Scaling and Migration of a Method for Naphazoline Hydrochloride, Pheniramine Maleate and Associated Related Substances to an ACQUITY Arc System

Significance of the Topic

In pharmaceutical quality control, transferring analytical methods between different column particle sizes and chromatographic systems is essential to ensure consistent product quality and regulatory compliance. Scaling UPLC methods to HPLC systems broadens method accessibility in laboratories lacking low-dispersion sub-2 µm capabilities, while preserving resolution, throughput, and robustness.

Objectives and Study Overview

This study aimed to adapt a UPLC method for the determination of naphazoline hydrochloride and pheniramine maleate, along with their related impurities, from a 1.7 µm (2.1 × 100 mm) column on an ACQUITY UPLC H-Class Plus system to a 2.5 µm (4.6 × 150 mm) column on an ACQUITY Arc system. The goal was to demonstrate equivalent chromatographic performance, system suitability, and assay accuracy across platforms.

Methodology and Instrumentation

• Preparation of individual stock solutions in methanol (4.0 mg/mL) and mixture standards in 80:20 water/methanol containing APIs at 0.1 mg/mL and impurities at 10 µg/mL.
• Ophthalmic sample dilutions to 500 µg/mL pheniramine maleate and 40 µg/mL naphazoline HCl in 80:20 water/methanol.
• Mobile phases: 0.1% formic acid in water (A) and methanol (B); column temperature set at 42 °C; detection at 260 nm with 10 pts/sec sampling rate.
• Geometric scaling of flow rate, injection volume, and gradient profile from 1.7 µm to 2.5 µm columns using Waters Columns Calculator.
• Measurement of system dwell volumes and compensation of gradient delay on the ACQUITY Arc system using the Gradient StartSmart feature.
• Empower 3 FR5 SR5 software for data acquisition, processing, and report generation.

Main Results and Discussion

Chromatograms obtained on both systems exhibited matching peak shapes, relative retention times (RRTs), and USP resolution between critical impurity pairs. System suitability tests (six injections) showed excellent repeatability in retention time and peak area on the ACQUITY Arc system, comparable to the UPLC platform. API assay recoveries and impurity percentages in ophthalmic samples were equivalent across instruments, confirming successful method transfer.

Benefits and Practical Applications

• Efficient scaling and migration workflow enabled by software tools and automated gradient delay compensation.
• Preserved analytical performance when transferring methods between UPLC and HPLC systems, supporting diverse laboratory capabilities.
• Minimized manual method adjustments, reducing development time and error potential.

Future Trends and Applications

Emerging instrument designs with reduced dispersion, advanced software automation, and AI-driven method optimization promise further improvements in cross-platform method transfer. Real-time system diagnostics and adaptive gradient control will enhance robustness and accelerate pharmaceutical method development and implementation.

Conclusion

The geometric scaling of column dimensions and careful compensation for system dwell volume differences enabled equivalent chromatographic separation, system suitability, and assay accuracy when migrating a UPLC method for naphazoline HCl and pheniramine maleate impurities to an HPLC-based system. This approach ensures reliable method performance across varied laboratory platforms.

References

• Dong MW. Ultrahigh-Pressure Liquid Chromatography, Part III: Potential Issues. LC-GC North America. 35(11):2017.
• Hong H, McConville PR. Dwell Volume and Extra-Column Volume: What Are They and How Do They Impact Method Transfer? Waters White Paper 720005723;2018.
• Waters Columns Calculator. Waters Corporation;2017.
• Waters Corporation. Protocol for Gradient Delay (Dwell Volume) Measurement. Waters Application Notebook. 2013:67–69.