SHARC Separation Technology

Significance of the Topic

Liquid chromatography techniques have traditionally leveraged hydrophobic, ionic, dipolar or size-exclusion interactions to resolve complex mixtures. The SHARC™ (Specific Hydrogen-bond Adsorption Resolution Chromatography) stationary phase introduces a fundamentally new mode of separation, exploiting hydrogen-bonding interactions in nonaqueous eluent systems. By targeting analytes’ ability to donate or accept hydrogen bonds, this approach provides unique selectivity for polar and isomeric compounds that are challenging for conventional reversed-phase or normal-phase methods.

Objectives and Overview

This application note from SIELC Technologies presents the development, operational principles, and performance evaluation of SHARC-1 HPLC columns. Key goals include:

• Define the hydrogen-bonding retention mechanism.
• Detail optimized mobile phase systems using acetonitrile (weak solvent) and methanol (strong solvent).
• Illustrate separation of pharmaceuticals, nucleosides, organic acids, and surfactants.

Methodology

SHARC-1 columns are packed with a stationary phase bearing polarized hydrogen atoms capable of forming specific H-bond interactions with analyte functional groups. Retention is tuned by varying the ratio of acetonitrile to methanol:

• Acetonitrile (MeCN) serves as a weak eluent with minimal competition for H-bond sites.
• Methanol (MeOH) acts as a strong eluent, displacing analytes from the surface.
• Gradient or isocratic methods in MeCN/MeOH mixtures (e.g., 95/5 to 50/50) with formic acid and ammonium formate modifiers achieve optimal resolution.

Instrumentation Used

• SHARC-1 HPLC columns (3.2 × 100 mm, 3 µm; 4.6 × 150 mm, 5 µm).
• Standard HPLC or UPLC systems capable of 150–200 bar.
• UV detection at 270 nm.
• Pump and autosampler compatible with nonaqueous MeCN/MeOH mixtures.

Main Results and Discussion

The hydrogen-bonding mode effectively separated compounds according to their polar surface area (PSA) and the accessibility of hydrogen-bonding sites. Representative findings include:

• Pharmaceutical bases (pyrilamine, trimipramine, pindolol): elution order correlates with PSA values and number of accessible H-bond sites.
• Xanthines and nucleobases: isomeric and methyl-substituted derivatives resolved by differential retention based on substitution patterns.
• Aminopyridine isomers and phthalic acid isomers: differences in intramolecular H-bonding and site accessibility drive selectivity despite identical PSA.
• Surfactant oligomers (Triton X-100): retention increases with the number of ether oxygens available for hydrogen bonding.

Benefits and Practical Applications

SHARC chromatography offers multiple advantages for analytical and preparative workflows:

• Speed: Lower viscosity of MeCN/MeOH mixtures enables higher flow rates and UPLC-like performance, achieving 2–6× faster analysis.
• Solubility: Broad solvent compatibility dissolves polar and nonpolar analytes without aqueous restrictions.
• Preparative capability: Volatile eluents facilitate efficient evaporation and MS-directed fraction collection.
• High selectivity: Precise discrimination of isomers, homologs and closely related analogs based on H-bonding interactions.

Future Trends and Applications

Emerging opportunities for hydrogen-bonding chromatography include:

• Integration with high-resolution mass spectrometry for metabolomics and proteomics.
• Design of tailored stationary phases for specific analyte classes (e.g., carbohydrate isomers).
• Automation of method development using PSA and computational modeling of H-bond interactions.
• Expansion into industrial quality control, environmental analysis and biopharmaceutical characterization.

Conclusion

SHARC™ columns represent a novel nonaqueous hydrogen-bonding separation mode, enabling high-efficiency, high-selectivity analysis of polar and isomeric compounds. By manipulating MeCN/MeOH ratios and leveraging PSA concepts, analysts can achieve rapid, reproducible separations that complement existing chromatographic techniques.

References

• Grushka E., Grinberg N., Eds., Advances in Chromatography, Volume 54, CRC Press, Boca Raton, 2018, Chapter 3.