Chromatography:

It is a broad range of physical methods used to separate and or to analyze

complex mixtures. The components to be separated are distributed between two
phases: a stationary phase bed and a mobile phase which percolates through the
stationary bed.

How Does It Work?

A mixture of various components enters a chromatography process, and the
different components are flushed through the system at different rates. These
differential rates of migration as the mixture moves over adsorptive materials
provide separation. Repeated sorption/desorption acts that take place during the
movement of the sample over the stationary bed determine the rates. The
smaller the affinity a molecule has for the stationary phase, the shorter the time
spent in a column.
The Column
Although there are other types of chromatography (e.g. paper and thin layer),
most modern applications of chromatography employ a column. The column is
where the actual separation takes place. It is usually a glass or metal tube of
sufficient strength to withstand the pressures that may be applied across it. The
column contains the stationary phase. The mobile phase runs through the
column and is adsorbed onto the stationary phase. The column can either be a
packed bed or open tubular column.

Packed Bed Column

A packed bed column is comprised of a stationary phase which is in granular

form and packed into the column as a homogeneous bed. The stationary phase
completely fills the column.

Open Tubular Column

An open tubular column's stationary phase is a thin film or layer on the column
wall. There is a pasageway through the center of the column.

The Mobile and Stationary Phases

The mobile phase is comprised of a solvent into which the sample is injected.
The solvent and sample flow through the column together; thus the mobile phase
is often referred to as the "carrier fluid." The stationary phase is the material in
the column for which the components to be separated have varying affinities. The
materials which comprise the mobile and stationary phases vary depending on
the general type of chromatographic process being performed.
Gas Chromatography

The mobile phase in gas chromatography is generally an inert gas. The

stationary phase is generally an adsorbent or liquid distributed over the surface
of a porous, inert support.

Liquid Chromatography

The mobile phase in liquid chromatography is a liquid of low viscosity which flows
through the stationary phase bed. This bed may be comprised of an immiscible
liquid coated onto a porous support, a thin film of liquid phase bonded to the
surface of a sorbent, or a sorbent of controlled pore size.

Carrying out thin layer chromatography

Background

Chromatography is used to separate mixtures of substances into their

components. All forms of chromatography work on the same principle.

They all have a stationary phase (a solid or a liquid supported on a solid) and a
mobile phase (a liquid or a gas). The mobile phase flows through the stationary
phase and carries the components of the mixture with it. Different components
travel at different rates. We'll look at the reasons for this further down the page.

Thin layer chromatography is done exactly as it says - using a thin, uniform layer
of silica gel or alumina coated onto a piece of glass, metal or rigid plastic.

The silica gel (or the alumina) is the stationary phase. The stationary phase for
thin layer chromatography also often contains a substance which fluoresces in
UV light - for reasons you will see later. The mobile phase is a suitable liquid
solvent or mixture of solvents.

The Rf value

Rf is the retention factor, or how far up a plate the compound travels.