4A Plant Pigment Chromatography
Purpose
The purpose of this experiment was to use Chromatography paper to identify plant pigments in spinach cells. We were testing to see if certain types of pigments were in spinach and the solubility of these pigments (Rf values). The relationship between the pigment and the solvent is the Rf value.
Introduction
The different pigments plants contain are carotene, xanthophyll, chlorophyll a and chlorophyll b. Paper Chromatography is used to separate the molecules because of the varying solubility of molecules in a certain solvent. It does this by separating the pigments using capillary action. This happens because solvent molecules are attracted to the paper as well as each other. When the solvent moves up the paper, it carries the pigments with it. The distance the pigments travel is dependent on their solubility.
Methods
In this lab we began by cutting a strip of chromatography paper into a pencil shape. (A long strip with a pointed end). Next, we scraped spinach cells off of a spinach leaf with a coin and dragged them across the chromatography paper, making sure that this green line did not go past the pointed end of the chromatography paper. We then hooked the chromatography paper onto a hook on the bottom of a stopper that was to go in a 50-ml graduated cylinder. The graduated cylinder had 1 cm of solvent in the bottom of it. We then made sure that the solvent only touched the pointed end and did not emerge the strip portion. The rest of the lab we let this chromatography paper sit and waited to record our results when the period ended, after the chromatography paper had diffused all of the spinach's pigments up the paper. |
| Rubbing spinach cells onto chromatography paper |
Placing the paper in into the test tube that holds the solution
Waiting..
Waiting...
Still waiting....
Measuring the distances traveled!
Measuring the distance of each band number 6.3-.6
Data
Graphs and Charts
Discussion
Our results contained the Rf values for 4 specific pigments found in spinach leaves: Carotene, Xanthophyll, Chlorophyll a, and Chlorophyll b. These results were directly related to how far each of them traveled up the chromatography paper. Each pigment was able to travel different distances up the paper because each pigment had different solubility levels and the chromatography paper is able to show this. A trend found in our data was the relationship between the distance of each pigment traveled and the Rf values of each of the same pigments. As a result if this, a conclusion can be made that the further the distance a pigment travels up the chromatography paper, the higher it's solubility is. The ones who traveled the furthest had the lowest Rf values and vice versa. These results were accurate and support our answer. If we were to change anything or re-do certain aspects of this lab we would have correctly measured the pigment's distances during the actual lab. Instead we just took pictures of the distances and referred to them because we were running out of tine by the end of this lab. Other than this, our lab was successful. Our results support this conclusion.
Conclusion
To conclude this experiment, spinach contained these 4 pigments: Carotene, Xanthophyll, Chlorophyll a, and Chlorophyll b. The solubility of these pigments were a directly related to how far the pigment traveled. Chlorophyll b had the lowest Rf value and traveled the shortest distance while Xanthophyll had the highest Rf value and traveled the furthest. Our data correctly supports this answer.
References
4B Photosynthesis/ The Light Reaction
Purpose
The purpose of this experiment was to test whether light and chloroplasts are required in order for light reaction to occur.
Introduction
Many factors can either inhibit, help, or have no effect on Photosynthesis (light reactions). This lab was conducted to study this. DPIP, a compound posing as an electron acceptor, water, a phosphate buffer, and a spectrophotometer were used in this lab to reach the answer to this question. A dye-reduction technique is used in this experiment to study photosynthesis.
Methods
To begin in this lab, we added the correct amount of DPIP, distilled H20, and phosphate buffer to each of the 5 cuvettes. The only one that did not contain DPIP was the first one. We made sure to wrap cuvette 2 in tin foil because no light was supposed to enter this cuvette. Each of the 5 cuvettes were measured in the spectrometer right after their mixtures were created. After each of these results were recorded, a 5, 10, and 15 minute time period was measured for each cuvette again. While one cuvette was in the spectrometer, the other 4 (except for number 2) were placed in front of a heat sink filled with water that was placed in front of a light.The heat sink's purpose was to absorb the heat so the cuvettes would experience no heat. After each of the 5, 10, and 15 minutes of the cuvettes were finished, we recorded our results.
Measuring DPIP
Filling a cuvette with DPIP
Still filling
Exposing the cuvettes to light
All of the materials used
Unboiled chloroplasts
Data
% Transmittance
Graphs and Charts
Discussion
Unboiled/dark and Unboiled/light both had a significant increase in transmittance within the first five minutes of the experiment which shows that photosynthesis occurs in both light and dark environments while the chloroplasts are still alive. Contrarily, the chloroplasts that were boiled had very little growth as time went on which showed that the denatured chloroplasts hardly undergo photosynthesis. The change in transmittance that was apparent with the boiled chloroplasts may be due to some of the sample not being fully denatured. If this experiment were to be done again, it would be beneficial to make sure all of the chloroplasts are fully denatured so that there would be even less photosynthesis occurring within the boiled chloroplasts. At the same time, it may be inevitable to have a portion of the sample could survive being boiled. This comparison also shows that the light has much less of an impact on photosynthesis occurring than the nature of the chloroplasts, as those which were simply not exposed to the light still transformed, while the boiled chloroplasts were incapable of undergoing photosynthesis. The results we obtained are in sync with what we thought would happen because in order for plants to go through photosynthesis there needs to be a presence of light (in order to start the light/dark reactions) and chloroplasts, in order to absorb the light. If we conducted the experiment again, in order to see a more significant transmittance change, we would need to be more careful with how we mix or cuvettes as well as being more persistence with how much of each substance is being tested. Also, testing each cuvette between a certain time period rather than so close together so the time we collect our data is more exact, because that affects the transmittance measured.
Conclusion
To conclude this experiment, light does not have as big of an impact on photosynthesis as chloroplasts do. Our data generally supports this theory except for the slight differences in the transmittance levels of the boiled chloroplasts. In order for our data to completely support our theory, all of these levels should have been exactly the same.
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