ENZYME LABS
Introduction:
Without enzymes chemical reactions in cells would progress at a snails pace and life
would cease to exist. For example, without enzymes it would take six months to digest
one Hershey bar.
Learning expectations:
1. What is an enzyme?
2. Are enzymes specific?
3. Are enzymes used up in chemical reactions?
4. What are enzymes mostly made of?
5. How do enzymes work?
6. How will changes in the cells physical or chemical environment (temperature or Ph)
affect enzyme activity?
EXPERIMENT 1- SPIT LAB-WHAT ARE THE FUNCTIONS OF ENZYMES
AND HOW DOES HIGH TEMPERATURE AFFECT ENZYME FUNCTION?
PREDICT THE EFFECT OF HIGH TEMPERATURE ON ENZYME
FUNCTION: _______________________________________________
Introduction:
If you ever chewed bread or a cracker for about three minutes without swallowing you
should notice that the bread or cracker would begin to taste sweet. An enzyme, called
amylase, is found in saliva. Amylase speeds up the break down of starch into sugar.
Iodine is an indicator for starch. Iodine, a reddish-brown solution, turns black in the
presence of starch. Benedicts is a test for monosaccharides. When you mix a
monosaccharide with Benedicts and heat, the Benedicts will turn from blue to green,
yellow or orange depending on the amount of monosaccharide present.
Material:
8 test tubes, test tube rack, cup (for saliva), Iodine, Benedicts solution,
2% starch, glucose, saliva (of labs group own making) and goggles. (1000
ml beaker with water on a hot plate in central location.)
Procedure:
A. Obtain eight test tubes and label as follows:
1. Starch-iodine
2. Starch-benedicts
3. Starch-saliva-iodine
4. Starch-saliva-benedicts
5. Starch boiled saliva-iodine
6. Starch boiled saliva-benedicts
7. Glucose and benedicts
8. Glucose and iodine
B. Spit into a cup to obtain about 4-8 ml of saliva.
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C. Add spit to test tubes # 3,4, 5, and 6 (approximately 1-2 ml)
D. Boil spit in test tubes # 5 and 6 for three minutes
E. Add 1 ml of 2% starch to test tubes 1, 2, 3, 4, 5 and 6
F. Add 1 ml of glucose to test tube # 7 and 8
G. Let test tubes stand for six minutes
H. After six minutes add 4 drops of iodine to test tubes # 1, 3, 5, and 8. Record color
change
I. Add 10 drops of benedicts solution to test tubes # 2,4,6, and 7 and place tubes in the
boiling water for five minutes. Record color change
COMPARING THE EFFECT OF BOILED AND UN-BOILED SALIVA (AMYLASE) ON STARCH
TUBE #
CONTENTS
TESTED WITH
1
STARCH
IODINE
2
STARCH
BENEDICTS
3.
STARCHSALIVA
STARCHSALIVA
IODINE
5
STARCHBOILED SALIVA
IODINE
6.
STARCHBOILED SALIVA
BENEDICTS
7.
GLUCOSE
BENEDICTS
8.
GLUCOSE
IODINE
4.
RESULT
INTERPRETATION
BENEDICTS
Analysis:
1. What do enzymes do?
2. What is the action of the enzyme found in saliva on starch?
3. What was the purpose of test tubes 1, 2, 7, and 8?
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4. What substances do iodine and Benedict's solution test for?
5. What change takes place when starch and saliva are mixed, according to the results in
tubes 3 and 4?
6. Tubes 5 and 6 probably did not give the same results as tubes 3 and 4. In what way
were the contents treated that could account for this difference?
7. (a) Are your results consistent with the hypothesis that an enzyme in saliva
has changed starch to sugar?
(b) Do your results demonstrate that an enzyme in saliva has changed starch to sugar?
8. In what way do the results with tubes 5 and 6 support the enzyme hypothesis?
9. Do your experimental results rule out the possibility that (a) starch converts unboiled
saliva to sugar or (b) starch and unboiled saliva combine chemically to form sugar?
10. The starch molecule consists of a long chain of carbon atoms with oxygen and
hydrogen atoms attached. A sugar, such as glucose, has molecules consisting of six
carbon atoms with oxygen and hydrogen atoms attached. Using this information, suggest
a way in which sugar could be formed from starch. What part would an enzyme play in
this reaction?
