Equipment
- Delivery tube
- Gas syringe
- Stopwatch
- 5 Conical flasks
- Clamp stand
- Blender
- Knife
- Tile
- 2 Measuring cylinder
- Funnel
- Filter paper
- 500ml beaker
- 2 carrots
- Hydrogen peroxide
- Distilled water
- Bung
Risk Assessment
- Take caution as harmful chemicals will be used (hydrogen peroxide)
- Safety clothing (goggles, lab coat, sensible shoes) should be worn to ensure the protection of skin
- Glass equipment to be handled with care to avoid glass breakage
- Long hair should be tied back
Diagram
Method
- Chop 2 carrots into small pieces and blend with 100ml of water in the blender.
- Filter the blended carrots with filtering paper into a beaker
- Whilst the carrot solution is filtering, prepare 5 test tubes and label them: 20%, 40%, 60%, 80%, 100%.
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To prepare these different hydrogen peroxide concentration solutions, use 2 different 20cm3 measuring cylinder, one for water, and one for the hydrogen peroxide to ensure accuracy.
The table below shows the amount of hydrogen peroxide and distilled water need to prepare different glucose concentrations.
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Add one sample of the 20cm3 to a conical flask
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Measure out 20cm3 of the carrot filtrate
- Set clamp stand up with the gas syringe clamped in and connected through a delivery tube to the conical flask
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Pour the 20cm3 of carrot filtrate into the conical flask containing the hydrogen peroxide solution and immediately put the gas syringe bung on the conical flask.
- At the same time start the stopwatch.
- Time for 1 minute. As soon as it pass 1 minute take the bung out
- Measure the volume of oxygen obtained from gas syringe
- Repeat with the other samples of hydrogen peroxide
- Repeat experiment 4 more times
The experiment will be repeated 5 times in order to get 5 sets of readings for the results. This is also to ensure that the results can be as accurate as possible and to also be able to obtain an average. This is also to reduce the errors caused during the experiment thus improving the overall successfulness of the experiment.
Raw data results table showing the volume of oxygen gas produced from the catalase reaction from varying the concentrations of hydrogen peroxide solution
The table above is a draft up of the raw data results table I will be using for my final readings from the experiment. It includes columns with headings, concentration of hydrogen peroxide by volume of oxygen gas produced, 5 trials, units and uncertainties, average and standard deviation.
Data collection and processing
Raw data table to show the volume of oxygen gas produced from the catalase reaction from varying the concentrations of hydrogen peroxide solution
Calculations
Observations
From observations, when the catalase enzyme (carrot filtrate) is added to the hydrogen peroxide solution, bubbles are immediately produced and started rising up the conical flask whilst the gas syringe slowly moved outwards. I also noticed as the concentration of hydrogen peroxide increases, the amount of bubbles produced during the reaction also increases. There was no colour change during the experiments.
Analysis
The table of result shown above shows the volume of oxygen gas produced from the reactions between the different concentrations of hydrogen peroxide and with the same amount/concentration of catalase enzyme. The table below it contains evidence of my calculations for the averages and the standard deviation.
From calculating the standard deviation, I found out that my results are not significantly different as the standard deviation number is very small ranging from 0.63 to 1.46 in my results. Therefore, there is no point drawing error bars on the graph as the numbers are so small, it will be very difficult to draw the error bars. With standard deviation being so low, this shows that my data is concentrated around the mean, meaning my experiment data results are stable and is reliable with accuracy being very high with little chances of errors.
Conclusion and Evaluation
The objective of this experiment was to determine how different concentrations of a substrate (hydrogen peroxide) affect the catalase enzyme activity, which is measured by the volume of oxygen gas produced in the reaction. In conclusion, the results I have obtained has supported my hypothesis, stating that the higher the substrate concentration, the higher the enzymatic activity until the enzyme is fully saturated in which the reaction will not go up any further. A trend can be shown from the figures of my results. At 100% concentration, there is on average 70cm3 of oxygen given off while at 20% concentration, there is only 16.2 cm3 given off. This reinforces that at higher concentrations of substrate, more gas is produced and given off while at lower concentrations there is less gas. So there is a relationship between the substrate concentration and the volume of gas given off
However, the amount of oxygen gas given off evidently decreases as it progresses. This is supported by the graph as the line of the graph becomes less steep and level off in the experiment. In the graph, we can clearly see the line steeply rising all the way through (and especially steep during 60% to 80%) till around 80% to 100% where the increase in value is very little as it starts to level off. We do not clearly see the line leveling off into a plateau, which is an error, caused to my planning of the experiment and is considered a limitation. Also, from the data results, the difference of oxygen gas produced during the experiment from 40% concentration to 60% concentration is 15.4cm3, whilst from 80% concentration to 100% concentration the volume of gas given off is only 6.6cm3 proving that the reaction is slowing down.
In my results, I did not encounter any anomalous results, also reinforcing how my standard deviation is very small meaning there was a high degree of accuracy in my experiment and my results are reliable.
The biological explanation to support my hypothesis and my results is that as the substrate concentration increases, the amount of substrate per unit volume increases. So it is more likely that an enzyme molecule will collide with a substrate molecule, as more successful collisions are more likely to occur. This means enzyme activity increase and the rate of reaction also increases directly in a proportional rate, thus producing more oxygen in the reaction as catalase enzymes convert hydrogen peroxide into water and oxygen. (H2O2 → H2O + O2). This is very important to the cells in living organisms as hydrogen peroxide is a harmful strong oxidizing agent which may disrupt the balance of cell chemistry. Therefore the catalase enzyme is needed in cells to quickly catalyse the hydrogen peroxide into less harmful substances (water and oxygen) to prevent the hydrogen peroxide building up in our cells.
However, the reason the reaction levels off as the concentration continue to increase is because the enzyme active sites are becoming saturated. This causes a large amount of substrate to wait for the active site to come free. At the saturation point, the reaction will not speed up no matter how much concentrated substrate is added, thus the rate of reaction will be reduced causing the reaction/graph to level off and reach a plateau (as shown on a model graph on the left).
Weaknesses and improvements
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Chelikani P, Fita I, Loewen PC (January 2004). "Diversity of structures and properties among catalases". Cell. Mol. Life Sci. 61 (2): 192–208.
Ward, William, and Alan Damon. "Chaper 3- Chemistry Of Life." Pearson Baccalaureate: Higher Level (plus Standard Level Options) : Biology Developed Specifically for the Ib Diploma. Harlow, [England: Pearson Education, 2007. 68-69. Print.
Ward, William, and Alan Damon. "Chaper 3- Chemistry Of Life." Pearson Baccalaureate: Higher Level (plus Standard Level Options) : Biology Developed Specifically for the Ib Diploma. Harlow, [England: Pearson Education, 2007. 68-69. Print.
Gaetani G, Ferraris A, Rolfo M, Mangerini R, Arena S, Kirkman H (1996). "Predominant role of catalase in the disposal of hydrogen peroxide within human erythrocytes". Blood 87 (4): 1595–9.
Lehninger, A.L.; Nelson, D.L.; Cox, M.M. (2005). Lehninger principles of biochemistry. New York: W.H. Freeman.