Substances called catalysts can speed up many chemical reactions.

In my investigation I am going to study the enzyme amylase. This enzyme is used to breakdown the substrate starch to maltose. I am going to investigate the affect of temperature on the enzyme amylase.

Variables

Independent: The Temperature of the reaction. The range of temperatures I will use are

This should give a wide range of results to see the behaviour of the enzyme.

The dependent variable: The time taken for Iodine to stop turning blue black and remain orange.

Controlled Variable:

• Volume of amylase ( 2cm3)
• Volume of starch (2cm3 )
• Drop of Iodine
• Time of sample taken(2 mins)

Prediction and justification of prediction.

I predict that as the temperature increases, the speed of the reaction will increase. When a particular temperature is reached I believe the rate of reaction will dramatically decrease. I believe this because most chemical reaction happens faster when the temperature is higher. At higher temperatures molecules mover around faster, which makes it easier for them to react together. Usually, a rise of 10OC will double the rate of reaction. This is true for enzymes up to about 40OC. However at 40OC the enzyme begins to be damaged, so the reaction slows down. By 60OC the enzyme is completely denatured. This means that the active site has changed shape and the substrate molecules no longer fit in. I would also expect that the optimum temperature for this enzymes activity would be 37oC as it is a human enzyme, which would function best at body temperature

To investigate the effect of temperature on the activity of the enzyme amylase:

• Pour amylase solution into a test tube to a depth of 2cm.
• Half fill another test tube with a 4% starch solution.
• Cool both test tubes and maintain at 0OC using ice
• With a pipette place a drop of iodine into each dimple in a dimple tray.
  With a glass rod lift a drop of the starch solution from the test tube and mix it with the first drop of iodine in the first dimple in the tray. A blue/black colour should develop; this will be used as the control.
• Rinse the glass rod.
•  Add 2cm3 of water to starch solution (see below).
• Pour amylase solution into the test tube of starch and shake quickly (measure temperature, try to maintain at 0OC).
• Repeat steps 4 & 5 (for the amylase & starch solution mixture) every 40 seconds until a blue/black colour no longer develops.
• Record the results in a table.
• Repeat steps 1-9 increasing the temperature by 20OC then 10oC each time till the temperature is 60oC

Apparatus 

• Measuring cylinder
• Dropper
• White dimple tile
• Iodine
•  Starch
• Stirring rod
• Beakers
• Bunsen burner
• Ice

Strategy of results

I will record my results in a table

I will plot a line graph on the axis shown below:

Average time (S)

                                           

Temperature (oC)

I expect that as the temperature increases, the time will decrease, i.e. the temperature is inversely proportional to the average time.  

The enzyme will peak at its optimum temperature (this will be the shortest time) and after this both as temperature increases so will the time as the enzyme becomes denatured.

My graph will look similar to this:

Average time (S)

                                             Temperature (oC)

If I decide to draw a graph showing the rate of reaction as well it would look like this:

Rate of reaction (S-1)

        

                                             Temperature (oC)

                                           

I expect that as the temperature increases, the rate of reaction will also increase.  I expect the rate of reaction to double with every 10oC rise in temperature.  This is because the molecules, which are reacting, move faster and have more energy at higher temperatures.

To calculate the rate of reaction I will divide 1000 by the average time.

Obtaining evidence

The results that I obtained and recorded during the experiment, testing the relationship between temperature and the rate of the reaction of starch and the enzyme amylase, were as follows:

Interpretation

First of all the average time taken for the starch to be digested in each condition was calculated.  (see Table of results)

The results were then plotted on a graph (see Graphs 1 & 2) and the points joined together.
From graph 1, which is a graph of temperature against average time of reaction, I can see that as the temperature of the mixture increases, the time taken for the amylase to digest the starch decreases. This happens fastest at 40OC, however if the temperature continues to rise then the time taken for the amylase to digest the starch rapidly increases. By the time 60 OC is reached the amylase does not digest the starch. 

