Chromatography Practical

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Introduction

• Chromatography is an experimental technique that is used in order to separate mixtures.
• This is done through the different components of the mixture travel up the chromatography paper (or TLC plate) at different speeds. Because of this, the components begin to separate, allowing you to differentiate between different components. This makes it possible to calculate a retention factor value (RF value) for each component within the mixture.
• There are two types of chromatography: Paper Chromatography and Thin Layer Chromatography.
• In Paper Chromatography, the mixture of pigments passes through a piece of paper.
• In Thin Layer Chromatography, the mixtures of pigments pass through a thin layer absorbent, like silica gel, through which the mixture travels away from the origin, continuing until the mixtures separate.  

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Apparatus

• Acetone
• Capillary Tube
• Chromatography Solvent
• Distilled Water
• Filter Paper
• Leaf Sample
• Pencil
• Pestle and Mortar
• Ruler

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Method

• Approximately 1 centimeter above the bottom of the filter paper / TLC Plate, draw a straight line in pencil using a ruler.
• Retrieve a sample of a leaf and place it in a mortar.
• Add 20 drops of an extraction solvent, like acetone or propanone, to the mortar, and begin to grind the leaves with a pestle. This allows the pigments to be released.
• Using the capillary tube, extract some of the pigment and spot it on the middle of the line that was drawn previously.
• Carefully suspend the filter paper / TLC Plate into the chromatography solvent, ensuring that the solvent reaches below the pencil line. Leave the paper in the solvent until it reaches the top of the paper.
• Before it reaches the top of the paper, remove it from the solvent and draw a line with a pencil, indicating where the solvent ended.
• Once you have done this, you will be able to calculate the retention factor value / RF value.
• RF Value = distance travelled by the component / distance travelled by the solvent.

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Terms

• Chromatography

  An experimental technique used in order to separate and identify species within a mixture.

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• TLC

  Thin Layer Chromatography. It is a type of chromatography that can be used in order to separate and identify non-volatile mixtures.

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• Mobile Phase

  When the mixture is dissolved in a mixture.

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• Stationary Phase

  The phase after the mobile phase.

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• RF Value

  Retention Factor - It can be used in order to compare and identify compounds.

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• Solvent Front

  How far the solvent has travelled.

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• Extraction Solvent

  A solvent used to extract pigments from the plant tissue.

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• Origin

  The spot on which a solution of the sample is placed with the capillary tube.

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