COMBINED USE OF GC TOF MS AND UPLC-QTOF MS FOR INVEST IGAT IVE ANALYSIS OF HONEYBEE POISONING

Importance of the Topic

The honeybee is a key pollinator essential for agricultural productivity and biodiversity. Extensive pesticide use has led to widespread honeybee mortality, making the identification of toxic exposures critical for environmental protection and food security.

Objectives and Study Overview

This study employs two complementary high-resolution mass spectrometry techniques—GC-TOF MS and UPLC-QTof MS—to perform an untargeted screening for insecticide residues and their metabolites in honeybee samples collected from the Valencia region (Spain). The concentrations and identities of contaminants were to be detected and confirmed without prior knowledge of specific pesticides.

Methodology

Sample preparation followed an established protocol: 1.5 g of homogenized honeybee tissue mixed with sodium sulfate and celite underwent acetone extraction, followed by dichloromethane partitioning and concentration under nitrogen. Final residues were reconstituted in ethyl acetate (for GC-MS) or methanol and diluted in water (for LC-MS). A nectarine flower and leaf sample served as a related matrix reference.

Used Instrumentation

• Gas Chromatography–Time-of-Flight Mass Spectrometry (GC-TOF MS): Agilent 6890N GC with HP-5MS column, helium carrier, splitless injection; Waters GCT Premier mass spectrometer, 50–650 Da, resolution ~8500 FWHM; acquisition at 5 spectra/s.
• UltraPerformance Liquid Chromatography–QTof Mass Spectrometry (UPLC-QTof MS): Waters ACQUITY UPLC with BEH C18 column; gradient of water/methanol with 0.1 mM ammonium acetate; Waters QTof Premier, mass range 50–1000 Da, ~10 000–17 500 FWHM, MS/MS with 10–30 eV collision energy.
• Data processing via MassLynx v4.1 with TargetLynx, ChromaLynx, and MetaboLynx XS application managers.

Main Results and Discussion

Both GC-TOF MS and UPLC-QTof MS consistently identified the organophosphorus insecticide coumaphos in two of the dead bee samples. GC-TOF MS provided deconvoluted exact mass spectra and library matches, confirming four diagnostic ions. UPLC-QTof MS aided in detecting coumaphos and its transformation products through high-resolution MS/MS analysis. Key metabolites—potasan, CMHC, coumaphos oxon, and 4-methylumbelliferone—were observed, suggesting pathways involving hydrolysis, hydroxylation, sulfur oxidation, and dechlorination. High-resolution data and isotope patterns enabled differentiation between isobaric species and accurate elemental composition assignments.

Benefits and Practical Applications

Combining GC-TOF MS and UPLC-QTof MS offers comprehensive coverage of volatile and polar compounds in a single workflow. Exact mass measurements facilitate untargeted screening and structural confirmation, supporting environmental monitoring, pesticide regulation, apicultural health assessments, and quality control in agricultural products.

Future Trends and Opportunities

Advancements may include the integration of real-time in-field analyses, expanded exact-mass spectral libraries, and machine-learning algorithms for automated feature annotation. Coupling these platforms with multi-omic approaches could deepen insights into pesticide fate and biological effects.

Conclusion

This study demonstrates that GC-TOF MS and UPLC-QTof MS are complementary techniques for untargeted identification of pesticide residues and metabolites in honeybee matrices. High-resolution mass spectrometry, combined with robust data-processing tools, effectively elucidates contaminant profiles and transformation pathways.

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