Ultra-high Speed Analysis of USP methods conforming to the New USP General Chapter 621 Allowed Limits

Significance of the Topic

Pharmacopoeial compliance and high analytical throughput are critical in pharmaceutical quality control. The revised USP General Chapter 621 permits adjustment of column dimensions and particle size within defined limits without revalidation if system suitability is met. This flexibility enables transfer of traditional HPLC methods to ultra-high performance liquid chromatography (UHPLC) platforms, reducing analysis time, solvent use, and operational costs while maintaining required resolution and precision.

Objectives and Study Overview

This work demonstrates two applications of the new USP 621 allowed adjustments:

• Conversion of the sulfacetamide antibiotic impurities method from a 150×4.6 mm, 5 µm HPLC column to a 50×2.0 mm, 1.6 µm UHPLC column for ultra-high speed analysis.
• Evaluation of the timolol maleate ophthalmic solution assay on a UHPLC system using the original USP method parameters without modification.

Methodology

Reagents and standards for sulfacetamide and timolol maleate analyses were prepared according to USP monographs. The sulfacetamide method used an isocratic mobile phase of methanol, water and acetic acid, while the timolol assay employed a phosphate buffer–methanol mixture. System suitability and standard solutions were prepared at pharmacopeial concentration levels and analyzed under isocratic conditions.

Applied Instrumentation

The study utilized two chromatographic systems from Shimadzu:

• Prominence conventional HPLC: LC-20AD pump, DGU-20A5R degasser, SIL-20AC autosampler, CTO-20AC column oven, SPD-20AV UV detector, CBM-20A controller.
• Nexera X2 UHPLC: LC-30AD pump, DGU-20A5R degasser, SIL-30AC autosampler, CTO-30A oven, SPD-M30A PDA detector, CBM-20A controller.

Key Results and Discussion

For the sulfacetamide impurity method:

• Column change from 150×4.6 mm, 5 µm to 50×2.0 mm, 1.6 µm maintained L/dp ratio and resolution.
• Flow rate adjusted from 0.8 to 0.47 mL/min to preserve linear velocity.
• Analysis time shortened from ~7.5 min to ~0.75 min and solvent use reduced ~15-fold.
• System suitability metrics (resolution ≥ 5.0; tailing ≤ 1.5; RSD ≤ 2%) were met on the UHPLC system.

For the timolol assay:

• The original USP method (150×4.6 mm, 5 µm, 1.2 mL/min) was run unchanged on Nexera X2.
• Chromatographic performance (tailing factor < 2.0; plate count > 3600; RSD < 2%) matched conventional HPLC results.

Benefits and Practical Applications

The adoption of USP-compliant UHPLC provides:

• Significant throughput increase via >10× faster run times.
• Reduced solvent consumption and waste generation.
• No need for full method revalidation when using allowed adjustments.
• Direct compatibility with established pharmacopeial monographs.

Future Trends and Opportunities

Advances in UHPLC hardware and software, coupled with green chemistry goals, suggest broader adoption of high-speed pharmacopeial methods. Prospective developments include gradient compatibility under USP limits, integration with mass spectrometry detectors, automated method transfer tools, and data-driven optimization in QA/QC laboratories.

Conclusion

The revised USP Chapter 621 offers practical flexibility for chromatographic method transfers. This study confirmed that sulfacetamide and timolol maleate assays can be accelerated on UHPLC without loss of performance or revalidation burden, yielding substantial time and solvent savings.

References

• USP General Chapter 621, USP 37–NF 32, First Supplement.
• USP Monograph “Sulfacetamide,” USP 37–NF 32, First Supplement.
• USP Monograph “Timolol Maleate Ophthalmic Solution,” USP 37–NF 32, First Supplement.