Automated UHPLC method development and robustness test for mebendazole and related impurities

Significance of the Topic

The development of robust UHPLC methods for APIs such as mebendazole is crucial to ensure accurate quantification of the drug and its impurities in pharmaceutical quality control. Automated method development platforms can drastically reduce time, costs, and manual effort while enhancing method transferability between laboratories.

Objectives and Study Overview

This study aimed to implement a software-assisted, fully automated workflow for rapid UHPLC method development, optimization, and robustness testing targeting mebendazole and its related impurities. The goal was to achieve baseline separation of all components within a short analysis time and define operating ranges for critical parameters.

Methodology and Used Instrumentation

The workflow comprised three stages: method scouting, optimization, and robustness testing. Key chromatographic variables such as column chemistry, mobile phase composition and pH, organic solvent type, gradient profile, temperature, and flow rate were systematically screened and refined.
Used Instrumentation:

• Thermo Scientific Vanquish Flex Quaternary UHPLC system with Automated Viper Method Scouting kit
• Hypersil GOLD column (100 x 2.1 mm, 1.9 µm)
• Diode Array Detector operating at 250 nm
• ChromSword Chromeleon Connect integrated with Chromeleon 7.3 CDS for automated method scouting, optimization, and robustness evaluation

Main Results and Discussion

Method scouting identified the Hypersil GOLD column as the most promising choice, yielding at least eight peaks with resolution ≥1.5. Rapid optimization using a multi-step gradient delivered full separation of mebendazole and all impurities within 13 minutes, with critical peak resolution of 3.15 and acceptable tailing factors. Fine optimization transitioned to a linear gradient, achieving complete separation in under 11 minutes. Robustness testing via a Plackett-Burman design defined stable operating ranges: ±1.5% organic solvent, ±2 °C column temperature, and ±0.1 pH unit. Resolution remained >1.8 across this design space.

Benefits and Practical Applications

Implementing an automated UHPLC method development system:

• Significantly reduces total development time to five days with minimal analyst intervention
• Ensures method robustness and reproducibility across laboratories
• Facilitates rapid selection of optimal chromatography conditions

Future Trends and Potential Applications

Advances in automation and AI-driven optimization are expected to further accelerate method development. Integration with predictive modeling and machine learning will enhance the selection of chromatographic conditions. Expanded application to biologics and complex matrices will broaden the utility of automated platforms.

Conclusion

The automated UHPLC method development workflow demonstrated here enabled fast, unattended generation of a robust analytical method for mebendazole and related impurities. The approach reduced development time, lowered resource requirements, and delivered a highly reliable method suitable for routine pharmaceutical quality control.

References

No additional literature references were provided in the source document.