Catalase Lab

Title: The Effect of pH on the Rate of Reactions Catalyzed by Catalase

Purpose: To test the effects of pH on the rate of reactions catalyzed by catalase.

Background: Enzymes are proteins within cells that act as catalysts in biochemical reactions. The enzyme catalase speeds up the decomposition reaction of hydrogen peroxide (H2O2) into water (H2O) and oxygen gas (O2) by lowering the reaction’s activation energy. The equation of the reaction is:

2H2O2 → 2H2O + O2

Like all catalysts, catalase is not used up or changed in the reaction, and can be reused. One factor that can affect the rate of an enzymatic reaction is the pH of the reaction. A pH level that varies greatly from the optimal pH of the reaction often causes a slowed reaction rate. The optimal pH level for enzymatic reactions usually matches the standard pH for the organism in which the enzymatic reaction typically occurs. Some of the other factors that affect the rate of an enzymatic reaction include temperature, presence of inhibitors, concentration of enzyme, and concentration of substrate, which is the molecule on which the enzyme acts.

 The substrate of catalase is H2O2, which breaks down into water and oxygen gas significantly faster with catalase present. This enzyme is found in nearly all living cells, because catalase is needed to break down excess H2O2 within the cell. Otherwise, the H2O2 could accumulate, and this would be toxic to the cell and the organism of which the cell is a part. Catalase is particularly concentrated in the livers of animals.

Hypothesis: If pH has an effect on the rate of catalase reactions, then as the pH increases in acidity, the rate of the reaction will decrease.

Materials: See apparatus set-up diagram.

Materials Diagram

(Original Diagram)

Procedure (for standard pH of 4.7):

  1. Fill right-side-up graduated tube with 10mL H2O2 and add 350μL H2O. Leave the lid off.
  2. Fill beaker with water, and place the upside-down graduated tube with a hole in the lid into the beaker of water.
  3. Attach one end of the tube to the top of the upside-down graduated tube.
  4. Attach the other end of the tube to the lid of the right-side-up graduated tube.
  5. Clamp binder clips near each end of the tube.
  6. Add 100μL of the catalase mixture to the H2O2.
  7. Quickly screw on the lid of the graduated tube containing catalase and H2O2. Ensure that the tube is secure and not leaking.
  8. Place the graduated tube containing catalase and H2O2 into an empty beaker.
  9. Holding the upside-down graduated tube steady, remove the binder clips and start the timer. Be sure that both beakers and graduated tube stay level and do not change positions.
  10. Record the volume of water (mL) dropped for every time interval.
  11. Rinse equipment and repeat steps 1 – 10 twice for the second and third trials.

Data:

Graph Chart

(Original Chart and Graph)

Analysis: As the pH grew more acidic, oxygen was produced at a slower rate. When the pH of the catalase solution was 4.7, 5.50 mL of oxygen were produced over 160 seconds. When the pH of the catalase solution was 2.5, 0.00 mL of oxygen was produced over the entire 400 seconds.

Conclusion: The data collected supported the hypothesis that pH would have an effect on the rate of catalase reactions. The data indicated that as the pH grew more acidic, the rate of the reaction decreased. The reaction with a pH of 4.7 produced oxygen about 16 times faster than the reaction with a pH of 3.4. Additionally, the reaction with a pH of 2.5 did not produce any oxygen. Oxygen was not produced as quickly during the experiements using solutions of higher acidity because if the optimal conditions for an enzymatic reaction is altered too drastically, the enzyme will be denatured.

Denaturization of enzymes causes a change in structure, which then disables the substrate from attaching to the enzyme’s active site. The enzyme is then denatured and can no longer serve as a functional enzyme. The data collected supports the idea that the catalase was denatured when put into contact with highly acidic pH levels. The standard baseline pH level of this bovine catalase reaction is 4.7.

Some of the potential errors that may have been made in completing this lab include user errors, like misinterpreting the data and slightly inaccurate time readings. This misinterpretation of the water level in the graduated tube and of the time reading may have caused each group’s data to be shifted slightly off from the others by around 1 mL or 1 s.

Another potential error could have resulted from the fact that the data from the 4.7 pH was an average of multiple trials, and the 2.5 pH data and 3.4 pH data were each taken from individual trials. Data from one single trial could include outliers or other user errors, and without using an average of several trials, the data could possibly have been inaccurate or inconsistent with expected results.

In addition, the data from the 3.4 pH and the 4.7 pH did not contain all of the data points that pH 2.5 contained. This left room for estimation, and the estimations that were made could have potentially been inaccurate. If any of the data points that were collected were in any way inaccurate, the estimations that were based off of this data would therefore also be inaccurate. ✎

Works Cited:

Worthington, Krystal. “Catalase.”Worthington Biochemical Corporation. N.p., n.d. Web. 11 Oct. 2013. http://www.worthington-biochem.com/ctl/default.html

Scandalios, John, Lingqiang Guan, and Alexios Polidoros. “Catalases in Plants.”North Carolina State University. N.p., n.d. Web. 10 Nov. 2013. http://users.auth.gr/palexios/Publications/Pdf-Chapters/Catalases%20in%20Plants.pdf

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