Enzymes in Saliva
Overall Goal
This activity will allow students to study a simple and readily available enzyme, salivary
amylase. Amylase acts to partially degrade starch into simple disaccharide sugars (maltose),
smaller polysaccharides (dextrin), and simple sugars (glucose). None of these products give a
positive iodine test, but starch does. Students will prepare an enzyme solution and test its
ability to break down starch.
Background Information
The first stage of the digestive process occurs in the mouth were food is mechanically broken down and
mixed with saliva. Saliva contains an enzyme, amylase, which begins the digestion of starches and
converts them to sugar molecules. In the presence of iodine starch turns a deep purple but sugar has no
color. In this activity we are going to use iodine to observe the effect saliva has on starches.
Materials
1. Soluble Starch Solution
2. Iodine Solution
3. Buffer Solution
4. 3 Beakers, 250mL
5. 3 Droppers
6. 2 Graduated Cylinders, 50mL
7. Dextrose Solution.
Solutions
Soluble Starch Solution – Add 1g of soluble starch to 200mL of water. Heat to almost boiling.
Solution should become almost clear. Dilute to 800mL and allow to cool to room temperature.
Iodine Solution – Add 1g of iodine to 200mL of water. While stirring the solution heat to almost
boiling. When no more iodine will dissolve into solution allow to cool to room temp.
Buffer Solution – pH 6.8–7.2 Follow directions provided on buffer package.
Dextrose Solution - Add 1g of dextrose to 200mL of water. Heat to almost boiling. Solution
should become almost clear. Dilute to 800mL.
Procedure
1. Test For Starch.
A. Place a dropper full of starch solution in a test tune.
B. Add a drop of iodine solution to the starch.
C. Deep purple color confirms the presence of starch.
D. In a separate test tube add 10mL of the dextrose solution to a few drops or iodine
solution.
E. Sugars do not turn purple in the presence of iodine.
2. Test Your Saliva for Amylase.
A. Make a saliva solution.
Saliva solution A: Add 2mL of saliva to 10mL of buffer solution
Saliva solution B: Add 2mL of saliva to 20mL of buffer solution.
B. Add 20 drops of iodine solution to a beaker.
C. Add 30 drops of starch solution to the beaker and stir. The solution should be a light
blue color.
D. Add 2mL of Saliva Solution A. Record the amount of time it takes for the blue color to
disappear.
E. Repeats steps B – D using Saliva solution B.
3. Test the Effects of Vinegar on Amylase.
A. Make a saliva solution.
Saliva solution A: Add 1 drop of vinegar to 2mL of saliva.
Saliva solution B: Add 2 drops of vinegar to 2mL of saliva.
Saliva solution C: Add 3 drops of vinegar to 2mL of saliva.
B. Add 20 drops of iodine solution to a beaker.
C. Add 30 drops of starch solution to the beaker and stir. The solution should be a
light blue color.
D. Add Saliva Solution A. Record the amount of time it takes for the blue color to
disappear.
E. Repeat steps B-D using Saliva solution B and Saliva solution C.
4. Test the Effects of Temperature on Amylase.
A. Make 2 saliva solutions.
B. Heat 1 saliva solution on a hotplate.
C. Place the other saliva solution in an ice bath.
D. Add 20 drops of iodine solution to a beaker.
E. Add 30 drops of starch solution to the beaker and stir. The solution should be a
light blue color.
F. Add the heated saliva solution. Record the amount of time it takes for the blue
color to disappear.
G. Add the cooled saliva solution. Record the amount of time it takes for the blue
color to disappear.
Expected Results
1. Test Your Saliva for Amylase: Saliva solution A should turn the iodine solution clear in about
1-2min. The saliva solution B should turn the iodine solution clear in about 3-5min. The results
may vary greatly from individual and from trial to trial. The concentration of amylase varies
within an individual’s mouth.
2. Test the Effects of Vinegar on Amylase: Depending on the saliva samples collected from the
individuals the amount of time it takes for the iodine solution to become clear should increase
respectively to the amount of drops of vinegar added to the solutions.
3. Test the Effects of Temperature on Amylase: Since heat is known to destroy or denature enzymes
the iodine solution should not change to clear at all for the heated saliva solution. The cooled
saliva solution may turn the iodine solution clear but the cold will slow the rate of the enzyme
action so the time to turn clear should increase significantly.
Reactions
1. Starch reacts with iodine, I3-, to produce the blue starch-iodine complex:
I2 + I- → I3I3- + Starch → I3- and starch complex (blue)
2. When the enzyme in your saliva, amylase, reacts with the starch it causes the starch to be
broken down into simple sugars. Once the starch is broken down into simple sugars it
not longer forms a complex with the iodine and so the dark blue color no longer is
present.
Possible Questions
0. Why did it take longer for saliva solution B in Step 1 to change the starch-iodine solution
clear?
1. What foods do you think contain starch? How could you test for starch?
2. Why does it take several minutes for the blue to fade totally away?
3. Alcoholics do not produce the enzyme needed for digestion of alcohol. How will this
affect their ability to metabolize alcohol?
4. How does heating affect the action of the enzyme?
Notes for the Teacher
TEACHING TIPS
1. The saliva solution is a buffer solution with a pH of about 6.5. Enzymes are sensitive to
changes in pH. This enzyme, amylase, has optimum activity at room temperature at pH
6.5.
2. Amylase gets its name from amylose, one of the two long-chain polymers that make up
starch. The other chain, which is branched, is called amylopectin. Amylose forms a coiled
structure. Iodine, probably as the species I5-, fits inside this structure to form the blue
complex.
3. The iodine solution is made in KI because iodine is soluble in a KI solution but not in
water alone.
4. Some saliva has a very low concentration of amylase. Thus, some students may need to
use more saliva in their solutions.
5. The pancreas also produces amylase. This substance is called pancreatic amylase and is
part of pancreatic juice. It is responsible primarily for the digestion of starch in the small
intestine.
6. You may see some individual variation in reaction times within the class. Stress the
importance of controlling all other variables. You may wish to construct a normal
distribution curve with your students’ results.