Even small changes in pH can cause distress to organisms. The changes in hydrogen ion concentration in cells can change the
rates of some chemical reactions. For example, the pH of human blood is normally about 7.4 and must be in the range of 7.0-7.8
for survival. If the pH is in the 7.0-7.3 range the person will feel tired, have trouble breathing, and may even be disoriented. If
the pH of the blood is in the 7.5-7.8 range, the person will feel dizzy and rather agitated.
It is very important for organisms to be able to maintain a fairly constant internal environment (homeostasis). To prevent the
hydrogen ion concentration of the cytoplasm from changing too much, cells have chemical compounds called “buffers” that will
bind with hydrogen ions when their concentration increases too much. Buffers can also release bound hydrogen ions when their
concentration in the solution decreases too much. In our blood stream, carbonic acid (H2CO3 ) acts as the buffer that maintains
our blood pH within a normal range.
H2CO3 <----------> (HCO3)-
Products such as Alka Seltzer take up hydrogen ions and reduce the acidity of the stomach fluids. They consist of sodium
bicarbonate, citric acid, and salicylate analgesic. When placed in water, sodium citrate and carbon dioxide are produced.
Sodium citrate is a strong buffer.
In this laboratory, you will investigate how several materials respond to the addition of an acid and a
base to determine whether living materials have buffering capacity.
You will follow a generic procedure using a variety of biological materials and other substances, then try to figure out the
explanation for the results. Typically, when acid is added to water, the pH will drop quickly and then level out. Conversely, when
base is added to water, the pH will rise quickly and then level out. When acid or base is added to buffer, the pH barely changes.
Before you start, hypothesize about the buffering ability of each substance to be tested. List (on your paper, as Hypotheses) the
substances that will have the best capacity to act as buffers, moderate buffering capacity, and poor or no buffering ability
pH probe & LabPro setup
dH2O in wash bottle
50 mL container
30 mL graduated syringe
0.1 M NaOH in dropper
0.1 M HCl in dropper bottle
pH 4 buffer solution
Alka Seltzer solution
You will measure the changes in pH resulting from adding acid and base to plain tap water, each of the homogenates, the pH 4
buffer solution, and the Alka Seltzer solution. Make sure you record all of your pH measurements in your data charts.
1. Pour 25 mL of test substance into the 50 mL beaker. Measure the pH of the solution following directions for using the
LabPro with pH probe. Record in Table 1.
2. Add 0.1 M HCl one drop at a time, swirling after each drop. After you have added 5 drops, measure the pH again.
Record in Table 1.
3. Repeat step 2, recording the pH after each 5 drops, until a total of 30 drops have been added.
4. Rinse the beaker well and add another 25 mL of test substance. Rinse the pH probe with distilled H2O.
5. Follow steps 2-3 again, but this time use 0.1 M NaOH and record in Table 2.
6. Repeat the whole procedure (steps 1-5) for each of the test solutions.
7. Graph your results using Microsoft Excel. Use separate graphs for the response to HCl and the response to NaOH.
Laboratory Investigation: BIOLOGICAL BUFFERS
How do Biological Materials Respond to Acids and Bases?
Table 1: The pH of various substances after adding HCl
Table 2: The pH of various substances after adding NaOH
Calculate the average change of the acid and base trials combined. You do not need to graph this data.
Potato Egg White Yeast Broccoli
Analysis Questions – Type up your responses and hand in with your printed graph.
1. Examine your graphs. Are the responses of the biological homogenates to the addition of acid and base more similar to
the responses of the tap water or to the responses of the buffer solutions? Explain your observations.
2. Which of the biological homogenates (not tap water or pH 4 buffer) has the most buffering capacity? How did you
determine this? Why do you think this substance buffers so well?
3. Which of the biological homogenates (not tap water or pH 4 buffer) has the worst buffering capacity? How did you
determine this? Why might this substance have such a poor buffering ability compared to the other biological
4. What did you learn about the ability of living systems to regulate pH?
5. Explain why buffers are important to living systems. Give examples to support your ideas.
Test Substance 0 drops HCl 5 drops HCl 10 drops HCl 15 drops HCl 20 drops HCl 25 drops HCl 30 drops HCl
Buffer pH 4
Buffer pH 4
PROCEDURE – LAB PRO USE FOR “BIOLOGICAL BUFFERS”
1. Obtain and wear goggles and lab aprons.
2. Open the Vernier LoggerPro software on the computer.
3. Attach the pH probe to the interface. Connect the interface to the computer.
Logger Pro should auto-recognize the pH probe. If it does not, let your
4. Change the data table. Do this by following the next five steps.
a. Click on “Experiment”.
b. Click “Data Collection”.
c. Click the drop-down menu next to “Mode” and choose “Event with Entry”.
d. Change the column name to “Drops”.
e. Click “OK”. The data table should now display “Drops” instead of seconds.
Testing the effect of acid and base on water
5. Before each use of the pH probe, you need to rinse the tip of the electrode thoroughly with distilled
water. To do this, hold the pH electrode above a sink and use the rinse bottle to thoroughly rinse
the electrode tip.
Important: Do not let the pH electrode dry out. Keep it in a 250-mL beaker with about 100 mL of tap
water when not in use. The tip of the probe is made of glass—it is fragile. Handle with care!
6. Obtain 25 mL of the sample to be tested.
7. Rinse the pH probe thoroughly with distilled water, then place it into the beaker to be tested.
8. Click to begin making pH measurements.
9. Before you begin, determine the initial pH of the solution. Click , then type “0” in the text box
and press ENTER.
10. Add acid or base to the solution. Stir each solution thoroughly after addition. CAUTION: Handle the
hydrochloric acid with care. It can cause painful burns if it comes in contact with the skin. Sodium
hydroxide solution is caustic. Avoid spilling it on your skin or clothing.
11. When the pH readings are stable click . Enter the total number of drops of acid or base you
have added to the water in the beaker. Type “5” in the text box and press ENTER.
12. Repeat Steps 8 through 9, adding 5 drops at a time until each team has added a total of 30 drops.
13. Click when you have added a total of 30 drops.
14. Rinse the pH probe thoroughly and place the probe into the beaker of tap water.
15. Follow the procedures above for all the homogenates to be tested as assigned by your teacher.
16. Put your data on the class chart at the front of the room.
17. Clean up your lab station appropriately, paying special attention to properly putting away the
Vernier equipment and laptop.