Lab 4: Photosynthesis and Chromatography of Spinach Leaves

Lab 4: Photosynthesis and Chromatography of Spinach Leaves

Discussion:

The chromatography of the spinach leaves was taken to establish what pigments were present in the chloroplasts of the leaves. The solvent carried and separated the different pigments up the cellulose paper. The solvent moved up the paper because of capillary action because of adhesion of the solvent molecules and the paper. The pigments dissolved in the solvent also were carried up the paper and were separated because they do not all dissolve equally into the solvent, less soluble pigments appear lower on the paper. The pigments also were attracted to the paper at different levels. Each pigment has a specific Rf value which is the distance the pigment migrated divided by the distance the solvent migrates. The data shows that Chlorophyll a has the highest Rf value, and it did travel the farthest out of the pigments. The control in this experiment was the same spinach leaves used. Error was no Carotene detected. Carotene is the very abundant in plants, so the lack of it found was based on reading errors of the chromatography, or the confusion of Xanthophyll with Carotene because the Rf value of Carotene is 0.98, which was calculated for Xanthophyll. A spectrometer was used to measure photosynthesis by detecting how much light was able to pass through the cuvettes. The spectrometer was set to 100 percent transmittance of light in cuvette 1 because if photosynthesis occurred in the other cuvettes, they would transmit the same amount of light. DPIP was used to replace the final electron acceptor of photosynthesis, NADP. When DPIP accepts electrons from the processes of photosynthesis, it changes in color from blue to clear. The most light was able to pass through in cuvette 3 because it had unboiled chloroplasts that had light. The presence of light started the light dependent reactions and reduced the DPIP making it clear and allowing more light to pass through. The lowest percentage of light transmittance was in cuvette 4 with the boiled chloroplasts. The DPIP remained blue, and the boiled chloroplasts clumped together to further reflect light back instead of through. Controls in this experiment were the keeping of the cuvette 1 as 100% transmittance, the amount of chloroplasts used, the amount of DPIP, the amount of water, and phosphate buffer in the cuvettes. The error was in cuvette five, the values of the light transmittance should have been the same as cuvette 2. This error can be caused by faulty spectrometer readings, or fingerprints on the cuvette.