Chromatography is a common analysis technique based on the principle where molecules in mixture applied to the surface or into the stationary phase is separating from each other while moving with the aid of a mobile phase. The primary purpose of chromatography is to purify and extract one or more components of a sample, and its analytical goal is to determine the qualitative and quantitative chemical composition of the sample. Any chromatography technique contains three main parts: a sample, a mobile phase, and a stationary phase. The components in the mixture sample can be separated and purified depending on the different distribution rates of different components, which depends on the solubility, affinity, and interaction with the stationary phase.
The mobile phase flowing through the stationary phase can be a gas or a liquid substance. If the mobile phase is liquid, it is called liquid chromatography (LC), and if the mobile phase is a gas, it is called gas chromatography (GC). GC is applied for mixtures of gases, and volatile liquids and solid substances. LC is especially used for thermally labile and nonvolatile samples. LC can further be divided into ion exchange, separations based on size, and gel based electrophoretic techniques. A particularly common detector for GC and LC is mass spectrometry, which converts each analyte from a chemically neutral species into a positive cation, often breaking various bonds in the process. Detection of the mass of individual fragments allows determination of the chemical structure of the analyte.
In addition to LC and GC, other chromatography methods include planar and column chromatography, which depend on the shape of the stationary phase, and affinity, ion exchange, partition, adsorption, and size exclusion chromatography, which depend on the interaction between the stationary and mobile phases. Chromatography methods based on partition are very effective for the isolation and identification of small molecules such as amino acids, carbohydrates, and fatty acids. Affinity chromatography can be used to separate large molecules such as nucleic acids and proteins. Paper chromatography is used for protein separation and protein synthesis related research, while molecular-sieve chromatography is utilized to determine the molecular weight of proteins. Agarose-gel chromatography can be employed for the purification of RNA, DNA particles, and viruses. Thin layer chromatography can be used for the purification of macromolecules, the separation of complex drug ingredients, as well as the determination of pesticides.
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