The effect of different substrate concentrations of Hydrogen Peroxide on the rate of enzyme activity

Updated: Oct 24, 2019


An enzyme is a biological catalyst that catalyzes biochemical reactions in living cells, in other words, speeds up a chemical reaction without permanently changing itself.

The enzyme used in the experiment is “Catalase” and can be found in most living cells as well as in yeast. Catalase causes the Hydrogen Peroxide to break down into water and oxygen and the chemical formula follows :

Hydrogen Peroxide ⇒ Water + Oxygen

The aim of this experiment was to investigate the effect of different substrate concentration, which is, in this case, different concentrations of Hydrogen Peroxide on the rate of enzyme activity of catalase. Filter paper discs are dipped in the Catalase solution before they are submerged into solutions with Hydrogen Peroxide. Oxygen will be produced during this reaction will form all around the disc until it will be buoyant enough to then float upwards. By measuring the time taken for the filter discs to sink to the bottom and back up again in beakers with different Hydrogen Peroxide concentrations, will give us data to​ calculate reaction rate.


By changing the concentration of Hydrogen Peroxide, I expected that the rate of the enzyme activity would increase as the concentration increases, thus the circular filter disc floating faster up to the surface. Although, there is always a possibility for the rate of enzyme activity to get manipulated by the temperature, pH etc. However, I came to the conclusion that the rate of the enzyme activity will increase until the enzyme concentration becomes the limiting factor. When it becomes the limiting factor, all the enzymes will be saturated with molecules (Plateau) and no further increase in the substrate concentration can increase the enzyme activity.


Dependent - Time is taken for the filter discs to reach the surface

The stopwatch will start as soon as the filter disc hits the surface of the solution and the time will be taken until the filter disc hits the bottom and back up again.

Independent - Concentration of the hydrogen peroxide solution

Three different concentrations of Hydrogen Peroxide are prepared in different beakers, 0.5%, 1.5% and 3.0%.​

Tools 1.3


  1. With a hole punch, 30-40 filter paper discs were cut out and with a force, they have gently put on a piece of clean paper without any human contact to keep them clean.

  2. 40 ml of 0.5%, 1,5%, and 3% Hydrogen Peroxide solutions were poured into individual beakers.

  3. The yeast suspension was poured into a 50ml measuring cylinder and prepared for the discs.

  4. Each individual disc was dipped in yeast suspension with forceps and put on clean paper to soak extra yeast on the discs.

  5. Individual discs were picked up and put into one substrate solution with the timer immediately set when the disc would hit the surface.

  6. This technique was repeated seven times at each beaker with different Hydrogen Concentrations.

Data and analysis

Bubbles appeared immediately around the disc when it submerged into the solution and increased as the disc reached the bottom. Sometimes the disc floated upwards to the surface steadily and other times flipped around one or two times before reaching the surface. I believe that must be because of the different masses of oxygen bubbles on each side of the disc, causing it to get unstable, thus flipping around while moving upwards.

Limitations ⇒ Improvements

A stronger Hydrogen Peroxide solution, appreciated as high as 35.0% would have proved the hypothesis and fulfilled the aim of the experiment, showing the Plateau. The experiment would then have a wide enough parameter for its manipulated variable (Hydrogen Peroxide) to show its Plateau in the data set.

However, the Plateau could be shown in another way, by diluting the catalase solution. We would then have a lower catalase concentration, which would, in turn, make fewer catalase molecules to be available on the filter disc, thus less enzyme active sites being available for chemical reaction to occur. In this way, less Hydrogen Peroxide concentrations will be required hence, a lower a lower manipulated variable will be just enough to show the Plateau.

Conclusion - as well as evaluation

The result matches up to one part of the hypothesis, considering the rate of enzyme activity did increase the more concentrated the Hydrogen Peroxide became. However, when it came to the enzyme activity ceasing to increase when reaching the plateau, was not proven. Due to the limiting substrate concentration, many enzymes still carried empty active sites, thus not reaching the plateau. When the enzymes were in the 3% substrate concentration, many of the enzyme molecules active sites were not filled completely, which it could have but it would require a higher substrate concentration. The enzymes needed a higher concentration of 3% to reach the plateau, thus leading to a lack of information in the lab report.

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