Investigate how temperature affectd the actions of pepsin on albumin.

Back ground knowledge

Enzymes are catalysts that speed up the chemical reactions. Pepsin is an enzyme which breaks down protein into amino acids. Pepsin works best in strongly acidic conditions. Pepsin also works best at the optimum temperature above this the enzyme becomes denatured and works very slowly. The enzymes become denatured because the high temperature causes the shape of the enzyme to change causing the enzymes not to work as efficiently. In this diagram below this is what an enzyme which looks normal and works like a lock an key mechanism, however the second diagram shows the enzyme when high temperatures have denatured the enzymes, thus destroying the lock and key mechanism. This obvious why enzymes stop working or don’t work efficiently at high temperatures.

Diagram of normal and denatured enzymes

Apparatus

• Goggles
• Two test tubes
• Water bath
• Thermometer
• Stop clock
• Pipette
• Spot chart
• Measuring cylinder
• Pepsin 3ml3
• Albumin 3ml3

Diagram

Variables

I am keeping the amount of albumin and pepsin the same in all experiments. The only thing I am changing in each experiment is the temperature.

Fair testing

To make my investigation Experiments a fair test I will have to measure the pepsin, albumin and water the same amount every time. PH a very acidic the same pH scale in each test.

Risk assessment

 

To ensure the safety of my eyes I wore safety goggles. If any chemical was spilt on my hands I would wash it off immediately because the chemicals are biological enzymes. I put my hair back so it wouldn’t get in the way.

Preliminary plan

1. To begin with I did a preliminary experiment to try out the procedure and make it as accurate as possible.
2. First of all I decided to investigate which temperatures are best to give me reliable and accurate results. To start with I used temperatures of 5°C between all of my investigation experiments. This worked out to be a too small margin between the temperatures. I decided to use a 10°C margin instead.
3. To start with the volume I used was 1ml³ of each substance. However I decided that this amount would be too small and the reaction would go too quickly and I wouldn’t be able to use the stop watch and eventually plot the results on a clear graph. Then I thought to myself, I would use 3ml³ of each substance as this would then make the reaction last longer for me to observe and record using a stop watch and see the reaction take place to make my own observations.
4. To start with I only did 2 experiments for each temperature however to make the investigation have a more accurate results I decided to do 3 experiments for each temperature.
5. To check if the albumin and pepsin had gone translucent I would check it ever so often on the spot chart on number 2. I think this would make the results slightly inaccurate because it could have been clear for longer than you realised.

Results of my preliminary experiment

Method

1. Obtain all the equipment listed under apparatus.
2. Heat/cool a water bath to 20°C
3. Measure out the pepsin and albumin separately and put the two test tubes in the water bath separately leaving time for them to reach the correct temperature.
4. Once reached pour the albumin into the pepsin and mix.
5. Start the stop clock
6. Observe and check ever so often in the spot chart to see if you can see the colour at the bottom.
7. If so stop the stop clock and record time.
8. Repeat two more times.
9. Do the same procedure for each temperature.

Results

Conclusion

In conclusion I think that the optimum temperature is 80°C, however I was slightly suspicious of this results. I found out that each time I was putting the albumin and pepsin in the water bath I realised that I hadn’t left them in the water long enough for them to reach the correct temperature. Since I have done all the experiments that way all the results will have the same fairness. If I were to do the investigation again I would consider measuring the temperature of each liquid, therefore overall the results would be more accurate. I have discovered then that hotter temperatures are best. This is because oh the collision theory. The hotter the temperature the faster the particles collide, therefore the experiment overall would be quicker. My prediction was quite accurate especially about the collision theory from my back ground knowledge.

Evaluation

 I have realised that my experiment could have been more accurate if I had taken more care over the actual temperature of the albumin and pepsin before the mixing of the chemicals. The results on my graph show this. (Graph below text). The next time I do the experiment I would like to have more results so I can compare to all my other results this would average out even more and try to eliminate any anomalous results. If I had time I would have done a lot more different temperatures to find the optimum temperature. I would have made the margin smaller such as in 5’s or 2’s. Also there will always be some human error in the results because of our reaction times. There is no real alternative except to record more results. Another factor which could help the experiments to be fairer is the temperature change. If I were to do the experiment gain I would place my test-tube in a water bath which would keep the liquids at a constant temperature giving me more accurate results.