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  1. 1. Chromatography By- Bhavya, Harsh, Harshvardhan, Namrata, Ronit and Vidhatri
  2. 2. Contents <ul><li>Definition of chromatography </li></ul><ul><li>Different kinds of chromatography </li></ul><ul><li>Uses and applications of each </li></ul>
  3. 3. What is Chromatography? <ul><li>Chromatography is the science which studies the separation of molecules based on differences in their structure and/or composition. </li></ul><ul><li>The word was first derived from the Greek word chroma meaning “colour”. </li></ul>
  4. 4. Glossary… <ul><li>Chromatograph: Instrument employed for a chromatography. </li></ul><ul><li>Eluent: Fluid entering a column. </li></ul><ul><li>Eluate: Fluid exiting the column. </li></ul><ul><li>Elution: The process of passing the mobile phase through the column. </li></ul><ul><li>Flow rate: How much mobile phase passed / minute (ml/min). </li></ul><ul><li>Linear velocity: Distance passed by mobile phase per 1 min in the column (cm/min). </li></ul>
  5. 5. Different types <ul><li>Thin layer chromatography </li></ul><ul><li>Gas Chromatography </li></ul><ul><li>HPLC </li></ul><ul><li>Electrophoresis </li></ul><ul><li>Paper Chromatography </li></ul>
  6. 6. Thin Layer Chromatography <ul><li>TLC is using a thin, uniform layer of silica gel or alumina coated onto a piece of glass, metal or rigid plastic. </li></ul><ul><li>The silica gel (or the alumina) is the stationary phase. </li></ul><ul><li>The mobile phase is a suitable liquid solvent or mixture of solvents. </li></ul>
  7. 7. Uses <ul><li>Determination of the components a plant contains </li></ul><ul><li>Monitoring organic reactions. </li></ul><ul><li>Detection of pesticides or insecticides in food and water </li></ul><ul><li>Analyzing the dye composition of fibers in forensics, or </li></ul><ul><li>Identifying compounds present in a given substance </li></ul>
  8. 8. Gas Liquid Chromatography <ul><li>Involves a sample being vaporized and injected onto the head of the chromatographic column. </li></ul><ul><li>The sample is transported through the column by the flow of inert, gaseous mobile phase. The column itself contains a liquid stationary phase which is adsorbed onto the surface of an inert solid. </li></ul>
  9. 9. In the animation below the red molecules are more soluble in the liquid (or less volatile) than are the green molecules. Gas Chromatography
  10. 10. Uses <ul><li>Determination of volatile organic compounds (gases & liquids) </li></ul><ul><li>Determination of partition coefficients and absorption isotherms </li></ul><ul><li>Isolating pure components from complex mixtures </li></ul>
  11. 11. HPLC <ul><li>High Performance Liquid Chromatography. </li></ul><ul><li>Done at room temperature. </li></ul><ul><li>Advantageous for chemicals that are heat sensitive or volatile. </li></ul><ul><li>During this process, high pressure was used to generate the flow required for liquid chromatography in packed columns. </li></ul>
  12. 12. Uses <ul><li>It has the ability to separate, identify, and quantitate the compounds that are present in any sample that can be dissolved in a liquid. </li></ul><ul><li>HPLC can be, and has been, applied to just about any sample, such as pharmaceuticals, food, nutraceuticals, cosmetics, environmental matrices, forensic samples, and industrial chemicals. </li></ul>
  13. 13. Electrophoresis <ul><li>Similar to TLC, but the substance is separated through a gel by electric current. </li></ul><ul><li>It is reasonably easy and inexpensive. </li></ul><ul><li>Electrophoresis may be the main technique for molecular separation in today's cell biology laboratory. </li></ul>
  14. 14. Uses <ul><li>Used in cell biological laboratory for molecular separation. </li></ul><ul><li>A method of separating substances, especially proteins. </li></ul><ul><li>It is used for analyzing molecular structure based on the rate of movement of each component in a colloidal suspension while under the influence of an electric field. </li></ul>
  15. 15. Paper Chromatography <ul><li>Paper chromatography is an analytical chemistry technique for separating and identifying mixtures that are or can be colored, especially pigments. </li></ul><ul><li>In paper chromatography, the stationary phase is a very uniform absorbent paper. </li></ul><ul><li>The mobile phase is a suitable liquid solvent or mixture of solvents. </li></ul>
  16. 17. Uses <ul><li>They are used in many scientific studies to identify unknown organic and inorganic compounds. </li></ul><ul><li>They are used in crime scene investigation. </li></ul><ul><li>They are used for DNA and RNA sequencing as well. </li></ul>
  17. 18. R f Factor <ul><li>Retention Factor </li></ul><ul><li>It is a number that represents how far a compound travels in a particular solvent. </li></ul><ul><li>It is measured by measuring the ratio of the distance traveled by the compound and the distance traveled by the solvent. </li></ul>
  18. 19. Thank you…