Analysis of Antibiotics in Plasma for Clinical Research

Significance of the Topic

Rapid and reliable quantification of multiple antibiotics in plasma is crucial to advance clinical research in antibiotic therapy. Understanding pharmacokinetic and pharmacodynamic profiles supports dose optimization, resistance monitoring, and personalized treatment strategies.

Objectives and Study Overview

This study presents a streamlined protocol for simultaneous quantification of 16 antibiotics covering a broad polarity range in human plasma. Using a single sample preparation step and dual-run analysis, the aim is to deliver high-throughput, robust data for clinical research settings.

Methodology

• Sample preparation via protein precipitation: 50 µL plasma mixed with methanolic internal standard solution, vortexed and centrifuged.
• Supernatant diluted with 1% formic acid aqueous solution and subjected to UPLC–MS/MS analysis.
• Analytes split into two sets to accommodate meropenem stability: Set 1 analyzed first, Set 2 subsequently, within an 8-hour window.

Instrument Used

• LC System: Waters ACQUITY UPLC I-Class with FTN autosampler and precolumn heater (60 °C).
• Column: ACQUITY UPLC BEH C18, 1.7 µm, 2.1 × 100 mm.
• MS Detector: Xevo TQD triple quadrupole mass spectrometer with electrospray ionization and rapid polarity switching.
• Software: MassLynx v4.2 with TargetLynx for data acquisition and quantification.

Main Results and Discussion

• No detectable carryover at highest calibration levels across all 16 antibiotics.
• Chromatographic separation achieved within 5 minutes with clear peak resolution.
• Analytical sensitivity demonstrated by low concentration samples over five days: precision ≤20% CV and bias ≤15% at LLOQ levels.
• Overall precision and repeatability across QC levels (n=25) remained below 12.5% RSD.
• Calibration models linear or quadratic as appropriate, ensuring accurate quantification over the target range.
• Matrix effects effectively compensated by stable isotope internal standards, with recovery from common endogenous interferents between 85%–115%.

Benefits and Practical Applications of the Method

• Low sample volume requirement and straightforward preparation support high-throughput workflows.
• Single protocol covers a diverse panel of antibiotics, reducing analyst time and resource consumption.
• Rapid run time enables same-day results, valuable for clinical pharmacokinetic studies and therapeutic drug monitoring research.

Future Trends and Potential Applications

• Expansion to additional antibiotic classes and metabolites to broaden clinical utility.
• Integration with automated sample handling systems for increased throughput and reproducibility.
• Application of microflow and high-resolution mass spectrometry for improved sensitivity and selectivity.
• Coupling quantitative data with PK/PD modeling and machine learning for personalized dosing strategies.

Conclusion

The presented method demonstrates robust performance for quantifying a comprehensive panel of antibiotics in plasma. Its simplicity, speed, and precision make it a valuable tool for clinical research into antibiotic pharmacokinetics and dynamics.

References

• Application Note: Analysis of Antibiotics in Plasma for Clinical Research, Balloch et al., Waters Corporation, October 2021.