Ultra-High Speed Analysis of USP Methods Conforming to Permissible Limits in New USP General Chapter 621

Importance of the Topic

The revised USP General Chapter 621 introduces broader flexibility for adjusting HPLC method parameters without full revalidation, enabling significant acceleration of compendial assays. Faster analyses reduce solvent consumption, increase laboratory throughput, and streamline quality control of pharmaceutical products.

Objectives and Study Overview

This study demonstrates how to apply the new USP 621 permitted modifications to accelerate two compendial assays—impurity profiling of sulfacetamide and assay of timolol maleate—by using a Shim-pack XR-ODS III sub-2 µm column on a Shimadzu Nexera X2 UHPLC. The goal was to maintain system suitability while achieving ultra-high-speed separations.

Methodology and Instrumentation

The approach is based on keeping the ratio of column length (L) to particle diameter (dp) nearly constant (L/dp within –25 % to +50 %), and adjusting flow rate according to the formula:
F2 = F1 × [(dc2² × dp1) / (dc1² × dp2)]
where F1 and F2 are original and modified flow rates, dc is column internal diameter, and dp is particle size. This ensures constant linear velocity and compensates for changes in particle size.

• Analysis mode: isocratic only (gradient not covered by Chapter 621 modifications)
• Column change: 4.6×150 mm, 5 µm → 2.0×50 mm, 1.6 µm
• Flow adjustment: derived from cross-sectional area and inverse dp ratio
• System suitability criteria: USP resolution ≥5 (sulfonamide pair), tailing ≤2.0, efficiency ≥3600 plates, RSD ≤2 %

Used Instrumentation

• UHPLC system: Shimadzu Nexera X2 with SPD-M30A detector
• Conventional HPLC: Shimadzu Prominence with SPD-20AV detector
• Columns: Shim-pack VP-ODS (4.6×150 mm, 5 µm) and Shim-pack XR-ODS III (2.0×50 mm, 1.6 µm)

Main Results and Discussion

For sulfacetamide impurity profiling, switching to the 1.6 µm column reduced run times by approximately 90 % (from ~7.5 min to ~0.75 min) while preserving resolution (14.5 → 12.2) and tailing factors (1.09 → 1.04). Column efficiency increased by ~10 %. Solvent use dropped to about 1/15 of the original volume on a per-run basis.

In timolol maleate assay, identical chromatographic performance was obtained on Nexera X2 compared with Prominence under the monograph conditions, confirming seamless method transfer.

Benefits and Practical Applications

• Up to tenfold reduction in analysis time accelerates batch release and stability testing.
• Substantial solvent savings lower operational costs and environmental impact.
• Method adjustments remain within USP-allowed limits, avoiding full method revalidation.
• Unified use of Nexera X2 for HPLC and UHPLC workflows simplifies laboratory infrastructure.

Future Trends and Potential Applications

The trend toward smaller particle sizes (<2 µm) and shorter columns will continue to grow. Future opportunities include applying these principles to gradient methods under risk-based method transfer strategies, integrating core-shell particle technology, and leveraging multi-dimensional LC separations for complex mixtures.

Conclusion

By adhering to the updated USP Chapter 621 guidelines and maintaining the L/dp ratio, laboratories can significantly speed up compendial HPLC assays with sub-2 µm columns on UHPLC platforms. This approach offers equivalent chromatographic performance, reduced solvent use, and enhanced throughput without extensive revalidation.

Reference

1. USP General Chapter 621, USP 37–NF 32 First Supplement.
2. USP Monograph: Sulfacetamide, USP 37–NF 32 First Supplement.
3. USP Monograph: Timolol Maleate Ophthalmic Solution, USP 37–NF 32 First Supplement.