Analysis of Polyether Antibiotics in Animal Feeds Using Fluorescence Detection

Importance of the Topic

Polyether antibiotics such as lasalocid sodium are widely used as feed additives to promote animal health and growth. Rapid and reliable quantitation of these compounds in feed is essential for quality control, regulatory compliance and food safety monitoring. Traditional microbiological assays require lengthy culture periods, whereas chromatographic methods can deliver results in minutes with higher throughput and automation.

Objectives and Study Overview

This study aimed to establish a rapid, fully automated HPLC method with fluorescence detection for the analysis of lasalocid sodium in animal feeds. Key goals included validation of sensitivity, linearity, repeatability and recovery in accordance with Japan’s Feed Analysis Standards.

Methodology and Instrumentation

Sample Preparation and Extraction:

• Feed samples fortified with lasalocid sodium were extracted following the official protocol involving phosphate buffer and methanol.
• Resulting extracts were filtered and injected directly for analysis.

Chromatographic Conditions:

• Instrument: Nexera lite HPLC system
• Column: Shim-pack Scepter C18-120 (250 × 4.6 mm, 5 μm)
• Mobile Phase: 0.01 M phosphate buffer (pH 3.0) / methanol (1 : 9, v/v)
• Flow Rate: 1.0 mL/min; Column Temp: 40 °C; Injection Volume: 20 µL
• Detection: RF-20Axs fluorescence detector, Ex = 310 nm, Em = 420 nm

Key Results and Discussion

Linearity and Sensitivity:

• Calibration range: 1–15 µg/mL (potency basis), R² > 0.9992.

Repeatability:

• Six replicate injections at 0.1 µg/mL showed retention time RSD = 0.009 % and peak area RSD = 0.36 %.

Recovery in Feed Matrix:

• Spike level equivalent to 75 g potency/t feed yielded 96.9 % recovery without interference.

Analysis Time:

• Each run completed in under 10 minutes, significantly faster than microbiological methods.

Advantages and Practical Applications

• High throughput: Rapid analysis suitable for routine QA/QC
• Automation: From instrument startup to shutdown, reducing labor and human error
• Broad dynamic range: Reliable quantitation from low ppb to high ppm levels
• Regulatory compliance: Meets MAFF Feed Analysis Standards for lasalocid sodium

Future Trends and Potential Applications

• Integration with LC-MS/MS for multi-residue screening and confirmatory analysis.
• Miniaturized and portable HPLC-FLD systems for on-site feed monitoring.
• Automated sample preparation modules to further streamline workflows.
• Use of chemometric and AI tools to optimize method parameters and data interpretation.

Conclusion

The HPLC-FLD method using the Nexera lite platform provides a rapid, precise and fully automated solution for quantifying lasalocid sodium in animal feeds. It offers significant time savings over microbiological assays while maintaining excellent linearity, repeatability and recovery in complex feed matrices.

Reference

1. MAFF Public Notice No. 750 of July 24, 1976, Establishing Feed Additives Under the Feed Safety Act.
2. Feed Analysis Standards, Notice No. 19/14729 of April 1, 2008, Food Safety and Consumer Affairs Bureau, MAFF.
3. Ministerial Ordinance No. 35 of July 24, 1976, Specifications and Standards of Feeds and Feed Additives, MAFF.