Ultra-sensitive and rapid assay of neonicotinoids in honey by UHPLC-MS/MS

Significance of the Topic

Neonicotinoid insecticides are extensively used for crop protection due to their systemic activity and persistence. However their accumulation in pollen and honey has been implicated in honeybee colony decline. Accurate trace level monitoring of these residues in honey is therefore essential to safeguard pollinator health and ensure food safety.

Study Objectives and Overview

This work presents the development and validation of an ultra-sensitive and rapid assay for five neonicotinoids in honey using UHPLC-MS/MS. The aim was to establish a straightforward sample preparation with high recovery and reproducible performance across diverse honey matrices while achieving detection limits well below regulatory thresholds.

Used Instrumentation

• UHPLC system: Nexera X2
• Mass spectrometer: LCMS-8060 triple quadrupole with heated electrospray ionization in positive mode
• Analytical column: ACE SuperC18, 100 x 2.1 mm, 2 micron
• Data acquisition: multiple reaction monitoring (MRM)

Methodology

The procedure employs a modified QuEChERS extraction with dispersive SPE cleanup. Five grams of honey spiked with internal standards are mixed with water and acetonitrile, followed by salt addition and centrifugation. The organic phase undergoes dSPE with magnesium sulfate, PSA and C18 sorbents. One microliter of final extract is injected with a water buffering plug to enhance peak shape. Chromatographic separation is completed in six minutes using a water and methanol gradient both containing ammonia additive. Critical MS parameters and MRM transitions were optimized for each analyte.

Key Results and Discussion

Calibration curves prepared in solvent demonstrated linearity from 2.5 pg/mL to 5 ng/mL, with correlation coefficients above 0.9998. Method recoveries at 0.1 ppb spiking level ranged between 91 and 110 across eight different honey types, with repeatability below 5 percent RSD. Real honey samples contained measurable levels of acetamiprid, imidacloprid and thiamethoxam at concentrations down to 0.0022 ng/g, illustrating the method capability for ultra-low residue detection.

Benefits and Practical Applications

• High sensitivity enables quantification far below maximum residue limits
• Simple sample preparation reduces labor and avoids evaporation or dilution steps
• Fast analysis time supports high throughput in quality control laboratories
• Robust performance across varied honey matrices ensures data reliability

Future Trends and Opportunities

Advances may include expanding the assay to additional pesticide classes and matrices such as pollen or wax. Automated extraction platforms could further streamline workflow. Integration with high-resolution mass spectrometry offers potential for non-targeted screening of emerging contaminants. Green chemistry approaches may reduce solvent use and environmental impact.

Conclusion

A rapid UHPLC-MS/MS method for ultra-sensitive detection of neonicotinoids in honey was established. The validated protocol demonstrates excellent accuracy, precision and robustness across multiple honey types, providing a valuable tool for monitoring pollinator and food safety concerns.