investigating the effect of substate concentration on the activity of catalase

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Effect of Substrate Concentration on Catalase

Aim: An experiment to find out how the concentration of hydrogen peroxide affects the rate of reaction of the enzyme catalase.

Background Information:

Enzymes such as catalase are globular protein molecules with catalytic properties. A catalyst is a substrate which can alter the rate of reaction without itself undergoing any permanent change. As they are not changed my reactions which they catalyse, enzymes can be used over and over again. They are therefore quite effective in small amounts.
All enzymes are specific as each one performs a particular reaction.

Catalase is renowned as the fastest known enzyme. It is found in the liver where it speeds up the decomposition of hydrogen peroxide into water and oxygen, catalase has a turnover number (the number of substrate molecules which one molecule of enzyme turns into products per second) of 6 million. Its action can be demonstrated by dropping a sample of liver into hydrogen peroxide: the fizzing that ensues as oxygen is given off is a dramatic demonstration of an enzyme in action. I intend to investigate the reaction of the enzyme with the metabolic poison hydrogen peroxide and a factor affecting the rate of the reaction. They are:

1. Concentration of Enzyme

The more enzyme molecules there are in a solution, the more likely a substrate molecule is to collide with one. If the amount of substrate is limited there comes a point where there is more than enough enzyme to deal with all the available substrate, so the addition of more enzymes has no effect.

2. pH Conditions

All enzymes have an optimum pH value. Most work best at neutral pH 7, however there are certain exceptions. Enzymes in our stomach work best in acidic conditions. By altering the optimum pH the H+ and OH-ions found in acids and alkalis can mess up the ionic bonds that hold the tertiary structure in place. This makes the active site change shape, so the enzyme is denatured and it is therefore no longer able to be used. 3. Temperature Conditions

More heat means more kinetic energy, so molecules move faster. This makes the enzyme more likely to collide with the substrate. But, of the temperature increases beyond a certain point the rate of reaction decreases and ultimately stops. If the temperature goes above a certain point the vibrating enzymes break some of the bonds which are holding them together. The active site changes shape and the enzyme and substrate no longer fit together. At this point the enzyme is denatured and can no longer carry out is function.

4. Substrate Concentration

Substrate concentration affects the rate

I intend to investigate the effects on the rate of reaction upon altering the concentration of the substrate. In order to do this while also ensuring a fair test. I will aim to maintain all other variables constant. Prediction:

Apparatus:

Boiling Tube

Rubber Bung and Delivery Tube

Potato Samples each of 10cm in length

Hydrogen Peroxide Solution

Distilled Water

Digital Stop Clock

Scalpel

Plastic Ruler

Measuring Cylinder

Water Bath set at 40Oc

Method:

1. Measure the hydrogen peroxide and distilled water for each solution using a measuring cylinder and pour into each boiling tube. This should give a decent range and adequate repeats to come to a conclusion. I decided that boiling tubes are the easiest way to keep the solutions as they can be labelled easily and can be kept in a rack till use. Although only slowly, hydrogen peroxide decays in the absence of a catalyst this may affect the results.

2. Collect the samples of potatoes and check whether they are 10cm in length, then cut each potato sample into 0.5cm intervals using the scalpel and the ruler. It is important that the potato samples are cut into intervals as it increases the surface.

3. Firstly, take Solution A and check its temperature. Once it reaches
40oC add the pieces of potato into the boiling tube.

4. Put the end of the delivery tube into a gas syringe. Measure the amount of gas produced over a five minute duration.

5. Repeat the procedure with the other hydrogen peroxide solutions.

6. Repeat all the tests at least three times so that an average can be obtained. Table of Results:

Volume of Hydrogen Peroxide Volume of Water Volume of Oxygen produced per 5 minutes Average Oxygen produced in 1 min

1 2 3 Average

Solution A 20 0 13 13 13 13 2.6

Solution B 15 5 12 10.5 11 11.1 2.22

Solution C 10 10 11 9 10 10 2

Solution D 5 15 4 6 4.5 4.8 0.96

Solution E 0 20 0 0 0 0 0

Graph:

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Analysis of Graph

The overall trend represented by the graph suggests that as the concentration of the substrate is increased there is a gradual increase in the amount of oxygen which is produced. There is a steady increase in the rate of reaction because there are more substrate molecules available. Due to the increased chnace of a successful collsion caused by random thermal motion when there are more molecules present there is an increase in rate with higher substrate molecules present, there is an increase in rate with higher substrate concentration. For lower concentrations the rate of reaction is therefore directly proportional to the concentration of hydrogen peroxide in the solution. This is because if we double the amounts of substrate molecules that are in the solution, double the amount will find an enzyme molecule at the same time if all the substrate molecules are moving at relatively the same pace.

Evaluation of Evidence

Accuracy

Although the results have validated a satisfactory graph of rate against concentration, and established a conclusion that is acceptable, the results were obviously not perfect.

Anomalies

Improvements

Although the procedure used was up to satisfactory standards, there were a number of improvements and chnages that could have been done in order to further improve the accuracy of the entire investigation.