What effects will be observed by varying concentrations of yeast on the rate of reaction of the enzyme catalase when used in hydrogen peroxide solution.
Skelmersdale College
Biology Coursework
Experiment
What effects will be observed by varying concentrations of yeast on the rate of reaction of the enzyme catalase when used in hydrogen peroxide solution.
Introduction
A preliminary experiment was carried out to see what factors would affect the biological enzyme catalase.
Enzymes are protein molecules produced by cells to carry out specific functions. They are known as biological catalyst which are present in most living tissue. There are thousands of different enzymes each catalyzing just one specific reaction. Enzymes speed up the formation or break down of chemical bonds. For example catalase will only breakdown hydrogen peroxide.
A catalyst is a substance which speeds up the rate of a chemical reaction without being changed itself. A catalyst reduces the energy required for the reaction to occur. A biological catalyst, an enzyme, such as catalase accelerates the rate of reaction of the breakdown of hydrogen peroxide to water and oxygen.
Preliminary Investigation
Catalase breaks down Hydrogen peroxide into oxygen and water. Given the different materials being used in the preliminary investigation, I predict oxygen will be given off from the test tubes containing potatoes, as they are living organisms which will contain the enzyme catalase; therefore a reaction should be observed.
I also predict no reaction would be seen when the sand is introduced to the hydrogen peroxide because it is not a living organism, therefore no catalase will be present.
Apparatus: Test tubes, heat proof mat, Bunsen burner, tripod and gauze, potato, splint, mortar and pestle, sand, matches, spatula, conical flask.
Reagents: Manganese IV oxide, hydrogen peroxide, water.
Health & Safety: Lab coats and goggles to be worn at all times to protect clothing and eyes. Tie long hair back to protect from Bunsen burner and loose hairs contaminating other materials. Keep all work areas clean and tidy, washing off any spills with water. Ensure all chairs are under tables to prevent trips/accidents.
Method
Five test tubes each filled with 2cm of hydrogen peroxide 30% were placed into holders.
A spatula of manganese IV oxide was placed into the first test tube which fizzed immediately, smoked and turned black. A bung was placed over the test tube for a few minutes. A splint was lit and blown out, then the bung was taken out of the test tube and the unlit splint held inside it, the splint lit again whilst held in the test tube.
A spatula of sand was placed in the second test tube, and a bung was inserted in the test tube to seal it for a few minutes. There were no reactions to observe.
cm of potato was placed into the third test tube and a bung placed over the top of the test tube. Initially the potato sank to the bottom, but after a few minutes it bubbled and floated to the top of the test tube.
cm of potato was ground using a mortar and pestle and a spatula of sand added to it, this was then placed in the fourth test tube which was also bunged for a few minutes. The sand sank to the bottom of the test tube immediately but the potato foamed and floated to the top.
cm of potato was placed in a conical flask containing water and boiled using a Bunsen burner for ten minutes. The boiled potato was then placed in the fifth test tube, which again was bunged and left for a few minutes. No reaction was observed.
The preliminary experiment proved that manganese dioxide is an inorganic catalyst; when introduced to hydrogen peroxide it decomposed it rapidly into water and oxygen. We also discovered that there is no catalase present in sand as no reaction was seen when sand was introduced to the hydrogen peroxide. The potato proved ...
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cm of potato was placed in a conical flask containing water and boiled using a Bunsen burner for ten minutes. The boiled potato was then placed in the fifth test tube, which again was bunged and left for a few minutes. No reaction was observed.
The preliminary experiment proved that manganese dioxide is an inorganic catalyst; when introduced to hydrogen peroxide it decomposed it rapidly into water and oxygen. We also discovered that there is no catalase present in sand as no reaction was seen when sand was introduced to the hydrogen peroxide. The potato proved to contain catalase, as in test tube three, initially the potato sank to the bottom but after a few minutes it bubbled and floated to the top. Sand was used for a second time in test tube four, together with ground potato; the same conclusion can be reached about sand, no catalase present in the sand because this just sank to the bottom.
The potato that was boiled did not react when added to the hydrogen peroxide, this was because the catalase denatured when the potato was boiled. Boiling the potato will surpass the optimum temperature of the enzyme catalase causing it to change shape and become useless.
The ground potato revealed evidence that when ground, the rate of reaction was faster compared with the un-ground potato. As soon as the ground potato was introduced to the hydrogen peroxide a reaction was seen. Reason for this being, the grinding of the potato released/exposed the catalase, which reacted straight away to the hydrogen peroxide. The potato foamed and floated to the top of the test tube immediately.
