THE ORIGIN OF CHROMATOGRAPHY
It was only when the process of chromatography was devised that complex
separations as required in the above biochemical example could be achieved.The word chromatography is Greek in origin (chroma - colour; graph -writing). The technique of chromatography was first applied to the separationof coloured pigments and dyestuffs, as evident from the pioneering work of the Russian chemist Mikhail Tswett who in 1906 separated the pigments from leaves into coloured bands by passing a solution of the pigments down a glass tube packed with powdered chalk.
The separation of pigments that Tswett achieved was due, not to their colour,but to the differing capacities of the various pigments to attach themselves to the chalk (called ‘adsorption’).
The most highly adsorbed pigment remains close to the top of the column and the least adsorbed pigment is ‘washed through’ to form the lowest band in the column. This technique was named as column chromatography.However, in this unit we shall be studying the relatively cheap and the simple laboratory method of paper chromatography.
You will also do an experiment based on chromatography during practical sessions. Let us now study the language of these techniques.
TERMINOLOGY OF CHROMATOGRAPHY
In chromatography, various terms are frequently used and they form the language of chromatographic techniques. These are explained below:
i) ‘Chromatography’
The term chromatography is applied to the process which involves the distribution of a sample between a stationary phase and a mobile phase.The historical origins of the word chromatography have already been discussed.
All types of chromatography have certain basic features,although from the design of the apparatus, this may not be apparent.
The basic methodology involved is as follows:
The mixture to be separated is added to a moving ‘solvent’ which may be liquid or gaseous. This moving stream, now containing components, is passed over or through a fixed medium, which is specially designed to separate theindividual constituents in the original mixture. However, there are many variations and hence in many cases, it is difficult to see the common link between the various chromatographic methods.
ii) ‘Mobile Phase’ and ‘Stationary Phase’
The moving component in the chromatographic technique is called the mobile phase, which is normally a liquid or mixture of liquids, except in gas chromatography where a gas is employed.
The fixed medium is called the stationary phase. Thus, if the sample consisting of a mixture of components (usually referred to as solutes) is added to the mobile phase, then the components by a variety of physical process will be carried along in the moving
stream to varying degrees. We will study about it in more detail later in this Unit.
iii) Retention Factor
A quantitative measure of the movement of each component can be obtained by specifying its retention (or retardation) factor (or Rf value) which is defined by the following relationship:
Distance movedby thecentreof the solute spot
Rf = -------------------------------------------------------------
Distance movedby solvent front
TYPES OF CHROMATOGRAPHY
Table classifies some of the chromatographic methods. However, thenature of the process whereby separation takes place is rarely confined to one physical mechanism; and although the table indicates the nature of the distribution process in each case, this is merely the most important factor.
APPLICATIONS OF CHROMATOGRAPHY
Chromatography is used to separate the constituents (usually organic but not entirely exclusive) of a mixture. The green colouring matter of leaves may be separated into their constituents, namely chlorophyll and xanthophyl (the yellow pigment). You will be performing an experiment based on Chromatography in the lab during practical sessions. Chromatography is also used to isolate, purify or identify the components of complex mixtures in the laboratory both for analytical and preparative purposes.
Chromatography is used in labs right from the simple techniques such as paper chromatography to the sophisticated ones such as HPLC and Gas chromatography which find applications in research labs and industries.