Enzyme Labs 1-3
Background Information
Name:
Date:
Period:
Pineapples
Pineapple’s lush, tropical sweetness is reason enough to enjoy it any way you can, but
this fruit also contains vitamin C and manganese. This fruit’s most promising nutritional
asset, though, may be bromelain, a natural enzyme found in both the fruit and the stem.
Most of the pineapple consumed in the United States is canned (in the form of juice as
well as fruit), but fresh pineapple is much more flavorful, and, despite its tough bristly
shell, is easy to prepare.
The fruit probably first grew wild in parts of South America and then spread to the
Caribbean, where Columbus encountered it. By 1600, early European explorers had
carried pineapples as far as China and the Philippines. In the 18th century, pineapples
were taken to the Hawaiian Islands, eventually becoming the major fruit crop. Hawaiian
pineapple producers were the first to can the fruit.
Bromelain
The pineapple plant contains protein-digesting enzymes called, as a group, bromelain.
In the health world, these enzymes are regarded as useful in reducing muscle and tissue
inflammation (hence the joint pain and wound-healing possibilities), as well as acting as
a digestive aid. In the cooking world, on the other hand, bromelain is regarded as the
enemy of the gelatin dessert. If you use fresh pineapple in gelatin, the enzyme eats the
protein and the gelatin will not gel—in fact bromelain is measured in units called GDU,
or gelatin digesting units. The classic kitchen trick for getting around this pineapplegelatin incompatibility is to cook the pineapple, thus reducing the power of the
bromelain.
Recipes that would highlight the benefits of bromelain start with fresh pineapple (which
has two to three times the amount of bromelain as canned pineapple does), and is then
subjected to as little heat as possible.
Bromelain is used in meat tenderizers, in hill-proofing beer, manufacturing precooked
cereals, in certain cosmetics, and in preparation to treat edema and inflammation.
Gelatin
Gelatin, a familiar, ingredient in cooking, is obtained by boiling the skin, tendons, and
ligaments of animals. As a result, it contains protein called collagen (a primary
component of joints, cartilage, and nails), and various amino acids (histidine, lysine,
leucine, tryptophan, and valine, to name a few). Remember: amino acids are the
building blocks of proteins.
Gelatin has long been a key ingredient for providing support for “jelled” deserts, salads,
frozen drinks, and soft candies such as Gummi Bears. (In fact, the word gelatin is
derived from the Latin “gelatus”, meaning stiff or frozen.)
Scientists have been studying gelatin for centuries. It has no smell or taste of its own,
adapting to whatever it is added to. During the Napoleonic Wars, the French, desperate
for nutrition sources during the English blockade, reportedly first turned to gelatin as a
source of protein (albeit a weak one). Gelatin began its long run as a popular
consumable, however, in the 1890’s, when it was first developed and then heavily
promoted as a commercial product by Charles Knox, founder of the Knox Gelatin
Corporation.
In addition to its famous “jiggly” food uses, gelatin with its flexible, dissolvable
structure is also used to manufacture capsules (both hard and “soft-gel”) to hold
medications, vitamins, and other dietary supplements. It also has a range of industrial
and medical engineering applications: Gelatin is an ingredient in film coatings, medical
devices such as artificial heart valves, and in specialized meshes used to repair wounds,
to name a few.
Collagen
About one quarter of all the protein in your body is collagen. It is a major structural
material that forms molecular cables to strengthen the tendons and resilient sheets that
support the skin and internal organs. Adding mineral crystals to collagen makes Bones
and teeth. Collagen provides structure to our bodies, protecting and supporting the softer
tissues and connecting them with the skeleton. But, in spite of its critical function in the
body, collagen is a relatively simple protein.
Collagen from livestock animals is a familiar ingredient in cooking. Collagen is a
protein, and like most proteins, when heated, it loses all of its structure. The polymer
molecule unwinds. Then, when the denatured mass cools down, it soaks up all of the
surrounding water like a sponge, forming gelatin.
If you've ever eaten in a cafeteria, chances are good that your dessert options included
Jell-O. There are hundreds of different desserts that use Jell-O to create everything from
your basic institutional-style gelatin square to ornate designs that incorporate varied
Jell-O flavors, fruit, and whipped toppings. Jell-O consists of five basic ingredients:
Gelatin
Water
sugar/artificial sweetener
Artificial flavors food coloring
The gelatin in Jell-O is what lets you transform it into all sorts of different shapes.
What exactly is gelatin? Gelatin is just a processed version of a structural protein called
collagen that is found in many animals, including humans. Collagen actually makes up
almost a third of all the protein in the human body. It is a big, fibrous molecule that
makes skin, bones, and tendons both strong and somewhat elastic. As you get older,
your body makes less collagen, and individual collagen fibers become cross-linked with
each other. You might experience this as stiff joints (from less flexible tendons) or
wrinkles (from loss of skin elasticity).
