There are many types of chromatography like HPLC, Gas chromatography, paper chromatography, thin layer chromatography, etc.
But in this article we will discuss all the instrumental methods and theoretical classification. So go through the article till the end.
Chromatography is an analytic technique based on the separation of molecules of a sample over two phases. These phases are called as the stationary and the mobile phase.
In the early stages, it was developed as column chromatography. But its applications and efficiency were much limited and the system was rudimentary.
The time span required for analysis was too long like taking up to few hours or days for a run. Also the range of compounds that could be analyzed or separated was low and the sample quantity required was high.
Then the method was highly modified and developed with an ability to measure wide range of substances. That too small quantities of sample like in nano-grams or pictograms could be measured at faster rate.
We assume that you know what are the basic principles involved. If not please go through the article “What is Chromatography?“.
Different Types of chromatography
Based on Instrumentation and procedure we have 10 types as
1. Column chromatography: It is the most widely used chromatography which find daily use in research labs and industries. It is easy to operate and less expensive technique. As the name indicates, there is a lengthy column which is suspended in the air with the help of a stand.
Into the column, the stationary phase is packed and mobile phase is allowed to flow taking the sample through the solid column.
Similar to the picture shown below, you pour mobile phase from top of the column to flow through the sample present on stationary phase in the column to get separated.
2. High performance liquid chromatography (HPLC): This is similar to column chromatography but high pressure is employed for effective separation of compounds. The mobile phase is pumped into the column at a defined pressure and further the column particles are very small so the surface area is high and better separation takes place. See more about the HPLC principle and method in detail. The application of pressure reduces the time of run. The pressure is applied by use of pumps. For better separation, enhanced stationary phases and mobile phases are incorporated. Due to this modification large molecules like proteins, fats and small molecules like monoamines can be separated efficiently.
3. Gas chromatography (GC): Here gas is used as mobile phase and a solid or liquid layer as the stationary phase. For details see gas chromatography principle.
4. Ion-exchange chromatography: Here the mobile phase is charged and sample molecules with similar charge present on the charged stationary phase get eluted out as the mobile phase molecules with charge displace them.
You can read more about ion exchange chromatography.
5. Size exclusion chromatography. Here the column is loaded with charged some gel having pores. Sample particles when poured along with mobile phase have to pass through the sieve like net work of the stationary phase. In doing so, the larger particles elute out first and smaller ones last. The reason is the smaller ones take longer path in the column stationary phase while larger particles take short path to elute out. Find more about gel permeation chromatography.
6. Thin layer chromatography (TLC): Here the stationary phase is a thin layer and plate like. The plate is immersed in a solvent up to a small height so that the mobile phase travels up leading to separation of compounds.
7. High performance thin layer chromatography (HPTLC): Similar to TLC but more efficient.
8. Paper chromatography: The sample is marked on the whatmanns paper and then placed in the solvent vessel to allow it to percolate through the paper. In doing so, it separates the sample over the paper surface which is then subjected to color formation for identification.
9. Super critical fluid chromatography: It is a normal phase chromatography with instrumentation similar to HPLC. Here mobile phase is mostly a super critical fluid like carbon dioxide.
10. LC-MS: Liquid chromatography combines with Mass spectroscopy (detector)
GC-MS: Gas chromatography combined with Mass spectroscopy (detector)
The types of chromatography mentioned are instrumental based. But theoretically the classification can be done based on other facts.
Different types of chromatography based on
- The physical states of stationary phase and mobile phases.
- Based 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.
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.
Based on the principle of separation used
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 solvent.
♠ 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 are 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.
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. The chromatography affinity for the sample is decided by both stationary phase and mobile phase combination.
♠ 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. Read the details on Normal Phase Chromatography.
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 reversed phase chromatography is highly used in routine analysis as most of the substances like drugs etc. used in daily life are polar in nature.
Ex: Reverse phase HPLC systems.
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.
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.
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 the molecules 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 the above mentioned types of chromatography are theoretically classified. But practically we have only 10 types of chromatography.