Factors in this experiment varied. Temperature of the enzyme was altered in test tube five and the surface area of the potato was changed when it was ground using a mortar and pestle. There were also many different materials used including Manganese dioxide and sand. The factors where varied to enable the discovery of the effects on the enzyme catalase and how other materials compared with the catalytic reaction of catalase.
Catalase is a naturally occurring enzyme, specifically produced to catalyze hydrogen peroxide into water and oxygen.
2H2O2 Catalase H2O & O2
Factors which affect the processes of enzymes include temperature, ph balance, surface area and concentration.
Different enzymes have different optimum temperatures. The human body has an optimum temperature of 98.6 degrees Fahrenheit therefore enzymes that work in the body will also have an optimum temperature in this range. If body temperature is above or below 98.6 degrees Fahrenheit then enzymes in the body will become unstable which can be life threatening for the individual. Too low a temperature will slow or stop the enzymes catalyzing reactions and too high a temperature can de-nature the enzymes, which changes their composition, they become dysfunctional. Many "genetic diseases" are the result of faulty enzyme systems.1
Different enzymes function better at different pH's. Enzymes in your stomach may prefer an acidic environment with a low pH while enzymes elsewhere may not2. Surface area and concentration levels are also said to affect the process rates of enzymes.
An example can be given to explain an enzyme at work and that is the enzyme present in milk. Whilst the milk is refrigerated the enzyme is slowed down or stopped. If you leave the milk in a warm room the enzyme will become active because it is at its optimum temperature, results are seen when the milk has turned sour and curdled.
Many household products contain enzymes, such as detergents where the enzymes break down grease, blood, sweat etc. (Proteases attacks proteins and lipases attack fats3). Enzymes also assist in the brewing industry. The enzymes in the yeast breaks down sugars into alcohol (ethanol) and carbon dioxide gas.
The preliminary experiment confirms catalase is only found in living tissue. It has an optimum temperature which if exceeded will de-nature the enzyme rendering it useless and it is extremely fast when directly exposed to its substrate matter. The preliminary also informs us that catalase is not the only substance that can decompose hydrogen peroxide as the inorganic catalyst Manganese dioxide also does this. The gas given off the hydrogen peroxide proved to be oxygen, as the splint re-lit when put in the test tube.
I am now interested to see if a change in other factors will affect the enzyme, catalase. I believe if the quantity of potato was increased, the rate of reaction would be much faster because more catalase is present. My research informs me that liver and banana have high catalase levels, I would expect if these were used, an even greater rate of reaction would be seen. Scientifically this means because they have a high content of catalase, more collisions between the active site of this enzyme and the hydrogen peroxide molecules (substrate). If the concentration of the catalase was so high that all the hydrogen peroxide was decomposed then the reaction should cease.
I will investigate further using yeast as I know this contains catalase. Changing the concentration factor of the yeast, to see how different concentrations affect the rate of reaction of the enzyme catalase using hydrogen peroxide as the substrate material.
I will measure, observe and record the levels of oxygen given off from five different concentrations of yeast solution when mixed with hydrogen peroxide under the conditions described below. All other variables will remain the same, i.e. concentration of hydrogen peroxide, temperature conditions, ph conditions and surface area.
Prediction
I think higher concentrations of yeast will increase the rate of reaction dramatically. More collisions (lock & key theory) will occur, that is between the molecules of hydrogen peroxide (substrate), and active site of the catalase (enzyme). The rate of reaction will be quicker, releasing oxygen molecules from the hydrogen peroxide solution in a shorter time.
I chose the following apparatus for accuracy and ease of measurement. The 250 ml measuring cylinder was easy to read and results could be seen immediately as oxygen was given off from the hydrogen peroxide. A pipette allowed accuracy and ease of measurement. The conical flask was a practical way to contain the mixed yeast solutions and the hydrogen peroxide; the bung stopped any air escaping except through the tube and into the measuring cylinder.
Apparatus 250 ml cubic measuring cylinder, pipette, water trough, water bath, bung, thermometer, conical flask, 5 test tubes, test tube rack, stopwatch and labels.
Reagents Hydrogen peroxide, yeast solution and water.
Health & Safety Lab coats and goggles to be worn at all times to protect clothing and eyes. Hydrogen peroxide can be dangerous and goggles must be worn to protect the eyes at all times when using this chemical. Keep all work areas clean and tidy, washing off any spills with water. Ensure all chairs are under tables to prevent trips/accidents.
Method
0 ml Yeast solutions were placed in 5 test tubes. Solutions made using 1ml yeast 9ml water, 2ml yeast 8ml water, 3ml yeast 7ml water, 4ml yeast 6ml water, 5ml yeast 5ml water, all test tubes were labelled correspondingly and placed in the test tube holder.
