Evaluation of XBridge™ BEH Amide Columns for the Analysis of Five Sugars in Food and Beverages Using an Arc HPLC, Coupled to a 2414 RI Detector

Significance of the Topic

Accurate quantification of common sugars in food and beverages is essential for product development, regulatory compliance, quality control, and authentication. Monosaccharides and disaccharides not only influence sweetness and texture but also play roles in preservation and fermentation. Analytical methods that provide reliable, high-throughput sugar profiling support consistency and safety across the food industry.

Objectives and Study Overview

This study compares two XBridge BEH Amide columns differing in particle size and length for the isocratic separation of five sugars (fructose, glucose, sucrose, maltose, lactose). Using Waters Column Calculator, conditions from a 4.6×250 mm, 3.5 µm column were scaled to a 4.6×150 mm, 2.5 µm column to reduce analysis time and solvent usage while maintaining performance.

Methodology and Instrumentation

Standards of each sugar were prepared in water and diluted in acetonitrile:water (50:50). Food and beverage samples (juices, sports drinks, milk, bread extract) were diluted, vortexed, centrifuged, and filtered before injection. Two columns were evaluated on an Arc HPLC system with a 2414 refractive index detector under isocratic conditions (75:25 acetonitrile:water, 0.2% triethylamine) at 35 °C. Flow rates were adjusted to 1.0 mL/min for the longer column and 1.4 mL/min for the shorter column.

Main Results and Discussion

Scaling to the shorter column reduced run time by 57%, with lactose eluting at 6.0 min versus 6.9 min on the longer column. Estimated solvent consumption per injection decreased by approximately 6 mL. Both columns achieved excellent linearity (R²>0.999) over 0.08–5 mg/mL. Precision for retention time and peak area showed %RSDs below 0.5%. The shorter column exhibited slightly lower resolution (e.g., Rs ≈1.3 for maltose/lactose), but remained acceptable for samples lacking coeluting sugars. Analysis of commercial products yielded sugar contents consistent with label claims.

Benefits and Practical Applications

• Rapid sugar profiling with reduced analysis time and solvent use.
• Simple isocratic method minimizes method complexity and maintenance.
• Sufficient resolution for routine quality control of foods and beverages.
• Flexibility to adapt column selection based on throughput and resolution requirements.

Future Trends and Potential Applications

Advances in particle technology and column hardware will continue to drive faster separations with improved efficiency. Coupling HILIC to alternative detectors or mass spectrometry may expand sensitivity and selectivity for complex matrices. Automated sample preparation and real-time monitoring could further integrate sugar analysis into process control and supply chain authentication.

Conclusion

The study demonstrates that transferring an HILIC method from a 4.6×250 mm to a 4.6×150 mm XBridge BEH Amide column delivers significant time and solvent savings while preserving analytical performance for five key sugars. The approach offers a robust solution for routine food and beverage testing.

References

1. Institute of Food Science & Technology. Sugars. IFST; 2017.
2. Waters Corporation. Columns Calculator Version 2.0.
3. Waters Corporation. XBridge Amide HPLC Columns Applications Notebook; 2010.
4. Waters Corporation. Beginners Guide to UPLC: The Promise of Small Particles.
5. FDA Center for Drug Evaluation and Research. Reviewer Guidance, Validation of Chromatographic Methods; November 1994.