Lab 4: Plant Pigments and Photosynthesis
Introduction:
State the purpose of part one to separate plant pigments using chromatography.
State the principles that allow pigments to move up. Predict which colors you would see
closest to the solvent and which farthest from the front and why they would be in that
order. (Chlorophylls, xanthophylls, Beta carotene) State which pigments you saw and
the order they appeared in relation to the solvent front.
State the purpose of part two to measure the rate of photosynthesis in isolated
chloroplast using the dye DPIP. State the predictions that were made for the three tested
conditions (unboiled dark, unboiled light, boiled light). State the actual results that were
found in the lab.
Procedure:
In paragraph form summarize the procedure for Part 1 found on page 46 of your
lab.
In paragraph form summarize the procedure for Part II found on page 49 & 50 of
your lab.
Data and Results
Data Table 1: Expected Results Distance Moved by Pigment Band (millimeters)
Band Number
Distance (mm)
Band Color
1
17
Yellow green
38
2
Olive green
61
3
Yellow
85
4
Yellow
178
5
yellow
Distance Solvent Front Moved for Expected Results: 190 mm
Data Table 2: Rf Values for Expected Results
Rf value (mm)
Pigment
Yellow to Yellow (Xanthophyll)
Yellow Green to Olive Green (Chlorophyll b)
Data Table 3: Individual Data Distance Moved by Pigment Band (millimeters)
Band Number
Distance (mm)
Band Color
1
2
3
4
5
Data Table 4: Rf Values for Individual Results
Rf value (mm)
Pigment
Yellow to Yellow (Xanthophyll)
Yellow Green to Olive Green (Chlorophyll b)
Bright Green to Blue Green (Chlorophyll a)
Analysis Questions
1. What factors are involved in the separation of the pigments?
2. Would you expect the Rf value to be the same if a different solvent were used?
Explain.
3. What type of chlorophyll does the reaction center contain? What are the roles of the
other pigments?
Data Table 5: Expected Data % Transmittance of Light under Various Conditions
Cuvette
0
5
10
15
Unboiled/Dark
31.3
32.5
35.5
34.8
Unboiled/Light
32.7
54.5
63.7
65.1
Boiled/Light
32.7
32.9
33.1
32.5
No
31.3
31.3
31.3
31.3
Chloroplast/Light
Data Table 6: Individual Data % Transmittance of Light under Various Conditions
Cuvette
0
5
10
15
20
Unboiled/Dark
Unboiled/Light
Boiled/Light
No
Chloroplast/Light
Graph 1: % Transmittance of Light for Expected Class Data
X Axis: Time (mm)
Y Axis: % Transmittance
Key
: unboiled/Dark
: unboiled/Light
: boiled/ Light
: no chloroplast
Graph 2: % Transmittance of Light for Individual Class Data
X Axis: Time (mm)
Y Axis: % Transmittance
Key
: unboiled/Dark
: unboiled/Light
: boiled/ Light
: no chloroplast
Analysis Question
1. What is the function of the DPIP in this experiment?
2. What molecule found in chloroplast does DPIP “replace” in this experiment?
3. What is the source of the electrons that will reduce DPIP?
4. What was measured with the spectrophotometer in this experiment?
5. What is the effect of darkness on the reduction of DPIP? Explain.
6. What is the effect of boiling the chloroplasts on the subsequent reduction of DPIP?
Explain.
7. What reasons can you give for the difference in the percentage of transmittance
between the live chloroplasts that were incubated in the light and those that were kept in
the dark?
8. Identify the function of each of the cuvettes:
a. Cuvette1:
b. Cuvette 2:
c. Cuvette 3:
d. Cuvette 4:
e. Cuvette 5:
Conclusion:
Paragraph 1: Summarize the purpose and predictions for part 1 and part 2. Summarize
the results found (briefly).
Paragraph 2: Discuss the results of plant pigment chromatography. Which pigments
were you able to identify and measure? How did your pigment Rf values compare to the
expected pigment values? Explain why your pigments traveled different distances. What
properties make pigments move different distances?
Paragraph 3: Discuss the results of photosynthesis/light reaction. Which cuvette had the
highest rate of photosynthesis? How did this compare to your prediction and to the class
data? Explain which cuvette should have the highest rate of photosynthesis and why.
Why should the boiled cuvette not do anything? Why should the dark cuvette not do
anything? Why did we have a control? If your control changed (no chloroplast) how can
you explain this?
Paragraph 4: Discuss errors that happened in the lab. What factors may have affected
errors in the chromatography lab? What factors may have affected errors in the
photosynthesis lab? How can you improve this next time?
Paragraph 5: Make a new test for this experiment. Change something in the experiment
or design a more in-depth experiment that you can test. What would you design?
Explain your tests.
Paragraph 6: Relate this lab to the major theme of AP Biology of ENERGY
TRANSFER. How does the principle of photosynthesis relate to this? Why is
photosynthesis so important and why would it be important to measure the rate of
photosynthesis?