Chromatography, types, techniques, ppt.pptx

Chromatography

1. Practical Lecture: Chromatography – Principles, Techniques & Applications Dr Showkat Ahmad Wani

2. Introduction to Chromatography Chromatography is a laboratory technique used to separate mixtures into their individual components. The term originates from the Greek words chroma (color) and graphein (to write), reflecting its early use in separating plant pigments. Developed by Mikhail Tsvet in the early 20th century, chromatography has become an essential tool in analytical chemistry for both qualitative and quantitative analyses.

3. Fundamental Principles Phases Involved : Mobile Phase : The solvent that moves through the system, carrying the mixture's components. Stationary Phase : The material that remains fixed inside the column or on a surface, interacting with the mixture's components.​

4. Types of Chromatography A. Based on Physical State of Mobile Phase Gas Chromatography (GC) : Mobile Phase: Inert gas (e.g., helium, nitrogen). Used for volatile compounds.​ Wikipedia+2Wikipedia+2Wikipedia+2 Wikipedia+1Wikipedia+1 Liquid Chromatography (LC) : Mobile Phase: Liquid solvent. Suitable for a wide range of compounds.

5. B. Based on Separation Mechanism Adsorption Chromatography : Separation based on adsorption of components onto the stationary phase.​ Wikipedia Partition Chromatography : Separation based on partitioning between two liquid phases.​ Ion-Exchange Chromatography : Separation based on ionic interactions between charged analytes and the stationary phase.​ Wikipedia Size-Exclusion Chromatography : Separation based on molecular size, with larger molecules eluting first.​ Affinity Chromatography : Separation based on specific interactions between an analyte and a ligand attached to the stationary phase

6. Common Chromatographic Techniques Paper Chromatography : Simple technique using paper as the stationary phase. Used for separating pigments and small molecules.​ Wikipedia+2Wikipedia+2Wikipedia+2 Thin-Layer Chromatography (TLC) : Uses a thin layer of adsorbent material on a glass or plastic plate. Allows for rapid qualitative analysis.​ Wikipedia Column Chromatography : Involves a column packed with stationary phase material. Used for purifying individual chemical compounds.​ Wikipedia+1Wikipedia+1 High-Performance Liquid Chromatography (HPLC) : Advanced form of LC with high resolution and sensitivity. Widely used in pharmaceutical and biochemical analysis.​ Gas Chromatography (GC) : Highly efficient for separating and analyzing compounds that can be vaporized.

7. Applications of Chromatography Pharmaceutical Industry : Drug purity testing and quality control.​ Environmental Analysis : Detection of pollutants in air, water, and soil.​ Food Industry : Analysis of food additives, preservatives, and contaminants.​ Clinical Diagnostics : Detection of biomarkers and metabolic profiling.​ Forensic Science : Analysis of substances in criminal investigations.

8. Practical Demonstration: Thin-Layer Chromatography (TLC) Objective : To separate and identify components in a mixture using TLC.​ Materials : TLC plates (silica gel-coated). Sample mixture (e.g., ink or plant extract). Solvent system (e.g., ethanol:water ). Developing chamber. Capillary tubes. UV lamp (for visualization).

9. Procedure : Draw a baseline on the TLC plate about 1 cm from the bottom. Apply small spots of the sample mixture onto the baseline using capillary tubes. Place the TLC plate in the developing chamber containing the solvent, ensuring the spots are above the solvent level. Allow the solvent to rise up the plate until it is about 1 cm from the top. Remove the plate and mark the solvent front immediately. Visualize the separated spots under a UV lamp and mark their positions.

10. Thank You