Chromatography Investigation

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Introduction

Chromatography is a highly regarded technique used to separate the components of a mixture.  It is based on the principle that each component possesses a unique affinity for a stationary phase and a mobile phase. The components that are more inclined to enter the mobile phase will migrate further on the chromatogram and distinguish themselves from the other components.  The type of solvent used in chromatography is known to directly affect the separation of the mixture.  In this experiment, thin-layer and column chromatography will be utilized to separate the numerous chlorophyll and carotenoid pigments of a spinach extract.  The optimal solvent that will maximize the resolution of the components on the chromatogram will be exhibited.

Procedure

The procedure is described in the Chemistry 282 Laboratory Manual (Department of chemistry, 2002).  A minor change to the experiment was the use of magnesium sulfate to dehydrate the organic layer instead of sodium sulfate.

Discussion

         Thin-layer chromatography (TLC) is a technique that involves the movement of the components of a mixture up a plate with the use of a solvent.  This process begins by spotting the mixture near the bottom of a TLC plate, which is composed of a thin layer of a polar adsorbent spread over the surface of a piece of thin plastic.  The separation process takes place in a closed jar containing a filter paper and the desired solvent.  The filter paper absorbs the solvent and functions to help the atmosphere inside the jar become saturated with the solvent.  This environment will promote the uniform permeation of the solvent across the adsorbent and help the experiment proceed more quickly.  When the bottom edge of the plate contacts the solvent, the adsorbent acts like a wick, slowly drawing the solvent up the plate by capillary action.  As the solvent moves past the spotted mixture, two opposing forces created by the solvent and the adsorbent influence the mixture.  Each component can either dissolve in the mobile solvent or remain adsorbed to the stationary adsorbent.  This process generates an equilibrium, as some components are adsorbed and others are dissolved and transported with the solvent until they are readsorbed further along the plate.  The different tendencies of each component to comply with the subjected forces may result in a successfully separated mixture on the plate.  

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The type of solvent used will determine the extent of separation of each component.  The concept that like constituents are attracted to each other and are miscible plays an essential role in determining the migration distances of various components of the mixture.  Each component has the option of entering either the mobile or stationary phase and the polarity of the solvent will influence its pathway.  For example, a more polar solvent will compete with the polar components for positions on the polar adsorbent and also transport its counterpart along the plate until it finally becomes adsorbed.  As a result, the ...

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