Organic Impurity Analysis of Levofloxacin Drug Material Following USP Monograph

Importance of the topic

Monitoring trace organic impurities in active pharmaceutical ingredients is critical for ensuring drug safety, efficacy and compliance with regulatory monographs.

Study objectives and overview

This work implements the United States Pharmacopeia (USP) monograph 41 method for impurity profiling of levofloxacin drug substance on a modern high performance liquid chromatograph. The goal is to demonstrate system suitability, sensitivity and identification of organic impurities in accordance with official requirements.

Methodology and instrumentation

The method was based on an isocratic reverse phase separation following USP 41. Key steps included preparation of levofloxacin standard solutions at 1.0 mg/mL and 0.3 µg/mL for system suitability and sensitivity tests. Mobile phase A consisted of ammonium acetate, cupric sulfate pentahydrate and L-isoleucine in water; mobile phase B was methanol. Chromatography was performed at 45°C with UV detection at 360 nm and a flow rate of 0.8 mL/min. The run time was 25 minutes, with the main levofloxacin peak eluting at 14.7 minutes.

Instrumentation used

• Shimadzu i-Series LC-2050 high performance liquid chromatograph
• Shim-pack GIST C18 column (250 mm × 4.6 mm, 5 µm particle size)
• UV detector set at 360 nm

Main results and discussion

System suitability results met USP criteria: signal-to-noise ratio of 14.4 (minimum 10), repeatability (%RSD) for peak area and retention time below 0.12% (maximum 1.0%), and tailing factor within 0.5–1.5. Impurity profiling of the 1.0 mg/mL solution revealed six trace impurities. Four impurities were matched to known USP listed impurities via relative retention time and two additional unknown peaks were observed at RRT 0.55 and 0.67. The total impurity level was 0.29%.

Benefits and practical applications

This validated method provides high sensitivity and selectivity for routine quality control of levofloxacin API. It can be transferred easily to pharmaceutical laboratories for batch release testing and stability studies, ensuring compliance with regulatory standards.

Future trends and applications

Emerging developments may include coupling with mass spectrometry for structural identification of unknown impurities, method miniaturization for higher throughput, and automation of data analysis. Advanced stationary phases and greener mobile phase formulations can further enhance selectivity and environmental sustainability.

Conclusion

The USP monograph 41 method for levofloxacin impurity analysis was successfully implemented on the Shimadzu i-Series LC-2050 system. All system suitability criteria were met, and six impurities were detected with four positively identified. This approach offers a robust solution for pharmaceutical quality control.

References

1. United States Pharmacopeia USP 41 May 1, 2018 Official Monograph Levofloxacin 2395