Chromatography is an analytical technique used in labs to separate different components individually from a sample mixture. The separation of the components involves distributing them into two phases:
The factors that influence the separation of the components include different molecular characteristics such as partition (liquid-solid), adsorption (liquid-solid), and affinity or molecular weight differences.
They determine whether molecules will stay longer in the stationary phase, move slower, or move faster in the systems. The time each component takes in its elution from the stationary phase is known as the retention time.
There are different types of chromatographic separation techniques available, including column chromatography, high-performance liquid chromatography (HPLC), gas-liquid chromatography, and thin-layer chromatography (TLC).
They all serve different purposes, as gas-liquid chromatography is used to separate esther, alcohol, lipid, and amino groups from a complex mixture, while the molecular-sieve chromatographic method has applications in determining the molecular weights of proteins.
The applications of chromatography include drug development, detecting trace amounts of chemicals or components of a mixture, protein purification, leaf pigment separation, and testing the quality of water and air.
In this article, we will review more about the different types of chromatographic separation methods.
Different chromatographic techniques involve different instrumentation based on their working mechanisms. However, two common systems in each system include:
Various types of chromatographic techniques have been developed to serve a range of applications.
It is also known as gas-liquid chromatography. It involves equipment like packed or capillary columns, a chromatogram, an autosampler, and a detector. It has a liquid stationary phase and a mobile phase composed of inert gas, which are passed through the chosen column at high pressure for effective separation of components.
The technique is used in the food industry for the quantification of contaminants and compounds, such as carbohydrates and vitamins, in food samples. Further, it’s also used in air quality testing, drug testing, pesticide detection, and product safety testing.
The technique is also known as liquid-liquid chromatography. In this method, a cellulose-made thick filter paper acts as a solid support, water drops settled in its pores act as a stationary phase, and a fluid placed in the developing tank acts as a mobile phase.
The technique is extensively utilized in the separation of plant pigments (such as photosynthetic pigments) and proteins from a complex mixture.
It’s also known as solid-liquid chromatography. Here, the stationary phase is a glass plate coated with a solid adsorbent substance, such as silica gel, alumina, or cellulose. It’s a versatile technique used in labs to simultaneously separate multiple samples.
The sample mixture placed in the stationary phase gradually starts moving upward through capillary action driven by the organic solvent used in the process. The movement rate of components depends on the solid phase, the polarity of the material, and the solvent
The TLC technique has uses in identifying natural products, such as fixed oil and waxes, and the separation of metabolites from a range of samples, including blood plasma and serum.
The chromatographic technique works on the principle of separation of molecules based on the difference in their partition coefficient between two liquid phases. The other chromatographic technique that works on similar principle includes Reverse phase partition, Liquid-liquid partition, and Thin-layer chromatography.
These methods are crucial for separating and identifying various biomolecules, such as fatty acids, amino acids, and carbohydrates.
The separation of biomolecules from a sample mixture based on their macromolecular binding interaction with the substrate is known as affinity chromatography. The interactions widely exploited in labs for research workflows include interaction between:
The technique is used to purify many biomolecules, including hormones, nucleic acids, antibodies, and specific proteins.
The technique is also known as gel filtration. During the process, the molecules are separated based on their size by filtration through the gel. The small molecules move into pores, while large molecules remain outside and get washed out in the column's empty space. Therefore, they are separated and eluted based on their decreasing molecular weight.
It’s an effective technique for analyzing different proteins, studying the properties of biomolecules, and checking the quality of samples.
The basic principle of ion exchange chromatography is the exploitation of the interaction between solid support matrix and charged protein groups. The proteins are separated either based on the ionic strength or pH change of a buffer solution. The two types of ion-exchange chromatography include Cation exchange chromatography and anion-exchange chromatography.
The technique is widely used in labs to separate biomolecules such as nucleotides, amino acids, and peptides from a sample mixture.
Chromatography is an analytical technique used in labs to separate and study individual components from a mixture of the sample. The technique involves solid support and a stationary and mobile phase for the separation of biomolecules based on their different properties.
Different types of chromatography include affinity, column exchange, and gas chromatography techniques. They all have different work mechanisms and serve different purposes. If you’re looking for an affordable option to enhance chromatography precision, consider leasing chromatography equipment through Excedr.
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