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Chromatography is one of the best techniques for separating substances found in blood or cells or components of a single substance. For the first time by chromatography it was shown that bacteria have more than 1000 different number of proteins and most of these proteins were isolated by chromatography. Once a single protein has been isolated using chromatography, we can hydrolyze it and then use chromatography to separate the amino acids from each other.
Many types of chromatography share the same basic mechanism. This mechanism is the simultaneous exposure of two different substances, such as a solid adsorbent, to two unmixed solvents. Substances adhering to the surface (molecules, molten solid or liquid) are called adsorbents. Each molecule of each solvent in the mixture is propagated backwards and forwards between the two substances, the relative affinity of the soluble molecule to these two solvents, on average, determines how long each will remain in each. For example, if the substance A, which is highly soluble in water and is slightly dissolved in phenol, is rinsed with water and phenol in a jar (not mixed), each molecule of A will share its time between these two solutions, so that it remains in water for a longer time.

Paper Chromatography

Paper chromatography is the simplest type of chromatography. Multiple drops of unknown molecules are dropped into the bottom edge of a filter paper moistened with water. The bottom edge of the paper is then immersed in a water-free solution such as phenol. As the solvent emerges upward with capillary action, the solvents in the mixture with more affinity than the water to the solvent run upwards along with the solvent in the filter paper. On the other hand, solutes with greater affinity for moisture in the filter paper cannot go away; but instead they are transferred very rapidly from the flowing solvent and connected to the fixed water layer on the filter paper. Compared to water, materials with moderate affinity for the solvent can proceed to moderate distances. After a period of time, the various solvents in the original mixture are located in different places along the filter paper. Often, the second solvent is used for further separations and is achieved by moving the solvents in different directions. When this is the case, the technique is called biaxial chromatography.

electrophoresis

Another popular way of separating substances is bir electrophoresis Mad. After an chromatographic separation along an axis, an electric field is installed along the other axis; molecules with different ionic strength and polarity move from one end to another at different speeds.


Column Chromatography

Column chromatography is like simple paper chromatography, using only a glass column filled with some hydrofoil adsorbent material such as starch and silica instead of paper. The mixture to be tested in the water-free solvent is dropped from the top of the column and filtered downwards (or pulled down by a pump). That is, when substances with high affinity for the solvent are flowing directly, those with affinity for particles will remain behind.
When the solvent is spilled from the column, each one is removed at different times from the bottom of the column and collected in separate containers for further analysis.

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