Effects of Temperature on the action of Diastase on a Starch Suspension.

Hypothesis:
The practical being carried out is to observe the effects of temperature on starch break down using a synthesized version of salivary amylase, this being Diastase. The starch will be placed into the Diastase and water and then placed in baths of water of different temperatures. The test tube containing water will have little or no reaction at all. However, the test tube containing the Diastase will have several different times to which the starch will break down. Once the Diastase reaches between 30 – 40 degrees Celsius, the time taken for the starch to break down should decrease and be at it’s lowest to do so. However, in cold solutions the starch will take longer as it will in temperatures beyond 40 degrees. Once it reaches this point, the break down will either take a very long period of time or have no reaction at all as enzymes are denatured at a certain point.

Materials:

4 x test tubes

5mL Diastase
5mL Water
10mL 2% Starch Suspension.
Pipette
2 x Spotting tiles
Large Beaker filled with water of assigned temperature
Thermometer
Iodine
Marker

Method:

The test tubes were collected and labelled W, S, A and B.
To the tube labelled W, 5mL of water were added and to S, the Diastase. In tubes A and B, 5mL each of 2% Starch Suspension was added.
Water was heated to 40oC in a kettle and added to a large beaker.
The four test tubes were placed in the bath of water for 5 minutes to adapt.
Two clean spotting tiles were collected, and in each indentation a drop of iodine was added.
The contents of A and B were poured into tubes S and W, respectively. These tubes were then placed back into the water bath, which has been kept constant by adding several drops of warm water to it.
As soon as this took place the procedure was timed. The pipette was used to place drops from test tube S into the iodine filled indents on the spotting tiles.
The same step was repeated using the contents of test tube W, after cleaning the pipette of its former contents.
Steps 7 and 8 were repeated every 30 seconds until the solution in the iodine did not turn black, thus indicating that the starch has been completely broken down.
The time was taken for the Diastase to break down and recorded in order to share with the class.

Diagram:

Results Table:

The following results are ones that would be expected if both classes had successful outcomes. Neither classes obtained proper results, so the following will be used instead to demonstrate what should have taken place.

Discussion:

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Diagram:

Results Table:

Discussion:

Enzymes are specific. A model that helps illustrate this is the Lock and Key Model. The model demonstrates that only certain “keys” (substrates) will fit into the correct “lock (enzymes). The practical carried out illustrates that starch is broken down by Salivary Amylase, which in this case has been substituted with Diastase, a synthesized version of the original substance. Enzymes are affected by the concentration of the enzyme, the pH of the medium in which the reaction is taking place and the presence of certain ions or non-protein molecules. However, the main factor affecting the rate of reaction in this examination is temperature.

Temperature influences enzyme activity. Most chemical reaction rates heighten as the temperature increases. The optimum temperature of an enzyme is the temperature at which it works best. So, although reaction rate may increase, as does the temperature, once it reaches its optimum temperature the enzyme is denatured. That is, due to the fact that enzymes are proteins, they will change shape once they exceed 45-50 degrees Celsius. The optimum temperature of most enzymes in the human body is between 30 and 40 degrees Celsius.

The table above shows that at 10 degrees, the starch took 18 minutes to completely break down. Due to the fact that the temperature is low, the enzyme is far from its optimum temperature and will not work as fast or efficiently.

It can be noticed that as the temperature increases, the time taken for the starch to break down decreases. It decreases from 18 minutes, to nine at 20 degrees, then to 4.5 at 30 degrees and finally reaches its lowest time of two minutes once it reaches 40 and 50 degrees.

The table illustrates that the optimum temperature for Diastase is between 40 and 50 degrees Celsius. It takes the shortest amount of time to break down the starch, however, once it goes beyond this point, the reactions start to slow down again at 60 degrees when it takes 7 minutes to complete the reaction. This is due to the fact that the Diastase is changing its shape. Once the Diastase goes beyond 70 degrees where it takes 14.5 minutes to break down, no reactions take place.

Once enzymes go beyond the optimum temperature, the enzyme is denatured. This is the reason why at 80 and 90 degrees Celsius no reaction occurs. The Diastase had changed shape and the starch suspension (the key) did no longer combine with the Diastase (i.e. fit it wouldn’t fit into the lock)

Conclusion:

Once the Diastase reaches a specific temperature, it would be at its fastest reaction rate. The specific (optimum) temperature is between 40 and 50 degrees Celsius, as illustrated by the experiment. It was expected that it would denature and have little or no reaction once it goes higher or lower than this point. The practical demonstrated that below the optimum temperature there was reactions taking place but at a very slow rate, as it was when it went beyond it, and after it got too high, there was no reaction at all.

Evaluation:

There were many errors that took place in this practical. Both classes were unsuccessful in their attempts to finish the practical.

One possible reason for things going wrong is the fact that the temperature of the water was not kept constant. Due to the fact that most groups were preoccupied with whether or not the iodine was changing colour and their timing, most were not paying attention to the thermometer.

To correct this, we should have organised before doing the experiment to have one person do the drops, while the other watches the temperature and time.

Another reason could be the accuracy of the timing. There may have been mistakes during the proposed 30-second intervals. Also, some groups did not leave the four test tubes in the water bath for the recommended five minutes, which would have disrupted the solutions, being allowed no time to acclimatise to the new surroundings.

Once again, organisation would have improved this error. If there was accuracy and organisation within groups, timing may not have been a problem.