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Thin Layer Chromatography and Paper Chromatography Abstract

In order to study distinct components of a mixture, they must be first separated individually. This is achieved through this lab by the technique called chromatography. We used two types of chromatography- thin layer and paper. They both involve a stationary phase which is either the paper or the TLC strip as well as a mobile phase, which could be one of two solvents used: iso-octane-acetone for TLC or ether acetone for paper. Chemicals in a sample separate due to their unique polarities. As a result, nonpolar substances move further than polar substances in this process.

First of all, in the Thin Layer Chromatography lab, my partner and I tested the plant pigments chlorophyll and carotenoid. About 1 cm from the bottom, spots were placed along the same horizontal plane using a capillary tube. After repeated spotting procedures, the TLC strip was placed into a glass developing chamber with the iso-octane solvent inside. The solvent was then absorbed and allowed to migrate until it reached about 1 cm from the top. We then made sure that we marked where the solvent front was, for after evaporation it would be invisible. Once the two plant pigments have undergone this method, the Rf, or the retardation factor may be calculated. This formula is represented by the distance a component travels divided by the distance a solvent travels. Consequently, observations could be made on both the chlorophyll and carotenoid TLC strips based upon Rf value and the color of each pigment. From the carotenoid chromatogram, the pigments lycopene, leutein, violaxathin, and neoxathin were obtained. From the chlorophyll chromatogram, a range of chlorophyll a, chlorophyll b, xanthophyll, and phaeophytin pigments was obtained. However, it is important to note that there are various factors that influence the reproducibility of chromatograms. These factors include the quality of absorbents, humidity, solvents, the developing chamber, and the spotting technique. Secondly, in the Paper chromatography lab, we tested plant pigment from spinach leaves. We drew a line 2 cm from the bottom of the strip and through the same process as above, we used the solvent ether acetone to create a chromatogram. We then obtained the Rf values with the solvent front marked and identified the pigments xanthophyll, chlorophyll b1 and phaeophytin b.

The main objective of these two labs was to separate plant pigments either by thin layer chromatography or paper chromatography. I have learned that the pigments within mixtures separate because of the solvent used and the polarities each pigment exhibits towards that solvent. The most nonpolar pigments will travel the furthest. In real life, chromatography is a tool for filtering and purification. It could be applied to bubble detection, for detecting protein concentration, and recognizing pH and conductivity. In the pharmaceutical industry, discrete substances may be completely separated from a drug so as to find its components. Chromatography could also distinguish between counterfeit drugs. Whenever the elements of a mixture are ambiguous or vague, chromatography provides the clarification.


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