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Tobramycin and vancomycin are the most commonly used antibiotics for the treatment of osteomyelitis. A sensitive and rapid method was developed for the analysis of tobramycin and vancomycin in human drainage tissue fluid. The procedure involved a simple liquid–liquid extraction of tobramycin, vancomycin and atenolol (internal standard) and separation by ultra-high performance liquid chromatography on an Acquity UPLC BEH C18 column (2.1 × 100 mm, 1.7 µm) with a mobile phase consisting of 0.1% (v/v) formic acid water solution and 0.1% (v/v) formic acid acetonitrile solution at a flow rate of 0.3 mL/min. Detection was performed by positive ion electrospray ionization in multiple reaction monitoring mode ( m/z  468 → 163 transitions for tobramycin;  m/z  725 → 144 for vancomycin;  m/z  267 → 74 for the internal standard). The retention times of tobramycin, vancomycin, and the internal standard were 0.68, 3.62 and 3.03 min, respectively. The total analysis time was less than 10 min. Excellent linear relationships (correlation coefficient &gt; 0.99) were demonstrated between the area under the peak ratios of tobramycin and vancomycin to the internal standard in the drainage tissue fluid, and the concentration ranges were 1.25–100.00 mg/L and 0.50–150.00 mg/L for tobramycin and vancomycin, respectively. The intra-day and inter-day accuracy and precision (coefficient of variation) acceptance criteria for each quality control was ≤ 7.8% and the mean accuracy values were &lt; 5.0% for tobramycin and &lt; 4.0% for vancomycin. All experiments suggested the high-throughput potential of the proposed method. The method was successfully applied to investigate local delivery of tobramycin and vancomycin in four calcaneal osteomyelitis patients who had accepted drug-loaded artificial bone implantation.

Rapid Quantification of Tobramycin and Vancomycin by UPLC–TQD and Application to Osteomyelitis Patient Samples