Column Chromatography

Chemistry is a subject of reasons that tells us why substance combines with different elements to form a new component. It is a science of invention and discovery. For many people, chemistry is a mystery, but for few, it is a subject for creating new things. One of the essential parts of chemistry is separation and formation of substance. And column chromatography is a part of such theory which we will know in detail in this article.


In the article, we will know about column chromatography, column chromatography principle, applications of column chromatography, and types of column chromatography, advantage and disadvantage of chromatography, and many more topics. Before diving into the big topic, let us understand from the basics.

Column Chromatography Definition 

Column chromatography is used by an organic chemist to separate liquids and solids from a solution. In a column of absorbents which is silica gel or it may be alumina gets loaded in a column filled with impurities when conducting an experiment. This mixture flows down the column, and the component of the sample receives separated by splitting between the mobile eluent and stationary packing material. In simple terms, column chromatography is the separation of substances from a solution to get the simpler substances from the complex elements. There are five types of column chromatography which are used in the process of separation. The process goes through two phases, that is the mobile phase and the stationary phase.

Types of Column Chromatography

Five types of chromatography can be applied to the basic principles of chromatography. Gas chromatography (GC), chiral liquid chromatography (LC), Ion exchange chromatography (IEC), and size exclusion chromatography (SEC), these are the five methods that use the column in column chromatography.

In the gas chromatography, the mobile phase is a gas, and in liquid chromatography, the liquid is used in the mobile phase. Ion exchange is used in the stationary phase to ionize and separate the ion and molecules. Size exclusion chromatography (SEC) and chiral chromatography are complex ways of separation of molecules in mixtures that are generally used for large samples.

Principle of Column Chromatography


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The Figure shows the working of Column Chromatography

Principle of Column Chromatography

A sample mixture is taken, which is set on the top of the column. This mixture is made to absorb on the top of the stationary phase.  The stationary phase is the next step after the mobile phase for the separation of the mixture in the column. The separation of the compounds happens due to the polarization of the molecules, and the speed rate moves the particles and separates them in the mixture. The polarization of the molecules in the mix initiates the separation of the components in the mixture. After the mobile phase, the molecules get collected in small fractions in a test tube, and then isolation and purification follow. Through a rotary evaporator, salt is separated from the mixture and another compound is separated. The process of column chromatography used for separation and purification of the compound in a mixture. It is a convenient method of separation and purification of substance in a mixture that is widely used by a chemist. One has to have all the focus and attention in the process. You can automate the process, but the cost increases, so the manual way is much preferred by the chemist.

Application of Column Chromatography

  • Column chromatography is used in the purification of compounds.

  • Separation of molecules for a mixture and using it in the formation of a new substance.

  • Used by a chemist to know the drug estimate in a drug solution.

  • It is used to separate diastereomers, isolate racemate and separate geometrical isomers

  • Used for the isolation of metabolic fluid from a biological fluid.

Advantages of Column Chromatography

  • The process involves the separation of any mixture

  • Separation of impurities from any mixture.

  • Any variety and quantity separation is possible.

  • The process involves low cost and simple understanding.

  • All varieties of solvents can be used in the process which will give you the required result. 

  • The process can be fully automated, which is done by big laboratories.

 

Disadvantages of Column Chromatography

  • It is a lengthy and time-consuming process.

  • Small quantities become inadequate in the separation process; the only substantial amount can be used for separation.

  • The process is expensive in comparison to the thin paper column chromatography process.

  • It is a long process, so attention and focus are required throughout the process.

  • If the process is automated, then the process becomes costly.

FAQ (Frequently Asked Questions)

1. How does Column chromatography improve separation?

There are five methods by which column separation is done. All these methods have their process advantage which means liquid chromatography can easily increase the method of polarization of substances in the impure mixture. And after that the stationary phase which is the phase of separation becomes easy. The polarization of the molecules in the mix initiates the separation of the components in the mixture. After the mobile phase, the molecules get collected in small fractions in a test tube, and then isolation and purification follow. Through a rotary evaporator, salt is separated from the mixture and another compound is separated. The process of column chromatography used for separation and purification of the compound in a mixture.


2. What is the duration of the process of the column chromatography?

It depends upon the separation you see on TLC. When you are good at it, it takes about 30-60 minutes, as I said though if you have a lousy separation on TLC, it can take longer. The longest time it took me was 12 hours, but that was with 40kg of silica reverse phase.