Paper Chromatography Principles

Principle

• Paper chromatography is a type of planar chromatography that uses a stationary phase of paper to separate, identify, and quantify components of a mixture.
• It is based on the principle of partition chromatography, where the components of the mixture are distributed between the paper and a solvent.

Apparatus

• A rectangular piece of filter paper (stationary phase)
• A solvent (mobile phase)
• A pencil or marker
• A ruler or straightedge
• A solvent reservoir
• A chromatography chamber (optional)

Procedure

1. Sample Application
   • A small amount of the mixture is applied to the paper as a spot or a line, about 1-2 cm from the top edge.
   • The sample should be applied in a minimal volume to prevent spreading.
2. Solvent Migration
   • The paper is placed in the chromatography chamber, and the solvent is allowed to migrate up the paper by capillary action.
   • The solvent front is marked with a pencil or marker as it migrates.
3. Separation and Detection
   • The components of the mixture separate based on their affinities for the paper and solvent.
   • The separated components can be detected using various methods, such as:
     • Visual observation (e.g., color, fluorescence)
     • Chemical testing (e.g., ninhydrin for amino acids)
     • Instrumental methods (e.g., densitometry)

Factors Affecting Separation

• Solvent composition: The choice of solvent affects the separation of components.
• pH: The pH of the solvent can influence the separation of ionizable compounds.
• Temperature: Temperature can affect the rate of solvent migration and component separation.
• Paper type: The type of paper used can influence the separation of components.

Applications

• Biochemistry: Paper chromatography is used to separate and identify amino acids, sugars, and other biomolecules.
• Forensic science: It is used in forensic analysis to identify and compare substances, such as ink, dyes, and drugs.
• Food analysis: Paper chromatography is used to detect and quantify food additives, contaminants, and nutrients.

Principle of Paper Chromatography

• Partition chromatography is the principle behind paper chromatography, where components of a mixture are distributed between the paper and a solvent.

Apparatus Used

• Rectangular piece of filter paper (stationary phase)
• Solvent (mobile phase)
• Pencil or marker
• Ruler or straightedge
• Solvent reservoir
• Chromatography chamber (optional)

Procedure of Paper Chromatography

• A small amount of the mixture is applied to the paper as a spot or a line, about 1-2 cm from the top edge.
• The sample should be applied in a minimal volume to prevent spreading.
• The paper is placed in the chromatography chamber, and the solvent is allowed to migrate up the paper by capillary action.
• The solvent front is marked with a pencil or marker as it migrates.

Separation and Detection

• Components of the mixture separate based on their affinities for the paper and solvent.
• Separated components can be detected using various methods, including:
  • Visual observation (e.g., color, fluorescence)
  • Chemical testing (e.g., ninhydrin for amino acids)
  • Instrumental methods (e.g., densitometry)

Factors Affecting Separation

• Solvent composition affects the separation of components.
• pH of the solvent influences the separation of ionizable compounds.
• Temperature affects the rate of solvent migration and component separation.
• Type of paper used influences the separation of components.

Applications of Paper Chromatography

• Biochemistry: separates and identifies amino acids, sugars, and other biomolecules.
• Forensic science: identifies and compares substances, such as ink, dyes, and drugs.
• Food analysis: detects and quantifies food additives, contaminants, and nutrients.