Partition Chromatography Principle and Applications

Partition chromatography

Partition chromatography. Image: Durfo via Common Wikimedia

Partition chromatography is a type of chromatography. It is one of the most useful biochemical lab manual used to separate different biomolecules like proteins, carbohydrates, lipids etc. It is based on differences in retention factor K as well as distribution coefficient Kd of the analytes using liquid for both stationary as well as the mobile phase. Partition chromatography can be divided into 1) liquid-liquid chromatography and 2) bonded-phase liquid chromatography.

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Partition Chromatography Types and Applications

Liquid-liquid chromatography uses liquid and the stationary phase is attached to a supporting matrix by physical means (i.e. no chemical bond). As for example cellulose, starch or silica matrix is used to support water stationary phase and all these have the ability to bind physically as much as 50% water (w/v) and remain free floating powder. There are some advantages of liquid-liquid chromatography as for example silica, cellulose starch used are cheap, have a high capacity with broad selectivity.

However, there are some disadvantages too like elution of the sample may gradually remove the stationary phase thereby altering the chromatographic condition. To overcome these problems, bonded phases are used like in bonded phase liquid chromatography. Silica is derivatized to the immobilized stationary phase by using reaction with an organochlorosilane. Thus, with elution stationary phase will not change.

Normal phase liquid chromatography uses polar stationary phase and relatively non-polar mobile phase. Alkylamine bonded to silica is most commonly used as a stationary phase while organic solvents like hexane, heptanes, dichloromethane or ethyl acetate are used as mobile phase.

These solvents are used in such a way to form an eluotropic series based on their polarity with increasing polarity. The order of elution of analytes is performed in such a way that the least polar analyst is eluted first and the most polar analyst eluted at last.

Reverse phase liquid chromatography has many similarities with hydrophobic interaction chromatography. In this chromatography, the stationary phase is nonpolar and the mobile phase is relatively polar than the stationary phase. Both these, normal-phase liquid chromatography and reverse-phase liquid chromatography are the type of bonded phase liquid chromatography.

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