Extraction of Chlorophyll from Spinach Leaves

The spinach plants were hole punched, mixed with 0.2% NaOH and dish detergent, placed in a syringe with the solution to have the oxygen

There are two main types of chlorophyll, chlorophyll a which absorbs wavelengths of 430nm (blue) and 662 (red) and is the main photosynthetic pigment, and chlorophyll b, which doesn’t directly participate in the photosynthetic process, but is capable of donating its energy to chlorophyll a

Lycopene and β-carotene are separated using thin layer chromatography (TLC). A mixture of tomato paste containing dissolved lycopene and β-carotene is first spotted on a TLC plate. The technique uses a stationary phase layer, on which the substances stick, and a mobile phase, which migrates upwards the plate and carries the components of the mixture with itself (Chemguide). Depending on their respective polarity, lycopene and β-carotene stick more or less to the stationary phase and so they migrate up the TLC plate at different rates (Chemguide). Thus, as the chromatography is completed, the two components will appear as distinct dots at different heights on the plate. Lycopene has a bright orange color, whereas β-carotene is light yellow.

The cell membrane (Plasma membrane) functions to provide cell support, cell stability and control entry and exit of materials from the cell. This study was conducted to test the effects of environmental conditions such as the on beet root cell membrane (Beta vulgaris). Five trials using varied pH concentrations were tested and absorbance rates were monitored. The experimental results showed that the protein function decreased sequentially when the pH decreased. This allowed the betacyanin dye to leak out which created the color that was needed to determine the intensity and therefore the effect of the circumstances. This supported the hypothesis that the more acidic or basic the environmental condition around the beet cell, the more permeable the, membrane indicated by color intensity. Pigment leakage in the solution was analyzed by using a spectrophotometer.

In the beginning of this experiment, our TA added water, salt, and 75/25 hexane/acetone to spinach leaves to a blender and blended the mixture to assume equal amounts for each group and to avoid erros if each student had to do the blending. The addition of the water to the mixture allowed the it to separate into a distinct organic layer after being run in a centrifuge, which was available to be collected at the top of the centrifuge. Salt reduces solubility, which will force the organic parts of the mixture (the desired pigments for example) to separate into the organic layer at the top. Lastly, 75/25 hexane/acetone is added because this is a moderately polar solvent and will useful for both the non-polar and polar pigments present within the spinach leaves. A mixed solution of hexanes and acetone must be used because acetone is very polar, while hexane in very non-polar, and the spinach leaves contain both non-polar and polar pigments in them that are important in the extraction and for further analysis. The mixture was placed in the centrifuge so the solids in the mixture (mostly cellulose) could be separated from the liquids into separate distinct layers for further extraction and testing. In the tube, the organic substances separated into the top layer, whereas the water layer remains at the bottom of the tube below the solid layer made up of mainly cellulose.

Abstract: The purpose of this lab is to separate and identify pigments and other molecules within plant cells by a process called chromatography. We will also be measuring the rate of photosynthesis in isolated chloroplasts. Beta carotene, the most abundant carotene in plants, is carried along near the solvent front because it is very soluble in the solvent being used and because it forms no hydrogen bonds with cellulose. Xanthophyll is found further from the solvent font because it is less soluble in the solvent and has been slowed down by hydrogen bonding to the cellulose. Chlorophylls contain oxygen and nitrogen and are bound more tightly to the paper than the other pigments.

This lab was conducted to explore the light energy, pigments and the rate of photosynthesis in magnolia leaves. In experiment one a magnolia leaf was used to see the separation of primary and accessory plant pigments using a process called paper chromatography. The importance of this process was to discover which pigment had the highest band along a piece of filter paper and identify various plant pigments in a magnolia leaf such as xanthophyll, chlorophyll a, chlorophyll b, and carotenoids that aid magnolia leaves during photosynthesis. Based on the conducted experiment, it can be concluded that chlorophyll a was the pigment that showed the highest band on the piece filter paper which means that chlorophyll a is the primary pigment in photosynthesis