From graph 2, which is a graph of temperature against rate I can see that as the temperature of the mixture increases the rate of reaction is increasing until a temperature of 40o C after this the rate decreases showing a value of 0 for 60 o C
Amylase speeds up the breakdown of long chain starch molecules into smaller chains of maltose. Enzyme molecules have a very precise three-dimensional shape. This includes a ‘dent’, which is called the active site. It is exactly the right size and shape for the enzyme’s substrate to fit into (in the case of amylase this is starch). When a substrate molecule slots into the active site, the enzyme ‘tweaks’ the substrate molecule, pulling it out of shape and making it split into product molecules. 

        Starch

        Active Site

        Amylase

        Maltose Molecules

        Active Site

        Amylase

High temperatures make enzymes inactive: this is because they are proteins, which are damaged by temperatures above about 40OC. In this investigation it was found that amylase operated fastest at 40 OC and was damaged above this temperature. The enzyme is damaged because the molecules are moving faster, these bombard the active site of the enzyme changing its shape; when it’s shape has been changed then the starch will no longer fit in the active site and therefore cannot be digested.

        Denatured

My original prediction was:

The rate of the reaction between the enzyme, amylase and the substrate, starch, will increase as the temperature increases, up until 40oC.  I predict that the rate of the reaction will then begin to decrease as the temperature rises, above 40oC, and eventually stop, probably at about 60oC.

Therefore I predict that the amylase will work best at 40oC as this is its optimum temperature.

My conclusion agrees with my original hypothesis as the rate of reaction did rise up and peaked at 40, and did decrease rapidly from that point onwards until 60 oC where the rate of reaction stopped as the enzyme became denatured, just as I had predicted.

Evaluating Evidence 

The method used in this experiment is

In my method I had to use bunsen burners and ice to set up the different temperatures, this was very difficult and I think could lead to anomolous results. I would have liked  to control the temperature of the water bath, by the use of thermostatically controlled water baths.  This meant I would  not have to worry about the temperature  and this would increase the accuracy of my results.

In my method I used 40 -second time intervals when adding the amylase and starch solution to the iodine. If I wanted to obtain more accurate results all I would have to do is use smaller time intervals such as every 10 seconds.

I believe that some of the results might be off because the enzyme was not given enough time at each particular temperature to be fully affected before it was added to the starch. Because of time restraints they were only left in the water bath for 10 minutes before starting the experiment. However, they should have been left in the water bath for about 30 minutes so that the amylase would be completely affected by the temperature before the experiment was started.

I will now evaluate the measuring instruments that I used during the experiment and rate how good they were.

1. Measuring cylinder- I used a Measuring cylinder to measure out the volume of starch and amylase solution.  I felt that a burette would have been a better choice of apparatus as I know from Chemistry this is a more accurate piece of apparatus

2. Stop clock- I think that a stop clock was a very good choice as it was very accurate.   The stop clock was digital and therefore very accurate to 2 decimal places.Any problem or inaccuracy must have been down to “human error.”  I aimed to start the stop clock the exact moment the starch and amylase were mixed together, but I may have been a second or two off.

3. Dropper- I used the dropper to measure out 2 drops of iodine.  I found this instrument quite accurate but found myself accedently putting in 1 or 2 more drops than required.

I think my results are reliable. I believe this because if you look at my table of results the replication of results are similar but not identical. The only replication, which would cause great concern, was the result at 20oC

When I plot my results on the graph there did not appear to be any anomalous results all points fell on the line of best fit.

Are my results good enough to convince other people?

My answer to this question would be yes.  I feel that I have enough evidence to convince other people.  In the experiment I obtained five separate, accurate readings of how temperature affected rate and I feel that this is an acceptable amount to support my conclusion above.  This conclusion proved that as the temperature of the water increased, the rate also increased up until 40 oC and then decreased rapidly.  My curved graph shows this trend.