It is a technique that separates and purifies components of mixtures of chemicals. Learn definition, types of chromatography and applications.
Introduction
Chromatography is a powerful analytical technique that separates and purifies complex mixtures of chemicals. Researchers in various fields, including pharmaceuticals, biotechnology, food science, and environmental analysis using this technique to obtain accurate and reliable results. In this guide, we will explore the basic principles, the different types and their applications.
What is Chromatography?
It is an analytic technique which separates components of a mixture based on their differential interaction with a stationary phase and a mobile phase. The stationary phase can be a solid, liquid, or gas, and the mobile phase can be a liquid or gas. As the mixture passes through the stationary phase, the components interact differently, causing them to separate and elute at different times.
Principle
The principle behind chromatography is that the components of a mixture will interact differently with the stationary phase and the mobile phase, causing them to separate and elute at different times. The separation occurs because the different components have different affinities for the stationary phase. And the mobile phase which carries the mixture through the stationary phase. The components that have a higher affinity for the stationary phase will spend more time in the stationary phase and elute later, while the components that have a lower affinity for the stationary phase will elute earlier. This differential interaction between the components of the mixture and the stationary phase results in the separation and purification of the mixture’s components.
Types of Chromatography
Liquid Chromatography (LC)
LC uses a liquid as the mobile phase and a solid or a liquid-coated stationary phase. In this technique, the mixture is dissolved in a liquid and passed through a column containing a stationary phase. The components of the mixture interact differently with the stationary phase, causing them to separate based on their chemical and physical properties. Researchers often use LC to analyze non-volatile and polar compounds in liquid samples, such as in the analysis of drugs, metabolites, and biomolecules.
Gas Chromatography (GC)
GC uses a gas as the mobile phase and a solid or a liquid as stationary phase. GC is similar to LC, but the mixture is vaporized and passed through a column containing a stationary phase. The components of the mixture interact differently with the stationary phase to separate based on their volatility and other properties. Researchers often use GC to analyze volatile and semi-volatile compounds in gas or liquid samples, such as in the analysis of environmental pollutants or the quality control of petrochemical products.
Ion-Exchange Chromatography (IEC)
In this technique, the stationary phase contains charged particles that attract or repel other charged particles in the mixture. Researchers often use IEC to purify and separate proteins, nucleic acids, and other biomolecules.
Size-Exclusion Chromatography (SEC)
Size-exclusion chromatography uses a porous stationary phase to separate molecules based on their size and shape. Larger molecules will exclude from the stationary phase and pass through the column faster than smaller molecules. Researchers often use SEC to analyze and purify macromolecules such as proteins, polysaccharides, and nucleic acids.
Affinity Chromatography (AC)
Affinity chromatography uses a specific ligand on the stationary phase to bind and separate a target molecule from a mixture. A ligand will attach to the stationary phase that specifically binds to the target molecule. It will retain target molecule while other molecules pass through. Researchers often use AC to purify and isolate proteins, antibodies, and other biomolecules.
Thin-Layer Chromatography (TLC)
The stationary phase is a thin layer of material (usually a solid) coated onto a flat surface, such as a glass plate. The mixture is spotted onto the plate and allowed to interact with the stationary phase as a solvent moves through the plate. Components of the mixture separate based on their interaction with the stationary phase and can be visualized by a variety of methods.
High-Performance Liquid Chromatography (HPLC)
HPLC is a more advance form of LC. It utilizes high-pressure pumps to increase the flow rate of the mobile phase and improve separation efficiency. It is commonly using for separation and analysis of complex mixtures in various industries, such as pharmaceuticals and food.
Applications
- Pharmaceutical Industry: Researchers use chromatography to analyze the purity and potency of drugs and to purify active ingredients.
- Biotechnology: To purify proteins, nucleic acids, and other biomolecules for research and clinical applications.
- Food Science: To analyze the quality and safety of food products, such as detecting food additives, contaminants, and allergens.
- Environmental Analysis: To analyze and monitor environmental pollutants, such as pesticides, heavy metals, and volatile organic compounds.
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