The purpose of this experiment was to take spinach leaves and extract the chlorophyll and carotenoid pigments by using acetone as the solvent. The chlorophyll and carotenoid pigments were extracted by using column chromography and alumina was used as the solvent. Solvents of different polarities were used, starting with the least polar, to extract the certain components from the leaves. They were then analyzed by using thin- layer chromatography. Procedure:
The first part of the experiment dealt with breaking down the spinach leaves in a mortar and pestle. Acetone was added to this to help with the breakdown of the spinach leaves. Once the spinach leaves were broke down enough that you could see the particles clearly, they were all put into a centrifuge tube. It was sometimes necessary to add a little bit more acetone if it evaporated. Next, 2 mL of hexane was added to the centrifuge tube. The tube then had 2 mL of water added to it. This was shaken up and then centrifuged for a couple of minutes so that the layers would separate and to get rid of the emulsion, which was the green layer.
Once the centrifuging process was completed, the bottom layer was removed using a pipet. This layer was the hexane layer. A column was assembled for the hexane layer to travel through. This column was a pipet with a little bit of cotton down at the bottom and .5g of sodium sulfate towards the top. The sodium sulfate was necessary because it allows for the hexane to dry, or remove and water from the solution. The column was clamped down in a vertical position and a test tube was placed under it which was labeled E. The hexane was transferred to the column using a pipet. Once it was completely drained, .5mL of hexane was added to the column so that it was known that all of the pigments were extracted from the sodium sulfate. Tube E was then put in a warm water bath with air flowing onto it to evaporate it. The residue that was left after the evaporation...
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