each one of the 3 boiling tubes where filled with 30ml of marvel milk. The measuring cylinder was then washed out thoroughly and 5ml of enzyme was measured out and added to each of the 3 test tubes in the second beaker.

Then dependent on what temperature the experiment was taking place at, the kettle was boiled and the same amount of water was added to each beaker (1 beaker with the 3 boiling tubes of milk in and the other with 3 test tubes of enzyme)

These were then left to equilibrate until the temperature was the same for all tubes. The temperatures where checked using a thermometer. When the temperatures were the same, the 3 test tubes containing the enzymes where added at exactly the same time to each of the boiling tubes, so all contained a solution of both enzyme and milk. As soon as they had been added a stopwatch was started, to time how long the rate of each reaction (if there was one) took.

When each tube reached its end point, the time was recorded in seconds in a results table.

The same was done for all 5 temperatures ranging from 12ºC-70ºC, and the only thing, which varied, was the temperature. To achieve the different temperatures different amounts of boiling water from the kettle were used.

Results tables:

Time taken for milk to go clear in seconds

Temp in ºC

in S

2 in S

3 in S

Average in S

Rate of reaction

(1/time -1)

2

20

24

36

27

0.007 (3dp)

20

66

71

76

68

0.013

30

52

57

60

42

0.016

40

40

47

52

39

0.019

50

33

37

39

36

0.026

60

34

40

37

40

0.027

70*

-

-

-

-

-

*At 70ºC no reaction occurred so we couldn't record any results, for this temperature.

You work out the rate of reaction by dividing 1 by the average time (1/time-1) This converts your graph to make it the right way round. The example of this is shown in my second graph were the rate of reaction is recorded.

Conclusion:

The first step in this experiment was to prove that the milk actually does need enzymes to be digested in the body. The control experiment proved this and the water in all 6 cases had no reaction with the milk. This also tells us that the body is reliant on enzymes to do their job, or our food would not be digested and broken down into the vital things in which all out bodies need.

In contrast the unusual thing about this experiment is that the optimum temperature was slightly higher then that of my prediction. My results table and graph show that the optimum temperature was around 40-50ºC. This suggests that this enzyme works at slightly higher temperatures then that of a normal enzyme, because the actually temperature at which enzymes work best is our body temperature (37ºC). Even though these results where not exactly the same as you would have expected, there is still an obvious link between temperature and the reactions of the milk and the enzymes.

* At 12ºC the reaction was very slow and took the longest of any of the temperatures to react and the solutions did not turn as clear as the others. This shows that this temperature is not a very suitable temperature for enzymes to be digested at

* At 20ºC the reaction had speeded up

* At 30ºC, the reaction had speeded up dramatically, this is possibly because 30ºC is close to body temperature.

* At 40ºC, this was only the second quickest reaction and is in the range of the optimum temperature, but it was the only temperature apart from 50ºC to turn fully clear

* At 50ºC, the enzyme worked the quickest and also the this solution was the clearest

* At 60ºC, even though this was the second quickest reaction it had taken a lot longer then 50ºC proving that the temperature was getting to hot for the enzyme and it was not reacting as efficiently

* At 70ºC, we waited 10 minutes for a reaction without any obvious sign in the change of state. So we came to the conclusion at this temperature the enzyme had been denatured

In conclusion I believe this experiment has proved that enzymes are dependent on specific temperatures and conditions, and really enzymes are very vital things in the complex structure of the human body, no matter what their size or their job is.

My prediction was actually very accurate even though the optimum temperature was slightly higher then expected.

Evaluation:

Overall the results we obtained, even though they weren't perfect, where enough to support the fact that enzymes are affected by temperature, and unless the temperature needed for the enzyme is right they will become denatured. We found this out when the temperature was around 70ºC and the enzyme actually became denatured.

Improvements could have been made to the experiment. We could have had more time because towards the end of the experiment out time was limited and we had to rush to finish.

With a longer time span not only could we have done the experiments more extensively, but also more accurately. We could have changed the concentrations of the milk and the enzymes, to see if that made a difference to the results. From what I have found out from this experiment it is likely that the higher the concentration of the enzyme, the quicker the reaction will take place, and the higher the concentration of the milk slower the reaction would take place.

This experiment is also controversial. Different groups participating in the experiment stopped and recorded their results at different times. This doesn't really make the experiment fair because there was different levels of clearness used. We waited to record the times until each tube had gone completely clear. We could have done a control experiment to prove that the marvel milk needs protease enzymes to work. We should also have used one stopwatch between all groups so we all had an accurate timing.

We could have even tried using a different enzyme, to prove the theory that the enzyme we used works at slightly higher temperature then that of a normal enzyme.