Separation of Maltooligosaccharides by HILIC

Significance of the Topic

Hydrophilic interaction chromatography (HILIC) has become a valuable approach for resolving highly polar carbohydrates that are challenging to separate by traditional reversed-phase methods. The ability to distinguish between closely related maltooligosaccharides is critical for quality control in food, biofuel, and pharmaceutical industries.

Objectives and Study Overview

This application note aims to demonstrate a simple, isocratic HILIC method for baseline separation of glucose (G1) through maltopentose (G5) using an amino-phase column and refractive index detection.

Methodology and Instrumentation

An HPLC system equipped with a refractive index detector was employed. Separation utilized a Eurospher II 100-3 NH2 column (100 × 3 mm ID) maintained at 30 °C. The mobile phase comprised 30 % water and 70 % acetonitrile, delivered isocratically at 1.5 mL/min. Sample injections (5 µL) contained 5 mg/mL of each maltooligosaccharide in 50 % acetonitrile.

Main Results and Discussion

Five discrete peaks were obtained in the order of increasing degree of polymerization: glucose, maltose, maltotriose, maltotetraose, and maltopentose. Each analyte exhibited sharp, symmetric peaks with no overlap, confirming the amino-phase HILIC column’s effectiveness for these compounds.

Benefits and Practical Applications

• Isocratic operation simplifies method development and routine use
• High resolution of non-UV-active saccharides with RI detection
• Reproducible retention times for consistent quantitation
• Applicable to carbohydrate profiling in food, beverage, and bioprocess monitoring

Future Trends and Applications

Coupling HILIC with mass spectrometry for enhanced detection sensitivity, adopting UHPLC formats for faster analysis, exploring novel polar stationary phases, and integrating multidimensional chromatography strategies are promising directions to further advance carbohydrate separation and characterization.

Conclusion

The described HILIC method offers a straightforward, reproducible solution for separating maltooligosaccharides, addressing common analytical challenges and supporting diverse applications in research and quality control settings.

Used Instrumentation

• High-performance liquid chromatography (HPLC) system with refractive index detector
• Eurospher II 100-3 NH2 column (100 × 3 mm ID)
• Binary pump and autosampler for isocratic mobile phase delivery