Ultra-High-Speed Analysis of Quinolone Antibacterial Agents

Importance of the Topic

Quinolone antibacterial agents play a critical role in veterinary medicine for preventing and treating infections in livestock, poultry, and aquaculture. Rapid and sensitive analysis of multiple quinolones is essential for quality control, residue monitoring, and regulatory compliance. Ultra-high-speed methods can enhance laboratory throughput and support timely decision-making in food safety and pharmaceutical research.

Objectives and Study Overview

This application note describes the development and evaluation of an ultra-high-speed simultaneous analysis of 11 quinolone compounds using a Shimadzu Nexera UHPLC system paired with an RF-20Axs fluorescence detector. The study compares conventional and fast-analysis columns to demonstrate significant reductions in run times while maintaining resolution and sensitivity.

Methodology and Instrumentation

The study employed three column formats:

• Conventional Shim-pack VP-ODS (150×4.6 mm, 5 µm)
• Shim-pack XR-ODS (75×3.0 mm, 2.2 µm)
• Shim-pack XR-ODS III (50×2.0 mm, 1.6 µm)

Gradient elution used 0.1 % formic acid in water (A) and 0.1 % formic acid in acetonitrile (B). Flow rates ranged from 0.8 to 1.3 mL/min depending on column dimensions. Injection volumes were optimized (0.4–22 µL), and column temperature was held at 65 °C. The RF-20Axs detector’s cell-temperature control ensured stable fluorescence measurement at excitation/emission wavelengths of 299/455 nm for peaks 1–8 and 325/365 nm for peaks 9–11.

Key Results and Discussion

All 11 quinolones (norfloxacin, ofloxacin, ciprofloxacin, danofloxacin, enrofloxacin, orbifloxacin, sarafloxacin, difloxacin, oxolinic acid, nalidixic acid, flumequine) were baseline-separated under each condition. Peak elution times were:

• Conventional column: ~27 min total run time.
• XR-ODS (2.2 µm): ~4.5 min run time (≈6× faster).
• XR-ODS III (1.6 µm): ~2.25 min run time (≈12× faster).

Ultra-high-speed conditions generated narrower peaks with minimal sensitivity loss, despite increased system backpressure (up to 55 MPa). The fluorescence detector’s temperature control minimized baseline drift under rapid gradients.

Advantages and Practical Applications

• Greatly increased sample throughput for routine residue analysis.
• High sensitivity and reproducibility for trace-level detection.
• Reduced solvent consumption and analysis costs per sample.
• Applicable to veterinary drug monitoring, QA/QC laboratories, and regulatory testing.

Future Trends and Potential Applications

Continued miniaturization and particle-size reduction are likely to push analysis times below two minutes for multi-component assays. Integration with automated sample preparation and multi-detector systems (e.g., UV, MS) can further enhance method versatility. Emerging detector technologies and advanced data processing will enable real-time monitoring of veterinary drugs in complex matrices.

Conclusion

The combination of Nexera UHPLC with fast-analysis Shim-pack XR columns and RF-20Axs fluorescence detection achieves ultra-high-speed separation of 11 quinolone antibacterial agents. The approach offers significant benefits in throughput, sensitivity, and operational efficiency, making it highly suitable for modern analytical laboratories.

Reference

Shimadzu Corporation. Ultra-High-Speed Analysis of Quinolone Antibacterial Agents. Application Data Sheet No. 10. Issued December 2010.