Studying the Action and Activity of Amylase on Starch Digestion

Principle
In order to find out the action of amylase on starch, amylase solution can be added to starch solution. After some time, the presence of starch in the mixture can be tested by iodine solution. The positive test for the starch is dark blue. If there is no starch, the iodine solution remains brown. If there is less starch or no starch in the mixture, it can be concluded that amylase can change starch into other chemicals. This shows that amylase can have some action on starch. To study the activity of amylase on starch, the amount of starch in the mixture can be estimated by observing the color change in the iodine solution at regular time intervals. The color changes from dark blue to brown indicating the disappearance of starch in the mixture. If the color changes very rapidly, it shows that the activity of amylase is high. If the change is slow, it shows that the activity of amylase is slow.

Procedure

1. Put two drops of iodine solution into each depression in a spotting tile.
2. Add 5 mL of 1% starch solution into Test Tube A with a 5-mL pipette.
3. Add 0.5 mL of 1% amylase solution into the same tube with a 1-mL pipette.
4. Mix the solution thoroughly.
5. At 30-second intervals, take 2 drops of mixture of Test Tube A and transfer them to each depression of the spotting tile.
6. Mix the iodine solution and the mixture into the depression with a glass rod.
7. Observe and record the color of the iodine solution. 8. Repeat steps 5 to 7 until the color of the iodine solution remains brown.

Explanation of Results
Starch molecules can combine with the active sites of the amylase. Amylase can then speed up the digestion of starch into maltose. When amylase is mixed with starch, starch is digested. Since less starch is present in the mixture, if we test the presence of the starch by iodine solution, the iodine is less dark blue. When there is a complete digestion of starch, the iodine solution will remain brown. The reaction rate is fast at first since the amount of starch in the mixture is high. However, the reaction rate is slower after 1.5 minutes, since the concentration of substrate decreases.

Precautions

1. The experiment should be carried out at the same temperature.
2. The solutions must be well-mixed since the starch and amylase are not completely dissolved.
3. Adding too much iodine solution and mixture into the depressions must be avoided. This may cause the over flow of the solution from the depressions. Mixing of the solutions in the depressions is not easily done.

Error Sources

1. The volume of each drop of solution may not be the same.
2. The amylase or starch solution left on the inside surface of the test tube may cause an inaccuracy in the amount of solution added.
3. The amylase and starch are not completely soluble.

Improvements

1. The color change is not easy to observe. If a colorimeter and data-logging device are used, the results can be quantitized and easily compared.
2. A shorter time interval can be used. This makes the result more clearly observed. Of course, this may be difficult when the process is done by 1 person