Adsorption Chromatography-Concept of Adsorption Chromatography, Factors affecting, Types -Thin Layer Chromatography, Column Chromatography, Application, Practice Problems, FAQs, 

Do you know that a plant leaf contains different coloring agents called pigments? Some of the pigments are xanthophylls (yellow), carotene (orange), and chlorophyll (green). How do separate them?

To separate the different pigments present in plant leaves the technique "adsorption chromatography" was used for the first time by Mikhail Tsvet, a Russian-Italian botanist, a century ago. The word "chromatography" means “drawing colors”, and the word "adsorption" means sticking to the surface. 

This article will help you understand more about adsorption chromatography and its importance.

Table of content:

• Adsorption Chromatography
• Key Concept of adsorption Chromatography
• Adsorbent
• Apparatus required
• Demonstration
• Factors affecting chromatography process
• Thin layer Chromatography
• Column chromatography
• Application of adsorption chromatography
• Practice Problems
• Frequently asked questions

Adsorption Chromatography

Adsorption chromatography is a method used for the separation of components from a mixture using their differential adsorption on a stationary component applied over a stationary solid surface.

The process was invented by Russian botanist Mikhail Tsvet. He used this procedure to separate chlorophyll and Carotenoid pigment. He passed a leaf sample through a wall of calcium carbonate/, alumina/, and sucrose and then rinsed it using a petroleum mixture.

Two atoms or molecules can interact through strong chemical or weak physical forces like van der walls or coulombic forces. The physical forces of interaction depend on the nature of the interacting molecules. So, when a mixture of substances is passed over a fixed (stationary) surface, each of them interacts with the fixed surface with differing strength - strong force of interaction makes them adsorbed on the top of the column. Molecules are adsorbed subsequently down the column as per their decreasing forces of interaction with the solid surface. On passing a suitable solvent, the least adsorbed molecule at the bottom of the column comes out first, followed in series by the next components facilitating separation of them

Adsorption chromatography involves several materials with unique terms as follows-

ADSORBENT

The adsorbent is a material that is porous and has a large surface area for the absorption of substances by intermolecular forces. Some examples are silica gel, cellulose microcrystalline, etc. To understand the procedure of the adsorption chromatography, let us understand the various components involved in this method. Since this is fixed it is called a STATIONARY PHASE.

MOBILE PHASE: Either a liquid or gas solvent is used as the mobile phase. It is made to pass through the column and the solutes based on their compound rate of interaction get separated from the solvent.

Factors affecting the adsorption chromatography

• Choice of the adsorbent
• Selection of the solvent for the mixture
• The rate of flow of the solvent
• The temperature of the system
• The column height for the procedure.

Types of Adsorption Chromatography

There are majorly four types of adsorption chromatography.

1. Thin Layer Chromatography
2. Solid Liquid Chromatography
3. Gas-Liquid Chromatography
4. Column Chromatography

Thin Layer Chromatography

In this method, the adsorbent used is a thin plate, and the adsorption takes place by virtue of the process of differential migration that takes place when the solvent moves upward along the surface of powdered spread on the glass plates.

Demonstration

• A clean and dry chromatographic jar is taken.
• The environment in the chromatographic jar is saturated using solvent vapours, and a paper soaked in the mobile phase is attached to the walls of the jar.
• The jar is then closed after adding the mobile phase into it.
• The equilibrium in the system is maintained throughout.
• The baseline is marked on the adsorbent.
• The sample is applied to the TLC plate with the help of a capillary tube, and it is then left to dry.
• The plates are then placed in the jar, and the lid of the jar is closed.
• The solvent is allowed to move from the baseline, and then the TLC plates are taken out and allowed to dry.

Apparatus Required

• Chromatography jar used to maintain the required temperature for smooth functioning of the process.
• Chromatography plate: A thin-layered Borosilicate glass plate of sizes 20cm 20cm; 20cm 10cm; 20cm 5cm
• Mobile Phase: The solvent (gas or liquid depending on the type of chromatography to be performed)
• Stationary phase: The adsorbent.

Solid-Liquid Chromatography

This method involves the use of a liquid as the mobile phase in the system.

Gas-Solid Chromatography

This method involves the use of gas as the mobile phase in the setup.

Column Chromatography

This method involves the use of a column composed of walls laden with the adsorbent material. The solvent is made to pass through this column and the solutes get adsorbed in different layers based on their affinity toward adsorption. The material with the strongest affinity gets adsorbed at the top of the column and those with lesser affinity get adsorbed at the bottom of the column.

Application of Adsorption chromatography

• Used for the isolation of antibiotics
• Used in the separation of amino acids
• Used to identify various carbohydrates
• Used to identify fatty acids and fats
• Used to isolate peptides and proteins.

Practice Problems

Q1: In adsorption chromatography, which of the following can be used as an adsorbent?

A. Alumina
B. Silica
C. Both A and B
D. None of the above

Solution: C)

The differential adsorption of distinct components of a mixture on a suitable adsorbent such as silica gel or alumina is the basis of the adsorption chromatography method.

Q2: Which statement is incorrect? 

A. Colorless compound spots are invisible to the naked eye
B. Under UV light, spots can be detected
C. In a closed jar of iodine crystals, spots are detected
D. None of the above

Solution : D)

The spots of colourless compounds are not visible to eyes. So UV light or crystals of iodine are used to identify spots.

Q3: Which expression is used for the relative adsorption of mixtures? 

A.  Flow rate
B. Acceleration factor
C. Retardation factor
D. Capillary action

Solution: C)

Retention factor is the ratio of the distance moved by a substance from the baseline to the distance moved by a solvent from the baseline.

Q4: Which spraying agent is used to detect amino acids? 

A. NaCl solution 
B. Ninhydrin solution
C. Ferric chloride
D. Berlin blue reagent

Solution: B)

Ninhydrin solution is used as a spraying agent in TLC to detect the presence of amino acids.

Q5: Which colour spots are seen in the iodine chamber? 

A. Purple
B. Red
C. Brown
D. Pink

Solution: C)

Yellowish-brown colour spots are seen because of the complex formation of iodine.

Frequently asked questions

Question 1. What is the Retention factor?
Answer: The retention factor is the ratio of the distance moved by a substance from the baseline to the distance moved by a solvent from the baseline.

Question  2. Why is the retention factor value always less than 1? 
Answer: Rf value close to 1 indicates that both substance and the solvent rise at the same distance from the baseline which is unreliable. It is also difficult to separate these substances. When the solvent is too polar for the sample, the ratio will be close to 1.

Question  3. What is the difference between adsorption and partition chromatography?
Answer: When the separation is based on the adsorption is adsorption chromatography and in partition, chromatography separation is done based on partition.

Question  4. Why TLC is preferred over column chromatography?
Answer: Thin layer chromatography takes less time and less quantity of sample and solvent but column chromatography takes more time and more quantity of solvent.

Question  5. What are the disadvantages of adsorption chromatography?
Answer: The adsorption chromatography technique is more costly and complicated. Some of the samples require retention time to separate and the major disadvantage is reproducing some results given by adsorption chromatography.

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