Name: _____________________________________ Class: ________________ Date: ____________

Effect of pH on Pepsin
Pepsin is an enzyme that catalyzes the breakdown of large protein and polypeptide molecules into smaller peptides and amino acids. This reaction is called hydrolysis, because a molecule of water is needed to break each peptide bond.

Pepsin is secreted by chief cells in the stomach. As an enzyme, pepsin is sensitive to many environmental factors, such as temperature and pH. The purpose of this lab is to determine the optimal pH for pepsin to cleave the peptide bonds within a sample protein.

Materials
Boiled egg white (protein) Four test tubes Graduated cylinder Digital Scale

Pre-Lab Questions
1. What exactly is pH a measurement of?

2. The pH scale runs from 0-14. Give a sample pH that you would expect from a strong acid, weak acid, strong base, weak base, and perfectly neutral solution.

3. Why must proteins be broken down into amino acids in the digestive system, and not simply absorbed whole?

4. Hypothesis: Given what you know about the human digestive system, what pH do you predict will be ideal for hydrolysis of protein by the pepsin enzyme?

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Name: _____________________________________ Class: ________________ Date: ____________

Procedure
1. Select four test tubes. Fill each about half-way with one of the different solutions of HCl or distilled water. Mark the pH on each test tube. 2. Take a piece of egg white and measure the mass. Try to keep the mass as close to 1.0 grams as possible. 3. Add the piece of egg white to one of the test tubes. Record the pH and mass in your data table. 4. Repeat for the other three test tubes and pH solutions. 5. Allow the egg proteins to sit in the solution overnight. 6. Drain off as much of the liquid in each of the test tubes as possible. Remove the remaining egg white and place it on a paper towel to dry. 7. Weigh each egg white, and record in your data table.

Data
Table 1 - Effects of pH on Hydrolysis of Egg Protein (Individual Data)

pH

Mass of Protein Before Reaction (g)

Mass of Protein After Reaction (g)

Mass Change (g)

Percent Change

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Name: _____________________________________ Class: ________________ Date: ____________

Table 2 - Effects of pH on Hydrolysis of Egg Protein (Class Data)

pH

Mass Change (g)

Percent Change

pH

Mass Change (g)

Percent Change

pH

Mass Change (g)

Percent Change

pH

Mass Change (g)

Percent Change

Table 3 - Class Average Egg Protein Mass Change and Percent Change

pH

Mass Change (g)

Percent Change

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Name: _____________________________________ Class: ________________ Date: ____________

Figure 1- Bar Graph of Egg Protein Average Percent Change vs. pH

Post-Lab Questions
8. Conclusion: Do the results support your hypothesis? Be specific.

9. Is it possible that the presence of acid itself was partially responsible for the breakdown of the egg protein? What experiment could you do to answer this question?

10. The chief cells of the stomach actually release an inactivated form of pepsin, called pepsinogen. The enzyme is not activated until it is exposed to hydrochloric acid released by the parietal cells of the stomach. Why is the enzyme is not simply active all the time, like amylase in the saliva?

11. Pepsin becomes inactive again when it leaves the stomach and enters the duodenum. What chemical change occurs in the duodenum to cause this inactivation?

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