Transfer of an EP method for mebendazole from a Waters Acquity UPLC system to a Vanquish Horizon UHPLC system

Importance of the topic

Liquid chromatography method transfer is essential in analytical laboratories to ensure consistent impurity profiling across different instruments and maintain compliance with pharmacopeial monographs. Transferring an EP method for mebendazole impurity analysis illustrates the challenges of matching chromatographic performance when system configurations and temperature controls differ.

Objectives and study overview

This study demonstrates the straightforward transfer of a European Pharmacopoeia HPLC method for mebendazole and its impurities from a Waters Acquity UPLC system to a Thermo Scientific Vanquish Horizon UHPLC platform. It also evaluates the impact of column oven and preheater settings on retention time alignment and explores downscaling the method to faster UHPLC conditions.

Methodology

Samples were prepared following the EP monograph by dissolving 5 mg of mebendazole reference standard (containing impurities A–G) in 5 mL N,N-dimethylformamide. A base-deactivated C18 column was used with mobile phases of 7.5 g/L ammonium acetate in water (A) and acetonitrile (B). The original HPLC gradient on a 4.6×100 mm, 3 µm column was scaled to UHPLC on a 2.1×50 mm, 1.9 µm column using an online calculator. Five replicate injections on each system assessed precision and alignment.

Used instrumentation

• Waters Acquity UPLC: Binary Solvent Manager, Sample Manager with 10 µL loop, High Temperature Column Heater, Tunable UV detector (10 mm, 500 nL), standard data rate (10 Hz).
• Thermo Scientific Vanquish Horizon UHPLC: Binary Pump H, Split Sampler HT (25 µL or optional 10 µL loop), Column Compartment H, Variable Wavelength Detector F (7 mm, 2.5 µL), active preheater, higher data rate (50 Hz).

Main results and discussion

Relative retention times and peak areas for mebendazole and its impurities aligned closely between the two systems, with RSDs below 0.5% and slightly higher signal-to-noise ratios on the Vanquish Horizon due to narrower peaks. Initial absolute retention time shifts (up to –7%) on the Vanquish system were linked to differences in gradient delay volume and effective column temperature. Adjusting the Vanquish column oven or preheater to match the Acquity’s thermal conditions reduced retention time deviations to under 0.5%.

Benefits and practical applications

• Minimal method modifications were needed beyond thermal fine-tuning.
• Flexible thermostatting on the Vanquish Horizon platform allowed precise emulation of original conditions.
• Higher data acquisition rates and narrower peaks improved resolution without altering the EP gradient.

Future trends and applications

Future work will focus on integrated temperature mapping, automated method translation tools across varied LC platforms, and potential updates to pharmacopeial guidelines to formally permit accelerated UHPLC scaling. Predictive software may further streamline method transfers and reduce experimental optimization.

Conclusion

The EP monograph method for mebendazole impurity analysis was successfully transferred from a Waters Acquity UPLC to a Thermo Scientific Vanquish Horizon UHPLC system. Careful matching of gradient delay volume and column temperature controls ensured equivalent chromatographic performance. Scaling to UHPLC conditions achieved 84% reduction in run time and 90% solvent savings without compromising separation quality.

References

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