Catalase Activity Lab

Abstract

The catalase activity lab performed was intended to demonstrate the effects of variations

in substrates, temperature, enzyme concentration, and pH on the rate and effectiveness of the

enzyme catalase. Enzyme activity was tested in 4 separate experiments using catalase to break

down hydrogen peroxide into oxygen and water. For experiment 1 hydrogen peroxide, water, and

sucrose solution were mixed separately with catalase in 3 separate test tubes. Experiment 2 saw

the effects of temperature change on the rate of catalase activity by placing a test tube in an ice

bath, a desk in room temperature, and an incubator. In experiment 3, the effects of changes in

enzyme concentration in a catalase + hydrogen peroxide reaction were tested and recorded by

changing the amount of catalase added to 3 different test tubes. In experiment 4, acidic, neutral,

and basic solutions were mixed in 3 separate test tubes with catalase to study the effects of

changes in pH on enzyme activity. It was found that the test tubes with the tallest bubble columns

were those that were (experiment 1) mixed with hydrogen peroxide and catalase (18.6 mm),

(experiment 2) mixed with hydrogen peroxide and catalase then placed in an incubator (40.2

mm), (experiment 3) mixed with hydrogen peroxide and more catalase than used previously

(68.8 mm), and (experiment 4) which was mixed with an acidic solution (20.4 mm). Catalase

plays the role of breaking down hydrogen peroxide protecting our cells from its damaging

effects.
Results

Catalase activity was determined in experiment 1 (figure 1) by performing three separate

trials, represented by test tubes 1-3. Averages and standard deviations were calculated for each

trial using Excel. For test tube 1, where 1 mL catalase was mixed with 4 mL hydrogen peroxide,

the average height of the oxygen bubbles produced was determined to be 18.6 mm, with a

standard deviation of 7.9 mm. There was no measured activity in the second test tube, where 1

mL catalase was mixed with 4 mL water. Test tube 3 had an average bubble column height of 2.8

mm with a standard deviation of 2.7 mm, where 1 mL catalase was mixed with 4 mL sucrose.

Figure 1.Catalase activity was investigated using three different substrates (1 = Hydrogen
peroxide, 2 = water, 3 = sucrose). The averages (blue circles) and standard error (black bars)
were calculated for each trial. Catalase activity was determined by measuring the height of the
bubble column in mm, 1 minute after the enzyme and substrates were mixed for 20 seconds.
 The effects of temperature on enzyme activity were studied in experiment 2 (figure 2) by

performing three separate trials, represented by test tube 1-3. Averages and standard deviations

were calculated for each trial using Excel. The average height of the bubble column measured

after 1 mL of catalase was left to cool in a 3 degree celsius ice bath for 15 minutes and then

mixed with 4 mL of hydrogen peroxide, was determined to to be an average of 13.8 mm with a

standard deviation of 4.7 mm. For trial 2 where 1 mL of catalase was left to sit at 22 degrees

celsius for 15 minutes then mixed with 4 mL of hydrogen peroxide, the average height of the

bubble column was measured to be 23.8 mm with a standard deviation of 10.7 mm. For trial 3,

the average height of the bubble column measured after 1 mL of catalase was left in a test tube

and placed in a 50 degree celsius incubator for 15 minutes, then mixed with hydrogen peroxide

was 40.2 mm with a standard deviation of 13.4 mm.

Figure 2. The effects of temperature were studied by observing and measuring a catalase +
hydrogen peroxide reaction under different temperatures (1 = Ice water bath (3°C), 2 = Room
temperat (22°C), 3 = Incubator (50°C)). The averages (blue circles) and standard error (black
bars) were calculated for each trial. For each trial, the height of the bubble column was measured
(mm) and recorded after 1 mL of catalase was left to sit for 15 minutes in a different
environment. It was then mixed with hydrogen peroxide, swirled for 20 seconds, and left to rest
for another 1 minute.

The effects of variations in enzyme concentration we observed and recorded in

experiment 3 (figure 3) by performing three separate trials, represented by test tube 1-3.

Averages and standard deviations were calculated for each trial using Excel. The height of the

bubble column measured for trial 1 after 1 mL of catalase was mixed with 4mL of hydrogen

peroxide was determined to to be an average of 19.7 mm with a standard deviation of 5.8 mm.

For trial 2, a bubble column with an average height of 43.0 mm and standard deviation of 8.7

mm was measured after 2 mL of catalase was mixed with 4mL of hydrogen peroxide. In trial 3,

where 3 mL of catalase was mixed with 4mL of hydrogen peroxide, the average height of the

bubble column was measured to be 68.8 mm with a standard deviation of 15.2 mm.
Figure 3, The effects of changes in enzyme concentration were examined using three different
concentrations of Catalase (1 = 1 mL , 2 =2 mL, 3 = 3 mL). The averages (blue circles) and
standard error (black bars) were calculated for each trial. The effects of enzyme concentration on
catalase activity were determined by measuring the height of the bubble column after different
catalase concentrations were mixed with hydrogen peroxide for 20 seconds and left to rest for 1
minute.

