Simple screening of natural sugars for detecting adulteration in honey

Significance of the Topic

Honey adulteration threatens product authenticity and consumer trust. Given honey’s high sugar content and widespread consumption, ensuring purity is critical for regulatory compliance and quality assurance. Reliable analytical methods to quantify main sugars (fructose, glucose, sucrose) offer a straightforward approach to detect adulteration, as deviations from established sugar profiles indicate addition of cheaper sweeteners.

Objectives and Study Overview

This study aimed to develop a robust, cost-efficient HPLC method with refractive index detection (RID) for quantifying glucose, fructose and sucrose in honey, meeting global regulatory criteria. The method was evaluated for precision, linearity, accuracy and transferability across laboratories to support routine honey quality control and adulteration screening.

Methodology and Instrumentation

Two chromatographic approaches were compared: a polymer-based HyperREZ XP Carbohydrate Pb2+ column with 100% aqueous mobile phase and a silica-based Hypersil GOLD Amino column with 80:20 acetonitrile–water. A refractive index detector (Vanquish RID) integrated with Chromeleon CDS enabled universal detection of non-UV active sugars. Calibration standards ranged from 5 to 200 mg/L for sucrose and 25 to 1 000 mg/L for glucose and fructose. Samples were prepared by diluting honey solutions 1:500 or 1:1 000 prior to injection.

• Standards: Stock solutions at 50 000 mg/L diluted to six levels
• Chromatographic conditions:
  • HyperREZ XP Pb2+: water mobile phase, 0.6 mL/min, 70 °C, 25 min run
  • Hypersil GOLD Amino: 80/20 ACN–water, 1.2 mL/min, 45 °C, 10 min run

Main Results and Discussion

Using the HyperREZ XP column, baseline separation of fructose, glucose, sucrose and maltose was achieved within 25 min. Precision for retention times was below 1 % RSD, and peak area RSDs were under 2.2 %. Calibration curves exhibited excellent linearity (R2 > 0.9999). Accuracy assessed via certified honey quality controls showed deviation within ±9 % for sucrose and <2 % for hexoses. Method transfer between two labs and column lots confirmed robustness.
The faster Hypersil GOLD Amino column reduced run times to 10 min, maintaining clear resolution by optimizing flow rate and column temperature. Both methods reliably quantified major sugars, with fructose predominant in all tested honey samples. Sucrose levels remained within Codex and EU limits, while maltose was consistently detected as a minor component.

Benefits and Practical Applications

• Cost-effective, reagent-efficient analyses with 100 % aqueous mobile phase option
• High throughput via fast run times and automated eWorkflows in Chromeleon
• Meets regulatory standards for honey quality and adulteration detection

Future Trends and Opportunities

Advances in column chemistries and detector sensitivity may further reduce analysis time and lower detection limits for minor sugars and adulterants. Integration with automated eWorkflows and data analytics will streamline routine testing and support traceability. Adoption of eco-friendly mobile phases aligns with green chemistry principles, promoting sustainable laboratory practices.

Conclusion

The Vanquish HPLC-RID approach provides a robust, sensitive and efficient solution for routine honey sugar profiling and adulteration monitoring. Both the HyperREZ XP Pb2+ and Hypersil GOLD Amino methods deliver reliable results, allowing laboratories to select protocols based on run time and environmental considerations. This analytical strategy supports quality control and regulatory compliance in the honey industry.

Reference

1. Food Fraud Database, Decernis
2. European Parliament Committee report on food fraud (2013/2091(INI))
3. Codex Alimentarius Standard for Honey CXS 12-1981 (Revised 1987, 2001; Amended 2019)
4. Council Directive 2001/110/EC relating to honey, Official Journal L10/47-52 (2002)
5. Thermo Fisher Scientific Carbohydrate Testing Information
6. International Honey Commission (IHC) platform