Chromatography is a laboratory technique for separation of mixtures. The mixture is dissolved in a fluid referred to as the mobile phase, which carries it through a structure holding another material called the stationary phase. The various constituents of the mixture travel at different speeds, causing them to separate. Chromatography was invented by the Russian botanist, Mikhail Tsvet. In chemistry, the process is used to identify unknown substances by separating them into the different molecules that make them up.
What Is Paper Chromatography?
Paper chromatography is a type of chromatography procedure which runs on a piece of specialized paper. It is used in both qualitative and quantitative analysis of both organic and inorganic samples. In this technique, it is necessary for the different chemicals in the solution to have different properties such as molecule size or a different ability to dissolve in a solvent. The stationary phase will absorb or slow down different components of the tested solution to different degrees creating layers as the components of the solution are separated.
The technique uses two types of substances:
- Mobile phase: A gas or liquid that transports the solution being tested through the other substance (water, rubbing alcohol are examples).
- Stationary phase: The liquid or solid through which the tested substance is carried (coffee filter paper, paper towel are examples).
Modes/Types Of Paper Chromatography
- Ascending chromatography
- Descending chromatography
- Ascending-descending mode
- Radial mode
- Two-dimensional chromatography
Uses Of Paper Chromatography
- Used for the separation of a mixture having polar and non-polar compounds
- It is used to determine organic compounds in urine.
- It is widely used in separating multi-component pharmaceutical formulations.
- Used for evaluation of inorganic compounds like salts and complexes.
What You Need To Know About Paper Chromatography
What Is Thin Layer Chromatography?
Thin-layer chromatography (TLC) is a chromatography technique used to separate non-volatile mixtures. Thin-layer chromatography is performed on a sheet of glass, plastic or aluminum foil, which is coated with a thin layer of adsorbent material, usually silica gel, aluminum oxide (alumina) or cellulose. This layer of adsorbent is also referred to as the stationary phase. On completion of separation, each component appears as spots separated vertically. TLC is one of the fastest, least expensive, simplest and easiest chromatography techniques.
Thin-layer-chromatography (TLC) is a very commonly used technique in synthetic chemistry for identifying compounds, determining their purity and following the progress of a reaction. It also permits the optimization of the solvent system for a given separation problem.
Uses Of Thin Layer Chromatography
- Used to check the purity of samples
- Identification of compounds like acids, alcohols, proteins, alkaloids, amines, antibiotics etc.
- Helps in isolation of most of the compounds.
- It is used in food industry to separate and identify colors, sweetening agent and preservatives.
- Used in biochemical analysis such as separation or isolation of biochemical metabolites from its blood plasma, urine, body fluids, serum etc.
What You Need To Know Thin Layer Chromatography
What Is Column Chromatography?
Column chromatography is a technique which is used to separate a single chemical compound from a mixture dissolved in a fluid. It separates substances based on differential adsorption of compounds to the adsorbent as the compounds move through the column at different rates which allow them to get separated in fractions. This technique can be used on small scale as well as large scale materials that can be used in future experiments.
Types of Column Chromatography
- Ion exchange column chromatography
- Gel column chromatography
- Partition column chromatography
- Adsorption column chromatography
Uses of Column Chromatography
- It is used to isolate active ingredients
- It is used to remove impurities
- It is used in isolation of metabolites from biological fluids
- Used in separating compound mixtures
What You Need To Know About Column Chromatography
Also Read: Difference Between Gas Solid Chromatography And Gas Liquid Chromatography
Difference Between Paper, Thin Layer And Column Chromatography In Tabular Form
| BASIS OF COMPARISON | PAPER CHROMATOGRAPHY | THIN LAYER CHROMATOGRAPHY | COLUMN CHROMATOGRAPHY |
| Description | Paper chromatography is a chromatographic technique used to separate compounds based on the liquid-liquid adsorption and solubility of the compound. | Thin Layer chromatography is a chromatographic technique based on solid-liquid adsorption of molecules. | Column chromatography uses a column packed with a matrix that is used to separate molecules mainly based on their size, affinity or its charge. |
| Stationary Phase | The stationary phase of paper chromatography is the water trapped in the cellulose filter paper. | The stationary phase of the thin layer chromatography is the glass plates coated with silica gel. | A column packed with a suitable stationary phase is used as the stationary phase in the column chromatography. |
| Movement Of Components | The compound which is more soluble in stationary phase will travel slower. | The components move according to relative affinities. | The components move according to relative affinities. |
| Affinity & Movement Of Compound | The compound which is more soluble in mobile phase travels faster. | The compound which has less affinity towards stationary phase travels faster. | The compound which has less affinity towards stationary phase travels faster. |
| Principle Of Separation | The principle of separation is partition. | The principle of separation is adsorption. | The principle of separation is adsorption. |
| Mobile Phase | Running solvent is the mobile phase of the paper chromatography. | Running solvent is the mobile phase of the thin layer chromatography. | Wash buffer is the mobile phase of the column. |
| Solvent Requirement | Requires less amount of solvent. | Requires less amount of solvent. | Column chromatography requires more amount of solvent. |
| Absorption | Paper chromatography is based on solid-liquid absorption. | Thin layer chromatography is based on solid-liquid absorption. | Column chromatography is based liquid-solid absorption. |
| Sensitivity Of Detection | Sensitivity of detection is less. The spots are easily diffused. | The sensitivity of detection of the fraction on the plate is high. The spots are less diffused. | Sensitivity of detection is less. |
| Evaluation Under UV-Light | Paper chromatography cannot be evaluated under UV. | Thin layer chromatography can be evaluated under UV light. | Column chromatography can be evaluated on Ultra-visible (UV) light if the compounds are colorless. |
| Reagents | Paper chromatography does not use strong reagents to identify different components of the mixture. Corrosive/strong reagents destroy the paper. | Thin-layer chromatography uses strong reagents to identify the different components of the mixture. This is because the plate can withstand strong solvents and color creating agents. | Column chromatography does not use strong reagents to identify different components of the mixture. |
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