Chromatography:Chromatography is an analytic technique developed long ago. In early stages its applications and efficiency was much limited and the system was rudimentary. Therefore the chromatography systems were greatly modified and developed by the use of available technology to meet the growing demands of scientific research and analysis. With these modifications carried over, there is an improved efficiency in the separation and also the quantification. Further the range of and type of substances which could be analysed has been greatly increased due to advancement in chromatography techniques.
Types of chromatography:
The differentiation is based on techniques of chromatography or principle or physical changes used
- The physical states of stationary phase and mobile phases.
- The on principle of separation used.
- The chemical nature of stationary phase and mobile phases used (polarity).
- Based on the shape of stationary phase employed.
- Based on purpose of chromatography experiment.
- Based on physical or chemical character of the stationary phase.
1. Based on Physical state of both phases: These are broadly classified as homogeneous or heterogeneous. The chromatography systems differ based on the physical states of the phases used.
♠ Homogeneous techniques have both stationary phase and mobile phase as liquid. Ex: Liquid-liquid chromatography.
♠ Heterogeneous techniques employ different stationary and mobile phases. Ex: Solid-liquid chromatography, Solid-Gas chromatography, Liquid-Gas chromatography etc.
Here the principle used in separation is considered i.e adsorption method or partition method.
♠ Adsorption chromatography: Here the sample molecules get separated due to greater affinity to adsorb the solid stationary phase compared to that of mobile phase. This principle works when stationary phase is a solid and mobile phase is liquid.
♠ Partition chromatography: Here the samples molecules get separated due to relative differences of dissolution and partition into different phases/layers. The molecules with greater partition or dissolution into mobile phase is separated faster while that with partition into solid phase liquid moves slower or latter. Here both stationary phase and mobile phase are liquid in nature or liquid as stationary phase and gas as mobile phase. The liquid on stationary phase exists as a thin layer on a solid background.
3. Based on the chemical nature of stationary phase and mobile phase:
This differentiation is based on chromatography column i.e the nature of stationary phase inside the column
♠ Normal phase chromatography: Here the stationary phase is polar in nature and mobile phase is in non-polar nature. Hence on elution, non-polar compounds are eluted first and polar compounds later as they have greater affinity to stationary phase. Mostly used in column chromatography technique
Ex: Normal phase column chromatography.
♠ Reverse phase chromatography: This is reverse to the above method. The stationary phase is non-polar and mobile phase is polar in nature. In practice this is highly used to analyse as most of the substances like drugs etc. used in our daily life are polar in nature.
Ex: Reverse phase HPLC systems.
4. Based on the shape of stationary phase: The shape of stationary phases depends on the support used to place the stationary phase. Hence based on the shape of stationary phase, there are two types like
Column chromatography and planar chromatography.
♠ Columnar chromatography is one where stationary phase is column shape. It is widely used in types like High pressure liquid chromatography (also medium pressure liquid chromatography), Column chromatography, Gas chromatography etc. Development of chromatogram occurs in volume aspect.
♠ Planar chromatography is one wherein a stationary phase is flat. The development occurs on the planar surface (only area).
This type of chromatography is used in Thin layer chromatography (TLC), High Pressure thin layer chromatography(HPTLC) and Paper chromatography.
5. Based on purpose of chromatography experiment: This is one of the types of chromatography Here the idea of experiment different. This can be done on both planar type and columnar type of chromatography. The types are
♠ Preparative chromatography: The amount of sample injected or applied is very large and the separated and pure component is collected for use. The desired component of sample is not disposed off. This is also exclusively applied in column types as preparative column chromatography.
♠ Analytic chromatography: Here the sample size applied or injected is very small and the intention is aimed to identify the components in the sample and also their individual concentrations in the sample. The eluted sample from the outlet is disposed off.
6. Based on physical or chemical character of the stationary phase:
This is especially followed in columnar chromatography where in the stationary phase used has specific character like being porous or charged.
♠ Size exclusion chromatography: Here the stationary phase has pores in its matrix. When molecules are allowed to pass through, those with large size travel a short path under mobile phase influence and pass out of column first and vice-verse.
♠ Ion exchange chromatography: Here the stationary phase has a definite charged ions. When the sample is passed through, it retains all the molecules with opposite charge and leaves off molecules with same charge. So to elute the bound molecules, you need to pass another mobile phase with similar charge to stationary phase to recover the bound molecules. (like molecule displacement method).
So over all available chromatography techniques for regular analysis include.
a) Cloumn chormatography
b) High performance liquid chromatography (HPLC)
c) Gas chromatography (GC)
d) Ion-exchange chromatography
e) Size exclusion chromatography.
f) Thin layer chromatography (TLC)
g) High performance thin layer chromatography (HTLC)
h) Paper chromatography.
i) Affinity chromatography.