Hydrophilic Interaction Liquid Chromatography HILIC

In those cases where the lack of analyte retention is due to one or multiple charged groups, retention is easily accomplished by ion-exchange chromatography 

Handbook of Ion Chromatography, Fourth Edition. Joachim Weiss. 

the most rational way to address the separation of very hydrophilic and uncharged compounds in HPLC is by utilizing hydrophilic interaction liquid chromatography (HILIC) [4-7], which is considered to be a variant of 

Separation Mechanism in Hydrophilic interaction Liquid Chromatography 

HI Lie materials can, in principle, he any one of the polar stationary phases, from silica to ion exchangers, making use of polar interactions between the analyte and the stationary phase with mohUe phases consisting of a mixture of water and acetonitrile or methanol or ethanol. Zwitteiionic materials (ZIC-HILIC) constitute a prominent type of HIUC columns. 

The HILIC mode starts with a high amount of organic solvent in the mohUe phase hydrophobic analytes are eluted first, while hydrophilic (polar) analytes are retained. By increasing the water content of the mobile phase, the polar analytes will be eluted. 

A HILIC gradient may start with 80% and go down to 20% acetonitrile in a 50 mM aqueous buffer. 

Typical applications for the HILIC mode are compounds with charged groups and high water solubility, giving little retention with reversed-phase columns. However, never inject large volumes of aqueous sample solutions in a HILIC column. Since water is the strongest eluent in HILIC, broad peaks are the inevitable result. 

A special issue on HILIC and mixed modes can be found in Ref [5]. 

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Direct injection of plasma or supernatant after protein precipitation on a short column with a high liquid flow rate is a common method for reducing analysis time in the pharmaceutical industry. The direct injection of a sample matrix is also known as the dilute-and-shoot (DAS) approach.62 DAS can be applied to all types of matrices and approaches and is the simplest sample preparation method with matrix dependency. Direct injection can also be approached through the extraction of eluent from PPT, SPE, and LLE onto a normal phase analytical column. The procedure is called hydrophilic interaction liquid chromatography (HILIC)70110111 and it avoids the evaporation and reconstitution steps that may cause loss of samples from heat degradation and absorption. 

Hydrophilic interaction liquid chromatography (HILIC) can handle the matrix effects nonvolatiles, which may reduce the evaporation of volatile ions during the electrospray process. Ion-exchange methods that require the use of salts could otherwise interfere with MS. 

One weakness of the dominant reverse phase separations mechanism has been the poor retention of highly polar analytes, and hydrophilic interaction liquid chromatography (HILIC) has emerged as an alternative. In HILIC, a polar stationary phase such as silica gel is used to retain highly polar analytes. Mobile phases components similar to those described above for reverse phase separations are used, but the proportions of aqueous vs. organic are changed. Analytes are retained under conditions of relatively low water content, and eluted using increased water content. 

Antidepressant separation was usually performed by reversed-phase chromatography with typical C8 or C18 alkyl chain columns, although phenyl [30, 59] or cyano [48,64, 84] stationary phases were also employed. As an exception, hydrophilic interaction liquid chromatography (HILIC), a variation of normal phase chromatography, was employed in two analytical methods for duloxetine [38] and... 

Nygren, Y., Hemstrdm, P., Astot, C., Naredi, P., Bjorn, E. Hydrophilic interaction liquid chromatography (HILIC) coupled to inductively coupled plasma mass spectrometry (ICPMS) utilizing a mobile phase with a low-volatile organic modifier for the determination of cisplatin, and its monohydrolyzed metabolite. J. Anal. At. Spectrom. 23, 948-954 (2008)... 

Application of LC-MS to the analysis of aminoglycosides has also been problematic, due to the low retention of the parent compounds on the reversed phase columns commonly used. To fry to resolve this problem, ionpairing reagents, such as fluorinated acids, have been used. The introduction of hydrophilic interaction liquid chromatography (HILIC) has provided an alternative means for the retention and separation of aminoglycosides prior to mass specfromefric determination (see Chapter... 

Wang, P.G. He, W. Hydrophilic Interaction Liquid Chromatography (HILIC) and Advanced Applications. Taylor Francis Boca Raton, FL, 2011. 

Karson, G. (2011) Development and application of methods for separation of carbohydrates by hydrophilic interaction liquid chromatography, in Hydrophilic Interaction Liquid Chromatography (HILIC) and Advanced Applications (eds P.G. Wang and W. He), CRC Press, Boca Raton, p. 491. 

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