Excellent separations with ZIC^®-HILIC HPLC columns

SeQuant^® HILIC HPLC columns have documented excellent selectivity for straightforward separation of polar hydrophilic compounds in HPLC and LC-MS methods. These columns carry densely bonded, truly zwitterionic functional groups with a charge balance of 1:1. Separation is achieved by hydrophilic partitioning combined with weak ionic interactions for maximum selectivity, high loadability and easy optimization of methods.

The comprehensive product range of SeQuant^® ZIC^®-HILIC columns covers dimensions from capillary through analytical to semi-preparative. Its favorable retention and robust performance have made SeQuant^® ZIC^®-HILIC one of the most popular HILIC columns all over the world.

“In particular, the zwitterionic phase ZIC-HILIC showed better selectivity for the test samples than the other phases.”
Kawachi et al., J. Chromatogr. A 1218 (2011) 5903-5919

This citation is taken from an article published by distinguished Japanese scientists. They performed comprehensive screening to compare the performance of 15 different HILIC columns. Their conclusion was that the selectivity of SeQuant^® ZIC^®-HILIC was higher and more versatile than that of all other stationary phases.

Alternative choices for separation of polar hydrophilic compounds include SeQuant^®  ZIC^®-cHILIC with complementary selectivity and SeQuant^® ZIC^®-pHILIC for higher pH stability. Find out more on the corresponding webpages giving an overview of SeQuant^® HILIC columns.

Read on: What is HILIC? – Product Range – Applications – Technical Support

What is HILIC?

With HILIC you combine the benefits of aqueous mobile phases like in reversed-phase HPLC with separation based upon normal phase principles where more polar compounds respond with higher retention resulting in more efficient separation. More details are available on our webpages giving an overview of SeQuant^® HILIC columns.

Visit the HILIC Technology Tutorial or request your free copy of ‘A Practical Guide to HILIC’ to learn more about the options offered by the HILIC technique and its advantages in separating polar hydrophilic substances.

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Product Range

SeQuant® Column	Charge arrangement and functional group	Features	Base particle	Particle size	Pore size	Column types
SeQuant® ZIC®-HILIC	Sulfobetaine	Well-known for high-performance selectivity and robustness	silica	3.5 µm, 5 µm	100 Å, 200 Å	analytical, capillary, semi-prep, guards

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Applications

The use of SeQuant^® ZIC^®-HILIC has been recommended by US FDA (United States Food and Drug Administration) in its methods for the analysis of melamine and related potential contaminations.

The ZIC^®-HILIC column has been used in numerous methods for a variety of compounds such as acids and bases, anions and cations, carbohydrates, metabolites, metal complexes, amino acids, peptides, protein digests and many more.

Separation of cyanuric acid melamine food contaminants
Column	ZIC®-HILIC 150 x 2.1 mm, 5 μm, 200 Å (1.50454.0001)
Mobile phase	A:	95% acetonitrile  in 0.1% aqueous formic acid
	B:	50% acetonitrile  in 20 mM aqueous ammonium              formate
Gradient	Time 0 4.2 8.0 8.5 9.0 11.0 11.2 13.0 14.0	A 100 100 65 65 25 25 100 100 100	B 0 0 35 35 75 75 0 0 0	Flow rate [mL/min] 0.4 0.4 0.4 0.4 0.4 0.4 0.6 0.6 0.4
Detection	MS/MS, -ESI and +ESI
Temperature	30°C
Inj. volume	5 μL
Sample	Standard in mobile phase (7 ng/mL) equivalent to 1 μg/g
Courtesy of	David N. Heller, FDA Center for Veterinary Medicine, Laurel, MD 20708 USA				Reference	David N. Heller and Cristina B. Nochetto, Rapid Commun. Mass Spectrom., 22 (2008) 3624-3632. DOI: 10.1002/rcm.3779

Separation of amino acids
Column	ZIC®-HILIC 150 x 4.6 mm, 3.5 μm, 100 Å (1.50444.0001)
Mobile phase	80%	acetonitrile
	20%	ammonium acetate in water, 50 mM pH adjusted to 4.5 with formic acid
Flow rate	0.75 mL/min
Detection	Refractive index, cell 9 μL, 40°C
Temperature	40°C
Inj. volume	50 μL
Gradient	1. 2-Amino-5-Hydroxy benzoic acid 2. Tryptophan 3. Isoleucine 4. Methionine 5. Lycine 6. Threonine			100 ppm 100 ppm 100 ppm 100 ppm 100 ppm 100 ppm
Courtesy of	Gora Sharangi, Merck India Application Lab

Bioanalysis of methyl malonic acid
Column	ZIC®-HILIC 100 x 2.1 mm, 3.5 μm,100 Å (1.50444.0001)
Mobile phase	A:	80% (v/v) acetonitrile
	B:	20% (v/v) 100 mM NH4Ac buffer pH 4.5
Flow rate	MS-ESI- with SIM at 117.2 m/z            Drying-gas flow rate Drying-gas temperature Capillary voltage			10 L/min 300°C 3.0 kV
Detection	Refractive index, cell 9 μL, 40°C
Inj. volume	4 μL
Sample	Precipitation-treated plasma sample containing 0.137 mol/L MMA
Reference	H-A Lakso, P. Appelblad, J. Schneede Clin. Chem., 54 (2008) 2028 DOI: 10.1373/clinchem.2007.101253

 Some exemplary application notes using ZIC^®-HILIC:

• ZIC^®- HILIC Separation of Sucrose with RI detection
• ZIC^®- HILIC Separation of Fructose and Sucrose
• ZIC^®- HILIC Separation of tryptic peptides from Ovalbumin

More examples of how you can solve analysis problems with ZIC^®-HILIC HPLC columns can be found in the ‘Chromatography Application World’ or in the comprehensive collection of SeQuant^® application notes.

Glycomics	Peptide mapping

Technical Support

Use the free online tool to predict retention on SeQuant^® ZIC^®-HILIC columns.

For technical support on ZIC^®-HILIC products or questions regarding application development, please consult the SeQuant^® FAQ list or contact the SeQuant^® Support Team.

Detailed instructions for using ZIC^®-HILIC columns can be found in the documents listed below. More supportive documents can be found in the SeQuant^® download area.

• ZIC^®- HILIC Analytical HPLC Columns
• ZIC^®- HILIC Nano, Capillary and Microbore HPLC Columns
• ZIC^®- HILIC Semi-Preparative HPLC Columns

Every HPLC separation column by EMD Millipore is individually tested and delivered with a certificate documenting its performance.

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