12. What were the control(s) in this experiment? Explain
13. After performing experiment and analyzing data do you agree or disagree with your
prediction? Why?
EXPERIMENT 2 – BELLY ACHE LAB-WHAT ARE THE FUNCTIONS OF
ENZYMES? ARE ENZYMES VERY SPECIFIC? CAN AN ENZYME BE
REUSED? WHAT IS THE AFFECT OF HIGH TEMPERATURE ON ENZYME
REACTIONS?
PREDICT: ARE ENZYMES VERY SPECIFIC? CAN ENZYMES BE REUSED?
_________________________________________________________________
Pre-lab:
Answer the questions below before starting experiment:
1. What type of carbohydrate is lactose? _______________________
2. What food group is lactose found in ? _______________________
3. What causes people to be lactose intolerant? _________________
4. What type of carbohydrate is sucrose? ______________________
Background:
Most foods that are ingested are composed of large, complex molecules. These molecules
are not useful to cells unless they are first broken down. In humans and many
multicellular animals, digestion is dependent on various enzymes produced by organs of
the digestive system. These enzymes convert large molecules into smaller molecules in a
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reasonable amount of time. Six months to digest a Hershey bar is not a reasonable
amount of time. Biurets is a test for proteins. In the presence of proteins Biurets goes
from blue to lavender.
Material:
Lactaid (contains the enzyme lactase), milk (contains the sugar lactose), sucrose,
Benedicts solution, Biuret solution, 7 test tubes, test tube rack, 2 500 ml beakers,
(1000 ml beaker with water on a hot plate in central location.)
PROCEDURE:
A. Preparing solutions
1. In a beaker, dissolve one Lactaid tablet in 0.2 liters (must convert to ml) of
distilled water.
2. Stir until dissolved and label beaker lactase (the enzyme in Lactaid).
3. In a second beaker, add 2000 milligrams (must covert to grams) of sucrose to
200,000 micro liters (must convert to ml) of distilled water.
4. Stir until dissolved and label beaker sucrose.
B. Testing for monosaccharides in milk
1. Place 500 micro liters (convert to ml) of milk in a test tube
2. Dilute with 1ml water so you can see the color of Benedicts solution
3. Add 5 drops of Benedicts solution
4. Boil test tube in boiling water and observe and record any color changes
C. Setting up and performing experiment
1. Label six test tubes as follows:
a. 1 ml of milk with 1 ml of lactase
b. 1 ml of milk with 1ml of water
c. 1ml of sucrose with 1ml of lactase
d. 1 ml of sucrose with 1ml of water
e. 1 ml of lactase
f. 2 ml of milk with 1 ml of lactase
2. Add the appropriate ingredients to each test tube and then add 1 ml of distilled
water to each tube.
3. Take only test tube e and boil for five minutes and then add 1 ml of milk & 1
ml of distilled water..
4. Let all six test tubes sit over night
5. Next day add 10 drops of Benedicts solution to each of the first five test
tubes (a-e BUT NOT F)
6. Boil test tubes a-e BUT NOT f and observe and record any color changes in
each test tube.
7. Take 1 ml from test tube f and add it to a new test tube and label it g.
8. Add 1 ml of fresh milk & 1 ml of distilled water to test tube g and let it sit
overnight.
9. Next day add 10 drops of Benedicts solution to test tube g.
10. Boil test tube g and observe and record any color changes.
D. Determining what organic compound of life enzymes are
1. Add 1 ml of distilled water to a test tube
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2. Add 3 drops of Biurets and record color
3. Add 1ml of Lactaid
4. Record your before and after color
5. Form a conclusion based on your data
6. After performing experiment and analyzing data do you agree or disagree with
your prediction? Why?
CREATE A DATA TABLE FOR YOUR RESULTS
EXPERIMENT 3 - ENZYME SALAD LAB-WHAT ARE THE FUNCTIONS OF
ENZYMES? WHAT AFFECT DOES HIGH TEMPERATURE HAVE ON
ENZYME ACTIVITY?
Pre-lab:
Answer the question below before starting experiment:
1. What organic compound is gelatin? _____________________
Background:
Have you ever noticed the warning on a Jell-O package not to use fresh or frozen
pineapple? The enzymes in some tropical fruit can break down proteins. For example,
meat tenderizer has the enzyme papain obtained from papaya fruit. Pineapple also has a
digestive enzyme, bromelin that breaks down protein.
