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Separation principle and characteristics of HILIC column

HILIC column is a hydrophilic interaction column. Its retention mode is similar to that of normal phase, and it breaks through the solvent limitation of normal phase mode.
Separation principle: the reverse phase mobile phase system is used, and the peaks are in the normal phase sequence. Through the hydrophilic interaction, the strong polar compounds which are difficult to be separated due to the weak retention capacity can be effectively separated from the reversed-phase chromatographic column. It is a kind of separation mechanism, and there are many kinds of bonding phases with this separation mechanism. In HILIC, the stationary phase is polar, and the water phase in the mobile phase is a strong eluent, which is completely contrary to the traditional reversed phase chromatography.
The characteristics of HILIC column are as follows
At the same time, it has weak electrostatic interaction with anions and cations, which has different selectivity to the target;
It provides multiple interactions of hydrogen bonds, molecules and electrostatic attraction;
It can provide suitable retention for the target with strong polarity;
Using water-soluble organic phase as mobile phase can significantly improve the solubility of samples in the mobile phase;
Separation of polar and hydrophilic small molecular targets and basic drug molecules.
The mechanism of HILIC column was as follows
It can be considered that reversed phase liquid chromatography (HPLC) can be operated by using polar stationary phase. The mobile phase used is a high proportion of organic phase containing a small amount of water / buffer salt or other polar solvents. The water in the mobile phase forms a rich water layer, which is adsorbed on the surface of the stationary phase, and the polar substances first enter the rich water layer to be retained. Polar compounds are retained through a complex mixing mode, including hydrophilic distribution between rich water layer and mobile phase, hydrogen bonding between polar functional groups and stationary phases, and electrostatic interaction of ionic functional groups. Of course, hydrophilic distribution is the main retention mode. From the point of view of retention mechanism, HILIC is quite different from RP column.
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