The effects of changes in pH on catalase activity were determined in experiment 4 (figure

4) by performing three separate trials, represented by test tube 1-3. Averages and standard

deviations were calculated for each trial using Excel. In trial 1 when 1mL of catalase was mixed

with 2 mL of pH 4 solution and 4 mL of hydrogen peroxide, an average bubble column height of

20.4 mm with a standard deviation of 7.8 mm was measured. For trial 2, when 1 mL of catalase

was mixed with 2mL of water (〜pH 7) and 4 mL of hydrogen peroxide, an average of 20.0mm

with a standard deviation of 6.1 mm was found. An average of 26.8 mm with a standard

deviation of 11.3 mm was found for trial 3 when 1 mL of catalase was mixed with 2mL of pH 10

solution and 4mL of hydrogen peroxide.

Figure 4, The effects of changes in pH were investigated by adding solutions of different pH to
the catalase in order to raise or lower it. (Tube 1 = pH 4, Tube 2 = pH〜7, Tube 3 = pH 10). The
averages (blue circles) and standard error (black bars) were calculated for each trial. The effects
of different pH levels were determined by measuring the height of the bubble column in mm 1
minute after the substrate, pH solution, water, and catalase were mixed for 20 seconds. 

Discussion

Enzymes are proteins that act as catalysts to accelerate biochemical reactions in living

organisms. Enzymes do this by applying pressure to a substrate to lower a chemical reaction's

activation energy. In this experiment, the enzyme tested was catalase. Catalase is an enzyme that

catalyses the reaction where hydrogen peroxide breaks down into water and oxygen. In

experiment 1, it was determined that catalase and hydrogen peroxide were the most reactive,

with an average measured bubble column height of 18.6 mm and a standard deviation of 7.9 mm.

This can be explained by catalase being the enzyme that is specific to hydrogen peroxide. For

trial 2 where water was mixed with catalase, there was no measurable activity. This is due to an

enzyme’s characteristic of having an active site that is (in most cases) specific to a particular

substrate. In trial 3, there was a small bubble column measured with an average height of 2.8mm
and a standard deviation of 2.7 mm. This is probably a result of human error as catalase can only

react with hydrogen peroxide due to its active sites specificity. Experiment 3 tested the effects of

variations in enzyme concentration on enzyme activity. The results of this experiment show that

if more catalase is used, the reaction will take place faster. This can be attributed to an enzyme's

ability to speed up chemical reactions, so, if more enzyme is used, the reaction will occur

quicker. What is not clear with this data is that more enzyme does not mean more substrate will

be catalysed, it only means that the reaction will happen faster. If the test tubes were allowed to

rest until the entire reaction was complete, the same amount of oxygen would have been given

off. For both experiments 2 and 4, the effects of denaturation could be observed and measured.

Denaturation involves the breaking of the weak linkages, or bonds, within a protein molecule

responsible for its ordered structure, resulting in inactivity. This can be caused by changes in pH,

as well as temperature. The effects of temperature on enzyme activity were studied in experiment

2 by subjecting catalase to 3 different environments then making it react with hydrogen peroxide.

Trial 1 saw the smallest average bubble column height likely due to the denaturation of the

catalase after being exposed to cold temperatures (3 degrees celsius). Cold denaturation of

proteins is thought to be a result of the hydration of polar and non-polar groups of proteins which

typically occurs at low temperatures. In experiment 4 the effects of changes in pH on catalase

activity were studied. The results were very similar for the 3 trials conducted on pH. Typically,

enzymes will denature in a pH that is too acidic or too alkaline, suggesting that the different pH’s

used in this experiment were likely not extreme enough.

Conclusion
 This experiment was designed to test the effects of variations in substrates, temperature,

enzyme concentration, and pH on the rate and effectiveness of the enzyme catalase which

functions to protect cells from oxidative damage. Another enzyme found in humans is amylase.

Amylase is an enzyme that breaks down amylose (a starch) into simple sugars and is found and

produced in the mouth. This is the first chemical step of digestion. Amylase can be put into 3

main classes of amylase enzymes; Alpha-, beta- and gamma-amylase, and each act on different

parts of the carbohydrate molecule. In conclusion, enzymes can be found in most living

organisms and play the vital role of catalysing biochemical reactions that make life possible.

Without them, humans and many other organisms would not exist.