Diagram
Pilot Test
After taking the information above into account, I expect to see froth to appear of the surface of the solution if I use pure hydrogen peroxide. But I only want to measure the amount of oxygen given off. In order for me to measure the oxygen only I will need to dilute the hydrogen peroxide. As a result of this I should only see a very same amount of froth appearing on the surface. To measure the amount of froth given off I will simply place a 1g piece of liver into a test tube with the allocated amount of hydrogen peroxide and water. A rubber tube will take the froth into the measuring cylinder, which is placed in water upside down, and I will be able to measure the amount of froth given off in one minute.
Diagram
Results
As a result of this pilot experiment, I will use 20 ml of hydrogen peroxide and 10 ml of water when I test to see how the a increase in temperature effects the amount of gas given off in one minute, using liver. I decided not to use 20 ml of hydrogen peroxide and 15 parts water because the hydrogen peroxide is too dilute. Therefore when I come to do my experiment it will be hard to see a difference in the amount of gas given off as it would be so little as a result of the hydrogen peroxide being too diluted.
Method
I plan to measure the amount of gas given off by the reaction between the liver enzymes breaking down the substrate, in this case the hydrogen peroxide. Firstly I will set up the equipment as shown in Diagram 1 below. I then gathered the necessary measurements of 10 ml of water, 20 ml of hydrogen peroxide and the 1 gram piece of liver. I then heated the beaker that contained the water that subsequently heated the hydrogen peroxide up the certain temperature that I was about to test. When the temperature was precisely at the temperature I was aiming to test I placed the 1 gram piece of liver into the test tube that contained the hydrogen peroxide. As soon as the liver first made contact with the solution I started the stop – watch and placed the rubber bung over the test tube. The oxygen gas given off through the reaction made its way through the rubber tube and into the measuring cylinder, that was submersed in water. When 1 minute was reached I pulled the rubber tubing from underneath the measuring cylinder and took the reading. I carried out the same procedure each time I was testing and only changed one variable that was the temperature of the hydrogen peroxide.
Diagram 1
Whilst carrying out this experiment I will keep constant the following:
- The same mass of the liver, 1g
- The same volume of hydrogen peroxide (20ml) and the same amount of water (10ml)
- The same equipment set up
- The same time span for the reaction to take place, 1 minute.
- Use the same method for each experiment so there won’t be any major differences. Only altering the temperature.
If I follow the above my experiment will be fair.
Whilst carrying out the experiment I will only change one variable each time I carry out
the experiment. I will only alter the temperature for the reaction to take place. I will do this strategically using the temperatures:
- 25 C
- 30 C
- 35 C
- 40 C
- 45 C
- 50 C
In order for my experiment to go to plan I will be as accurate as possible so I will measure to the correct measuring size:
- Measure the 1-gram of liver on the scales.
- Take the measurements of hydrogen peroxide and water at eye level and on an even surface and with a 90-degree angle to the lab bench from the measuring cylinder.
- Take each reading three times to ensure that there isn’t and anomalous result. Three is a good number to use as you can see the if there is an anomaly whereas if I was to take each reading twice I wouldn’t be able to distinguish the anomaly. Also I am able to obtain an average.
To make sure I am carrying out a safe experiment I will make sure that I am wearing safety glasses, as the hydrogen peroxide is corrosive and an irritant. I will also carry out any other safety precautions e.g. hold hot beaker with tongs.
Prediction
Taking into account my research I believe that the amount of gas given off by the reaction will increase as I increase the temperature that the reaction takes place in. The amount of gas given off will start to decrease after 37.5C and any other temperature above that. This is because the enzyme is denaturing. Reason being that enzymes are proteins and their structure is three dimensionally. Increasing the temperature disturbs the intra – molecular bonds that hold the 3D shape. As a result of this the shape has been altered. Enzymes have an active site. This fits into the substrate molecular. If the active site is altered the substrate will no longer fit in and so the enzyme doesn’t work properly.
The rise of reaction rate is also to do with the increase in temperate and the kinetic theory. The higher the temperature, the faster the particles move. As a result the particles of hydrogen peroxide and liver will collide more often the higher the temperature is. The more they collide the higher the amount of oxygen gas is given off. This happens but only to an optimum of 37.5C. After this temperature the enzyme will start to denature. If you were to look at a graph, the curve leading up to the optimum point would be gradual but as it reaches the optimum point it would fall dramatically. The reason being that the active site is destroyed therefore no reaction can take place as there is only one specified active site per substrate.
