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Garett Rikard

The Rate of Reaction of the Enzyme Catalase

Introduction

All enzymes are proteins. Enzymes serve as catalyst to control the types of chemical changes and the rates at which they occur. Enzymes act by binding substrates, forming a complex. The complex stresses chemical bonds forming a transition state. This makes the substrate more reactive. Energy is needed to form this state and it is provided by the enzyme. The enzyme's site of attachment and the parts that stress the substrate's bonds is known as the active site. Enzymes can be greatly affected by certain factors which can cause “denaturation with a complete loss of catalytic activity.” High temperature, pH, and ion concentration are three factors that change the shape of the enzyme. When the shape of an enzyme is changed, the enzyme will not be able to perform its function.

The reaction studied in this lab is the breakdown of hydrogen peroxide into oxygen and water when a catalase is added to speed up the reaction. This reaction can be written as 2H2O2 + catalase O2 + 2H2O. This experiment was conducted to test the effect of different factors on the rate of an enzymatic reaction. Hydrogen peroxide is toxic in the body, so it is useful to know the rate of the reaction. It can damage DNA, protein and lipid membranes and may even be a causative factor in cancer. Therefore, it is essential to know how quickly the toxin can be broken down so that it is not harmful. The effect of enzyme concentration, substrate concentration, pH, temperature, and an inhibitor will be tested on an enzymatic reaction.

My hypothesis I concluded from this information is that a high temperature and a low pH will have the greatest affect on the enzymatic reaction. The high temperature will destroy the active site, therefore, no reaction can take place as there is only one specific active site per substrate. A low pH is too acidic and will change the shape of the enzyme.

Materials and Methods

Material used:

U-shaped glass delivery tube

10 ml graduated cylinder

Rubber stopper

Erlenmeyer flask

0.6 % hydrogen peroxide

0.3 % hydrogen peroxide

pH 6.8, regular enzyme

pH 3.5, regular enzyme

When testing the effect of an enzyme concentration on an enzymatic reaction, a baseline concentration was used. First fill a beaker with water. Next fill a graduated cylinder with water. Put your thumb over the top of the graduated cylinder and invert it into the beaker of water. Make sure that no air bubbles got into the graduated cylinder and that it is full of water. Attach one arm of a U-shaped glass delivery tube to a rubber stopper, while the other arm should be inserted into the graduated cylinder. The rubber stopper in connect to a 50 ml Erlenmeyer flask.

Baseline was used as the control. Add 10 ml of a regular enzyme with pH 6.8 and 10 ml of 0.6% hydrogen peroxide to the Erlenmeyer flask. Place the rubber stopper of the flask. Swirl the flask and time the reaction for the first bubble that entered the graduated cylinder until 10 ml of water has been displaced, or until five minutes have passed (whichever on comes first). Record the time (min) to produce 10 ml of oxygen. To get the reaction rate you divide the ml of O2 by minutes it took to produce 10 ml of oxygen.

Repeat this procedure for low enzyme concentration, low substrate concentration, low pH, low temperature, high temperature, and co-factor inhibitor. Use the chart below to determine which pH to level, percent of hydrogen peroxide, and temperature to be used:

Experiment

Enzyme pH & concentration (10ml)

Hydrogen peroxide (10ml)

Temperature (degrees Celsius)

Baseline

pH 6.8, regular

0.6%

22

Low enzyme concentration

pH 6.8, low

0.6%

22

Low substrate concentration

pH 6.8, regular

0.3%

22

Low pH

pH 3.5, regular

0.6%

22

Low temperature

pH 6.8 regular

0.6%

2

High temperature

pH 6.8, regular

0.6%

100

Co-factor inhibitor

pH 6.8, regular

0.6%

2

 

 

 

Results

Experiment

Enzyme pH & concentration (10ml)

Hydrogen peroxide (10ml)

Temperature (degrees Celsius)

Time (min)

Reaction rate

Baseline

pH 6.8, regular

0.6%

22

0.444 min

22.531

Low enzyme concentration

pH 6.8, low

0.6%

22

0.939 min

10.648

Low substrate concentration

pH 6.8, regular

0.3%

22

1.235 min

8.099

Low pH

pH 3.5, regular

0.6%

22

No reaction

Low temperature

pH 6.8 regular

0.6%

2

0.653 min

15.322

High temperature

pH 6.8, regular

0.6%

100

5 min

2ml/ 5 min

Co-factor inhibitor

pH 6.8, regular

0.6%

2

No reaction

 

 

The average rate of reaction was found to be higher when the enzyme catalase was used in the reaction. A low pH had no affect on the rate of reaction. The average rate of reaction was lowest when a high temperature was used in the enzymatic reaction. The average rate of reaction found when a high temperature was used was 2 ml/minute. Low temperature had the greatest rate of reaction with 15.322 ml/minute.

Discussion

The data that was collected from this experiment suggests that my hypothesis was half way correct. Low enzyme concentration also had a big affect on the rate of enzymatic reaction. High temperature caused the reaction to have a low rate of reaction, while low pH has a high rate of reaction.

There are many errors in this experiment that could have caused the results to be incorrect. Some people may have used the same pipets to extract the different solutions. A difference in the speed at which the flasks were swirled would cause a difference in the results obtained. If one flask was swirled faster than the other, then more bubble would be produced and the rate of reaction would be increased. If the flask was held in the palm, then the temperature may have increased slightly to change the rate of reaction.

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Bibliography

McGuinness, Candace. 2003-2004. Lab Five: The Activity of the Enzyme Catalase. Biology 101 Lab Manual. page 5-1.

http://encarta.msn.com/encnet/refpages/RefArticle.sapz?refid=761552710

http://www.emc.maricopa.edu/faculty/farabee/BIOBK/BioBookEnzym.html