<p>Currently, the abuse of antibiotics is becoming increasingly severe, which leads to drug residues in food. Aminoglycoside antibiotics, as a traditional antibiotic, are widely used because of their wide antibacterial spectrum and low cost. Honey, highly valued for its nutritional benefits, is a common animal-derived food product. The current beekeeping practices are primarily characterized by scattered farming by rural households. Beekeepers often indiscriminately overuse aminoglycoside antibiotics to promote the growth of bees and treat larval diseases, resulting in drug residues in honey. However, the complex composition and numerous interfering substances in honey make the efficient extraction and analysis of aminoglycoside antibiotics a persistent challenge. Therefore, this study utilizes magnetic solid-phase extraction technology, employing the prepared magnetic graphene oxide composite material as an adsorbent, in conjunction with high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS), to establish an efficient method for the simultaneous analysis of aminoglycoside antibiotics—kanamycin sulfate (KAM) and tobramycin (TOM)—in honey. We conducted characterization of the materials and investigated both their static and dynamic adsorption properties. Under optimal experimental conditions, this method showed good linear range from 5 to 500&#xa0;µg·kg^− 1 (R² ≥ 0.998), the limits of detection were 0.61&#xa0;µg·kg^− 1 (KAM) and 0.75&#xa0;µg·kg^− 1 (TOM), good recoveries of spiked honey samples (94.0%-108.0%), and satisfied repeatability for intra-day and inter-day (KAM, RSD ≤ 8.62%; TOM, RSD ≤ 12.48%). These results indicate that the method has high sensitivity, good stability and can be successfully applied to the enrichment and analysis of KAM and TOM in actual honey samples.</p>