Plant Pigments and Photosynthesis

AP BIOLOGY LAB 2
MR. BAMBINO PERIOD 1/2
Benjamin Dynkin and Brandon Marvisi
 Introduction
In this lab we learned how to separate plant pigments using chromatography, and measure the rate of
photosynthesis in isolated chloroplasts using the dye DPIP. When electrons are transferred during the light
dependant reactions of photo synthesis DPIP is reduced changing the color from blue to colorless.

Exercise Four A: Plant Pigment Chromatography

Distance Moved by Pigment Band

Band # Distance (mm) Band Color
1 5 Green
2 5 Yellow

R(f) for xanthophyll (yellow) = .5

R(f) for chlorophyll b (yellow green to olive green) = .5

1. What factors are involved in the separation of the pigments?

The factors that may be involved in the separation of the pigments include the size of the particles, the
solubility of the particles, and any affinity the particles may have for the paper.

2. Would you expect the Rf value of the pigment to be the same if a different solvent were used? Explain.
We wouldn’t expect the Rf value of the pigment to be the same if a different solvent were used, because
there would be different variables in the experiment.

3. What type of chlorophyll does the reaction center contain? What are the roles of the other pigments?
The reaction center in the plant contains chlorophyll a; several other pigments collect different light
waves (such as p680 and p700) and transfer the energy to chlorophyll “a”.

Exercise Four B: Photosynthesis and The Light Reaction

Cuvette 0 5 10 15
2 unboiled/Dark 89% 91% 91% 91%
3 Unboiled/Light 86% 89% 89% 96%
4 Boiled/Light 48% 51% 50% 50%
5 no cloroplasts 88% 92% 91% 91%

1. What is the purpose of DPIP in this experiment?

DPIP is an electron acceptor in this lab.

2. What molecule found in chloroplasts does DPIP "replace" in this experiment?

DPIP substitutes for the NADP molecules, which help to transport the electrons.

3. What is the source of the electrons that will reduce DPIP?

The source of the electrons that will reduce the DPIP will come from the photolysis of water.

4. What was measured with the spectrophotometer in this experiment?

The spectrophotometer is used to measure the percentage of light transmittance through the lab cuvette
due to DPIP reduction.

5. What is the effect of darkness on the reduction of DPIP? Explain.

Darkness will not allow for reaction to occur, as it can be seen in the difference in results between the
cuvette with the tinfoil around it compared to the other cuvettes.

6. What is the effect of boiling the chloroplasts on the subsequent reduction of DPIP? Explain.
The process of boiling the substance denatures the protein molecules of the substance, and stops the
reduction process occurring with the DPIP.

7. What reasons can you give for the difference in the percent transmittance between the live chloroplasts that
were incubated in the light and those that were kept in the dark?
In the cuvette with the tinfoil covering it, there was no available light energy to help facilitate
photosynthesis; so therefore, there was no flow of electrons and no photolysis of water, rendering the
chloroplasts useless. However, the uncovered cuvette had chloroplasts where photosynthesis and photolysis
were thriving.

Conclusion:

The lab was designed to teach us about photosynthesis. During this lab we did plant pigment
chromatography to learn how to separate the plant’s pigments and we learned what changes in transmittance
when the different chloroplasts are used, boiled/ unboiled, and when there are variances in the light exposure.
We also learned about the proper procedure in doing photosynthesis incubation as well as chromatography.
Also if the spectrophotometer is not set correctly (at zero) then it will affect our readings. In addition to faulty
readings, the experimenters may have messed up the experiment by not following the procedures exactly. The
timing may have gotten off which could alter the data as well. After data was compared we were able to
understand the basics of photosynthesis such as darkness will not allow for reaction to occur. All these things
helped us reach our objective of the lab, to understand photosynthesis.