If this lipid soluble solvent is present, as opposed to the water soluble solvent, then the lipid soluble pigments will move up the chromatography paper instead of the water soluble pigments. Plants absorb light in order to carry out photosynthesis. Sometimes the motion is radial, sometimes descending, sometimes as in this experiment ascending. Xanthophyll is found further from the solvent front because it is less soluble in the solvent and has been slowed down by hydrogen bonding to the cellulose. Rf values are in the right column These results show that the Green leaf has mainly green pigment Chlorophyll , while the Red leaf has all pigments. This experiment gives insight into what pigments different colored leaves are composed of and how calculating the Rf value can indicate what pigment it is.
Rf value stands for the the Relative Mobility Factor. The pure solvents will readily distinguish between the polar and non-polar molecules. You can try this lab at home, check out to see how you can perform this experiment with a pen. The formula for an Rf value is migration distance of solvent front divided. The more soluble, the further it travels and vice versa. Purpose: Part A The lab groups were to separate pigments and calculate Rf values using plant pigment chromatography, describe a technique to determine the photosynthetic rate, compare photosynthetic rates at different light intensities using controlled experiments, as well as explain why rate of photosynthesis varies under different environmental conditions.
This gives a green line across the paper, which contains the pigments of the leave. It is a quick and simple method for determining the number of compounds in a mixture, identifying an unknown, testing purity, and monitoring reactions. A solvent is placed at the bottom of the paper. Chlorophyll b is another plant pigment. Using a different part of the leaf, roll the penny again over the same line.
Through photosynthesis, plants synthesize their own food. Plants have many varieties of pigments, all of which absorb different colors of light. Pour 5 ml chromatography solvent into the test tube. Other than these there were few places for error in this lab. On this line 10 crosses were drawn which would be the starting point of the samples this paper was measured so that it would fit into the Chromatography tank. The spectrophotometer may have not been calibrated correctly because this was the first time this particular one had been used.
Considering the introductory remarks above and your answers to 1 and 2 , explain briefly the order of the pigments. There are three types: Column, Paper, and Thin Layer chromatography. The filter paper was then placed in the cylinder with the tip barely touching the solvent and none of the edges touching the sides. Chlorophyll b passes its exited electrons to chlorophyll a, which passes its excited electrons to other molecules, which store the energy as chemical potential energy. Examine the strip which was developed with pure 2-propanone acetone.
Conclusion Precautions To obtain a clear and accurate chromatogram the students were advised to use gloves. The chlorophyll in the leaf of the plant contains different pigments which are chlorophyll a, chlorophyll b, xanthophyll, phaeophytin and carotene. Bend a paper clip and attach it to a cork stopper. In this test it was solubility. Analysis These questions should be answered in your laboratory notebook following your data and observations. Five cuvettes were numbered respectively and then wiped with lens tissue. To measure light transmittance in chloroplasts a spectrophotometer will be used.
Carotene is the most soluble of the pigments and as a result will be carried the farthest by the solvent. The Spinach in acetone S in a has a dark green line chlorophyll a and b in the middle of the chromatography paper. Lastly, some chloroplasts had shown transmittance of chloroplasts solution when there should not have been any. A straight line is drawn 1cm away with a pencil from the edge on a strip of filter. The two chlorophylls found in green plants are chlorophyll a and chlorophyll b. The distance from the bottom of the paper to this line is the distance traveled by the solution 9. Discussion Questions: Part A: If the primary pigment color of the leaf was purple the results would be similar since retinal acts very similar to chlorophyll.
Describe what each of your chromatography strips looked like. Anthocyanin is another accessory pigment that absorbs bright red colors. The colors of these pigments are because of the double bonds in their structure. Cut a strip of filter paper or chromatography paper so that it just fits inside a 15-cm or larger test tube. Three drops of boiled chloroplasts were added to cuvette 4, and the transmittances were recorded at the same times.
The first step was to cut a point on one end of the filter paper and draw a pencil line 1. The purpose of this paper is to help analyze or compare pigments from two organisms. Hypothesis: In this experiment it is hypothesized that the cuvette with boiled chloroplasts and the cuvette kept in the dark containing unboiled chloroplasts will have very slight changes in light transmittance, whereas the cuvette containing unboiled chloroplasts that have been exposed to light will have an increasingly higher % transmittance over the course of time. Rf is a characteristic that is specific to a certain substance and is calculated by Dunknown divided by Dsolvent, which stands for Distance solute traveled divided by Distance solvent traveled. The ratio of the distance traveled by a compound to that of the solvent front is known as the Rf value; unknown compounds may be identified by comparing their Rf's to the Rf's of known standards. During the autumn, however, the chlorophyll starts to break down, causing the carotenes and xanthophylls to show their bright red, orange and yellow colors. It is very important that the bottom of the filter strip is in the acetone, but the green spot is not in the liquid.
This light is absorbed by chloroplasts in plant cells. These compounds are present in relatively large amounts in ivy leaves and they also have characteristic colors which are easily seen in a simple chromatogram. These tests occur on chromatographic sheets, commonly constructed of alumina or silica pasted on a metal backing. Chlorophyll a, Chlorophyll b, and carotenoids capture light energy and transfer it to chlorophyll a at the reaction center. In the light-dependent reactions pigments trap energy from light, and this energy is used to split water molecules photolysis. When the solvents rose about 6 cm they were removed. Using your ruler, measure 2 cm from the end and draw a line and draw a ball in the middle of the line.