CHM260 - Separations Methods

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Transcript of CHM260 - Separations Methods

  • Chromatography Is a technique used to separate and identify the components of a mixture. Works by allowing the molecules present in the mixture to distribute themselves between a mobile and a stationary phase. mobile phase = solvent or gas stationary phase = column packing material 2
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  • How separation occur? Chromatography is a powerful separation method that is used to separate and identify the components of complex mixtures. Works by allowing the molecules present in the mixture to distribute themselves between a stationary and a mobile phase to varying degrees. Those components that are strongly retained by the stationary phase move slowly with the flow of mobile phase. 5
  • How separation occur? In contrast, components that are weakly held by the stationary phase travel rapidly (fast). As a consequence of these differences in mobility, sample components separate into discrete bands that can be analyzed qualitatively and/or quantitatively. 6
  • Classification of Chromatographic Methods1. Based on physical means The way stationary and mobile phases are brought into contact.2. Based on the types of mobile phase Either gas, liquid or supercritical fluid.1. Based on the kinds of equilibria involved in the in solute transfer between the phases Interaction of analyte between stationary and mobile phases. 7
  • Chromatographic Methods based on physical means Column Planar chromatography chromatographystationary phase is stationary phase isheld in narrow tube; supported on a flat platemobile phase moves by or in the interstices of apressure or gravity paper; mobile phase moves through capillary action or gravityExample: Example:Gas chromatography (GC) Thin-layer chromatographySupercritical-fluid (TLC)chromatography (SFC) Paper chromatography (PC) 8
  • Chromatography based on types of mobile phase Mobile Phase Gas (SupercriticalGas Chromatography fluid) Supercritical-fluid Chromatography (Liquid) Liquid Chromatography 9
  • Chromatography based on interaction of the analyte with stationary phase1. Adsorption - of solute on surface of stationary phase; for polar non-ionic compounds.2. Ion Exchange - attraction of ions of opposite charges; for ionic compounds. 1. Anion - analyte is anion; bonded phase has positive charge. 2. Cation analyte is cation; bonded phase has negative charge. 10
  • Chromatography based on interaction of the analyte with stationary phase3.Partition - based on the relative solubility ofanalyte in mobile and stationary phases. a. Normal phase stationary phase polar, the mobile phase nonpolar. b. Reverse phase stationary phase nonpolar, the mobile phase polar.3.Size Exclusion separate molecules by size;sieving- stationary phase is a porous matrix. 11
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  • Classification of Chromatographic Methods Chromatography Partition Adsorption Ion-exchange Size-exclusionLiquid- Gas-liquid Liquid-solid Gas-solid Liquid-solid Liquid-solidliquid 13
  • AdsorptionChromatography 14
  • Adsorption Chromatography Components of the mixture selectively adsorb (stick) on the surface of a finely divided solid stationary phase. As mobile phase (gas/liquid) carries the mixture through the stationary phase, the components of the mixture stick to its surface with varying degrees of strength and thus separate. Stationary phase: solid Mobile phase: gas or liquid 15
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  • PartitionChromatography 17
  • Partition Chromatography Accomplished by selective and continuous transfer of the components of the mixture back and forth between a liquid stationary phase and a liquid mobile phase as the mobile phase liquid passes through the stationary phase liquid. Stationary phase: liquid Mobile phase: liquid or gas 18
  • Partition Chromatography Partitioning is a distribution (by dissolving) of the components between 2 immiscible phases. Seperations of the components will be based on relative solubilities of the components in the mobile and stationary phase. Example of partitioning using polar stationary phase. Polar components will retain longer than the non-polar components. Non-polar components will move quickly through stationary phase and will elute first before the polar components and vice-versa. 19
  • Partition Chromatography The stationary phase actually consists of a thin film adsorbed (stuck) on or chemically bonded to the surface of a finely divided solid particles. If the mobile phase is gas, the volatility (vapor pressure) and solubility in stationary phase plays an important role. 20
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  • Ion-exchangeChromatography 22
  • Ion exchange Chromatography Method for separating mixture of ions. Sample is aqueous solution of inorganic ions or organic ions Stationary phase are small polymer resin beads usually packed in a glass tube. a. These beads have ionic bonding sites on their surfaces which selectively exchange ions with certain mobile phase compositions as the mobile phase penetrates through it. 23
  • Ion exchange Chromatography Ions that bond to the charged site on the resin bead are separated from organic or inorganic ions aqueous solution. The process is repeated several times by changing of the mobile phase composition. 24
  • Ion exchange Chromatography The process begin with initially running the analysis using a mobile phase with all the ions in the mixture. The mobile phase is then change for several times in a stepwise fashion so that one kind of ion at a time is removed. The process is repeated until complete separation achieved. 25
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  • Size-exclusionChromatography 27
  • Size exclusion ChromatographyAlso called gel permeation chromatography.Separation technique of dissolved species is based on the size of the components.Stationary phase: porous polymer resin particles (molecular sieves).The components to be separated enter the pores of these particles and are slowed down from progressing through this stationary phase. 28
  • Size exclusion Chromatography Separation depends on the sizes of the pores relative to the sizes of the molecules to be separated. Small particles are retarded to a greater extent than large particles (some of which may not enter the pores at all) and separation occurs. Particles with size bigger than the pore size will be eluted first from the column. 29
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  • Terminologies in chromatography 31