Partition chromatography principle and applications

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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 procedure of high use. Biochemistry and chemistry researchers use partition chromatography to separate different biomolecules such as proteins, carbohydrates, and lipids. Partition chromatography involves in differences in the retention factor (K) as well as distribution coefficient (Kd) of the analytes. Both, stationary phase as well as the mobile phase, use liquid solvents. Partition chromatography is of two types; 1) liquid-liquid chromatography and 2) bonded-phase liquid chromatography.

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Partition Chromatography Types and their Applications

Liquid-liquid chromatography uses liquid solvents and a supporting matrix to support the stationary phase by physical means. The supporting matrix involves the use of either of the components; cellulose, starch or silica. They support water stationary phase because of their ability to bind physically up to 50 % of water (w/v) and remains free floating powder. There are some advantages of liquid-liquid chromatography. For example, silica, cellulose, and starch, these are all cheap and have a high binding capacity and wide selectivity.

However, there are some disadvantages too such as, elution of the sample may gradually remove the stationary phase thereby altering the chromatographic condition. Bonded-phases liquid partition chromatography can overcome these problems because they utilize bonded-phases. Silica is derivatized to the immobilized stationary phase by using reaction with an organochlorosilane. Thus, with elution, the stationary phase will not change.

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Normal phase liquid chromatography uses polar stationary phase and relatively non-polar mobile phase. For the stationary phase, use an alkylamine bonded to silica while for the mobile phase, use organic solvents such as hexane, heptanes, dichloromethane or ethyl acetate.You must use these solvents based on their polarity in a way that forms an eluotropic series with an increase in the polarity. Perform the elution of analytes in such an order that the least polar analyte elutes first and the least one elutes at the last.

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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 of these chromatographic procedures; normal-phase liquid chromatography and reverse-phase liquid chromatography are the types of bonded phase liquid chromatography.

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