Material:
4 test tubes, test tube rack, gelatin (100 ml/1 packet), canned pineapple crushed into juice,
fresh pineapple crushed into juice, meat tenderizer, ice water bath, graduated cylinder
Procedure:
1. Label four test tubes 1-4
2. Pour 10 ml of gelatin into each of the four test tubes
3. Place 3 ml canned pineapple juice into test tube #1
4. Place 3 ml fresh pineapple juice into test tube #2
5. Place meat tenderizer into test tube #3
6. Gently mix the contents in each test tube
7. Place all four test tubes in an ice water bath (may have to refrigerate over night)
8. Every few minutes check to see if the gelatin is setting in the tubes.
9. Record your observations
10. Form a conclusion based on your data
Analysis:
1. Explain how meat tenderizer softens meat. __________________________
_______________________________________________________________
________________________________________________________________
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EXPERIMENT 4-FUN WITH MILK & EGGS-WHAT ARE THE EFFECTS OF pH &
TEMPERATURE ON THE STRUCTURE OF PROTEINS (ENZYMES)?
Background
Most people think of milk as a liquid. Yes, it is a liquid, but milk is really a mixture of fat
and protein molecules in a watery solution.
As we discussed in class, proteins are large organic molecules that are built as a chain (or
polymer) of amino acids. The behavior and function of the protein is caused by the
specific amino acids that are linked together in the chain. These amino acids react with
each other and cause the protein chain to twist and fold up into a large 3-D shape,
forming a globular protein.
The “R” groups (or side groups) of each amino acid can be either hydrophobic (waterfearing) or hydrophilic (water-loving). When a section of the protein is made of
hydrophobic amino acids, it will cause that part of the protein to try to stay away from
water. When a section of the protein is made of hydrophilic amino acids, it will cause that
part of the protein to try to stay in water.
The protein molecules in milk, called caseins, are very hydrophobic. They try to get away
from the watery liquid of the milk, so they fold in on themselves a lot to hide from the
water. This folding makes the milk protein molecules into globules in the milk. You can’t
see them because even though they are large molecules, molecules are still too small to
see with the human eye.
Because pH (the acidity of a liquid) and high temperature both disrupt chemical bonds,
they can affect how a molecule forms or how it behaves. This is especially true for
proteins, since how they are shaped directly controls how well they function. When a
protein loses its 3-D shape and unravels back into a long chain, it is called “denaturing.”
We will investigate these concepts and effects in this lab.
PART A: SAY “CHEESE” AFFECT OF pH
Materials:
•
600 mL beaker
•
hot plate or water bath 37 C
• 10 mL of vinegar
•
stirring rod
•
500 mL of milk (4%)
PREDICT: What do you think will be the affect of lowering the pH on enzyme
activity? _________________________________________________________
Procedure:
1.Place milk in the beaker and warm on hot plate or water bath to a little above 37 C.
37°C is roughly body temperature, so the milk should be just hot to the touch.
2. Add vinegar and remove from hot plate. Stir very gently only once and then let sit.
Coagulation, or curdling, occurs quickly.
3.Drain excess liquid through a filter. This is the whey.
You may let the curds (the solids) sit in the filter (or cheese cloth) until well-drained, for
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a drier cheese.
4. Make observations and record
5. After performing experiment and analyzing data do you agree or disagree with your
prediction? Why?
PART B: EGGS-ACTLY WHAT IS THE AFFECT OF TEMPERATURE?
The protein in egg whites is called albumen. The albumen proteins are also long chains of
amino acids that are folded up onto themselves in a compact, 3-D, globular shape. Each
protein molecule is held in this shape by various kinds of bonds between different parts
of its chain.
The bonding that shapes the egg protein molecules is very easily disturbed by changes in
temperature (or pH or salt). These changes can cause the protein molecules to bond
together into a solid mass — to coagulate.
Materials:
• 400 mL beaker
•1 uncooked egg
•hot plate
•300 mL water
Procedure:
1.Heat water to almost boiling. Swirl water in a fast vortex.
2.Crack raw egg and drop into vortex of hot water and leave undisturbed.
3. Make observations of changes and record
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