Plant Pigments and Photosynthesis

By: Laura Cameron, Sarah Soppe, and Zahra Naseer

Purpose for 4a; Chromatography

The purpose of the lab was to separate the various pigments in spinach leaves through the process of chromatography.

Class Data for 4a

Pigment Distance Traveled Rf Value

Carotene 12 cm 0.923

Xanthophyll 2 cm 0.154

Chlorophyll a 7 cm 0.538

Chlorophyll b 5 cm 0.385

Distance Solvent Front Moved: 13 cm

Presentation of Data for 4a

Purpose for 4b; Photosynthesis

The purpose of the lab was to identify the impact of boiling chloroplasts and the impact of darkness on the rate of photosynthesis of chloroplasts extracted from spinach leaves using a spectrometer.

Set Up of the Lab

IV: The type of chloroplasts (boiled or unboiled) and the amount of light during incubation

DV: The transmittance of the cuvettes

Levels of IV: Boiled, unboiled, light, or dark

Number of Trials: 4 trials using the class data set

Constants: - The time in which the chloroplasts in the light are placed in front of the lamp

- The heat transferred to the cuvettes was limited by the tank in front of the cuvettes

- The same blank cuvette was used to calibrate the spectrometer before each reading

Variables in the Spectrometer Lab

Hypothesis

Our null hypothesis is that each of the boiled, unboiled, light and dark cuvettes will have a transmittance equal to 100%, the transmittance of the baseline cuvette.

Class Data for 4b

Time 0 min 5 min 10 min 15 min

Cuvette #

1 100% 100% 100% 100%

2 55.85% 54.16% 58.27% 55.19%

3 56.38% 55.00% 57.90% 55.72%

4 64.40% 63.27% 62.27% 62.90%

5 54.30% 54.77% 57.00% 54.77%

Percent Transmittance Over Time

Presentation of the Data for 4b

Debrief Questions

Factors impacting the separation of pigments include the varying attraction of the solvent molecules to the chromatography paper and the varying attraction of solvent molecules to one another.

In addition, the pigments travel up the paper at different rates because they are not equally soluble in the solvent and some pigments are more attracted to the chromatography paper than others because of the formation of intermolecular bonds between the paper and certain pigments. For instance, xanthophyll forms forms hydrogen bonds with the chromatography paper and therefore travels less than the other pigments.

1. What could be some factors in separating pigments in chromatography?

Yes, the Rf values would likely change if a different solvent was used as the pigments would have a different solubility in a different solvent, causing the distance traveled by the pigments to be different and therefore the Rf values would be different.

2. Would you expect the Rf values to change if a different solvent was used?

The reaction center contains chlorophyll a. The other pigments collect light energy and pass it to the reaction center. Carotene and xanthophyll also protect the photosystem from the damaging effects of ultraviolet light because they are carotenoids.

3. What type of chlorophyll does the reaction center contain? What are the roles of the other pigments?

The function of the DPIP is to change from blue to clear as photosynthesis proceeds and DPIP is reduced, allowing the spectrometer to measure the rate of photosynthesis through the clarity of the DPIP solution.

The DPIP replaces the NADP+ molecules in photosynthesis.

The source of the electrons is the water.

4. What was the function of DPIP in the experiment? What molecules is it replacing? What is the source of the electrons?

The spectrometer measured the percent light transmittance through the cuvette containing DPIP and the chloroplasts. The more light transmitted through the cuvette, the greater the reduction of the DPIP molecules and the greater the rate of photosynthesis of the chloroplasts.

5. What was measured with the spectrometer in this experiment?

In the absence of light, the light reactions in photosynthesis can no longer occur and therefore DPIP cannot be reduced. The percent light transmittance through DPIP will stay low throughout the five minute time intervals.

6. What is the effect of darkness on the reduction of the DPIP?

Boiling the chloroplasts prevents photosynthesis from occurring by causing proteins to denature. Therefore, DPIP can no longer be reduced.

7. What is the effect of boiling the chloroplasts on the reduction of DPIP?

Chloroplasts in the light have the light energy required for photosynthesis while chloroplasts in the dark can not photosynthesize. Therefore, chloroplasts incubated in the light reduced DPIP, changing it from blue to colorless and increasing the percent transmittance of the solution. The chloroplasts in the dark cannot reduce DPIP and its percent transmittance will remain low.

8. What reasons can you give for the difference in the percentage of transmittance between live chloroplasts that were incubated in light and those that were kept in the dark?

9. Identify the function of each of the cuvettes.

The function of cuvette 1, the blank cuvette, is to calibrate the spectrometer by setting it to 100% transmittance in between measurements.

The function of cuvette 2, the unboiled chloroplasts in the dark, is to identify the effect of light or the absence of light on the rate of photosynthesis by comparing its data to cuvette 3.

The function of cuvette 3, the unboiled chloroplasts in the light, is to identify the effect of light by comparing its data to cuvette 2 and the effect of boiling the chloroplasts by comparing its data to cuvette 4.

The function of cuvette 4, the boiled chloroplasts in the light, is to identify the effect of boiling chloroplasts on the rate of photosynthesis by comparing its data to cuvette 3.

The function of cuvette 5, the cuvette in light with no chloroplasts, is used as the control in order to isolate the variables that are causing trends in the data.

The amount of sunlight reaching the plants, the availability of carbon dioxide in the atmosphere, and the temperature of the environment may impact the net rate of photosynthesis.

10. What environmental factors might affect the net rate of photosynthesis?

The surface area of a plant and the placement of a plant in relation to other plants may impact the amount of light it can absorb.

Also, the waxy surface of the leaf, or the cuticle, helps retain water, affecting the rate of photosynthesis. Also, the varying amounts of phototropism, the orientation of a plant in response to light, allows some plants to absorb more sunlight than other plants.

11. What features of a plant might affect the net rate of photosynthesis?

Thank you!