Chromatography

  Analytical Techniques


 

In this series of lecture, we will study various types of Chromatography.

Before introducing chromatography we should know that it is one of the analytical technique out of many techniques available for testing, seperating or purifying any component.

Various broad categories of analytical techniques are listed below:

Gravimetric techniques : Gravimetric analysis is a technique through which the amount of an analyte (the ion being analyzed) can be determined through the measurement of mass.Gravimetric analyses depend on comparing the masses of two compounds containing the analyte

Titrimetric analysis : Method of analysis in which a solution of the substance being determined is treated with a solution of a suitable reagent of exactly known concentration. The reagent is added to the substance until the amount added is equivalent to the amount of substance to be determined

Spectroscopic techniques : employ electro-magnetic radiation (light) to interact with matter and thus probe certain features of a sample. Colorimetry, in which a sample absorbs visible light, is one example of a spectroscopic method of analysis

Electrochemical techniques : methods in which a concentration of analyte is measured by measurement of potential, current, or charge in an electrochemical cell which serves as the analytical signal

Chromatographic Techniques : separation of a sample into its constituent parts because of the difference in the relative affinities of different molecules for the mobile phase and the stationary phase used in the separation

  Chromatography


Definition

Chromatography is an analytical technique commonly used for separating a mixture of chemical substances into its individual components, so that the individual components can be thoroughly analyzed

Chromatography is a combination of two words;

  * Chromo – Meaning color

  * Graphy – representation of something on paper

 

 

It is a physical method of separation in which the components of a mixture are separated by differences in their distribution between two phases, one of which is stationary (stationary phase) while the other (mobile phase) moves through it in a definite direction. The substances must interact with the stationary phase to be retained and separated by it

 

Basic Terminology in Chromatography

Phases in Chromatography

Analytical Separation

  • By the physical state of the mobile phase and stationary phase;
  • By the method of contact between the mobile phase and stationary phase;
  • Or by the chemical or physical mechanism responsible for separating the sample’s constituents

The mobile phase is usually a liquid or a gas, and the stationary phase, when present, is a solid or a liquid film coated on a solid surface

Chromatographic techniques are often named by listing the type of mobile phase, followed by the type of stationary phase

Thus, in gas–liquid chromatography the mobile phase is a gas and the stationary phase is a liquid. If only one phase is indicated, as in gas chromatography, it is assumed to be the mobile phase

  Techniques by chromatographic bed shape


Two common approaches are used to bring the mobile phase and stationary phase into contact

In column chromatography, the stationary phase (bed) is placed in a narrow column through which the mobile phase moves under the influence of gravity or pressure. The stationary phase is either a solid or a thin, liquid film coating on a solid particulate packing material or the column’s walls

In planar chromatography the stationary phase (in a plane) coats a flat glass, metal, or plastic plate and is placed in a developing chamber. A reservoir containing the mobile phase is placed in contact with the stationary phase, and the mobile phase moves by capillary action.

 

Type of Chromatography based on polarity of phase

Normal-phase chromatography

implying that our stationary phase is polar (hydrophilic) in nature and our mobile phase is non-polar (hydrophobic) in nature

Reverse-phase chromatography

where the scenario is reversed i.e. the stationary phase is non-polar while the mobile phase is polar

 

Basic Principle of Chromatography : Differential Affinities

 

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