The gelatin you eat in food products comes from the collagen in cow or pig
bones, hooves, and connective tissues. To make gelatin, manufacturers grind up these
various parts and pre-treat them with either a strong acid or a strong base to break down
cellular structures and release proteins like collagen. After pre-treatment, the resulting
mixture is boiled. During this process, the large collagen protein ends up being partially
broken down or denatured, and the resulting product is called gelatin. The gelatin is
easily extracted because it forms a layer on the surface of the boiling mixture.
Terms, Concepts, and Questions to Start Background Research
Gelatin, Protein, Collagen, Protease, Bromelain, Papain, and Protein denaturation.
Side Questions that will help you create a strong introduction
• What is gelatin?
• How does gelatin solidify?
• What does proteases do?
• Which fruits contain papain, bromelain, or another protease?
• Why are Papain and bromelain often called natural “meat tenderizers?” Explain
in detail.
• What are commercial meat tenderizers, like the ones in the spice aisle of the
grocery store, made out of?
The purpose of these lab experiments is to be to answer the three questions below.
1. What form of pineapple juice contains enzymes that digest protein?
2. What effect does temperature have on an enzyme?
3. What effect does pH have on enzyme activity?
Enzyme Pre-Lab
Name ___________________________Date __________ Period____
Directions: Read the background material provided to you. Using this material,
your notes, and text, answer the following questions.
1. By definition, what is an enzyme?
_________________________________________________________________
_________________________________________________________________
2. (a) Describe what happens when a protein denatures.
_________________________________________________________________
_________________________________________________________________
(b) Is the denatured protein still able to function?
_________________________________________________________________
_________________________________________________________________
3. (a) What is the name of the enzyme we are using in this lab?
_________________________________________________________________
(b) What fruit is it found in?
4. (a) What is the protein that creates the protein we are using in this lab?
_________________________________________________________________
(b) For this lab, what is our food source for the protein?
____________________________________________________________________
5. In your own words, describe what happens to collagen when it is heated.
_________________________________________________________________
_________________________________________________________________
_________________________________________________________________
_________________________________________________________________
6. Read the procedure for Enzyme Lab 1. Make a hypothesis regarding which of the
three forms of pineapple (fresh pineapple, canned pineapple, concentrated
pineapple juice) will allow the gelatin to become a semi solid and which will
remain a liquid when you examine your results on Day 2.
What form of pineapple juice contains enzymes that digest protein?
_________________________________________________________________
_________________________________________________________________
7. Why is water used in Enzyme Lab 1?
________________________________________________________________
Read the directions for Enzyme Lab 2. Make a hypothesis as to the nature of the
effect that temperature will have on the contents of each test tube at Day 2, which
test tube’s contents will be solid and which will be liquid? (Higher or Lower
temperatures?)
What effect does temperature have on an enzyme?
________________________________________________________________
_________________________________________________________________
8. Why do you hypothesize this?
_________________________________________________________________
_________________________________________________________________
Read the procedure for Enzyme Lab 3. Make a hypothesis as to which test tube’s
contents (A, B, or C) will be liquid at Day 2 and which will be solid.
What effect does pH have on enzyme activity?
9. _________________________________________________________________
_________________________________________________________________
10. Why did you hypothesize this?
_________________________________________________________________
_________________________________________________________________
Enzyme Lab 1
What form of pineapple juice contains enzymes that digest protein?
Materials:
For every 3 groups
1 envelope Knox gelatin
Measuring cup
1 spoon
100 mL graduated cylinder
For each group
4 test tubes
1 test tube rack
4 (1mL) disposable pipettes
10ml graduated cylinder
Marking pen
For entire class
water
pineapple juices (below)
hot plate to heat water
Procedure:
1. Number and label the test tubes “1-4” and use the initials of a group member to identify
your group.
2. Find 2 other groups and prepare 1 package of gelatin in the measuring cup using 90.0
mL of boiling water and 30 mL of cold water. Stir well with a spoon until the gelatin is
dissolved.
3. Place 3 mL of the designated pineapple juice into each test tube. Use a separate pipette
for each type of juice. Failure to do so may result in mixing of the juice types and
inaccurate results.
o Tube 1: water only
o Tube 2: fresh pineapple juice
o Tube 3: Canned pineapple juice
o Tube 4: Concentrated pineapple juice (thawed)
4. Add 10 mL of gelatin mixture to each test tube. Shake well to ensure proper mixing and
place your samples in the refrigerator overnight using a test tube rack.
5. On Day 2, check the contents of each test tube for solidification of the contents and
record your observations.
Enzyme Lab Data Sheet
Enzyme Lab 1
Test Tube
Juice
1
Water
2
Fresh Pineapple Juice
3
Canned Pineapple Juice
4
Concentrated Pineapple
Juice
State of test tube contents on Day 2
1. Was your hypothesis supported by the data? Why or why not?
2. What caused the contents in the test tube to not gel and why?
Enzyme Lab 2
What effect does temperature have on an enzyme?
