Digestion and Enzyme Activity
Background:
The digestive system is made up of the digestive tract – a series of hollow organs joined in a
long, twisting tube from the mouth to the anus- and other organs that help the body break down and
absorb food. Organs that make up the digestive tract are the mouth, esophagus, stomach, small
intestine, large intestive – also called the colon – rectum, and anus. Two accessory digestive
organs, the liver and pancreas, produce digestive juices that reach the intestines through small
tubes called ducts. The gallbladder stores the liver’s digestive juices until they are needed in the
intestine.
The digestive glands that act first are in the mouth – the salivary glands. Saliva produced by
these glands contains an enzyme called amylase that begins to digest the starch from food into
smaller molecules. An enzyme is a substance that speeds up chemical reactions in the body.
The next set of digestive glands is in the stomach lining. These produce hydrochloric acid
and an enzyme called pepsin that digests protein. A thick mucus layer coats the mucosa and helps
keep the acidic digestive juice from dissolving the tissue of the stomach itself. After the stomach
empties the food and juice mixture into the small intestine, the juices of two other digestive organs
mix with the food. One of these organs, the pancreas, produces a juice that contains a wide array of
enzymes such as amylase to break down the carbohydrate, lipase to break down fat, and trypsin to
break down protein.
The second organ, the liver, produces yet another digestive juice—bile. Bile is stored
between meals in the gallbladder. At mealtime, it is squeezed out of the gallbladder, through the
bile ducts, and into the intestine to mix with the fat in food. The bile acids dissolve fat into the
watery contents of the intestine, much like detergents that dissolve grease from a frying pan.
Purpose: The goal of this lab will be to observe how digestive enzymes are able to break
down macromolecules and how environmental changes affect their functioning.
Digestion and Enzyme Activity
Materials:
Spot Plate
10 Stirring Sticks
11 pH Strips
Water
Corn Oil
Lipase
Dishsoap
Starch Solution
Amylase
Potassium Iodine
Albumin
HCl
Pepsin
Biuret
Boiled Amylase
Procedure:
+++ All data tables are on Pg 3+++
PART A: Fat Digestion
The corn oil represenst the lipid polymer, and lipase is the enzyme responsible for breaking the
lipid down. A change in pH will demonstrate that the enzyme is active. The soap works similarly
to bile, and will break up the oil so lipase can work more easily.
1.
2.
3.
4.
2. Add 10 drops of water to wells 1-3.
Add 3 drops of Corn oil to wells 1-3.
Use separate stirring sticks to mix each well thoroughly.
Use the forceps to dip a separate pH strip into each of the wells 1-3.
Compare the color change of each strip to the pH strip chart to determine
the pH of the mixture in each well. Record your results in Data Table 1.
5. Add 5 drops of lipase to well 2 and 3.
6. Add 2 drops of liquid soap to well 3.
7. Re-stir the mixture in each well, and allow them to sit for 20 minutes.
8. Retest the pH and record your results in Data Table 1.
9. Record your observations of each of the mixtures in Data Table 1.
10.
Rinse out your spot plate and dry it off with a paper towel.
PART B: Carbohydrate Digestion
In part B starch is a polysaccharide, and amylase is the enzyme responsible for breaking starch
down into monosaccharides. Since the potassium iodine tests for starch, you will be looking for
a negative test to determine whether amylase actually broke down the starch.
1.
2.
3.
4.
Add 15 drops of 1% Starch solution to wells 1-3.
Add 5 drops of 2% Amylase solution to well 2.
Add 5 drops of 2% Boiled Amylase solution to well 3
Use separate stirring sticks to mix each well, and allow them to sit for 5-7
minutes.
5. Add 1 drop of Potassium iodine to wells 1-3. A positive test for starch will turn
dark blue- black. If a well is positive for starch, put a + in the Data Table 2. If
the test is negative for starch put a –.
6. Record your observations of each of the mixtures in Data Table 2.
7. Rinse out your spot plate and dry it off with a paper towel.
PART C: Protein Digestion
Digestion and Enzyme Activity
In part C the albumin represents a polypeptide protein. Pepsin is the enzyme needed to break
down the albumin into amino acids, but can only work within a certain pH range. The Biuret
will turn pink with smaller protein chains and will turn purple in the presence of large
proteins.
1.
2.
3.
4.
5.
6.
7.
8.
Place 5 drops of 2% Albumin in wells 1-4.
Add 5 drops of 1% HCl to wells 2 and 4
Add 5 drops of 3% Pepsin to wells 3 and 4.
Use separate stirring sticks to mix each well, and allow them to sit for 5-7
minutes.
Use the forceps to dip a separate pH strip into each of the wells 1-4.
Compare the color change of each strip to the pH strip chart to determine
the pH of the mixture in each well. Record your results in Data Table 1. I
will provide you with the data for HCl
Add 2 drops of Biuret to wells 1-4. A positive test for protein breakdown will
turn pink. If the well is positive, put a + in Data Table 1. If the well is
negative put a -.
Record your observations of each of the mixtures in Data Table 1.
Rinse out your spot plate and dry it off with a paper towel.
++++ Clean up your area and complete the analysis questions for all parts ++++
Data Tables:
TABLE 1
Well
Contents
1
Oil + Water
2
3
Initial pH
Observations
Oil + Water +
Lipase
Oil + Water +
Lipase + Soap
TABLE 2
Well
1
Contents
Starch
2
Starch + Amylase
3
Starch + B. Amylase
TABLE 3
Well
Contents
1
Albumin
2
Final pH
Albumin + HCl
Starch Test
pH
Protein Test
Observations
Observations
Digestion and Enzyme Activity
3
Albumin +
Pepsin
Albumin +
Pepsin + HCl
HCl
4
5
Analysis:
Answer on a separate sheet of paper
PART A:
1. How does the pH of each solution show that fat digestion has occurred?
2. Which well showed the greatest degree of fat digestion? Explain.
3. What effect would fat digestion have on the pH of the surrounding solution?
Explain.
4. What do you predict would happen to the pH of a corn oil solution if we added
Pepsin. Explain.
PART B:
1. What are the subunits that make up carbohydrates?
2. How could you tell that carbohydrate digestion had taken place?
3. What do you think may happen if HCl was added to the starch + amylase
mixture? Explain your answer.
4. What do you predict would happen to a starch solution if we added lipase?
(i.e.What would be the results of an iodine test?)
PART C:
1. What are the subunits that make up a protein?
2. How could you tell that protein digestion took place?
3. Which of the wells showed the greatest degree of protein digestion? Why?
4. Does the effectiveness of pepsin depend on pH? Explain.
5. What can you conclude about the optimal pH for pepsin?
6. What do you predict would happen to an albumin solution if we added amylase?
(i.e. What would be the results of a Biuret test?)
Digestion and Enzyme Activity
Conclusion:
Explain, in detail and with examples, how this experiment demonstrates that enzymes
are highly specific to both substrates and environmental conditions.