Photosynthesis Lab

Photosynthesis Lab

Purpose

As photosynthesizers, the chloroplasts in plant cells need a consistent input of light in order to fully function. In this experiment, we examine the effects on several samples of chloroplasts’ rates of photosynthesis as certain factors are manipulated. The chloroplasts’ light exposure and functionality are altered to experimentally display the most favorable conditions for the productivity of chloroplasts. The productivity level is measured using a colorimeter and the chemical 2,6-dichlorophenol-indophenol, or DPIP, which acts as a replacement for the electron acceptor NADP+. As DPIP is reduced (in place of the reduction of NADP+ to NADPH), it turns from dark blue to colorless; thus, as the light reactions in the chloroplasts progress, the mixture in the cuvette will become clearer, and the transmittance values measured with the colorimeter will increase.

Introduction

In a greater perspective, the chloroplasts, the plant-specific organelles which contain the prior mentioned pigments, use light energy in the process of photosynthesis. The pigment chlorophyll is found embedded in the membranes of the thylakoids. Chloroplasts are extremely sensitive to the presence of light because sunlight is so necessary to the production of sugar, the stored source of energy for these organisms. Live chloroplasts are able to use light energy to offset this process, while chloroplasts that have been damaged may be able to transmit light to some degree, but certainly not as effectively. The longer chloroplasts are exposed to light energy, the more the photosynthetic processes are able to produce.

  

Methods

To better understand the concept of photosynthesis, we test the hypothesis; for light reactions to occur light and chloroplasts need to be present. In order to test this hypothesis we use a dye-reduction technique. To set up the lab a flood light is placed, and turned on, directly in front of the heat sink. The heat sink is a large beaker filled with water in order to minimize the heat capacity that the cuvettes would take on. On the other side of the heat sink is where the cuvettes stood through the time intervals of 5, 10, 15, and we also measured 20 minutes.

Our colorimeter was set up too close to the flood light, creating a skew in our data. (Further explained in the conclusion)

Each cuvette contained a specified mixture of phosphate buffer, water, DPIP, and boiled or unboiled chloroplast. The liquids were put into test tubes with droppers, estimating every ten drops to equal one milliliter. The chloroplasts were put in last because the amounts of each kind had to be exact.

Above: Before the chloroplasts were added.

Above: After the chloroplasts were added.

At each timed interval the cuvettes were taken away from the light individually and places into the colorimeter to measure the percent transmittance of each solution and then placed back into the light.

Above: At the five minute interval

Data

Graphs & Charts

Discussion

The distinct differences between the cuvettes were the presence of chloroplasts, the use of boiled or unboiled chloroplasts, and the exposure to light. The first cuvette was the phosphate buffer that was used to set transmittance in the colorimeter to zero. The second cuvette contained live, or unboiled, chloroplasts and it was covered in foil to keep its insides from being exposed to light. The third cuvette contained live chloroplasts and it was not wrapped in foil. The fourth cuvette contained dead, or boiled, chloroplasts which were exposed to light, and the last one contained a solution with no chloroplasts that was exposed to light.

The heat sink of the flask in front of the heat lamp was used to decrease the intensity of the lamp’s heat reaching the chloroplasts. This was necessary so the chloroplasts wouldn’t be harmed by the heat. Water has a high specific heat, so the light passing through it did not pass as great an amount of heat to the chloroplasts.

In all four sets of data, the transmittance generally increased. Whether the chloroplasts in the cuvette were boiled or unboiled, present or not, the transmittance detected by the colorimeter increased over time. However, this is a visible disparity between the transmittance percentages of the unboiled chloroplasts kept in the dark. This shows the importance of having at least live chloroplasts or the presence of light in a plant organism.  

Conclusion

Our experiment proved that the presence of light and favorable conditions for chloroplasts’ survival (e.g. they shouldn’t be boiled) are imperative for the proper functioning of chloroplasts. Unfortunately, our second cuvette was exposed to light too frequently due to insufficient coverage with aluminum foil in the first attempt and exposure to light during the colorimetry process (our colorimeter was set up near the lamp and the door on the machine needed to remain open for it to work properly, so the cuvette was exposed to light as we ran the tests), so our data for cuvette #2 is inaccurate. However, the transmittance values for cuvette #2 were still lower than #3's, which means that the chloroplasts were still at a lower level of productivity with the limited light, and the results still support our hypothesis to a degree. Additionally, cuvettes #4 and #5 should realistically have little to no increase in transmittance, as #4 has boiled (dead) chloroplasts that should not be functioning very much, if at all, and #5 has no chloroplasts to change the blue DPIP to its colorless, reduced form.

References

• The procedures, graphs, and charts used here were taken from "Lab Four: Plant Pigments and Photosynthesis" by CollegeBoard.