Materials:
For every 2 groups
For each group
For entire class
1 envelope Knox gelatin
6 test tubes
water
Measuring cup
1 test tube rack
pineapple juices (below)
1 spoon
1 (1mL) disposable pipette
100 mL graduated cylinder 10mL graduated cylinder
Marking Pen
Hot plate
Procedure:
1. Prepare the Knox gelatin by mixing 1 package of gelatin in the measuring cup using 90.0 mL of
boiling water and 30.0 mL of cold water. Stir well with a spoon until the gelatin is dissolved.
2. Number the test tubes from 1-5. Label the remaining test tube as “RT” (room temperature).
3. Each group will be assigned their own temperature gradient ranging from 40◦C – 100 ◦C. Each
group must record the test tube numbers and the corresponding test temperatures for each test
tube. Record the temperature of the room for the temperature for the “RT” test tube.
4. Next, add 3.0 mL of pineapple juice to each test tube.
5. Then heat each test tube to the appropriate temperature as assigned. Leave the test tube “RT” at
room temperature. (HINT: start with all the test tubes in cool water in a glass beaker water
bath. Gradually increase the temperature withdrawing the numbered test tubes in 2◦C
increments in order as the appropriate temperature level in the bath is reached.)
6. After the test tubes have been pulled from the water bath, add 10 ml of Knox gelatin (prepared
in step 1) to each test tube and mix well.
7. Finally, place the test tubes in the refrigerator overnight.
8. On day 2, check each test tube for solidification or liquidity of the contents and record your
observations.
Enzyme Lab 2
Test Tube Temperature
1
2
3
4
5
Room Temperature
6
Higher temp. Close to 100oC
State of test tube contents on Day 2
(Possible descriptions)
Firm Gel
Semi firm Gel
Loose Gel
Thick liquid (pulp trapped in thick liquid)
Loose liquid (pulp can move slowly)
All liquid (pulp moves freely)
3. Graph the results of class date to help determine when the gelling process was
affected. Was your hypothesis supported by the data? Why or why not?
4. What caused the contents in the test tube to not gel and why?
Enzyme Lab 3
What effect does pH have on enzyme activity?
Materials:
For every 3 groups
1 envelope Knox gelatin
Measuring cup
1 spoon
100 mL graduated cylinder
For each group
3 test tubes
1 test tube rack
4 (1 mL) disposable pipettes
10 mL graduated cylinder
Marking pen
For entire class
goggles
apron
water
pineapple juice
1M HCl
1M NaOH
Procedure:
1. Prepare the Knox gelatin by mixing 1 package of gelatin in the measuring cup
using 90 mL of boiling water and 30 mL of cold water. Stir well with a spoon
until the gelatin is dissolved.
2. Label 1 test tube “A” for acid, 1 test tube “B” for base, and the last test tube “C”
for control.
3. Place 3 mL of pineapple juice into each of the labeled test tubes. Transfer 1 mL of
base, 1 mL acid, and 1 mL of water into the appropriate test tubes of pineapple
juice. HINT: Use a different pipette for each test tube to avoid contamination.
4. Add 10 mL of gelatin mixture to each test tube. Mix well, being careful to not get
any of the acid or base on your skin.
5. Refrigerate the test tubes overnight and on day 2 check each test tube for
solidification or liquidity of the contents. Record your observations.
Enzyme Lab 3
Test Tube Contents
A
Acid
B
Base
C
Control (water)
State of test tube contents on Day 2
5. Was your hypothesis supported by the data? Why or why not?
6. What caused the contents in the test tube to not gel and why?
Graph
The independent variable is temperature at which the gelatin was subjected to and the
gelatin’s ability to gel is the dependent variable.
Possible terms to place on the y-axis, assign numbers to the descriptions and then you
will have two numerical values to graph against each other.
2-Firm Gel
4-Semi firm Gel
6-Loose Gel
8-Thick liquid (pulp trapped in thick
liquid)
10-Loose liquid (pulp can move slowly)
12-All liquid (pulp moves freely)
Create your Data Chart
Temperatures place on the x-axis
Temperature (oC)
20
30
40
50
60
70
80
90
100
Gel description # value on y-axis
12
10
8
6
4
2
http://en.wikipedia.org/wiki/Bromelain
Temperature stability[edit]
At the optimum temperature, the enzyme (bromelain) acts the fastest, but (at least the fruit variant) is
destroyed within few minutes. After an hour at 50 °C (122 °F), 83% of the enzyme remains, while at 40 °C
(104 °F), practically 100% remains.[9] As a result of this, the optimum temperature for maximum cumulated
activity over time is 35-45 °C. At room temperature, the enzyme can survive at least a week even under
multiple freeze-thaw cycles.[10]