Determination of Aminoglycoside Drugs Residual in Bee Products by LC-MS/MS

Importance of the Topic

Monitoring aminoglycoside antibiotic residues in bee products is essential to ensure food safety and consumer health. Aminoglycosides can cause ototoxicity, nephrotoxicity and other severe side effects. Regulatory standards such as GB 31650-2019 set maximum residue limits for these drugs in food, highlighting the need for sensitive and reliable analytical methods.

Objectives and Study Overview

This study aimed to develop and validate a robust liquid chromatography–tandem mass spectrometry (LC-MS/MS) method for simultaneous determination of 11 aminoglycosides in honey. Key objectives included optimizing sample preparation, chromatographic separation without mobile-phase ion-pairing reagents, and establishing method performance parameters.

Methodology and Instrumentation

Sample Preparation:

• Extract 5 g honey with phosphate buffer (5% TCA, 0.4 mM EDTA).
• Centrifuge, re-extract, combine supernatants to 20 mL.
• Divide extract: purify one portion on MCX SPE (200 mg/6 mL), elute with ammonia–methanol; second portion on WCX SPE (150 mg/3 mL), elute with methanol–acetate.
• Evaporate, reconstitute in 0.3% acetic acid–HFBA (99:1) and filter.

Chromatography and Detection:

• LC system: Shimadzu Nexera LC-40 X3, column Shim-pack Scepter C8 (100×2.1 mm, 1.9 µm) at 35 °C, 5 µL injection.
• Mobile phase A: 0.5 mM ammonium acetate + 0.1% formic acid in water; B: acetonitrile; gradient from 5% to 90% B in 7.5 min.
• MS: Shimadzu LCMS-8050, ESI positive, MRM transitions optimized for each compound.

Instrumentation Used:

• Nexera LC-40 X3
• Shim-pack Scepter C8 column
• LCMS-8050 triple quadrupole mass spectrometer

Main Results and Discussion

The method achieved strong retention of aminoglycosides on the C8 column without ion-pair agents. Calibration curves (5–500 ng/mL) displayed excellent linearity (R² > 0.996). Limits of detection ranged from 0.04 to 0.90 ng/mL and limits of quantification from 0.12 to 2.74 ng/mL. Intra-day precision (RSD) was below 9.5% for retention time and peak area at three concentration levels. Recovery studies at 25 and 50 µg/kg spiking levels yielded recoveries between 68.5% and 93.4% with RSDs under 9.5%.

Benefits and Practical Applications

• High sensitivity and specificity for 11 aminoglycosides in a single run.
• No need for high-concentration salts or ion-pairing in the mobile phase simplifies maintenance and improves MS signal.
• Suitable for routine quality control and regulatory compliance testing in bee products.

Future Trends and Potential Applications

Expanding this approach to other food matrices such as milk or meat can streamline multi-residue analysis. Integration with high-resolution MS may allow broader suspect screening. Development of portable LC-MS platforms may enable on-site rapid testing in apiaries.

Conclusion

A fast, sensitive and accurate LC-MS/MS method was established for aminoglycoside residue analysis in honey. It meets regulatory requirements and can support routine monitoring in food safety laboratories.