Definitions and Separation Mechanisms - Basics

Lehrstuhl für Siedlungswasserwirtschaft Lehrstuhl für Siedlungswasserwirtschaft
Ingenieurfakultät Bau Geo Umwelt Ingenieurfakultät Bau Geo Umwelt

Technische Universität München Technische Universität München
Definition and Separation Mechanisms - Basics
PD Dr. J. Graßmann; PD Dr. T. Letzel


Definition: Chromatography is a physical method of separation in which the
components to be separated are distributed between two phases, one of which
is stationary (stationary phase) while the other (the mobile phase) moves in a
definite direction.
Mobile phase
Component mixture
Stationary phase
Separated components


Chromatographic methods can be classified according to
 the shape of the chromatographic bed
 the pyhsical state of the mobile phase
 the mechanism of separation

Shapes of the bed

Column Chromatography
A separation technique in which the stationary bed is within a tube. The particles
of the solid stationary phase or the support coated with a liquid stationary phase
may fill the whole inside volume of the tube (Packed Column) or be concentrated
on or along the inside tube wall leaving an open, unrestricted path for the mobile
phase in the middle part of the tube (Open-Tubular Column.
Planar Chromatography
A separation technique in which the stationary phase is present as or on a plane.
The plane can be a paper, serving as such or impregnated by a substance as
the stationary bed (Paper Chromatography, PC) or a layer of solid particles
spread on a support, e.g., a glass plate (Thin Layer Chromatography, TLC).
Sometimes planar chromatography is also termed Open-Bed
Chromatography.

Mobile Phases
Liquid Chromatography (LC): A separation technique in which the mobile phase
is a liquid. Liquid chromatography can be carried out either in a column or on a
plane.
Gas Chromatography (GC): A separation technique in which the mobile phase

is a gas. Gas chromatography is always carried out in a column.
Supercritical-Fluid Chromatography (SFC): A separation technique in which the

mobile phase is a fluid above and relatively close to its critical temperature and
pressure.

Separation Mechanism
• Adsorption Chromatography (Normal Phase and Reversed Phase)
• Partition Chromatography (or Liquid-Liquid-Chromatography)
• Size Exclusion Chromatography
• Ion Chromatography: Ion Exchange, Ion Pair, Ion Exclusion Chromatography
• Affinity Chromatography

• Adsorption Chromatography (Normal Phase and Reversed Phase)
• The retention mainly is based on the hydrophobicity of an analyte molecule


• Partition Chromatography (or Liquid-Liquid-Chromatography)
• The retention mainly is based on the hydrophobicity of an analyte molecule

• Size Exclusion Chromatography
• The retention mainly is based on the size of an analyte molecule

• Ion Chromatography: Ion Exchange, Ion Pair, Ion Exclusion Chromatography
• The retention mainly is based on the charge of an analyte molecule

• Affinity Chromatography
• The retention mainly is based on biological interactions

Schematic representation of an HPLC unit
(1) Solvent reservoirs, (2) Solvent degasser, (3) Gradient valve, (4) Mixing vessel for delivery
of the mobile phase, (5) High-pressure pump, (6) Switching valve in "inject position", (6')
Switching valve in "load position", (7) Sample injection loop, (8) Pre-column (guard column),
(9) Analytical column, (10) Detector (11) Data acquisition, (12) Waste or fraction collector.
von WYassineMrabetTalk Diese Vektorgrafik wurde mit Inkscape erstellt [GFDL (http://www.gnu.org/copyleft/fdl.html) oder CC BY-SA 3.0
(http://creativecommons.org/licenses/by-sa/3.0)], via Wikimedia Commons

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Lehrstuhl für Siedlungswasserwirtschaft Lehrstuhl für Siedlungswasserwirtschaft

Ingenieurfakultät Bau Geo Umwelt Ingenieurfakultät Bau Geo Umwelt

Definition and Separation Mechanisms - Basics

PD Dr. J. Graßmann; PD Dr. T. Letzel

Definition and Separation Mechanisms - Basics

PD Dr. J. Graßmann; PD Dr. T. Letzel

Definition and Separation Mechanisms - Basics

 the shape of the chromatographic bed

 the pyhsical state of the mobile phase

 the mechanism of separation

PD Dr. J. Graßmann; PD Dr. T. Letzel

Definition and Separation Mechanisms - Basics

Shapes of the bed

PD Dr. J. Graßmann; PD Dr. T. Letzel

Definition and Separation Mechanisms - Basics

Gas Chromatography (GC): A separation technique in which the mobile phase

Supercritical-Fluid Chromatography (SFC): A separation technique in which the

PD Dr. J. Graßmann; PD Dr. T. Letzel

Definition and Separation Mechanisms - Basics

PD Dr. J. Graßmann; PD Dr. T. Letzel

Definition and Separation Mechanisms - Basics

PD Dr. J. Graßmann; PD Dr. T. Letzel

Definition and Separation Mechanisms - Basics

PD Dr. J. Graßmann; PD Dr. T. Letzel

Definition and Separation Mechanisms - Basics

PD Dr. J. Graßmann; PD Dr. T. Letzel

Definition and Separation Mechanisms - Basics

PD Dr. J. Graßmann; PD Dr. T. Letzel

Definition and Separation Mechanisms - Basics

PD Dr. J. Graßmann; PD Dr. T. Letzel

Definition and Separation Mechanisms - Basics

Schematic representation of an HPLC unit

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