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A Plan for a Practical to Determine the Effect of Temperature Change on the Operation of Bacterial Amylase

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A Practical to Determine the Effect of Temperature Change on the Operation of Bacterial Amylase Planning: The object of this experiment is to determine the effect of temperature change on the operation of bacterial amylase. Bacterial amylase is an enzyme found and used in bacterial digestive processes which breaks down starch to maltose. Starch is a polysaccharide thus bacterial amylase catalyses the hydrolysis of the 1-4 glycosydic bonds in order to form the disaccharide maltose. There are a number of variables which affect the efficiency of the bacterial amylase. A very high or very low pH may denature the bacterial amylase due to the alteration of the tertiary structure of the bacterial amylase, changing the 'shape' of the bacterial amylases active site preventing the starch from binding to the active site in order to form an enzyme-substrate complex and thus degrade the starch into maltose, the pH will be controlled with the introduction of a pH buffer into each test tube before the experiment is started in order to keep the pH constant at 7, keeping it a fair test. ...read more.


In order to measure the operation of the bacterial amylase under different temperatures, the solution must be tested with iodine at regular intervals at the specified temperature until there is no more dark blue colour in the solution. Temperature will be kept constant as the independent variable with the thermostatically controlled water baths; pH will be controlled by the use of a pH buffer while the volume will be controlled using pipettes. Before the experiment is begun safety glasses and a lab coat must be worn in order to prevent damage to eyes and clothes from spillages. 7 thermostatically controlled water baths will be set up at temperatures 20�C, 25�C, 30�C, 35�C, 40�C, 45�C, 50�C. 5�C increments were used in order to obtain a broad enough range of readings in order to better determine the effect of temperature on the operation of the bacterial amylase. A second thermometer will also be added to the water bath in order to ensure that the temperature of the water bath is as close as possible to the temperature being tested. 7 test tubes will then be selected and 5ml of a 1% starch solution will be added to each of them, making sure that there is no air bubbles present in the syringes while the 5ml is being measured out. ...read more.


A pipette will then be taken and 3 drops of the solution will be placed on a spotting tile every 20 seconds in order to allow sufficient progress in between each reading, as lower intervals would be impractical while higher intervals will be a lot less accurate and iodine added to the solution after it had been placed on the spotting tile. If there is starch present then the solution will go a dark blue colour, while if all of the starch has been broken down into maltose then the solution will go a orange-yellow colour as that is the original colour of the iodine solution. Once this orange-yellow colour has been achieved, the time taken to achieve it will be recorded This process will then be repeated for each of the other test tubes and the times noted for the temperatures used in each of them. After this has been completed for each temperature, the 7 test tubes will be cleaned thoroughly with distilled water and the process repeated again for each temperature so that an average time could be calculated for each temperature in order to reduce the effect of anomalies on the final time. ...read more.

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