investigating the optimum pH for the breakdown of starch

outline

using buffers to provide different pH environments, testing the samples of an amylase starch solution at 10 seconds intervals to determine when all the starch has broken down. iodine is used as the indicator and the absence of a blue-black colour one the amylase-starch solution has been added indicates the end point. the rate of the enzyme controlled reaction can be calculated using the 1/time.

health and safety

• all enzymes have the potential to be allergens → spillage should be cleaned immediate + avoid skin contact

• pH buffer solutions risk can varies based on the pH and manufacturer

• iodine solution low risk once made up

equipment

• water bath set at 35c

• amylase solution starch solution

• iodine solution

• stopwatch

• spotting tile x2

• dropping pipette

• 5cm3 syringes

• pH buffers

• test tubes

method outline

1. place the starch and amylase solution into the water bath and allow for acclimatisation

2. use the dropping pipette to place one drop of iodine into the each well of the spotting tile

3. use separate syringes to place 2cm3 amylase and 1cm3 of the pH buffer into the a test tube and return to water bath

4. simultaneously add 2cm3 starch to the amylase/buffer solution and start the stop watch. ensure the mixture is combined thoroughly

5. after 10 seconds remove one drop of mixture and test if with the iodine solution to check for completion. repeat this every 10 seconds until the end point is reached

6. carryout any repeats to ensure a full spread of data

data analysis

• calculate mean and the use it to determine the rate of reaction: 1/time

• use graph to estimate the opium pH of amylase

hypothesis

Amylase will break down starch fastest at an optimal, near-neutral pH (approx. pH 6.7–7.0), because extreme pH levels denature the enzyme's active site, restricting its ability to bind starch. Consequently, iodine tests will show the quickest disappearance of starch at this optimal pH, while alkaline or highly acidic environments will show slower digestion.

is there a difference between activity if amylase at any pH

no significant difference

evaluation

errors:

• though mixing of the enzyme substrate buffer solution due to the narrow nature of the reaction vessel and ensure the aliquots are taken out exactly 10 seconds so that reaction time is not over or under estimated

describe how the changes in pH effect the activity of amylase

• as the pH increases the rate of the reaction increases

• peak value → graph

• after the optimum value as the pH increases the rate of reaction decreases

variables - independent

pH of the solution

variables - dependent

time taken for strach to be fully broken down