Lecture 5

Lecture 5 - Introduction to Chromatographic Separations Why...

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1 Introduction to Chromatographic Separations Why separation? What is the molecular basis for separations? How are separations achieved? Why Separations • Analytical and preparative needs • For analysis LOD to lower concentrations selectivity b structural elucidation – Eliminate interferents in analysis – Purify pure compounds at small quantity – … Chromatographic Separation of a Mixture of A and B What do you observe?
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2 Molecular basis for separations Fundamental molecular properties – Physical properties • Size, charge, shape – Chemical properties • Reactivity – Specific molecular recognition • Affinity Nonbonding molecular interactions – Important • Separations of molecules • Governing solubility, volatility – Types • H-bonds • Electrostatic interactions – Charge, permanent dipole, transient dipole • Hydrophobic forces How are separations achieved? • Differentiate molecules according to their properties/interactions at a particular scale (i.e., a separation domain) – Differential migration rate in space • Molecules migrate at different rates • Result in spatial difference in concentration • It is important to understand the molecules to be separated, so that an appropriate method can be selected or several methods can be selected and assembled Differential migration rate in space • Molecules migrate at different rates – Result in spatial difference in concentration • Molecular properties to use – Charge: electrophoresis, ion exchange – Charge-size: zone electrophoresis, gel electrophoresis – Size: filtration, size exclusion – Hydrophobicity: reversed-phase
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3 Partitioning between two phases • Liquid-Liquid – Extraction • Solid-liquid – Solid-phase extraction • Liquid-gas – Distillation Phase A Phase B W G G W W G Partitioning between phases + Phases move relatively to each other • Liquid chromatography (solid-liquid partitioning) • Gas chromatography (Liquid-gas partitioning) • Micellar electrokinetic chromatography Two Important Processes in Chromatographic Separation • Differential migration • Band broadening
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Two Ways to Improve Separation • Increase differential migration (b) • Decrease band broadening (c) Basis of Retention and Differential Migration • Partition coefficient – K = C s /C m • The rate of analyte migration υ = u · R = (L/t m ) [1/(1 + K (V s /V m )] Mobile phase B m B s A m A s Stationary phase Retention • Retention time, t R – Time to travel through the column for analytes partitioning between mobile phase and stationary phase • Dead time, t
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This note was uploaded on 10/01/2009 for the course CHEM 334 taught by Professor Lei during the Spring '09 term at UConn.

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Lecture 5 - Introduction to Chromatographic Separations Why...

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