The apparatus was set up as shown above. 20 ml Hydrogen peroxide was put in the conical flask together with the contents of test tube 1, results of oxygen released where measured and noted every 10 seconds, upto and including 60 seconds using the stopwatch. This process was carried out for the remainder of the different concentrations of yeast solutions, all measurements noted.
All other factors were kept constant, that is, temperature conditions, concentration of hydrogen peroxide (substrate), ph balance and surface areas.
This experiment was undertaken twice, to obtain more reliable results.
st Data Collection Table
0cm³ Yeast solution
Hydrogen peroxide
Oxygen given off
0 secs
20 secs
30 secs
40 secs
50 secs
60 secs
cm³ yeast/9cm³ water
20 cm³
20 cm³
26 cm³
30 cm³
34 cm³
37 cm³
40 cm³
2cm³ yeast/8cm³ water
20 cm³
20 cm³
28 cm³
38 cm³
40 cm³
44 cm³
50 cm³
3cm³ yeast/7cm³ water
20 cm³
20 cm³
34 cm³
39 cm³
60 cm³
68 cm³
76 cm³
4cm³ yeas/6cm³ water
20 cm³
39 cm³
58 cm³
76 cm³
98 cm³
10 cm³
28 cm³
5cm³yeast/5cm³ water
20 cm³
48 cm³
80 cm³
15 cm³
40 cm³
64 cm³
86 cm³
Results from 1st Data Collection Table
2nd Data Collection Table
0cm³ Yeast solution
Hydrogen peroxide
Oxygen given off
0 secs
20 secs
30 secs
40 secs
50 secs
60 secs
cm³ yeast/9cm³ water
20 cm³
20 cm³
26 cm³
30 cm³
34 cm³
38 cm³
40 cm³
2cm³ yeast/8cm³ water
20 cm³
21 cm³
29 cm³
38 cm³
42 cm³
48 cm³
51 cm³
3cm³ yeast/7cm³ water
20 cm³
21 cm³
39 cm³
48 cm³
60 cm³
68 cm³
76 cm³
4cm³ yeas/6cm³ water
20 cm³
40 cm³
60 cm³
78cm³
98 cm³
12 cm³
28 cm³
5cm³yeast/5cm³ water
20 cm³
49 cm³
81 cm³
16 cm³
40 cm³
64 cm³
86 cm³
Results from 2nd Data Collection Table.
Analysis
In my experiment I have been trying to establish if higher concentrations of yeast will effect the rate of reaction of the enzyme catalase.
My prediction proved to be accurate. The line graphs show the results observed. They show a clear indication that the higher concentrations of yeast solution caused more collisions to occur between the active site of the enzyme catalase and the substrate, hydrogen peroxide. More oxygen released at a quicker rate.
The results also indicate at the lower levels of yeast concentration, little difference is seen in oxygen production until the 20th second mark, then an increase is more evident.
Results of oxygen given off at the higher levels of concentration, that is 4 ml and above, show clear evidence that the rate of activity is greatly increased because of the higher levels of yeast present. Greater volumes of oxygen were given off in a shorter time.
Both sets of data prove undoubtedly that the greater the concentration of the yeast solution used, the steeper the line of ascent. Proving a greater rate of activity when concentration of the enzyme catalase is increased.
Evaluation
Although my prediction proved to be correct, I am not fully satisfied with the results of the experiment.
It appears there could be errors with the readings. I am not fully confident that the reading taken in the 1st data collection table for the 3cm³ solution are reliable. The rate of activity does not seem consistent. At the 30 seconds mark the oxygen given off only increased by 5cm³ compared with the 2cm³ solution which increased its oxygen given off by 10 cm³. Comparing the two sets of data tables for the 2cm³ and 3cm³ yeast solutions at the 30 second mark, I think there may be an element of human error as the readings differ significantly.
Although care was taken in all areas of the experiment to try and maintain a fair test keeping factors under control there may have been human errors, for example: maybe the bung was not put on the conical flask quick enough, allowing oxygen produced to escape into the room. Other human errors that may of occurred include, inaccuracy when measuring solutions and gas given off, soiled equipment contaminating solutions leading to inaccurate results and human reflexes may have been inconsistent when timing.
I would like to repeat this experiment a few times again to record further, taking the average results to give more reliability. I would try to minimize human errors with the use of more refined equipment (e.g. measuring in mm rather than cm) and ensure all measurements are precise.
In further investigations I am also going to use greater concentration values to see proof that at saturation point, the rate of reaction remains constant.
http://wsrv.clas.virginia.edu/~rjh9u/enzychar.html
2 http://library.thinkquest.org/28751/review/biochem/7.html
3 http://www.crocodile-clips.com/absorb/AC4/sample/LR1507.html
Sylvia Callaghan
Biology practical assignment 17 Dec 2004
