Overview
Many polar compounds exhibit little or no retention in conventional reversed phase HPLC methods. HILIC (Hydrophilic Interaction Liquid Chromatography) has become a widely accepted approach for analyzing these compounds using organic-rich mobile phases and stationary phases that provide enhanced retention of polar analytes.
Cogent™ TYPE-C™ silica hydride columns offer a unique approach to HILIC separations and are particularly effective for compounds that are often difficult to retain using traditional reversed phase methods.
Typical applications include:
- Polar pharmaceuticals
- Metabolites
- Amines
- Organic acids
- Zwitterions
- Biomolecules
- Environmental contaminants
- Food and beverage analytes
What Is HILIC Chromatography?
HILIC is an industry-recognized chromatographic mode used for the separation of polar compounds.
Typical HILIC methods utilize:
- High organic mobile phases
- Acetonitrile-rich solvent systems
- Low percentages of water
- MS-compatible additives
These conditions often provide improved retention for analytes that elute near the void volume in reversed phase chromatography.
Comparing HPLC Separation Modes
Normal Phase Chromatography
Typical characteristics:
- Polar stationary phase
- Non-polar mobile phase
- Useful for highly polar compounds in organic solvent systems
Reversed Phase Chromatography
Typical characteristics:
- Hydrophobic stationary phase
- Aqueous-organic mobile phase
- Ideal for non-polar to moderately polar analytes
HILIC Chromatography
Typical characteristics:
- Organic-rich mobile phases
- Enhanced retention of polar compounds
- Excellent compatibility with LC-MS
- Suitable for highly polar and water-soluble analytes
HILIC Performance with TYPE-C™ Columns
TYPE-C™ silica hydride columns provide selectivity for polar compounds while operating under mobile phase conditions commonly associated with HILIC methods.
Advantages include:
- Strong retention of polar analytes
- Excellent LC-MS compatibility
- Reproducible retention behavior
- High organic mobile phase compatibility
- Effective separations of acids, bases, and zwitterions
These characteristics make TYPE-C™ columns useful alternatives for many polar compound analyses.
Benefits for LC-MS Applications
HILIC methods are frequently used with LC-MS because high organic mobile phases often enhance ionization efficiency.
Benefits include:
- Increased LC-MS sensitivity
- Improved desolvation
- Lower solvent load entering the mass spectrometer
- Enhanced detection of polar compounds
Common additives include:
- Formic acid
- Acetic acid
- Ammonium formate
- Ammonium acetate
These additives are widely used in mass spectrometry workflows.
Typical Mobile Phase Conditions
HILIC methods on TYPE-C™ columns often use:
- Acetonitrile-rich mobile phases
- Low water content
- Volatile buffer additives
- LC-MS compatible solvent systems
Gradient or isocratic conditions may be used depending on the analytes and method objectives.
Why Use TYPE-C™ Columns for Polar Compounds?
TYPE-C™ columns offer several advantages when developing methods for polar analytes.
Potential benefits include:
- Retention of compounds that exhibit little retention in reversed phase methods
- Excellent compatibility with LC-MS detection
- Effective separations using organic-rich mobile phases
- Broad applicability across many compound classes
- Reproducible chromatographic performance
These advantages have made TYPE-C™ columns useful tools for analytical laboratories working with challenging polar analytes.
Supporting Research
The use of silica hydride stationary phases for HILIC applications has been described extensively in the scientific literature.
One representative publication is:
Pesek, J.J. et al.
Hydride-based stationary phases: A rapidly evolving technology for the development of new bio-analytical methods.
Analytical Methods, 2014, Volume 6, Pages 4496-4503.
Key Takeaways
- HILIC is widely used for the analysis of polar compounds.
- TYPE-C™ columns provide unique selectivity for HILIC applications.
- Organic-rich mobile phases are commonly used.
- Excellent compatibility with LC-MS detection.
- Useful for acids, amines, zwitterions, metabolites, and other polar analytes.
- Often provides retention where reversed phase methods may not.