Results
I decided to test the temperature of 37.5C as I expect this to give off the most amount of oxygen gas in one minute. This decision is supported in my prediction.
Graph of results : This graph shows the amount of gas (oxygen) given off by the reaction between the liver and the hydrogen peroxide
Conclusion
The most amount of oxygen gas given off was at the temperature of 37.5C (28ml). The least amount of oxygen gas given off was at the temperature of 50C (1.2ml). Therefore I conclude that enzymes work best at 37.5C.
From my results it appears that catalase works best at 37.5C, and starts to denature as the temperature increases from then onwards. I expected this, as the human body temperature is 37.5C. After taking into account my background knowledge and research I can explain my results as follows. Kinetic theory states that particles, which gain heat energy move more quickly as temperature increases. In my case the reacting particles are the substrate (hydrogen peroxide) and the liver. As the temperature increases the hydrogen peroxide gain more energy therefore they collide with the liver more so increasing the rate in which the product is formed. However, at a certain temperature this is no longer the case. This is because enzymes are proteins and proteins can denature at high temperatures. This is because proteins have a 3D shape that the substrate fits into. At high temperatures the active site on the enzyme is altered (due to the braking of bonds therefore changing structure) as a result stopping products from being formed. There is only one active site per enzyme.
From my graph you can see that there is a steady rise in the amount of gas given off between 20C (9.1 ml) and 37.5C (28 ml). From this point onwards the graphs shows a dramatic reduction in the amount of gas given off, 28ml at 37.5C to 1.2ml at 50C. This is because the enzyme is denaturing. This supports my prediction that there would be a steady curve up to the optimum temperature then a dramatic fall due to denaturalisation.
It’s exactly as I expected.
Evaluation
Overall I thought my experiment was successful. My results were as I expected and as I predicted. I also only recorded one slight anomaly; this shows that the accuracy precautions that I took were justified. The anomaly is ringed on the graph of results. From the line of best fit on my graph of results, it is clear that one of the points do not fit exactly. This is my anomalous result. Although it is a very slightly inaccuracy, they’re an indicator of possible errors in the investigation. These may have occurred due to measuring inaccuracies. Therefore my results could have been effected as the quantities of enzyme and substrate would have varied (In my plan I stated that I would only change one variable). Another possibility is that the temperature that I was testing was not precise. The temperature could have fallen due to the time that it took for me to place the liver into the solution. I didn’t retake this result as it was only a very slight anomaly and if I had rested it the difference would have been marginally minimal. My graph of results was as I expected to see; therefore this proves my predictions were correct. My results were reliable, as each reading was similar. I also made sure that I carried out the same method procedure each time that I tested and measured the volumes as accurately as possible. To make my results even more reliable I was able to calculate an average, as I took three readings. Overall I have sufficient evidence to support my conclusion as my predictions compliments it. I know the predictions are correct as I used primary and secondary sources of evidence to compile my predictions
Suggestions and Improvements:
If I were to carry out the same experiment again I would change the following:
- I would use a water bath to heat the hydrogen peroxide solution to ensure the temperature was precisely the temperature that I wanted to test.
- I would take longer to carry out the experiment and achieve more results, therefore improving the accuracy of my results.
- As well as changing the temperature at which the enzyme works I would also change the weight of the liver piece. I would investigate if the change in temperature has more of an effect than the amount of enzyme present.
- I would maybe take more results (5 tests) to give me even more reliable evidence and a more accurate average.
- I would see how temperature effects other enzymes such as pepsin or trypsin, which bring about the digestion of meat
- I would investigate the effect of enzymes in different conditions (changing pH levels).
There isn’t really another way of carrying out this experiment, except for using the suggestions and improvements that are above. What would be the most interesting extension to this work is to see what pH conditions the liver enzyme works best in.
Diagram
I would use different pH levels and measure have much oxygen gas is given off. I would use the same procedure for the method above, but I would use different pH levels instead of the hydrogen peroxide. I wouldn’t apply heat at first I would just see which pH level the liver enzyme works best in and maybe apply heat to that. I wonder if I would see the same effect?