The purpose of this experiment is to determine the specific pigments that are found in each of the chosen plant leaves, as well as, discover the intermolecular forces present in each leaf. This is accomplished by using a technique called chromatography, which splits a mixture or solution into its different parts based on the mixtures ability to dissolve in a chosen solvent. Chromatography works by placing a strip of chromatography paper, that has a small amount of mixture on it, into a cup of the specific solvent chosen. In this experiment, one hundred percent acetone was chosen to be the solvent. Once the small amount of the mixture is aligned with the solvent, the mixture will begin to separate and rise by capillary action based on the solubility,

Each spot Rf value was a ratio and the data obtained can be found in the table connected. During the lab, once the plate was placed into the beaker, the eluent was seen traveling up the plate. The spots were not visible until the UV light was used, but under the light the plate was and eon green the spots were pink.

This experiment demonstrates the effects of pH on the rate of photosynthesis by examining the behavior of leaf disks in different pH solutions under light. In this experiment, we used five different pH levels: pH 5, pH 6, pH 7, pH 8 and pH 9. These solutions were created using a combination of hydrochloric acid and sodium hydroxide. Spinancia olcerea or spinach, leaves were used in the experiment to examine the effects of pH on the rate of photosynthesis. The rate of photosynthesis was measured by counting the number of leaf disks that rose to the surface of the solution after each minute. In acidic solutions, the rate of photosynthesis increased while in basic solutions, the rate of photosynthesis decreased.

The isolation of β-carotene from lycopene via column chromatography was accomplished, but results were inconsistent. Many students were unable to isolate the first elution compound, β-carotene; however, lycopene isolation was very obtainable; due to the structural similarity of these two molecules, it was intuitive that a separation would be difficult. UV/VIS and TLC analysis of the compounds confirmed that it is possible to isolate β-carotene from lycopene, due to differences in dipole moments of these chemicals. In future experiments, it is suggested to use a mobile phase less polar than 15% ethyl acetate and 85% hexanes such as 5% ethyl acetate and 95% hexanes, for this would have potential to create a better separation. It was

After wearing the gloves we obtained a chromatography vial from professor and label it with my and my peer initials. We dried up the chromatography vial in fume hood and added 1 ml of chromatography solvent to the vial. Then we took a chromatography strip and measure it 1.5 cm with ruler from one end of the strip and drew a line with pencil we cut two small pieces below the pencil line to form a pointed end. We applied spinach on the strip using quarter to rub the spinach leaf on the line that we drew on the strip and put it into the chromatography vial and placed that in fume hood. We observed as the solvent was moving up the chromatography strip by capillary action. When the solvent was reached approximately 1 cm from the top of the strip then we removed the cap from the vial. We took out the strip from the vial using forceps and marked up the location of the solvent front because it evaporates quickly. We measure out the distance as well as the pigment in order to find out the rf value. Moreover we compared rf values to the one in reference list in order to identify the

For lab 12, it is hypothesized that chlorophylls a and b are present in a plant leaf and contribute to the starch production in photosynthesis. Also, products of photosynthesis will be present in leaf tissue exposed to red and blue light wavelengths for several days, but a decreased presence in leaf tissue exposed to green and black light wavelengths. In lab 13, it is expected that since chlorophylls a and b are more polar and smaller molecules than the anthyocyanins and carotenoids, they will travel higher up the chromatography paper than the other pigments.

The next step was to place the strip of chromatography paper on a paper towel. Then dip a capillary tube into the plant pigment extract (spinach pigment extract) provided by the teacher. The tube will fill on its own. We applied the extract to the pencil line on the paper, blew the strip dry, and repeated it three to four times until the line on the paper is a dark

The particles that were collected for the lab were very concentrated prior to being mixed into the medium. Once the particles where mixed into the medium they was a decreased in their concentration as the medium allowed for separation by filling the spaces between them. Once the first streak was made it was slightly less concentrated and this allowed for the particles to be spread out over the surface. The concentration of particles that were spread with each streak continued to decrease. In the final streak, as seen in picture 7, the particles were separated almost to the point of being able to individually identify each color of particle. Much as the same results that you might see with a mixed culture that was completed in a lab and final results reviewed under a microscope.