Enzyme concentration and rate of reaction This experiment is carried out to investigate the effects of the increase in the

enzyme concentration on the rate of reaction. It is done in order to determine how the rate of reaction will be altered, whether it will increase, decrease or remain constant when the different concentration of enzymes added. The aim of this practical is to investigate the effect of a reduction in enzyme concentration on the rate of reaction by measuring the initial rate of reaction. In this case the reaction investigated is the breakdown of Albumin by pepsin enzymes. My Hypothesis is at low enzyme concentration there is great competition for the active sites and the rate of reaction is low. As the enzyme concentration increases, there are more active sites and the reaction can proceed at a faster rate. Eventually, increasing the enzyme concentration beyond a certain point has no effect because the substrate concentration becomes the limiting factor.

1. Syringe 1ml of the protein suspension (egg albumin) into a test tube. 2. Syringe 1ml of the protease into the test tube. Mix thoroughly and immediately start the stop clock. 3. Record the time taken for the protein solution to clear. 4. Repeat procedures 1, 2 and 3 of the experiment at a range of different enzyme concentrations ensuring that other conditions are unchanged. 5. Present your results in the most appropriate way 6. Identify any trends in your results. 7. Explain any trends or patterns, supporting your statements with evidence from your data and using biological knowledge. 8. Propose modifications to your procedure which would improve the validity of the results. The dependant variable will be the rate of reaction. The reaction must be independent of the substrate concentration. Any change in the amount of product formed over a specified period of time will be dependent upon the level of enzyme present. In the experiment the enzyme concentration is the independent variable. This is because the concentration is being changed to see the effect of different concentration of enzymes on the rate of reaction. The Substrate (1ml of Albumin), amount of enzyme (1ml), temperature, and PH are maintained the same throughout the whole experiment. The control in the experiment will be the protein and it will be kept the same in all the tubes. This is because the rate at which the proteins react will be dependent on the enzymes. By controlling the amount of proteins present it will prove if the hypothesis that the greater the concentration of enzyme the faster the reaction is correct.

In this experiment we will investigate how the concentration of enzyme will affect the rate of reaction. For the experiment egg white (substrate) and the enzyme Pepsin will be used. Egg white contains the protein albumen. When Egg white is cooked it appears opaque. If the enzyme protease is added to egg white it is broken down and the solution clears. Enzymes are proteins that act as catalysts. They speed up chemical reactions which would otherwise take a very long time. The advantages of enzymes are that they do not get used up in the reaction but stay unchanged. Enzymes are protein molecules which are made up of amino acids. They have a three- dimensional shape because of the way the amino acids are arranged o the protein. A few of these amino acids on the surface of the molecule fold inwards making what is called the active site. Within the enzyme the active site is where the reactant molecule (in this case the egg white) attaches to the enzyme by random collision. The substrate binds to the active site of the enzyme where the reaction takes place. A reaction is able to takes because the amino acids around the active site hold the substrate in place. This forms an enzyme-substrate complex when the enzymes active site binds with the substrate. This is known as the lock and key theory as the substrate attaches to the active site of the enzyme fitting like a key in a lock. Therefore enzymes are very specific as they will only function correctly if the shape of the substrate matches the active site. After the enzyme substrate complex is formed the enzyme changes the substrate into a product molecule. These product molecules then break down and the product is released. The enzymes active site is now free and can be re-used by another molecule to take part in another reaction. This will cause the rate of reaction to increase but eventually it level off because all the active sites are filled.

Discussion (a)From drawing up the results as a lined graph a clear correlation between enzyme concentration and rate of reaction can be seen. There is a positive correlation. As the enzyme concentration increases the rate of reaction also increases linearly. This shows that as there are more enzymes present the reaction is faster as the enzyme are able to catalyse more of the proteins. With a lower concentration of enzymes there are not enough active sights for the reaction to take place quicker. The graph supports the fact that higher concentration causes more collisions between the molecules as, as the concentration increased the reaction time decreased. The initial prediction of how the graph would look is therefore correct.

Although an acceptable set of results was acquired there are some limitations with the results. Improvements could be made to make sure that the experiment is conducted in a better way and that no mistakes are made with the procedure. To ensure random errors are not made a standardized procedure must be followed. Also care and concentration should be kept in mind when conducting the experiment to ensure a valid and reliable set of results. If the experiment is to be repeated the same procedure must be followed. Not knowing exactly where the end point is, because only small concentrations were used in the experiment the time when the enzymes stopped working could not really be identified. Larger concentrations for example 10, 20, 30 could be used. This way the point at which the enzymes started to denature could have been identified. However the results cannot be guaranteed absolutely reliable, as the interpretation of when the solution goes clear could be different for all the tubes. The results would have been more valid and accurate if the experiment had been repeated at least three times. Because there was not enough time the experiment was only conducted once. Averaged results are more accurate than a single experiment. This will ensure that there is a much more reliable set of results. All the test tubes need to be clean before the experiment is conducted as any substance left over in the test tubes could affect the validity of the results. This is because it could cause the solution to appear less clear. The tubes should also be dry and not contain any water as this could cause the proportion of water to be used unequal. Therefore all equipment to be used and the workbench will need to be fully clean before the start of the experiment The test tubes for all the different concentrations must be equal in size. This is because a bigger test tube compared to a small one will have a bigger surface area for the enzymes to work. Using different sizes could affect the results because the solution may not be as much concentrated if using as small tube compared to a big one. It could be possible that the stop clock was stopped or started earlier. This could have affected the results, as the stop clock could have been started at a wrong time. Human Error. For an accurate result shaking must be at an equal pace at all times. When doing the experiment we must use the same type of egg all the time for an accurate result. Safety Precautions: All enzymes are potential allergens and eye and skin contact should be avoided. Wear eye protection and avoid rubbing your eyes in case you have enzyme solution on your hands.

Conclusion: The results show that as the concentration of the enzyme was increased the time taken for the reaction decreased. It took less time for the solution to go clear. So it is write to conclude that the prediction that the rate of reaction would become faster is correct. After studying the graph it is clear that they support the hypothesis and although the results are not perfect they support a valid conclusion. The line of a best fit shows a clear trend. Although there is one anomalous result it is not very far away from the line of best fir therefore the results are still valid.