Question

Describes the chromatography of hydrophobic interactions.

Answer 1

Chromatography ;

it is a technique for the separation of a mixture of any organic compound ( Mobile phase )by passing it in solution ( Stationary phase) or suspension through a medium ( media ) in which the components move at different rates according to the various characters do fond in the separating compound. Chromatography is a method used by scientists for separating organic and inorganic compounds so that they can be analyzed and studied. .Chromatography is used in many different ways like chromatography to find out what is in a solid or a liquid. It is also used to determine what unknown substances are by different criteria of calculating.

Chromatography of Hydrophobic Interaction

simply Hydrophobic Interaction Chromatography is a separation technique that on the basis of the hydrophobicity of the mixture compounds to separate proteins from one another. In this type of chromatography, hydrophobic groups such as phenyl, octyl, or butyl, are attached to the stationary column which can further interact with the protein bypassing the mixture by the column and proteins bind. as hydrophobic amino acid side chains on the surfaces of the proteins, it interacts with the hydrophobic group in the column.

by the mechanism, the stationary phase is designed to form hydrophobic interactions with other molecules. there are three major theories that may help explain the mechanism behind HIC – (1) the salting-out theory, (2) the thermodynamic theory, and (3) the surface tension or Van der Waals forces theory. by the theories, we can find out that These interactions are too weak in water and that's, why to increase interaction the addition of salts to the buffer, is needed. The following is a list of salts that increase hydrophobic interactions in the order of their ability to enhance interactions:

1. Na2SO4
2. K2SO4
3. (NH4)2SO4
4. NaCl
5. NH4Cl
6. NaBr
7. NaSCN

the actual mechanism of the hydrophobic interaction chromatography is the hydrophobicity property of molecules which can do the crucial role of separation of molecules The salt in the buffer reduces the solvation of sample solutes thus as solvation decreases, hydrophobic regions that become exposed are adsorbed by the medium. The more hydrophobic the molecule, the less salt needed to promote binding. To elute the proteins, the salt concentration is gradually decreased, eluting bound species in order of increasing hydrophobicity. Additionally, elution can also be achieved through the use of mild organic modifiers or detergent.

fig under high salt conditions there is increase in the hydrophobic interactions due to this regions of hydrophobic get aggregated.

in a nutshell, Hydrophobic interaction chromatography separates out protein molecules using the properties of hydrophobicity. In this method, proteins containing both hydrophilic and hydrophobic regions are applied to an HIC column under high salt buffer conditions.