How do different pH values affect the enzyme catalase's
activity in the decomposition of hydrogen peroxide
Ananya Punyamurtula
Catalase is a enzyme present in most organisms, and aid in the decomposition of
hydrogen peroxide into water and oxygen, mitigating its toxic effects on organisms.
(Augustyn, 2017) Hydrogen peroxide is a known chemical that causes damage to the
body. (Watt et al., 2004) Therefore, catalase's role is highly crucial. The level of pH in
the environment catalase is present in, is a factor that contributes to its activity, and
therefore either decreases, or increases the rate of reaction.
The aim of the experiment was to investigate the effects of the pH value on catalase's
enzyme activity, and thus the rate of reaction, in the decomposition of hydrogen
peroxide, by measuring the oxygen gas released by the reaction.
it was hypothesized that a pH above or below pH 4 would result in lower enzyme
activity. This is because catalase is a digestive enzyme, and digestive enzymes tend to
exist in acidic environments.
The experiment measured the amount of oxygen gas released in the decomposition
reaction of hydrogen peroxide into water, with catalase acting on the reaction as an enzyme,
while in different pH environments (3, 4, 5, 6, 10, 11) . The gas tubing was first set up using a
measuring cylinder filled with water, upside down in a tub. Next, 6 chicken liver pieces were
cut into 2cm cubes and weighed to 8.8 grams. These pieces were each places into a test
tube, and the test tubes placed in ice, and checked with a thermometer to be below 25
Celsius. 4mL of different solutions added to the chicken liver pieces and used to change the
pH of the catalase. These included, water, 0.1M Hydrochloric acid. 0.05M Hydrochloric acid,
0.1M Sodium hydroxide and 0.05M Sodium hydroxide. One group was left without any
solution, acting as a negative control. One at a time, 6mL of 3% Hydrogen peroxide was
added to each chicken liver piece, and then gas tubing was used to measure the amount of
oxygen released in five minutes. To obtain the result, how far the water level decreased in
the measuring cylinder, was measured.
Augustyn, A. (2017). Catalase | Function & Applications. In Encyclopædia Britannica.
https://www.britannica.com/science/catalase
Gillespie, C. (2018, May 21). PH Levels of Catalase. Sciencing. https://sciencing.com/ph-
levels-catalase-6826245.html
Watt, B. E., Proudfoot, A. T., & Vale, J. A. (2004). Hydrogen Peroxide Poisoning. Toxicological
Reviews, 23(1), 51–57. https://doi.org/10.2165/00139709-200423010-00006
As can be seen on the raw data table, the optimal pH for catalase is pH 6-8, with a the highest rate of reaction
at pH 6. The most amount of oxygen releases, and thus the highest rate of reaction and enzyme activity, was at
pH 6, releasing 120 mL of oxygen gas. This was followed by a pH of 4, producing 54 mL of oxygen gas. The least
amount of oxygen released, and the lowest rate of reaction, is at pH 3, releasing 1mL of oxygen gas. This does
not support the hypothesis, as catalase was found to work best at pH 6, not 4. This supports scientific theory,
as the liver sustains at a pH of 7, and catalase must work optimally at the pH range of its environment (Gillespie,
2018).
The designed methodology was valid and supports that pH changes the enzyme activity of catalase, and thus
impacts the rate of reaction. Most variables that had the potential to affect the dependant variable, the enzyme
activity of catalase, was controlled. As indicated by the methodology, for each IV group, the pH impacting
solutions, the temperature was kept constant under 25C° using ice, as after this temperature, catalase begins
to slowly denature. The concentration and amount of substrate was kept constant at 6mL of 3% hydrogen
peroxide. The surface area and amount of chicken liver was also controlled by measuring equal pieces of
catalase and then weighing the pieces to ensure consistency. Furthermore, a negative control, where no
solution was added, was used to measure how much the pH impacted the catalase. However, to further
improve validity, the surface area of liver could be better controlled by cutting the pieces into 0.5cm cubes
instead of 2cm cubes.
The methodology was not reproducible. Instructions on how to perform a serial dilution was not provided and
could be interpreted differently. Furthermore, the methodology does not inform where to place the
thermometer clearly, which could lead to inaccurate measurement, and control of temperature. The
methodology should be more reproducible by instructing how to perform serial distillations and where to place
the thermometer. The methodology is repeatable as there where four trials. The high number of trials is
sufficient to ensure that trials with errors would skew to an average data point which can be repeated. If given
more time, the methodology could be made more repeatable by instructing to conduct 8 trials.
As no additional trials were conducted, precision cannot be evaluated for the experiment.
The data collected was not accurate. The graph's data points for pH 5 and 11 do not match
the expected trend as the rate of reaction fluctuated instead of having a single peak point.
.This may have occurred due to underestimation. The gas tubing used may have lead to
inaccurate results due to systematic error. The tubing fell out of the measuring cylinder,
and therefore less gas release may have been recorded. To improve accuracy, a gas
syringe could be used instead.
The experiment investigating the effects of pH on the enzyme activity of
catalase, in the decomposition of hydrogen peroxide, suggests that
catalase's most optimal pH is pH 6. From the raw data table, it can be
understood that catalase's optimal pH range is 5-7. A pH outside these
values, significantly decreases the rate of reaction. While a pH of 6 produces
120 mL of oxygen gas, a drop to pH 3 only produces 1 mL. Increasing the pH
does not show as drastic changes, compared to decreasing the pH. A pH of
10 produces 35 mL of oxyegn gas. Therefore, it is suggested that by
decreasing the pH, the enzyme catalase would denature more, compared to
increasing the pH. However, this is not investigated by solutions, and only
predicited by trends on the graph. To further investigate the effects of pH on
catalase enzyme activity, solutions between pH 5 and 10 should be used in a
similar experiment.
I would like to acknowledge the lab technicians, the teachers and staff who supported
me including Ms. Lam, and Keerthana and Krish who all partook in this experiment. I
would also like to acknowledge the traditional owners of this land, the Bunurong and
Wadawurrung peoples.